Liver Cirrhosis - Dr. Falk Pharma GmbH

Transcription

Falk Symposium 162
Liver Cirrhosis: From Pathophysiology
to Disease Management
October 13– 14, 2007
International Congress Center
Dresden
Abstracts
Poster Abstracts
© 2007 Falk Foundation e.V
All rights reserved.
Abstracts of Invited Lectures
Poster Abstracts
Falk Symposium 162
LIVER CIRRHOSIS: FROM PATHOPHYSIOLOGY
TO DISEASE MANAGEMENT
Dresden (Germany)
October 13–14, 2007
Scientific Organization:
J. Bosch, Barcelona (Spain)
A.K. Burroughs, London (Great Britain)
F. Lammert, Bonn (Germany)
D. Lebrec, Clichy (France)
T. Sauerbruch, Bonn (Germany)
CONTENTS
Page
Session I
Pathomechanisms of fibrogenesis (1):
Initiation
Chair:
S.L. Friedman, New York
D. Häussinger, Düsseldorf
Apoptosis and oncotic necrosis: The common pathways
of liver injury
H.W. Jaeschke, Kansas City
Entero-hepatic lymphocyte homing and liver
manifestations of IBD
D.H. Adams, B. Eksteen, A. Grant, Birmingham
Immune responses to chronic hepatitis C virus infection
U. Spengler, Bonn
Cytokines: Interleukin-6 and TNF signalling
C. Trautwein, Aachen
19
20 – 21
22
23 – 24
Session II
Mediators
Chair:
D.A. Brenner, La Jolla
A.M. Gressner, Aachen
Growth factors in experimental liver fibrogenesis
R. Weiskirchen, E. Borkham-Kamphorst, E. Kovalenko,
S. Meurer, O. Scherner, W.N. Vreden, O.A. Gressner,
J. Herrmann, F. Drews, A.M. Gressner, Aachen
27 – 28
Genetic determinants of liver fibrogenesis
F. Lammert, Bonn
29 – 30
Molecular mechanisms of fibrosis progression in
non-alcoholic steatohepatitis (NASH)
C. Hellerbrand, Regensburg
31 – 32
3
New role of bile acid metabolism in bacterial translocation
A. Moschetta, Bari
33 – 35
Session III
Cell responses to chronic liver injury
Chair:
D. Lebrec, Clichy
M. Pinzani, Florence
Role of hepatic progenitor cells in regeneration
T. Roskams, Leuven
39 – 40
The hepatic stellate cell: A progenitor cell
C. Kordes, I. Sawitza, A. Müller-Marbach, N. Ale-Agha,
V. Keitel, H. Klonowski-Stumpe, D. Häussinger, Düsseldorf
41
Role of non-parenchymal liver cells in portal hypertension
V. Shah, Rochester
42
Role of angiogenesis in portal hypertension
M. Fernández, Barcelona
43 – 44
The relative roles of lymphocytes, Kupffer cells, and
myofibroblasts in hepatic fibrosis
E. Seki, S. De Minicis, Y. Osawa, T. Kisseleva, R. Schwabe,
D.A. Brenner, La Jolla
45 – 46
Hepatic fibrogenesis and carcinogenesis:
Krüppel-like factors and beyond
S.L. Friedman, New York
47 – 48
Session IV
Clinical management of progressive
liver fibrosis
Chair:
G. Ramadori, Göttingen
C. Trautwein, Aachen
Non-invasive assessment of liver fibrosis (1):
Serum surrogate markers
D. Thabut, Paris
4
51 – 52
Non-invasive assessment of liver fibrosis (2):
Fibroscan
M. Beaugrand, Bondy
53 – 54
The clinical evaluation of disease progression in CLD:
Towards an integrated system?
M. Pinzani, Florence
55
Fibrosis regression and innovative antifibrotic therapies:
From bench to bedside?
H.E. Wasmuth, Aachen
56
Session V
Clinical management of portal hypertension (1):
Preprimary and primary prophylaxis
Chair:
J. Heller, Bonn
F. Wong, Toronto
Dynamic increase of intrahepatic vascular resistance
in cirrhosis
J.C. Garcίa-Pagán, Barcelona
Remodeling portal hypertension: Preprimary prophylaxis
G. Garcia-Tsao, New Haven
59
60 – 62
When and how to scope in portal hypertension
R. de Franchis, Milan
63
Prevention of first variceal bleeding
P. Calès, Angers
64
Session VI
Complications of cirrhosis
Chair:
J. Bosch, Barcelona
R. Wiest, Regensburg
Management of acute variceal bleeding
A.K. Burroughs, London
67 – 69
5
Prevention of variceal rebleeding
D. Lebrec, Clichy
From sodium retention to therapy for refractory ascites:
The role for new drugs
F. Wong, Toronto
71 – 72
The role of TIPS in the treatment of refractory ascites
M. Rössle, Freiburg
73 – 75
Spontaneous bacterial peritonitis: A disease of the gut?
Therapeutic implications
A. Cárdenas, P. Ginès, Barcelona
76
Hepatorenal syndrome in cirrhosis
A.L. Gerbes, Munich
77
Infections and hepatic encephalopathy
D. Häussinger, Düsseldorf
Liver and lung – Treatment of hepatopulmonary diseases
P. Schenk, Vienna
78 – 79
80
Allocation in liver transplantation: Is MELD the solution?
North American perspective
D.M. Heuman, Richmond
81 – 82
European perspective
A.O. Rahmel, Leiden
83 – 84
Emerging future therapies for portal hypertension
J. Bosch, Barcelona
List of Speakers, Moderators and Scientific Organizers
6
70
85
87 – 90
Poster Abstracts
1.
Hepatic cirrhosis in the elderly in a Hispano-American population
E. Aravena, F. Contesse, R. Muñoz, H. Iturriaga (Santiago de Chile, RCH)
2.
Liver affection in different types of glycogen storage disease in children
E.S. Dublina, E. Uvarova, T.V. Strokova, A.S. Potapov (Moscow, R)
3.
Wilson disease in childhood: Results of a single center experience
M. Georgieva-Shakola, D. Konstantinova-Kanazireva, I. Krasnaliev,
A. Atanassova, S. Tomova, A. Savov, V. Tzaneva (Varna, Sofia, BG)
4.
Liver cirrhosis in cystic fibrosis
K. Kalinova, K. Todorova (Stara Zagora, BG)
5.
Niemann-Pick disease-type B
D. Konstantinova-Kanazireva, M. Georgieva, V. Sinigerska, V. Tzaneva,
L. Angelova (Varna, Sofia, BG)
6.
Clinical features of pediatric liver cirrhosis resulting from alpha1-antitrypsin
deficiency
I. Lastauka, M. Halabarodzka (Minsk, WR)
7.
US scanning data in liver cirrhosis in infants of first 3 years old
T. Raeuneva, M. Halabarodzka (Minsk, WR)
8.
H63D heterozygotes with alcoholic liver disease are proned to hypercholesterolemia: A single centre, prospective study
J. Raszeja-Wyszomirska, I. Zawada, J. Suchy, G. Kurzawski, J. Lubinski,
P. Milkiewicz (Szczecin, PL)
9.
Natural history of primary sclerosing cholangitis in patients with inflammatory
bowel disease: Our experience
P. Svorcan, D. Bojic, M. Protic, B. Bojic, S. Djorovic, J. Djordjevic, N. Jojic,
S. Kazic, B. Dapcevic (Belgrade, SRB)
10. Copper and ceruloplasmin in sera of patients with alcoholic liver cirrhosis
E. Uhlikova, V. Kupcová, M. Szántová, I. Vozár, L. Turecky (Bratislava, SK)
11. Hepatic stellate cells and liver regeneration in chronic viral hepatitis C
S. Abdulhakov, N.A. Cheremina, A.A. Gumerova, M.A. Titova, A.P. Kiassov,
L. Fatkheeva, T. Smetannikova, D.I. Andreeva (Kazan, R)
12. TGF-β and ethanol: Two substances that exponentiate each other leading to
increased liver damage
K. Breitkopf, H. Gaitantzi, L. Ciuclan, P. Godoy, S. Dooley (Mannheim, D)
7
13. Cholestasis but not steatosis potentiates liver injury by activated Kupffer cells:
Evidence for a role for cyclooxygenase 2
M. Bystron, C.J. Steib, J.M. Härtl, F. Roggel, M. Bilzer, A.L. Gerbes (Munich, D)
14. Expression of C-kit in liver after partial hepatectomy in rats
I.M. Gazizov, M.S. Kaligin, D.I. Andreeva, G.O. Pevnev, G.R. Burganova,
A.A. Gumerova, A.P. Kiassov (Kazan, R)
15. Perisinusoidal cells after partial hepatectomy in rats
I.M. Gazizov, M.S. Kaligin, D.I. Andreeva, G.O. Pevnev, G.R. Burganova,
A.A. Gumerova, A.P. Kiassov (Kazan, R)
16. Hepatic overexpression of RhoA- and Rho-kinase and elevated Rho-kinase
activity in livers from rats with CCI4-induced micronodular cirrhosis
M. Hennenberg, J. Trebicka, A. Eckhardt, T. Sauerbruch, J. Heller (Bonn, D)
17. The association of intestinal iNOS expression and oxidative stress in the
intestine in experimental cirrhosis
M.R. Mas, B. Savas, R. Ocal, I. Tasci, A. Ensari, B. Comert, S. Yilmaz, A.T. Isik,
N. Mas, E. Erden, Ö. Uzunalimoglu (Ankara, TR)
18. Large-conductance calcium-activated potassium channels modulate vascular
tone in experimental cirrhosis
A. Rodriguez-Vilarrupla, M. Graupera, V. Matei, R. Bataller-Sifre, J.G. Abraldes,
J. Bosch, J.-C. Garcia-Pagán (Barcelona, Valencia, E)
19. Kupffer cell (KC) activation in fibrotic livers by β-glycans: The COX-2 inhibitor
Parecoxib reduces portal pressure (PP) incerase in vivo
C.J. Steib, M. Bystron, J.M. Härtl, C. Opelz, I. Liss, B. Göke, A.L. Gerbes
(Munich, D)
20. Stem cell properties of hepatic stellate cells
A.A. Gumerova, A.P. Kiassov, S. Abdulchakov, M.S. Kaligin, I.M. Gazizov,
D.I. Andreeva (Kazan, R)
21. Effects of pentoxifylline on established liver fibrosis and hepatic stellate cells
after bile duct ligation in rat
M.R. Mas, R. Ocal, G. Bilgin, Y. Karslioglu, B. Comert, L. Ozsari, I. Tasci,
N. Mas, A.T. Isik, Ö. Uzunalimoglu (Etlik, Ankara, TR)
22. CD95 ligand induces proliferation and CD95-tyrosine nitration in quiescent rat
hepatic stellate cells
R. Reinehr, A. Sommerfeld, S. Becker, D. Häussinger (Düsseldorf, D)
23. TGF-β signaling in hepatocytes promotes EMT and liver fibrogenesis, whereas
hepatocyte-specific expression of TGF-β antagonist Smad7 suffices to protect
from liver damage and fibrogenesis
H. Weng, L. Ciuclan, P. Godoy, S. Kanzler, R. Heuchel, E. Ueberham,
R. Gebhardt, J. Hamzavi, P. Mertens, K. Breitkopf, S. Dooley
(Mannheim, Mainz, Leipzig, D; Uppsala, S)
8
24. The role of ADH and ALDH gene polymorphism in alcohol liver cirrhosis
H. Cichoz-Lach, K. Celinski, J. Partycka, M. Slomka (Lublin, PL)
25. Effect of copy number repeats of the CCL3L1 gene one susceptibility to HCV
infection and fibro-inflammatory response
F. Grünhage, O. Gressner, H. Keppeler, H. Wasmuth, M. Odenthal, U. Drebber,
H.P. Dienes, C. Hellerbrand, T. Sauerbruch, F. Lammert
(Bonn, Aachen, Cologne, Regensburg, D)
26. Genome wide scans for fibrosis susceptibility genes in experimental crosses of
inbred mouse strains
R. Hall, S. Hillebrandt, K. Hochrath, F. Grünhage, T. Sauerbruch, F. Lammert
(Bonn, D)
27. Immunogenetic factors in the development of HCV-related liver cirrhosis
A.O. Romanov, T.V. Belyaeva, E.V. Esaulenko (St. Petersburg, R)
28. Hypersecretory haplotype of interleukin-1 gene cluster increases the risk of
alcoholic liver cirrhosis
J. Sperl, J. Petrasek, S. Reznakova, J.A. Hubacek, F. Stickel, P. Trunecka,
T. Berg, J. Spicák, M. Jirsa (Prag, CZ; Bern, CH; Berlin, D)
29. Beta-glucosidase 2 (GBA2) deficiency leads to zonal glucosylceramide
accumulation in the liver
Y. Yildiz, H. Matern, S. Weber, S. Matern, D.W. Russell, T. Sauerbruch,
F. Lammert (Bonn, Aachen, D; Dallas, USA)
30. Impact of peginterferon α-2b and ribavirin combined therapy on serum gastrin
and progastrin concentrations in Helicobacter pylori (HP) infected patients with
chronic hepatitis C (CHC)
W. Bielanski, W.W. Mazur, Z. Gonciarz, M. Gonciarz, S.J. Konturek,
M. Magiera, J.F. Rehfeld (Cracow, Chorzów, Sosnowiec, PL; Copenhagen, DK)
31. Researches on the immunologic-inflammatory answer at the patients chronically
infected with the hepatitis virus C
M. Deac, M. Boitan, R. Mihaila, A. Fraticiu, C.E. Rezi, A.V. Zaharie, A. Olteanu,
D. Dascalu, L. Tiurean, P.-J. Porr, C.C. Cipaian, D.V. Vulcu, O. Petrascu,
C.B. Beca, A.B. Boicean, L. Chicea, D. Orga-Dumitriu, C. Bidlo, A. Catana,
R. Mihaila (Sibiu, RO)
32. Detection of SEN virus in chronic hepatitis C virus infected patients genotype 4
and sequencing in Egypt
S.A. Girgis, A. Monis (Cairo, ET)
33. Clearance of HCV improves insulin resistance and early insulin secretion of
pancreas in chronic hepatitis C patients
R. Narita, M. Hiura, S. Abe, A. Tabaru, M. Otsuki (Kitakyushu, J)
9
34. Relationship of serum interleukin-6 and alpha tumor necrosis factor levels with
the metabolic syndrome in NASH patients
C.E. Rezi, M. Boitan, R. Mihaila, A. Fraticiu, A.V. Zaharie, A. Olteanu,
D. Dascalu, L. Tiurean, M. Deac, P.-J. Porr, C.C. Cipaian, D.V. Vulcu,
O. Petrascu, C.B. Beca, A.B. Boicean, L. Chicea, D. Orga-Dumitriu, C. Bidlo,
V. Crangaciu, A. Catana, R. Mihaila (Sibiu, RO)
35. Correlation between development of liver fibrosis and intrahepatic bile ducts'
state in chronic viral hepatitis and PBC
L. Fatkheeva, A. A. Goumerova, A.P. Kiassov, A.S. Sozinov, M. Fatkheeva
(Kazan, R)
36. Serum fibronectin as marker of liver regenerative process
L.S. Gotia, S.R. Gotia, D. Verdes, L.M. Susan (Timisoara, RO)
37. Is serum hyaluronic acid a marker of liver fibrosis in acute and chronic liver
disease?
J. Keiss, V. Sondore, N. Sevastjanova, B. Rozentale, L. Viksna (Riga, LV)
38. APRI in chronic hepatitis C patients with AST levels equal or below 1.5 of upper
limit of normal. To do or not to do?
O. Kosseva, E. Pophristova, K. Antonov, D.G. Adjarov, Z. Krastev (Sofia, BG)
39. Comparison of three non-invasive methods for the diagnosis of liver fibrosis in
chronic hepatitis B
O. Kosseva, E. Pophristova, E. Kaneti, S. Bocheva, D.G. Adjarov, Z. Krastev
(Sofia, BG)
40. Assessment of cholestasis and fibrogenesis in patients with primary biliary
cirrhosis
V. Kupcová, L. Turecky, M. Valková, M. Szántová, E. Uhlíková, S.Z. Zigraiová
(Bratislava, SK)
41. Serum gammaglutamyltranspeptidase, steatosis and fibrosis in chronic virus C
hepatitis
B. Mihai, C. Mihai, C. Cijevschi, C. Lacatusu, P. Gogalniceanu
(Iasi, RO; London, GB)
42. Alpha-2-macroglobulin in sera of patients with chronic hepatopathies and effect
of liver transplantation on level of this marker
L. Turecky, V. Kupcová, M. Szántová, E. Uhliková (Bratislava, SK)
43. Is IGF-1 response to growth hormone administration in cirrhosis reflect more
hepatocellular dysfunction rather than portal hypertension and malnutrition?
N. Assy, Y. Pruzansky, Z. Hochberg, D. Gaitini, Z. Shen Orr, Y. Baruch
(Safed, Haifa, IL)
44. Investigation of isoenzymes of gamma-glutamyltransferase and bile acids in
chronic liver diseases
V. Kupcová, L. Turecky, E. Uhliková, M. Szántová, A. Krsáková (Bratislava, SK)
10
45. Investigation of serum cytokines in chronic viral hepatitis after IFN-α treatment
V. Kupcová, L. Turecky, Z. Zelinkova, M. Valková, E. Seligová (Bratislava, SK)
46. Study of the relationship between chronic hepatitis C infection, non-alcoholic
fatty liver disease (NAFLD) and insulin resistance
N.M. Mashaal, A.A. Abdel Moety, E.A. Shaat, R.A.O El Sheikh (Alexandria, ET)
47. The study of correlation between IL-6 and TNF alpha and the biochemical liver
tests at the patients chronically infected with the hepatitis virus C
R. Mihaila, M. Boitan, A. Fraticiu, C.E. Rezi, A.V. Zaharie, A. Olteanu,
D. Dascalu, L. Tiurean, M. Deac, P.-J. Porr, C.C. Cipaian, D.V. Vulcu,
O. Petrascu, C.B. Beca, A.B. Boicean, L. Chicea, D. Orga-Dumitriu, C. Bidlo,
A. Catana, R. Mihaila (Sibiu, RO)
48. The study of correlation between IL-6, TNF alpha and erythropoietin with the
biochemical liver tests at the patients with non-alcoholic fatty liver disease
R. Mihaila, M. Boitan, R. Mihaila, C.E. Rezi, A. Fraticiu, A.V. Zaharie,
A. Olteanu, D. Dascalu, L. Tiurean, M. Deac, P.-J. Porr, C.C. Cipaian,
D.V. Vulcu, O. Petrascu, C.B. Beca, A.B. Boicean, L. Chicea, D. Orga-Dumitriu,
C. Bidlo, A. Catana (Sibiu, RO)
49. Serum apoptotic cytokeratin 18 neoepitope in acute and chronic liver damage
V. Sondore, J. Keiss, N. Sevastjanova, L. Viksna, B. Rozentale (Riga, LV)
50. Study of the correlation between nitric oxide profile, degree of liver injury and
structural vascular changes of the gastric mucosa in chronic hepatitis C virus
related liver diseases
M.Y. Taher, H. El Saed, N.M. Baddour, A.A. Abdel Moety, A. Zeid
(Alexandria, ET)
51. Serum selenium and carnitine concentrations of cirrhotic children
N. Uslu, I.N. Saltik-Temizel, H. Demir, F. Gürakan, H. Özen, A. Yüce
(Ankara, TR)
52. On the optimization of the 13C-alpha-ketoisocaproic acid breath test – An indepth study speaks for an extended breath sampling period
K. Jonderko, A. Kasicka-Jonderko, M. Kaminska, M. Bielecka,
B. Blonska-Fajfrowska (Sosnowiec, PL)
53. Band ligation vs. miniloops in the endoscopic treatment of esophageal varices –
Comparative pilot study
C. Banciu, L.M. Susan, O. Chirileanu, L. Marian, I. Romosan (Timisoara, RO)
54. Upper digestive bleeding in liver cirrhosis
C. Cijevschi, C. Mihai, V. Drug, P. Gogalniceanu (Iasi, RO; London, GB)
55. Upper digestive haemorrhage in cirrhosis - Prevalence and precipitating factors
D. Damian, M. Grigorescu, A. Berecz, M. Hogea, M. Rusu (Cluj-Napoca, RO)
11
56. Upper gastrointestinal bleeding in cirrhosis - Clinical aspects and evolution
D. Damian, M. Grigorescu, A. Berecz, M. Hogea, M. Rusu (Cluj-Napoca, RO)
57. Major complications in liver cirrhosis
C. Mihai, C. Cijevschi, B. Mihai, V. Drug, P. Gogalniceanu
(Iasi, RO; London, GB)
58. Use of clonidine in cirrhotic patients with large diuretic resistant ascitis –
Prospective pilot study
C. Banciu, L.M. Susan, V. Vacariu, C. Dascau, L. Marian, I. Romosan
(Timisoara, RO)
59. Superiority of the serum-ascites albumin gradient over the ascites total protein
concentration in diseases with ascites
N. Colic, D. Vrinic, A. Colic, S. Kazic, P. Svorcan, D. Necic, B. Dapcevic
(Belgrade, SRB)
60. The evolution of spontaneous bacterial peritonitis under medical treatment in
cirrhotic patients with ascites
A. Genunche-Dumitrescu, P. Mitrut, D. Badea, M. Badea (Craiova, RO)
61. E. coli lipopolysaccharides antibodies (CLA) and some species of chronic
hepatitis and liver cirrhosis
B.N. Levitan, A. Umerova, G. Levitan (Astrakhan, R)
62. Bacterial translocation in cirrhotic rats: Therapeutic role if iNOS inhibitor
S-methylisothiourea
M.R. Mas, S. Yilmaz, Ö. Uzunalimoglu, M. Ozyurt, I. Tasci, A.T. Isik, A. Ozcan,
B. Comert, N. Mas, C. Akay, H. Bozkaya, C. Yurdaydin (Ankara, Istanbul, TR)
63. Enzymes of purine degradation in lymphocytes of patients with liver cirrhosis
L. Turecky, V. Kupcová, V. Rendekova, M. Szántová, E. Uhliková
(Bratislava, SK)
64. Hepatorenal syndrome is a major complication of end stage liver cirrhosis and
has a poor prognosis
B. Appenrodt, J. Zielinski, J. Heller, T. Sauerbruch, M. Schepke (Bonn, D)
65. Assessment of the renal hemodynamics in liver cirrhosis with duplex-Doppler
ultrasonography
A. Caraba, L.M. Susan, A. Pacurari, I. Romosan (Timisoara, RO)
66. Predictive diagnosis of hepatorenal syndrome
P. Mitrut, A. Genunche-Dumitrescu (Craiova, RO)
67. Predictor factors in development of hepatopulmonary syndrome in patients with
liver cirrhosis
A. Genunche-Dumitrescu, P. Mitrut, D. Badea, M. Badea (Craiova, RO)
12
68. Pulmonary hypertension, a rare but dreaded complication of liver cirrhosis
M. Marin, O. Chira, S. Tanu, V. Mercea (Cluj-Napoca, RO)
69. Impaired cardiovagal autonomic function in patients with chronic hepatitis C:
Is there an association with sensory nerve function and cryoglobulinaemia?
J. Osztovits, T. Horvath, Z. Visnyei, T. Csák, G. Bekö, P.L. Lakatos, G. Lengyel,
E. Ibranyi, T. Toth, M. Abonyi, J. Feher, P. Kempler, M. Kollai, F. Szalay
(Budapest, H)
70. The etiological spectrum of portal hypertension in pediatric population –
Evolution and treatment (5 years retrospective study)
A. Constantinescu, C. Gheorghe, L. Gheorghe, C. Nicolae, I. Lupescu,
V. Herlea, A. Constantinescu, I. Popescu, C. Arion (Bucharest, RO)
71. Prehepatic portal hypertension in patients with portal thrombosis: Clinical and
laboratory characteristics
E. Kitsenko, E. Lukina, E.P. Sysojeva, E. Voronkova, E. Lubivy, G. Sukhanova,
S. Vasiljev (Moscow, R)
72. Myeloproliferative disorders in patients with prehepatic portal hypertension
E.P. Sysojeva, E. Lukina, E. Semenova, E. Voronkova, N. Khoroshko,
G.A. Frank (Moscow, R)
73. Minimal hepatic encephalopathy: The role of neuropsychological assessment in
the diagnosis
G. Jankovic, Z. Djuric, D. Mijac, E. Stefanova (Belgrade, SRB)
74. Efficacy of flumazenil administered in small repeated intravenous doses in
patients with hepatic encephalopathy
J.J. Jovic, N.J. Jovic, T. Pecelj-Brocic, B. Roganovic (Belgrade, SRB)
75. Prognostic significance of bacterial infection as precipating factors of chronic
hepatic encephalopathy in cirrhosis
S. Naumovski-Mihalic, M. Katicic, V. Colic-Cvrlje, B. Papa, T. Filipec-Kanizaj,
M. Ticak, B. Sabaric, B. Skurla (Zagreb, HR)
76. The role of Helicobacter pylori infection for development of portal
encephalopathy
D. Neagoe, G. Ianosi, A. Saftoiu, C. Vere, F. Racanel, D. Tudorascu
(Craiova, RO)
77. Hepatic encephalopathy does not correlate with depression and quality of life in
patients with liver cirrhosis: A single centre, prospective study
E. Wunsch, I. Zawada, M. Kaczmarczyk, E. Gebala, P. Milkiewicz
(Szczecin, PL)
78. UDCA in the treatment of primary biliary cirrhosis
K. Kalinova (Stara Zagora, BG)
13
79. The somatostatin analogue octreotide inhibits angiogenesis and VEGF
expression in the early, but not in advanced, stages of portal hypertension in
rats. Role of somatostatin receptor subtype 2 downregulation during the
evolution of the portal hypertensive syndrome
M. Mejias, E. Garcia-Pras, J. Bosch, M. Fernandez (Barcelona, E)
80. Comparison of two statins efficacy on a reversibility of experimental rat liver
fibrosis
O. Lukivskaja, Y. Popov, J. Zaks, D. Schuppan, V.U. Buko
(Grodno, WR; Boston, USA)
81. Effect's evaluation of the treatment with carvedilol versus propranolol and
isosorbide-5-mononitrate on hepatic haemodynamic parameters in patients with
liver cirrhosis
E. Tcaciuc, S. Matcovschi, A. Tcaciuc, N. Nacu (Chisinau, MOL)
82. The efficacy of the treatment with Losartan and Ursofalk® versus only Losartan
on hepatic haemodynamic parameters in patients with compensated liver
cirrhosis
A. Tcaciuc, S. Matcovschi, E. Tcaciuc, N. Nacu (Chisinau, MOL)
83. Atorvastatin inhibits fibrogenesis by inhibiting Ras and RhoA activation
J. Trebicka, M. Neef, K. Shir, M. Hennenberg, F. Lammert, J. Reichen,
T. Sauerbruch, J. Heller (Bonn, D; Bern, CH)
84. Urotensin II receptor antagonist palosuran lowers portal pressure by activation
of mesenterial RhoA/Rho-kinase signalling in liver cirrhosis
J. Trebicka, A. Eckhardt, M. Hennenberg, E. Biecker, F. Lammert,
T. Sauerbruch, J. Heller (Bonn, D)
85. Antioxidant taurine treatment in experimental liver fibrosis
Ö. Uzunalimoglu, M.R. Mas, S.A. Vural, S. Deveci, C. Akay, A.T. Isik,
M. Bozdayi (Ankara, Etlik, TR)
86. Nutritional state in alcoholic liver cirrhosis
J. Daniluk, E. Krzyzanowska, A. Zwolak (Lublin, PL)
87. Affective alterations: Determinant factor of sleep disorders in cirrhotic patients
M. González, J. Poniachick, M. Cumsille, A.M. Madrid, J. Brahm
(Santiago de Chile, RCH)
88. Impaired health-related quality of life in Romanian patients with hepatic cirrhosis
C. Pojoga, D.L. Dumitrascu (Cluj-Napoca, RO)
89. Fatigue, depression and functional disability in patients with liver cirrhosis
G. Stefanescu, C. Stefanescu (Iasi, RO)
90. Total paracentesis with albumin infusion in patients with ascites before
radiofrequency ablation (RFA) of hepatocellular carcinoma (HCC)
J. Genov, N. Grigorov, R. Mitova, B. Golemanov (Sofia, BG)
14
91. S-100 and GFAP-positive cells in human liver cirrhosis and in human
hepatocellular cancer
M.V. Gulubova, T. Vlaykova, V. Velev (Stara Zagora, BG)
92. Frequency of hepatocellular carcinoma in chronic viral infections
D. Svarc (Novi Sad, SRB)
93. Expression of the reactive oxygen species-detoxifying enzymes, GST-PI and
CU/ZN-SOD in cirrhosis and hepatocellular carcinoma
T. Vlaykova, M.V. Gulubova (Stara Zagora, BG)
94. Orthotopic liver transplantation for chronic hepatitis B
S. Frankova, P. Trunecka, J. Sperl, M. Adamec, J. Spicák, S. Vitko
(Prague, CZ)
95. Plasma value of endothelin-1 versus renal and liver function in patients after
orthotopic liver transplantation in long-term follow up
H. Gottfriedova, O. Schück, L. Karasová, M. Stollová, J. Maly, J. Skibová,
P. Trunecka, J. Spicák (Prague, CZ)
96. Absence of teratogenicity of sirolimus used during early pregnancy in a liver
transplant recipient
I. Jankowska, U. Oldakowska-Jedynak, Z. Jabiry-Zieniewicz, A Cyganek,
J. Pawlowska, M. Teisseyre, P. Kalicinski, L. Paczek, J. Socha (Warsaw, PL)
97. Recurrence of nonalcoholic steatohepatitis in transplanted liver in a 13-year-old
boy
I. Jankowska, P. Socha, J. Pawlowska, M. Teisseyre, D. Gliwicz, P. Kalicinski,
J. Cielecka-Kuszyk, J. Socha (Warsaw, PL)
98. Activation of latent TGF-β inside the hepatocyte leads to target gene expression
via autocrine, intracellular signalling – Relevance for the initiation of acute and
chronic liver injury
O.A. Gressner, B. Lahme, M. Siluschek, K. Rehbein, R. Weisskirchen,
A.M. Gressner (Aachen, D)
15
Session I
Initiation
17
Apoptosis and oncotic necrosis: The common pathways of
liver injury
Hartmut W. Jaeschke
Department of Pharmacology, Toxicology & Therapeutics; University of Kansas
Medical Center, Kansas City, KS 66160, USA
Apoptosis and oncotic necrosis are modes of cell death defined by their
morphological changes. Hallmarks of apoptosis are cell shrinkage, chromatin
margination and condensation, and formation of apoptotic bodies. The characteristics
of necrotic cell death include cell swelling, vacuolation, nuclear fragmentation
(karyolysis) and cell contents release. Over the last 10 years substantial progress
was made in the understanding of intracellular signaling mechanisms of cell death, in
particular apoptosis. However, more recent insight into mechanisms of oncotic
necrosis indicates that many of the apoptotic signaling mechanisms also apply to
necrotic cell death. In particular, mitochondrial dysfunction and opening of the
membrane permeability transition pore are integral features of both apoptosis and
oncotic necrosis. The considerable overlap between these mechanisms of cell death
and the fact that apoptosis can easily deteriorate into secondary necrosis, is the
basis for many misinterpretations and controversial discussions on the mode of cell
death in liver diseases.
Chronic liver injury and cell death can lead to fibrosis, which is a pathological process
of excessive extracellular matrix deposition by activated hepatic stellate cells. Most
mechanisms of oncotic necrosis involve reactive oxygen species and the generation
of lipid peroxidation products, which have been shown to activate stellate cells
directly and promote collagen formation. On the other hand, phagocytosis of
apoptotic bodies and necrotic cell debris activates Kupffer cells to generate profibrogenic mediators such as transforming growth factor-β and platelet derived
growth factor, both of which activate stellate cells. Furthermore, a number of proteins
released by necrotic cells induce pro-inflammatory cytokine formation in Kupffer cells
and stellate cells by binding to toll-like receptors. The cytokines promote an
inflammatory response with leukocyte recruitment, which propagates cell injury and
stellate cell activation. In addition, there is evidence that hepatic stellate cells can
phagocytose apoptotic bodies leading to activation of NADPH oxidase and collagen
formation in these cells. Thus, both apoptotic cell death and oncotic necrosis directly
and indirectly contribute to hepatic stellate cell activation and liver fibrosis through
multiple mechanisms and self-amplifying loops.
19
Entero-hepatic lymphocyte homing and liver manifestations of
IBD
David H. Adams, Bertus Eksteen & Allister Grant
Liver Research Group, MRC Centre for Immune Regulation, 5th Floor IBR, The
University of Birmingham Medical School, Wolfson Drive, Edgbaston, Birmingham,
B152TT, UK
The liver and gut axis, which is central to our ability to absorb and process nutrients,
requires immunological control to prevent the induction of immune responses to food
antigens while maintaining immunity to pathogens. Translocation of gut antigens to
the portal circulation means that antigens not eliminated directly in the gut are
brought to the liver, where further levels of immune regulation operate. This constant
exposure to gut antigens might explain why the human liver contains more than 1010
lymphocytes including memory T cells with specificity for persistent viruses. It is
possible that some memory lymphocytes that are activated in the gut recirculate
through the liver to allow memory of mucosal antigens to be shared across both
sites. Although there is currently no good evidence that mucosal memory T cells
show enhanced trafficking through the normal liver there is evidence that under
certain inflammatory conditions mucosal T cells are preferentially recruited to the liver
in for instance graft versus host disease and in patients with hepatic complications of
inflammatory bowel disease (IBD). The existence of such shared pathways could
help to explain the association between autoimmune liver disease and IBD.
Lymphocytes that are destined to traffic to the gut are primed by DCs in mesenteric
lymph nodes and Peyer’s patches which have the unique ability to imprint responding
T cells with the gut homing receptors CCR9 and α4β7 allowing them to respond to
CCL25 and MAdCAM-1 both of which are largely restricted to the gut.
But what occurs if tissue specificity is lost and MAdCAM-1 and CCL25 are aberrantly
expressed? In such a scenario mucosal lymphocytes might be recruited to
extraintestinal sites and thereby cause inflammation outside the gut. Most extraintestinal manifestations of inflammatory bowel disease IBD are driven by the
inappropriate recruitment of mucosal effector lymphocytes to extra-intestinal tissues
and resolve once the bowel inflammation is treated. However the autoimmune liver
diseases autoimmune hepatitis and primary sclerosing cholangitis (PSC) develop
when bowel inflammation is quiescent or after the bowel has been removed. We
have shown that these diseases are associated with the aberrant expression of
CCL25 and MAdCAM-1 on hepatic endothelium which results in the recruitment of
α4β7+CCR9+ mucosal T cells to the liver where they interact with hepatocytes and
biliary to mediate tissue damage. Evidence that these T cells are of mucosal origin is
provided by experiments in which DCs isolated from gut or normal and inflamed
human liver were used to activate naïve T cells and B cells and the induction of a gut
homing phenotype looked for in responding lymphocytes. In the absence of retinoic
acid only DCs isolated from the gut were able to imprint α4β7+CCR9+ on responding
T cells or IgA-secreting B cells suggesting that these cells are originally activated in
the mucosal immune system rather than the liver.
20
Thus we propose a novel model to explain the extra-intestinal complications of
inflammatory bowel disease in which long-lived mucosal T cells undergo enterohepatic recirculation between the liver and gut. Under normal conditions this will
provide immune surveillance across both sites. However, aberrant expression of gut
adhesion molecules in the liver can amplify the recruitment of mucosal lymphocytes
resulting in sustained hepatitis and liver injury.
21
Immune responses to chronic hepatitis C virus infection
Ulrich Spengler
Medizinische Klinik I, Universitätsklinikum Bonn, Germany
Hepatitis C virus (HCV) has a high propensity to persist and cause chronic liver
inflammation. HCV itself does not seem to be cytopathic, because the rise in
aminotransferases is usually linked to the onset of anti-viral immune responses
rather than viral replication. Thus, liver disease is commonly attributed to immunemediated mechanisms. This concept is further supported by the observation that
functionally relevant polymorphisms in immunoregulatory genes are associated both
with different outcomes of natural infection and divergent responses to anti-HCV
treatment.
It remains still unclear which pathways determine resolution of HCV infection. The
role of HCV antibodies still remains uncertain. Of note, antibodies in hepatitis C may
even augment liver damage by directing damage to virus-coated but un-infected cells
via antibody-dependent cellular cytotoxicity instead of being protective. Unlike
antibodies, mechanisms of innate and adaptive cellular immunity are considered
pivotal for control over HCV infection. However, in patients who progress to chronic
hepatitis C these mechanisms of cellular immunity obviously fail to eliminate the
virus. HCV proteins can interact with several pathways regulating the host’s immune
response. These interactions may lead to impaired innate immunity and altered
functions of natural killer cells, virus-specific CD4+ and CD8+ T cells. HCV-specific
T cells become increasingly unresponsive and apparently disappear over time.
Moreover, regulatory T cells are generated during chronic hepatitis which on one
hand further suppress antiviral immune responses but on the other hand can dampen
tissue damage. There is circumstantial evidence to suggest that cytotoxic
lymphocytes are crucial for HCV elimination. These effector cells kill their target cells
via perforin/granzyme and release of cytokines. Cytotoxic cytokines, however, can
cause damage also to uninfected cells and attract non-specific inflammatory cells to
invade the liver. These non-specific mechanisms expand liver damage and promote
fibrosis progression. Long-term chronic inflammation in hepatitis C is a risk factor for
disease progression towards hepatocellular carcinoma and lymphoma, probably
because cytokines induced by chronic hepatitis C also exert growth promoting effects
on a variety of different cells. Thus, the immune response in hepatitis C is a twoedged sword: It is needed to eliminate the virus in the acute phase of infection but
during hepatitis it is a major factor contributing to progressive liver disease.
22
Cytokines: Interleukin-6 and TNF signalling
Christian Trautwein
Medizinische Klinik III, Universitätsklinikum der RWTH Aachen, Germany
Intestinal translocation of bacteria and bacterial products results in its influx via the
portal vein into the liver. Therefore the liver represents a filter system for all potential
pathogens entering the body via the gastrointestinal tract. In order to protect the body
from systemic spreading the liver has developed major tasks in immune defense
especially by using mechanisms of innate immunity. In the liver residual
macrophages, Kupffer cells detect incoming bacterial products for example via Tolllike receptors. As a consequence these cells become activated and release proinflammatory cytokines like tumor necrosis factor alpha (TNF) and interleukin-6 (IL-6).
These cytokines initiate an immune response in the surrounding tissue which also
includes further attraction of cells of the immune system. However also in
parenchymal liver cells – the hepatocytes – these cytokines have a major impact on
gene expression and activate intracellular pathways. These pathways are of major
relevance not only for the immune response, but also for other functions in the liver,
e.g. metabolic tasks. Therefore the liver is the most essential organ mediating the
interplay between Gut-induced inflammation and organ-specific functions. We thus
developed in our laboratory specific mouse strains that are able to specifically
address the role of IL-6 and TNF-dependent pathways in hepatocytes and study their
in vivo relevance.
In hepatocytes IL-6 triggers intracellular pathways via the gp130 receptor and as a
consequence activates the acute-phase response (APR). Results of recent years
demonstrated that gp130-dependent pathways are protecticve for hepatocytes and
therefore in different models of liver injury lack of IL-6 expression results in more
injury. However several IL-6 family members do exist and have some redundancy in
their response during inflammation. Therefore we developed a hepatocyte-specific
gp130 knockout mouse (Alb/gp130-/-). These animals are more susceptible for LPSinduced liver injury and also show more bacterial translocation e.g after bile duct
ligation. Therefore these animals are a valuable tool to understand and test the
impact of inflammatory pathways on functions of the liver during metabolic insults
and stress conditions. Additionally, our recent results demonstrated a direct role of
the APR for arteriosclerosis progression as one of the major consequences of the
metabolic syndrome. Lack of APR in Alb/gp130-/- mice is associated with less
arteriosclerosis as acute-phase proteins like serum amyloid A are directly involved in
triggering arteriosclerosis progression. Therefore activation of the gp130 system in
hepatocytes links bacterial translocation in the bowel to systemic inflammation during
metabolic conditions.
TNF levels also increase after Kupffer cell activation in the liver. TNF binds its
receptor TNF-R1 on hepatocytes and induces distinct signaling cascades leading to
NF-κB, Jun Kinase and apoptosis activation. Especially NF-κB has been shown to
play an important role in hepatocytes as it mediates protection and inflammatory
mechanism in hepatocytes. NF-κB is held in the cytoplasm in its inactive form when
bound to IκB. Upon stimulation, the IKK complex (consisting of IKKα/IKK1, IKKβ/IKK2
and IKKγ/NEMO) phosphorylates I-κB, leading to its degradation and resulting in NFκB nuclear translocation and target gene transcription. Knockout models for IKK2 and
23
NEMO result in embryonal lethality due to hepatocyte apoptosis. Therefore we
generated hepatocyte-specific NEMO knockout (Alb/NEMO-/-) mice. Alb/NEMO-/animals show a spontaneous severe phenotype characterized by steatohepatitis,
which over time results in liver fibrosis and development of hepatocellular
carcinomas. Therefore this animal model at present is the only available model
showing the natural progression of steatohepatitis in mice as regularly found in
humans. Interestingly, preliminary data demonstrate that progression of
steatohepatitis in Alb/NEMO-/- animals is dependent on the health status of the
animals. Therefore bacterial infections have a direct impact on inflammation and
fibrosis in the liver of these animals.
Literature:
1. Klein C, Wustefeld T, Assmus U, Roskams T, Rose-John S, Muller M,
Manns MP, Ernst M, Trautwein C. The IL-6-gp130-STAT3 pathway in
hepatocytes triggers liver protection in T cell-mediated liver injury. J Clin Invest
2005; 115: 860–869.
2. Luedde T, Assmus U, Wustefeld T, Meyer zu Vilsendorf A, Roskams T, SchmidtSupprian M, Rajewsky K, Brenner DA, Manns MP, Pasparakis M, Trautwein C.
Deletion of IKK2 in hepatocytes does not sensitize these cells to TNF-induced
apoptosis but protects from ischemia/reperfusion injury. J Clin Invest 2005; 115:
849–859.
3. Pietrangelo A, Dierssen U, Valli L, Garuti C, Rump A, Corradini E, Ernst M,
Klein C, Trautwein C. STAT3 is required for IL-6-gp130-dependent activation of
hepcidin in vivo. Gastroenterology 2007; 132: 294–300.
4. Luedde T, Beraza N, Kotsikoris V, van Loo G, Nenci A, De Vos R, Roskams T,
Trautwein C, Pasparakis M. Deletion of NEMO/IKKgamma in liver parenchymal
cells causes steatohepatitis and hepatocellular carcinoma. Cancer Cell 2007; 11:
119–132.
5. Beraza N, Ludde T, Assmus U, Roskams T, Vander Borght S, Trautwein C.
Hepatocyte-specific IKK gamma/NEMO expression determines the degree of
liver injury. Gastroenterology 2007; 132: 2504–2517.
24
Session II
Mediators
25
Growth factors in experimental liver fibrogenesis
R. Weiskirchen, E. Borkham-Kamphorst, E. Kovalenko, S. Meurer, O. Scherner,
W.N. Vreden, O.A. Gressner, J. Herrmann, F. Drews, A.M. Gressner
Institute of Clinical Chemistry and Pathobiochemistry, RWTH University Hospital
Aachen, Germany
A broad array of cytokines and chemokines contribute to hepatic fibrogenesis, a
process in which quiescent hepatic stellate cells (HSC) become activated and
transdifferentiate into myofibroblast (MFB)-like cells. We have shown that the
inactivation of platelet-derived growth factor (PDGF) or transforming growth factor-β
(TGF-β) by soluble receptors or other antagonistic strategies is sufficient to block
ongoing hepatic fibrogenesis in various animal and cellular models [1, 2].
Furthermore, recent reports have demonstrated that the bone morphogenetic protein7 (BMP-7) antagonizes the strong fibrogenic effects of TGF-β1 during hepatic injury.
We here present data delineating the impact of the different PDGF-isoforms (PDGFA, -B, -C, -D), TGF-β, and BMP-7 for activation of HSC and hepatic fibrogenesis. We
show that at the cellular level the signalling pathways form an interconnected
signalling network influencing the activity of different target genes (e. g connective
tissue growth factor) in a cell type specific (HSC versus hepatocytes) and
activation/transdifferentiation-dependent manner [3]. This complex network of
cytokine activities is further influenced by accessory receptors (e. g. endoglin) and
binding proteins (e. g. LTBP-1) that modulate and fine tune the activity of respective
cytokine actions thereby influencing the fibrogenic response [4, 5]. In addition, we
present data showing that the different cytokines regulate potent transcription factors
(e. g. serum response factor) that control superior cellular functionalities of
profibrogenic HSC during cellular activation and differentiation [6]. The knowledge of
the different pathways, their interactions, and the cellular responses induced by
members of the PDGF and TGF-β superfamilies offer several steps of
pharmacological interferences in experimental hepatic fibrogenesis.
References:
[1]
Borkham-Kamphorst E, Herrmann J, Stoll D, Treptau J, Gressner AM,
Weiskirchen R. Dominant-negative soluble PDGF-β receptor inhibits hepatic
stellate cell activation and attenuates liver fibrosis. Lab. Invest. 2004; 84:
766–777.
[2]
Borkham-Kamphorst E, van Roeyen CR, Ostendorf T, Floege J, Gressner AM,
Weiskirchen R. Pro-fibrogenic potential of PDGF-D in liver fibrosis. J. Hepatol.
2007, in press.
[3]
Gressner OA, Lahme B, Demirci I, Gressner AM, Weiskirchen R. Differential
effects of TGF-β on connective tissue growth factor expression in hepatic
stellate cells and hepatocytes. J. Hepatol. 2007, in press.
27
[4]
Scherner O, Meurer SK, Tihaa L, Gressner AM, Weiskirchen R. Endoglin
differentially modulates antagonistic TGF-β and BMP-7 signaling. J. Biol. Chem.
2007, in press.
[5]
Drews F, Knoebel S, Moser M, Muhlack KG, Stoll C, Bosio A, Gressner AM,
Weiskirchen R. Disruption of the latent transforming growth factor-β binding
protein-1 gene causes alteration in facial structure and reduced TGF-β activity,
submitted for publication.
[6]
Herrmann J, Haas U, Gressner AM, Weiskirchen R. TGF-β up-regulates serum
response factor in activated hepatic stellate cells, submitted for publication.
28
Genetic determinants of liver fibrogenesis
F. Lammert
Department of Medicine I, University Hospital Bonn, University of Bonn, Bonn,
Germany
Hepatic fibrosis is the result of chronic viral, toxic, autoimmune, or cholestatic liver
injury. During the complex fibrotic response, distinct cell populations (e. g. hepatic
stellate cells, portal myofibroblasts, bone-marrow derived fibrocytes) might contribute
to the excess deposition of abnormal extracellular matrix (ECM), and interactions
between multiple cytokines, chemokines, receptors and ECM components modulate
the fibrotic phenotypes. Of note, course and extent of hepatic fibrosis display
significant variability among individual patients. These well-known differences in
progression of hepatic fibrosis have been attributed to age, gender, and exogenous
factors, e.g. coinfections or alcohol consumption. However, host genetic factors are
likely to modify liver fibrogenesis and to contribute to the overall variability in disease
progression. In the past decade single variants of human genes encoding
immunoregulatory proteins as well as pro- and antiinflammatory cytokines were
studied in association studies in patients with chronic hepatitis C virus infection,
alcoholic liver disease, and autoimmune liver diseases, yielding inconsistent results
(Österreicher et al. 2007). Recently new and potent high-throughput methods for
genome analysis have been implemented, in particular quantitative trait locus (QTL)
analyses in experimental mouse crosses and mouse-human comparative genomics
that allow the identification of genes that contribute to genetically complex diseases
such as liver fibrosis (Hillebrandt et al. 2005; Lammert 2007). Genetic networks that
control liver gene expression can be dissected through the combination of largescale quantitative mRNA expression analysis in reference populations such as BXD
recombinant inbred mouse lines (Gatti et al. 2007). Based on genome wide
association (GWA) analyses, a novel "gene signature" consisting of seven variants
has been proposed to identify the risk of progressive fibrosis in patients with chronic
hepatitis C virus infection (Huang et al. 2007). These genomic approaches represent
invaluable tools to identify unknown pathomechanisms, to investigate how systems of
genes interact, and to define therapeutic targets.
References:
Österreicher CH, Stickel F, Brenner DA. Genomics of liver fibrosis and cirrhosis.
Semin Liver Dis 2007; 27: 28–43.
Hillebrandt S, Wasmuth HE, Weiskirchen R, Hellerbrand C, Keppeler H, Werth A,
Schirin-Sokhan R, Wilkens G, Geier A, Lorenzen J, Köhl J, Gressner AM, Matern S,
Lammert F. Complement factor 5 is a quantitative trait gene that modifies liver
fibrogenesis in mice and humans. Nat Genet 2005; 37: 835–843.
Lammert F. Genetic determinants of complex liver diseases: mouse models and
quantitative trait locus analysis. In: Rodés J, Benhamou JP, Rizzetto M, Reichen J,
Blei A (eds.). Oxford's Textbook of Clinical Hepatology: From Basic Science to
Clinical Practice. Blackwell Publishing, Oxford 2007
29
Gatti D, Maki A, Chesler EJ, Kirova R, Kosyk O, Lu L, Manly KF, Williams RW,
Perkins A, Langston MA, Threadgill DW, Rusyn I. Genome-level analysis of genetic
regulation of liver gene expression networks. Hepatology 2007; 46: 548–557.
Huang H, Shiffman ML, Friedman S, Venkatesh R, Bzowej N, Abar OT,
Rowland CM, Catanese JJ, Leong DU, Sninsky JJ, Layden TJ, Wright TL, White T,
Cheung RC. A 7 gene signature identifies the risk of developing cirrhosis in patients
with chronic hepatitis C. Hepatology 2007; 46: 297–306.
30
Molecular mechanisms of fibrosis progression in non-alcoholic
steatohepatitis (NASH)
Claus Hellerbrand
University of Regensburg, Germany
Non-alcoholic fatty liver disease (NAFLD) is associated with obesity, type 2 diabetes,
dyslipidaemia and hypertension. These conditions have insulin resistance as the
common factor and cluster to form the metabolic syndrome. Most patients with
NAFLD have increased liver fat content alone (simple steatosis), but others develop
increasing hepatic inflammation known as non-alcoholic steatohepatitis (NASH) and
up to 20% of patients develop progressive hepatic fibrosis and may eventually
progress to cirrhosis or liver failure. In addition, the risk of development of
hepatocellular carcinoma in NASH-related cirrhosis is comparable to hepatitis C
infection. Considering the rising prevalence of obesity, NAFLD has to be considered
as (one of) the most frequent chronic liver disease in western countries.
A wealth of studies in animal models and humans with this disorder have increased
our understanding of the pathogenesis of NASH, however, still the underlying
mechanisms that influence that steatosis can progress to steatohepatitis (ie, the
association of lipid overload and inflammation) and fibrosis are largely unknown.
Currently, there is a "two hit" hypothesis: in addition to the "first hit” namely steatosis
characterized by an increase in lipid deposition in hepatocytes, a “second hit” is
required to induce inflammation. As one mechanism that promotes inflammation in
NASH lipid peroxidation and has been identified that induces increased DNA
oxidative damage and cell death.
Hepatic (necro)inflammation appears necessary but not sufficient for progression to
severe fibrosis, since progression from NASH to cirrhosis develops only in a
(relatively small) fraction of patients. This may reflect the requirement for additional
factors, a "third hit", to initiate and perpetuate fibrogenesis. Looking for such factors it
appears that in addition to histopathology of NASH that resembles that of alcoholinduced steatohepatitis (ASH) also common pathogenic mechanisms exist. For
example, similarly as in ASH, products of intestinal bacteria, particularly LPS, have
been identified to promote disease progression in NASH.
Furthermore, also the loss or reduced expression of (hepato)protective factors as
adiponectin may cause the "third hit". Adiponectin is specifically secreted by adipose
tissue and adiponectin serum levels almost always decrease in obesity. Interestingly,
hypoadiponectinemia has been identified as risk factor for fibrosis progression in
NASH independent of insulin resistance. Complementary, several studies identify
insulin resistance as a promotor of fibrosis in NASH, and insulin induces proliferation
and collagen expression in hepatic stellate cells, the main effectors of fibrosis in the
liver.
Additionally, it appears that also pure fatty liver without insulin resistance has the
potential to promote liver fibrosis. Increased hepatic lipid accumulation results in an
induction of CYP2E1, leading to an increased formation of reactive oxygen species,
which may directly contribute to the activation of hepatic stellate cells. Interestingly,
31
hepatocyte regeneration has been shown to be impaired in fatty livers, and steatosis
promotes hepatic fibrosis in experimental models. Of note, in addition to the amount
also the type of dietary fat seems to determine these effects. This and other findings
indicate that steatosis promotes fibrogenesis in liver diseases in general. For
example, hepatic steatosis is an independent risk factor for the progression of fibrosis
in patients with chronic hepatitis C infection, and weight reduction in these patients
reduces fibrosis and hepatic stellate cell activation. In summary, one might suggest a
second hypothesis: In addition to a "third hit", the "first hit", e. g. steatosis, may
already determine whether the liver will react with severe fibrosis in response to a
"second hit". If true, this would have important prognostic, diagnostic and therapeutic
implications for NAFLD and chronic liver disease in general.
32
New role of bile acid metabolism in bacterial translocation
Antonio Moschetta
Clinica Medica “Augusto Murri”, Dept. of Internal and Public Medicine, University of
Bari, Italy
Laboratory of Lipid Metabolism and Cancer, Dept. of Translational Pharmacology,
Consorzio Mario Negri Sud, Santa Maria Imbaro, Chieti, Italy
Bile acids are endogenous molecules synthesized in the liver as end product of
cholesterol metabolism. Secreted in bile, stored in the gallbladder in the inter-prandial
period, bile acids are released in the intestine in response to food in-take. After active
up-take in the ileum, bile acids return back to the liver through the portal vein, thus
completing their enterohepatic circulation. Classical physiologic functions of bile acid
include promotion of hepatic homeostasis and bile flow, support of the smooth
progression of lipid digestion and absorption, absorption of lipophilic vitamins,
cholesterol solubilization and disposal (1). Moreover, bile acids are directly involved
in preserving the physiologic commensal host-intestinal bacteria relationship. The
microbial flora hosted in the intestinal lumen is precious since it solves key metabolic
roles for nutrient processing, as well as correct development and tuning of the
immune system. The delicate host-intestinal bacteria balance is regulated to maintain
a fixed composition of microbial species, while preventing their excessive
proliferation. In pathophysiological conditions, the expansion of intestinal microflora
leads to migration of bacteria through the intestinal mucosal, up to mesenteric lymph
nodes and extraintestinal tissues. Bacterial overgrowth and translocation play major
pathophysiologic roles, especially in cirrhotic patients, with portal hypertension,
ascites, spontaneous bacterial peritonitis, and hepato-renal syndrome (2).
The bacteriostatic activity of bile acids has long being recognized: the first reports of
the inhibitory effects of bile acids upon intestinal bacteria proliferation date back to
1970 (3). Both in experimental models and humans, interrupted bile flow results in
hyper-proliferation of intestinal microflora, leading to inflammation of intestinal
epithelia, bacterial translocation, and systemic infection (4). Since biliary bile acid
output is strikingly reduced in cirrhotic patients, the disruption of the host-intestinal
microflora balance typically observed during the course of the disease has been
linked to bile acid depletion in the intestinal lumen (5). In line with this hypothesis,
therapeutic replenishment of conjugated bile acids by oral feeding in experimental
cirrhotic rats resolves bacterial over-growth and translocation, resulting in increased
survival (6). The mechanism whereby bile acids inhibit bacterial proliferation was
classically restricted to direct contact and detergency (7), and related to bile acid
hydrophobicity (8). Compelling new evidence links the bacteriostatic activity of bile
acids to a transcriptional mode. Farnesoid X Receptor (FXR) is the nuclear receptor
bile acid sensor (9,10) responsible for a coherent transcriptional regulation of a series
of genes that codify for proteins directly involved in the maintenance of lipid
homeostasis in the gut-liver axis, with relevance for disease such as cholesterol
gallstone disease (11). FXR knockout mice display increased levels of both aerobic
and anaerobic bacteria in the intestinal epithelia, deteriorated epithelial barrier,
greater bacterial population in mesenteric lymph nodes (12). At molecular level,
activation of FXR by bile acids ignites transcription of genes involved in antimicrobial
defense, innate immune response, mucosal fitness (mucus secretion, vascular tone,
maintenance of the epithelial barrier): such genes include inducible nitric oxide
33
synthase, angiogenin, carbonic anhydrase 12, interleukin 18. In line with these
findings, bile duct ligated rats treated with FXR agonists display inhibition of bacterial
overgrowth and reduction of intestinal mucosal damage. Enteroprotection by
synthetic FXR agonists is lost in FXR knockout mice.
These findings extend the signaling competence of bile acids: next to shielding the
liver from the toxic effect of bile acid overload, FXR also promotes intestinal health
(13). Consistent with the enteroprotective role, FXR is highly expressed in the
intestine, reaching highest levels in ileum and colon. Would FXR agonists be helpful
in the treatment and prevention of intestinal overgrowth? The road is open, but fewer
obstacles are still there. One of the main questions is: how is FXR expression and
function regulated in the intestine?
Reference list:
1.
Hofmann AF. Biliary secretion and excretion in health and disease: Current
concepts. Ann Hepatol 2007; 6: 15–27.
2.
Wiest R, Garcia-Tsao G. Bacterial translocation (BT) in cirrhosis. Hepatology
2005; 41: 422–433.
3.
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intestinal microflora – A function for simple bile acids? Gastroenterology 1971;
61: 228–233.
4.
Berg RD. Bacterial translocation from the gastrointestinal tract. Trends
Microbiol 1995; 3: 149–154.
5.
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Hepatic secretion of bilirubin and biliary lipids in patients with alcoholic
cirrhosis of the liver. Digestion 1983; 26: 80–88.
6.
Lorenzo-Zuniga V, Bartoli R, Planas R, Hofmann AF, Vinado B, Hagey LR, et
al. Oral bile acids reduce bacterial overgrowth, bacterial translocation, and
endotoxemia in cirrhotic rats. Hepatology 2003; 37: 551–557.
7.
Binder HJ, Filburn B, Floch M. Bile acid inhibition of intestinal anaerobic
organisms. Am J Clin Nutr 1975; 28: 119–125.
8.
Sung JY, Shaffer EA, Costerton JW. Antibacterial activity of bile salts against
common biliary pathogens. Effects of hydrophobicity of the molecule and in the
presence of phospholipids. Dig Dis Sci 1993; 38: 2104–2112.
9.
Makishima M, Okamoto AY, Repa JJ, Tu H, Learned RM, Luk A, et al.
Identification of a nuclear receptor for bile acids. Science 1999; 284:
1362–1365.
34
10.
Parks DJ, Blanchard SG, Bledsoe RK, Chandra G, Consler TG, Kliewer SA, et
al. Bile acids: natural ligands for an orphan nuclear receptor. Science 1999;
284: 1365–1368.
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Moschetta A, Bookout AL, Mangelsdorf DJ. Prevention of cholesterol gallstone
disease by FXR agonists in a mouse model. Nat. Med 2004; 10: 1352–1358.
12.
Inagaki T, Moschetta A, Lee YK, Peng L, Zhao G, Downes M, et al. Regulation
of antibacterial defense in the small intestine by the nuclear bile acid receptor.
Proc Natl Acad Sci USA 2006; 103: 3920–3925.
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Modica S, Moschetta A. Nuclear bile acid receptor FXR as pharmacological
target: are we there yet? FEBS Lett 2006; 580: 5492–5499.
Correspondence to:
Dr. A. Moschetta
Via Nazionale 8/A
66030 Santa Maria Imbaro (CH), Italy
Phone (39) 0872-570344
Fax (39) 0872-570299
E-mail: [email protected]
35
Session III
Cell responses to chronic liver injury
37
Role of hepatic progenitor cells in regeneration
Roskams, T., M.D., Ph.D.
Dept. Morphology and Molecular Pathology, KU Leuven, Belgium
Hepatic progenitor cells are immature epithelial cells that reside in the smallest
ramifications of the biliary tree in human liver. These cells are capable of
differentiating towards the biliary and the hepatocytic lineage and represent the
human counterpart of the oval cells in rodent liver. An increased number of
progenitor cells (referred to as ‘activation’) and differentiation of the same towards
hepatocytes and/or bile duct epithelial cells is a component of virtually all human liver
diseases. The extent of progenitor cell activation and the direction of differentiation
are correlated respectively with the severity of the disease and the type of mature
epithelial cell type (hepatocyte or bile duct epithelial cell) that is damaged. Anologous
to findings in animal models of hepatocarcinogenesis, human hepatic progenitor cells
most likely can give rise to hepatocellular carcinoma.
The factors that govern human hepatic progenitor cell activation and differentiation
are beginning to be indentified. Growth factors like hepatocyte growth factor,
transforming growth factor alpha and beta play a role. Recently it has been shown
that also the autonomic nervous system plays a role in liver regeneration. Since the
liver is denervated after liver transplantation this is of relevance for the regenerative
capacity of the transplanted liver. The microenvironment is essential for the
differentiation of progenitor cells. Interaction of progenitor cells with the surrounding
stroma has to be further explored; the stroma contains mesenchymal cells like
stellate cells/myofibroblasts, matrix components, growth factors. Using microdissection and microarray techniques, we could identify different stromal factors in
biliary diseases, compared to hepatitis with parenchymal damage, in which
progenitor cells differentiate into hepatocytes.
Animal data indicate that oval cells are activated when oxidative stress inhibits the
regenerative capacity of more mature hepatocytes. Inhibition of replicative activity of
mature hepatocytes is also shown in human liver diseases like alcoholic hepatitis and
viral hepatitis. Recently, Wiemann et al illustrated that hepatocyte telomere
shortening and senescence are general markers of human liver cirrhosis induced by
a wide variety of etiologies. A study of Falkowski et al very nicely confirms inhibition
of hepatocyte replication in the cirrhotic stage of chronic viral hepatitis and that this is
accompanied by an increased replication rate of the progenitor cell compartment.
This increase in progenitor cell activation has also been shown in (cirrhotic) alcoholic
and non-alcoholic fatty liver disease, conditions in which replication of hepatocytes is
also inhibited. Overall, oxidant-induced replicative senescence, as well as the
progenitor cell response, seems to be stereotypical, irrespective of the type of
underlying liver disease. Progenitor cell activation is particularly pronounced in the
cirrhotic stage of a variety of chronic liver diseases, the stage in which most
carcinomas arise.This should be kept in mind when progenitor cells are considered
as a therapeutical option in chronic liver diseases.
Several recent detailed immunohistochemical studies have shown that hepatocellular
carcinomas show a range of hepatocellular, but also progenitor cell features,
suggesting that at least part of the HCCs originate from progenitor cells. HCCs with
39
progenitor/biliary features have a faster and higher recurrence rate after surgical
treatment. Especially the independent prognostic value of Keratin19 in primary liver
carcinoma has been extensively documented. Its validity was proven in several
studies by diverse methodologies, in different centres and in different ethnic patient
groups. Since the current classification (hepatocellular carcinoma, cholangiocellular
carcinoma, mixed hepatocellular-cholangiocellular carcinoma, intermediate cell
tumors, cholangiolocellular carcinoma, collision tumors, …) is confusing and of little
prognostic value, an international study group proposes a new classification of
primary liver carcinomas based on the main cell type present in the tumour and the
expression and percentage of expression of CK19 in the liver cancer.
40
The hepatic stellate cell: A progenitor cell
Claus Kordes*, Iris Sawitza, Alexis Müller-Marbach, Niloofar Ale-Agha, Verena Keitel,
Hanne Klonowski-Stumpe, and Dieter Häussinger*
Clinic of Gastroenterology, Hepatology and Infectiology, Heinrich-Heine-University,
Moorenstr. 5, 40225 Düsseldorf, Germany
It is well known that isolated hepatic stellate cells (HSC) transform into myofibroblastlike cells during culture. The transformed HSC synthesize α-smooth muscle actin
(α-SMA) and extracellular matrix proteins such as collagen type 1. Therefore, it is
thought that HSC are involved in progression of liver fibrosis. Here we report, that
HSC are hitherto unrecognized progenitor cells of the liver. Freshly isolated HSC of
rat liver displayed an active β-catenin-dependent Wnt signaling pathway and
expressed the stem and progenitor cell marker CD133. This cell surface protein was
used in the present study to select HSC by magnetic cell sorting after density
gradient centrifugation. The selected CD133+ HSC synthesized molecular markers of
stem cells (c-kit, nanog, numb1/3, OCT4, PITX2c) and displayed the expression
pattern of endothelial progenitor cells (BMPER, CD31, CD105, FLK1, FLT1, RUNX1,
TIE2) as well as monocytes (CD14, CD49f). CD133+ HSC, which were cultured on
plastic and treated with platelet-derived growth factor-BB, transformed into α-SMA
positive myofibroblast-like cells. However, if the CD133+ HSC were cultured on
collagen and treated with an appropriate cytokine cocktail, they differentiated into
cells of hepatocyte or endothelial lineages. The cytokines fibroblast growth factor 4,
hepatocyte growth factor, basic fibroblast growth factor (bFGF), and interleukin-6
(IL-6) induced the hepatocyte markers albumin and α-fetoprotein in CD133+ HSC.
Endothelial cell markers such as endothelial nitric oxide synthase and vasoendothelial cadherin were induced after treatment of CD133+ HSC with vascular
endothelial growth factor 164, bFGF, erythropoietin, and IL-6. In addition, these
cytokines led to the formation of branched tube like structures. Cells that formed
these tubes synthesized numb similar to freshly isolated HSC. Numb synthesis was
also prominent in tubes and regenerating structures that appeared after treatment of
rats with carbon tetrachloride. The presence of stem and progenitor cell markers as
well as β-catenin-dependent Wnt signaling in HSC clearly indicate that stellate cells
are undifferentiated cells of the vertebrate liver. These findings in conjunction with the
capability of HSC to differentiate into hepatocyte- or endothelial-like cells and to form
tube-like structures points towards an important function of stellate cells in liver
regeneration.
*Corresponding authors:
E-mail: [email protected], Tel.: +49-211-81-14445, Fax: +49-211-81-10718
E-mail: [email protected], Tel.: +49-211-81-17569, Fax: +49-211
41
Role of non-parenchymal liver cells in portal hypertension
Vijay Shah
Mayo Clinic, GI Research Unit, Rochester, MN, USA
Liver sinusoidal endothelial cells (LEC) and hepatic stellate cells (HSC) contribute
importantly to the development of portal hypertension. This occurs through the role
they play in the process of increased intrahepatic resistance and pathological
remodeling of the sinusoids in cirrhosis. One of the major phenotypic changes in LEC
includes changes in the (nitric oxide) NO generation system that result in deficient
NO generation. Deficient NO generation results in a myriad of downstream
alterations in both LEC and HSC. These are most pronounced in HSC, which in the
absence of adequate regional NO levels, become more contractile, more migratory,
and more proliferative. This contributes to the “activation” phenotype of HSC and is a
major factor in the process of increased HSC coverage of the sinusoids and ensuing
increases in intrahepatic resistance. This lecture will cover the mechanisms that
contribute to these processes and their relevance to the pathophysiologic
phenotypes of portal hypertension.
References:
1. Nitric oxide and portal hypertension: interface of vasoreactivity and angiogenesis.
J Hepatol 2006; Jan; 44 (1): 209–216.
2. Lee JS, Semela D, Iredale J, Shah VH. Sinusoidal remodeling and angiogenesis:
a new function for the liver-specific pericyte? Hepatology 2007; Mar; 45 (3):
817–825.
42
Role of angiogenesis in portal hypertension
Mercedes Fernández, Ph.D.
Liver Unit, IDIBAPS, Hospital Clinic, University of Barcelona, Barcelona, Spain
Portal hypertension is a frequent syndrome in patients with chronic liver diseases.
Portal hypertension is characterized by a hyperdynamic splanchnic circulation and by
the formation of portosystemic collateral vessels and gastroesophageal varices
(which are responsible of most clinical consequences of portal hypertension). Despite
major advances in knowledge over the past two decades, the mechanisms
underlying the splanchnic hyperemia and the formation of collaterals are not fully
understood.
We investigated whether the development of hyperdynamic splanchnic circulation
and the formation of portosystemic collaterals are mediated through a VEGFdependent angiogenic process in rodent models of portal hypertension induced by
partial portal vein ligation. As anti-angiogenic strategies, we have used a monoclonal
antibody against VEGF receptor-2 (VEGFR-2) (1) and a selective and specific
inhibitor of VEGFR-2 autophosphorylation (2). These studies demonstrated the
presence of a significant VEGF-dependent splanchnic neovascularization in portal
hypertensive animals, and that the development of hyperdynamic splanchnic
circulation and the formation of portosystemic collateral vessels are angiogenesisdependent processes that can be markedly inhibited by blockade of the VEGF
signaling pathway (1, 2). These findings indicate that modulation of angiogenesis
may indeed represent a potential target in the treatment of portal hypertension.
We also studied the mechanisms modulating angiogenesis in portal hypertension.
Our results shown that oxidative stress (NADPH oxidase dependent) modulates
splanchnic angiogenesis in portal hypertension, directly by activating the VEGF
signaling pathway, and indirectly by stimulating the induction of heme oxygenase-1,
which increases VEGF expression (3, 4).
More recently, we have found that besides VEGF, portal hypertension development
is associated with a progressive overexpression of the proangiogenic growth factor
PDGF, which modulates de maturation of the newly formed vessels. Our results also
provide new insights into how angiogenesis regulates portal hypertension by
demonstrating that the maintenance of increased portal pressure, hyperkinetic
circulation, splanchnic neovascularization and portosystemic collateralization is
regulated by VEGF and PDGF in portal hypertensive rats. Importantly, these findings
also suggest that an extended anti-angiogenic strategy (ie, targeting both
VEGF/endothelium and PDGF/pericytes) may represent a novel approach in the
treatment of portal hypertension (5).
References:
1.
Fernandez M, Vizzutti F, Garcia-Pagan JC, Rodes J, Bosch J. Anti-VEGF
receptor-2 monoclonal antibody prevents portal-systemic collateral vessel
formation in portal hypertensive mice. Gastroenterology 2004; 126: 886–894.
43
2.
Fernandez M, Mejias M, Angermayr B, Garcia-Pagan JC, Rodes J, Bosch J.
Inhibition of VEGF receptor-2 decreases the development of hyperdynamic
splanchnic circulation and portal-systemic collateral vessels in portal
hypertensive rats. J Hepatol 2005; 43: 98–103.
3.
Angermayr B, Mejias M, Gracia-Sancho J, Garcia-Pagan JC, Bosch J,
Fernandez M. Heme oxygenase attenuates oxidative stress and inflammation,
and increases VEGF expression in portal hypertensive rats. J Hepatol 2006; 44:
1033–1039.
4.
Angermayr B, Fernandez M, Mejias M, Gracia-Sancho J, Garcia-Pagan JC,
Bosch J. NAD(P)H oxidase modulates angiogenesis and the development of
portosystemic collaterals and splanchnic hyperemia in portal hypertensive rats.
Gut 2007; 56: 560–564.
5.
Fernandez M, Mejias M, Garcia-Pras E, Mendez R, Garcia-Pagan JC, Bosch J.
Reversal of portal hypertension and hyperdynamic splanchnic circulation by
combined vascular endothelial growth factor and platelet-derived growth factor
blockade in rats. Hepatology 2007 (in press).
44
The relative roles of lymphocytes,
myofibroblasts in hepatic fibrosis
Kupffer
cells,
and
Ekihiro Seki, Samuele De Minicis, Yosuke Osawa, Tatiana Kisseleva,
Robert Schwabe and David A. Brenner
Department of Medicine, University of California, San Diego, School of Medicine, La
Jolla, CA, USA
Background: The pattern recognition receptor Toll-like receptor (TLR) 4 plays a
central role in innate immune responses. In hepatic fibrosis, portal level of the TLR4
ligand lipopolysaccharide (LPS) is elevated increased intestinal permeability. Both
Kupffer cells, the resident macrophages in the liver, and hepatic stellate cells (HSCs),
which play a crucial role in hepatic fibrosis, express TLR4. However, the precise
contribution of TLR4 in hepatic fibrosis is unknown.
Aim: (1) To define the role of TLR4 in hepatic fibrosis, (2) to identify the source of
TLR4 ligand during hepatic fibrosis (3), to determine the cellular target of TLR4 ligand
in the liver (4), to characterize molecular mechanisms by which TLR4 promotes HSC
activation and fibrogenesis.
Methods: Hepatic fibrosis was induced by bile duct ligation (BDL) of TLR4-intact
(C3H/HeOuJ), TLR4-mutant (C3H/HeJ), MyD88-/- and TRIF-/- mice. To deplete
intestinal micro flora, we treated the mice with an antibiotics cocktail (Ampicillin,
Metronidazole, Neomycin and Vancomycin) for 4 weeks prior to BDL. TLR4-chimeric
mice were generated by Cl2MDP injection, irradiation and bone marrow (BM)
transplantation of TLR4-mutant BM into TLR4-intact mice and vice versa. We
evaluated hepatic fibrosis by sirius red staining, measurement of hydroxyproline
content and profibrogenic gene expression (collagen α1(I), αSMA, TGFβ1, TIMP1) by
quantitative real time PCR (qPCR). Cellular targets of LPS in the liver were analyzed
by nuclear translocation of NF-κBp65 by immunofluorescence. To evaluate HSC
activation, we measured collagen promoter activity in HSCs isolated from collagenpromoter driven GFP transgenic mice and TGFβ transcriptional activity after
stimulation with TGFβ or co-culturing with Kupffer cells (KCs). LPS targets in HSCs
were analyzed by microarray and qPCR.
Results: Upon BDL, hepatic collagen and hydroxyproline content were less
increased in TLR4-mutant mice and antibiotics-treated mice compared with TLR4intact mice. Combination of Cl2MDP, irradiation and BMT completely reconstituted
KCs, but not HSCs, with BM-derived cells, generating TLR4-chimeric mice. Chimeric
mice containing TLR4-mutant BM/TLR4-intact HSC had normal fibrosis after BDL,
whereas chimeric mice with TLR4-intact BM/TLR4-mutant HSC had reduced fibrosis,
indicating HSCs are the main target of TLR4 ligand in hepatic fibrosis. After LPS
injection into KC-depleted mice, NF-κB was strongly activated in HSCs. LPS
treatment enhanced HSC activity by TGFβ- and KC-induced TGFβ transcriptional
and collagen promoter activity. Microarray analysis revealed 154 LPS-regulated
genes in quiescent HSCs including downregulation of the TGF pseudoreceptor
BAMBI. BAMBI was downregulated by LPS stimulation in WT-HSCs, but not in
MyD88-/- or IκB super-repressor-treated cells. TGFβ transcriptional and collagen
45
promoter activities were upregulated and downregulated in the HSCs overexpressing
of dominant negative BAMBI and full length BAMBI, respectively. Upon BDL,
profibrogenic gene expression was suppressed in MyD88-/- mice, but not in TRIF-/mice.
Conclusion: Hepatic fibrosis is induced by TLR4-dependent MyD88-dependent and
NF-κB-dependent manner. HSCs, but not KCs are the main targets of TLR4 ligands
in hepatic fibrosis. TLR4 enhances hepatic fibrosis by downregulating TGF
pseudoreceptor BAMBI and enhancing TGFβ signaling in HSCs. In the fibrotic liver,
myofibroblasts and fibrocytes are recruited from the bone marrow, but only constitute
a small component of the Type I collagen expressing cells.
46
Hepatic fibrogenesis and carcinogenesis: Krüppel-like factors
and beyond
Scott L. Friedman, M.D.
Fishberg Professor and Chief, Division of Liver Diseases, Mount Sinai School of
Medicine, New York, NY, USA
The understanding of hepatic fibrosis, or the liver’s scarring response has emerged
as a major focus of current research in hepatology. In particular, hepatic stellate cell
activation has emerged as a central event in fibrogenesis, and has contributed to a
detailed understanding of fibrosis pathogenesis. What is less clear, however, is why
the fibrotic milieu accelerates emergence of hepatocellular carcinoma
Hepatic stellate cells (HSC) are the primary source of extracellular matrix in normal
and fibrotic liver. They undergo an “activation” or transdifferentiation in progressive
stages yielding a cell type that is highly proliferative, fibrogenic and contractile.
Activation of HSCs is the central event of fibrogenesis. Exciting progress has been
made in understanding the molecular basis of this process. Major advances include:
(a) elucidation of effects of key cytokines on and their signaling pathways in HSC;
(b) understanding of transcriptional regulation of HSC activation; (c) characterization
of matrix proteases and their inhibitors; (d) demonstration of apoptosis as an
important event in resolution of hepatic fibrosis and identification of its mediators;
(e) elucidation of the complex and dynamic interaction between HSC and matrix;
(f) understanding of the role of other cellular elements in hepatic fibrosis and their
interaction with HSC. In addition, clinical studies have begun to identify host genetic
polymorphisms that may soon predict risk of fibrosis progression.
Our recent investigations of fibrosis have focused on the molecular regulation of
stellate cell activation, in particular, Krüppel like factor 6 (KLF6), which we previously
cloned by subtractive hybridization from activated stellate cells. For several years we
were puzzled by the paradoxical findings that KLF6 is growth suppressive and a
bona fide tumor suppressor gene, yet it was induced during a growth burst in this cell
type. The paradox has been resolved with the finding that KLF6 is alternatively
spliced, and studies to date indicate that early stellate cell activation is marked by
induction of splice forms of KLF6, which have dominant negative, growth-promoting
activities. Splicing is also enhanced by oxidant stress and occurs in vivo and in
culture. These findings point to a novel mechanism of transcriptional/posttranscriptional gene regulation in hepatic stellate cells, and add to the complexity
underlying regulation of this fascinating cell type’s behavior.
At the same time, we have explored the role of KLF6 in hepatic carcinogenesis, since
the molecule is ubiquitiously expressed, is upregulated during liver injury, and has
been established as a tumor suppressor gene in a number of cancers, including
HCC. Initial studies analyzing 41 HCCs of different etiologies confirmed a high
degree of KLF6 LOH but very few inactivating mutations. We next compared mRNA
expression of full length KLF6 and KLF6 splice form mRNAs between either
dysplasia or HCC and surrounding tissue in HCV-infected patients, as well as in a
cohort of HBV patients. Expression of KLF6 mRNA was decreased in 73% of HBVassociated HCCs compared to matched surrounding tissue (ST), with reductions of
47
~ 80% in one third of the patients. KLF6 mRNA expression was also reduced in
dysplastic nodules from patients with HCV compared to cirrhotic livers with an
additional, marked decrease in the very advanced, metastatic stage. An increased
ratio of KLF6SV1/ wt KLF6 was present in a subset of the HBV-related HCCs
compared to matched ST, but this was not associated with any specific histologic
subgroups in the HCV subset. Reconstituting KLF6 in HepG2 cells by retroviral
infection decreased proliferation and related markers including cyclin D1, and betacatenin, increased cellular differentiation based on induction of albumin, E-cadherin,
and decreased alpha fetoprotein. These data indicated that reduced KLF6
expression is common in both HBV- and HCV-related HCCs and occurs at critical
stages during cancer progression. Effects of KLF6 were attributable at least in part to
regulation of genes controlling hepatocyte growth and differentiation.
These findings underscore the dual role of a transcription factor (KLF6) in directing
the response to liver injury and contributing to hepatocarcinogenesis by divergent
regulation in different liver cell types. Moreover, the findings establish a paradigm
wherein alternative splicing may contribute to both the response to injury as well as
the progression of carcinoma. Other transcription factors may follow similar pathways
of dysregulation in cancer, in particular other Kruppel like factors, (e. g., KLF4).
It remains unclear whether these findings contribute to our understanding of how
fibrosis accelerates carcinogenesis in liver. Traditional explanations have included:
1) Enhanced survival signals released by fibrotic matrix that prevents death of
hepatocytes after DNA damage; 2) Reduced NK cell number and activity, leading to
diminished tumor surveillance as fibrosis progresses; 3) Decreased telomere length
during carcinogenesis.
Further advances in the understanding of the molecular biology of hepatic fibrosis are
critical not only to the development of effective, targeted antifibrotic therapies, but
also to our understanding of hepatocarcinogenesis. Effective antifibrotic therapies
may ultimately reduce the incidence of HCC but clinical trials are needed to establish
this possibility. Continued insights into signaling and gene regulation may ultimately
yield new therapies that are both antifibrotic and anti-neoplastic.
48
Session IV
Clinical management of progressive
liver fibrosis
49
Non-invasive assessment of liver fibrosis: Serum surrogate
markers
D. Thabut
Hôpital Pitié Salpêtrière, Hépato-Gastroentérologie, AP-HP Groupe, Paris, France
Recognition of the limitations of liver biopsy and cost pressures has led to an
increasing desire for non-invasive tests to assess the stage of liver fibrosis. Hence, in
the last five years, a large number of studies have been published, evaluating several
non-invasive ways to assess fibrosis. There are two categories of non-invasive tests:
serum markers and methodologies related to liver imaging technique (the most
innovative being the measurement of transient elastography).
Serum markers can be direct or indirect. Direct markers measure components of
extracellular matrix in serum (e. g. hyaluronic acid, collagen IV…) as well as
cytokines involved in the fibrogenetic process. Direct markers can be used singly or
may be combined. Further studies are needed to assess their performance in
diagnosing cirrhosis and changes in liver fibrosis. Moreover, they are expensive and
have similar accuracy to indirect non-invasive tests, which are less expensive.
Indirect markers comprise routinely available tests (Bonacini index, Forn’s index,
APRI, modified APRI and Fibroindex), sometimes combined with other components
(Fibrotest and PGAA index). The Fibrotest is the most validated non-invasive test for
liver fibrosis. Some scores combine direct and indirect markers, with an improved
accuracy as compared to markers used singly. Besides, serum markers can be
combined to liver imaging techniques, improving also their diagnostic performance.
The existence of several and different scores for non-invasive tests for liver fibrosis
shows that no single measure is sufficient. Moreover, a lot of serum markers are not
a surrogate for inflammation or steatosis, which have significant predictive value for
the progression of fibrosis. Finally, most markers are not able to identify the
intermediate stages of fibrosis, whereas the detection of advanced fibrosis or
cirrhosis is easier, with higher accuracy and less variability. Recent studies showed
disappointing results, with a rather poor correlation between non-invasive tests and
fibrosis stages. Some explanations can be raised to explain these discouraging
findings:
First, non-invasive tests have continuous scores, and have been correlated to
categorical variables (fibrosis stages), which are only descriptive, differ amongst the
various histological assessments, and do not have an arithmetical progression.
A solution to tackle this point would probably be to compare non-invasive tests to
quantitative measurement of liver fibrosis. Furthermore, the quality of the current gold
standard, i.e. liver biopsy, is far from perfect. Optimal evaluation of fibrosis can be
performed with liver samples of 25 mm length and/or 11 portal tracts or more, a goal
that is rarely achieved in most studies evaluating the performance of non-invasive
tests. Therefore, non-invasive tests could be wrongly considered unreliable because
the basis of their evaluation is flawed by inadequate biopsy specimens. In the future,
studies evaluating the performance of serum markers of fibrosis should include
patients with optimal liver biopsies, and comparing the collagen content of liver
samples, in order to provide the best reference standard.
51
To date, the comparison of different serum markers failed to identify which markers
are the best. However, before raising firm conclusions, one must check that the
comparison is possible. First, when evaluating non-invasive tests, one must wonder
which particular question is being asked: confirmation of cirrhosis, exclusion of
cirrhosis, confirmation of normal liver, confirmation or exclusion of portal
hypertension, or accurate staging of fibrosis. The clear definition of the endpoints is
crucial, as results have been extrapolated wrongly from one clinical setting to
another. Secondly, the diagnostic value of each test is generally determined by using
the discriminatory ability (AUROC) of the test, which depends on the relative
prevalence of fibrosis stages in the population studied. Hence, comparison of
different tests by comparing the AUROC should be allowed only if the prevalence of
each stage of fibrosis is the same in the population studied.
In conclusion, non-invasive tests for the detection of liver fibrosis have the potential
to become an important tool in clinical practice, providing that better validation is
performed, using optimal liver biopsies. Comparison between the different tests
should be performed based on solid statistical grounds. Combination of several noninvasive tests could increase their accuracy. We are in a period of transition that will
lead to non-invasive diagnostic procedures of better quality. Hopefully, the need for
liver biopsies will then be limited to the subset of patients in whom precise noninvasive diagnosis is not possible.
52
Non-invasive assessment of liver fibrosis: Fibroscan
M. Beaugrand
Liver Unit, Hopital Jean Verdier, Université Paris XIII, Bondy, France
Morbidity and mortality in patients with chronic liver diseases are linked to the
progression of fibrosis. Evaluation of fibrosis is therefore essential in the
management of these patients. It relied until recently on liver biopsy.
Limitations to liver biopsy have been widely emphasized. Additionally to its high cost,
morbidity and poor acceptance the assessment of liver fibrosis by this technique is
prone to errors due to the sampling effect and only large biopsies (> 25 mm) quoted
by an experienced pathologist could be considered as reliable for fibrosis staging.
Therefore the need for non invasive tools has been advocated. Serum markers and
liver stiffness measurement (LSM) by ultrasonography or more recently MRI are
alternative options that have been extensively studied these last years.
Fibroscan is a new device able to measure liver elasticity (stiffness) using the
principle of transcient elastography (1). Briefly this system is equipped with a probe
consisting in an ultrasonic transducer mounted on the axis of a vibrator. A low
frequency vibration is transmitted from the vibrator to the liver where it induces an
elastic shear wave followed by pulse-echo ultrasonic acquisitions. Its velocity is
directly related to liver stiffness and results are expressed in KPa. The final value
called liver stiffness measurement (LSM) is the median value of 10 successive
measurements (1). LSM has a fair reproducibility.
Results of Fibroscan for the assessment of liver fibrosis and the diagnosis of cirrhosis
Fibroscan has now been extensively studied in different types of liver diseases and
different parts of the world with homogenous results. The first large multicentric study
(2) concerned patients with HCV chronic hepatitis and compared LSM with fibrosis
grade according to the Metavir classification. The area under the ROC curves were
0.79 for F ≥ 2, 0.91 for F ≥ 3, and 0.97 for F = 4. The results in patients with the
larger biopsies were even more favourable. A comparison with Fibrotest (3) showed
a better diagnostic accuracy of Fibroscan for cirrhosis or extensive fibrosis. The
diagnostic accuracy of LSM by Fibroscan for the diagnosis of cirrhosis has been
confirmed by a larger study in 1257 patients with various causes of liver diseases (4).
Complementary studies with Fibroscan
LSM in patients with cirrhosis is correlated with portal pressure and is able to predict
the presence or not of oesophageal varices (5). Furthermore it seems to be
correlated with the area of fibrosis and could help in the future to monitor fibrosis
particularly during anti-viral or anti-fibrotic treatments.
53
Limitations of Fibroscan
LSM by Fibroscan is not usable in patients with ascites and it often fails in patients
with a thick chest wall such as obese people. The mean failure rate in patients
without ascites is approximately 7% highly correlated with BMI (2).
Fibroscan versus blood tests
Blood tests are usually inferior to LSM in the prediction of cirrhosis or extensive
fibrosis but most of all they suffer several limitations due (1) to their heterogeneity
and their absence of universality as the candidate composite indexes are regularly
increasing in number (2). Their insufficient validation (for many of them) in often
limited subgroups of patients such as those with chronic hepatitis C (3). The fact that
the variables used could be influenced by extra hepatic diseases. Nevertheless well
validated blood tests such has Fibrotest could be used complementarily to Fibroscan.
Fibroscan versus MRI transcient elastography
Limited data are available concerning the use of MRI to measure liver stiffness.
Although the method seems promising it will obviously be more costy and less
practical.
References:
1) Sandrin L, Fourquet B, Hasquenoph JM, et al. Transient elastography: a new
noninvasive method for assessment of hepatic fibrosis. Ultrasound Med Biol
2003, 29: 1705–1713.
2) Ziol M, Handra-Luca A, Kettaneh A et al. Noninvasive assessment of liver fibrosis
by measurement of stiffness in patients with chronic hepatitis C. Hepatology
2005, 41: 48–54.
3) Castera L, Vergniol J, Foucher J. Prospective comparison of transient
elastography, Fibrotest APRI and liver biopsy for the assessment of fibrosis in
chronic hepatitis C. Gastroenterology 2005, 128: 343–350.
4) Ganne-Carrie N, Ziol M, de Ledinghen V et al. Accuracy of liver stiffness
measurement for the diagnosis of cirrhosis in patients with chronic liver diseases.
Hepatology 2006, 44: 1511–1517.
5) Kazemi F, Kettaneh A, N'Kontchou et al. Liver stiffness measurement selects
patients with cirrhosis at risk of bearing large oesophageal varices. J Hepatol
2006, 45: 230–235.
54
The clinical evaluation of disease progression in CLD: Towards
an integrated system?
Massimo Pinzani, M.D., Ph.D.
Dipartimento di Medicina Interna – Center for Research, High Education and
Transfer DENOThe Università degli Studi di Firenze, Florence, Italy
Progressive hepatic fibrosis is the common consequence of most chronic liver
diseases. Significant achievements have been made in the understanding of the
cellular and molecular mechanisms of hepatic fibrogenesis and major “translations”
into clinical applications are currently awaited. Among other central issues, the need
for rapid, non-invasive, and reliable methods for the evaluation of the fibrogenic
progression of chronic liver diseases (CLD) has become an urgent clinical reality. In
recent years, several approaches such as serum markers, transient elastography
and a re-visitation of classical imaging techniques have been proposed as a
replacement and/or as an integration of liver biopsy. The large majority of the studies
is focused on surrogate measures of disease progression in chronic HCV infection.
At the present stage of development, all the proposed non-invasive methodologies
have been shown to be effective in predicting either very early or very advanced
fibrosis rather than being able to precisely assess the progression of fibrosis in a
step-wise fashion. However, since most of the available evidence relies on a
retrospective analysis, it is likely that more precise indications will derive from
ongoing longitudinal studies. Most of the proposed non invasive methods could be
instrumental in the staging of advanced fibrosis from F3/F4 and beyond, i.e. when the
patient is in a clinical stage defined as “compensated cirrhosis” and, subsequently,
when clinically significant portal hypertension and relative complications develop.
This transition, that may need several years or even decades, undeniably represents
a “grey area” where no effective diagnostic tools are available as an alternative or as
an integration to the measurement of the hepatic venous pressure gradient (HVPG).
Monitoring of disease progression in patients with patients with NASH and fibrosis
represents a key clinical challenge with an important impact on both public health
and health economics. Work in this area is still scarce and preliminary. Some of the
proposed methods are directed at predicting fibrosis in patients with fatty liver and
are designed by integrating parameters related to the clinical features of the
metabolic syndrome with those generically associated with advances stages of
fibrosis. Overall, it appears that these tests will allow to identify or exclude patients
with severe fibrosis, although a large proportion of the population is likely to fall in an
undetermined area.
Major efforts are currently directed at validating the proposed non-invasive methods
in larger and longitudinal studies. In addition, special attention should be
concentrated on the possible integration of different non invasive systems with the
actual gold-standards in order to gain a real diagnostic advantage.
55
Fibrosis regression and innovative antifibrotic therapies: From
bench to bedside?
Priv.-Doz. Dr. med. Hermann E. Wasmuth, M.D.
Medical Department III, University Hospital Aachen, 52057 Aachen, Germany
Liver fibrosis is the hallmark of all chronic liver diseases and is associated with
increased morbidity and mortality. Scarring of the liver due to continuous
hepatocellular damage is considered as potentially reversible since the late 1970’s.
Today, there is ample laboratory and clinical evidence showing the capacity of the
injured liver to resorb excess extra cellular matrix with regression of established
fibrosis.
In most animal models, cessation of the insult (e. g. the administration of CCl4) leads
to a delayed but significant reduction in fibrotic liver tissue. This resolution is
associated with decreased activity of tissue inhibitors of matrix metalloproteinases
(TIMPs) and increased expression of matrix metalloproteinases (MMPs). Therefore,
delivery of MMPs has already been evaluated as an approach to ameliorate
experimental fibrosis in animal models.
Hepatic stellate cells are considered as an important cells type in this context. While
these cells produce matrix proteins during the process of fibrogenesis, the undergo
apoptosis during fibrosis regression. Stellate cell apoptosis is driven by cytokines,
including nerve growth factor, while sustained TIMP-1 expression blocks apoptosis of
hepatic stellate cells. Notably, several other cell types can also degrade matrix in the
liver, including neutrophils and macrophages. This underscores the importance of the
immune system in fibrosis regression as well as fibrosis progression.
Although these in vivo and in vitro results have provided important insight in the
molecular mechanism driving fibrosis regression, innovative therapies based on
these data are still lacking. In human trials only a few defined compounds have been
evaluated. These include silymarin and the cytokines interferon-alpha/gamma and
interleukin-10. While trials with silymarin were negative, the treatment of patients
infected with the hepatits C with interferons or IL-10 have shown beneficial effects in
short termed studies. However, such non-specific interventions are less effective
than (rather) specific therapies for the underlying liver disease. With specific
therapies, evidence of fibrotic and cirrhotic regression has been documented in the
entire spectrum of chronic liver diseases, including autoimmune hepatitis, hepatitis B
and C, biliary obstruction, hemochromatosis and NASH. Nevertheless, whether the
improvement in fibrosis will also ameliorate portal hypertension or the incidence of
hepatocellular carcinoma remains to be proven.
56
Session V
Preprimary and primary prophylaxis
57
Dynamic increase of intrahepatic vascular resistance in
cirrhosis
Juan Carlos García-Pagán
Consultant in Hepatology, Hepatic Hemodynamic Laboratory, Liver Unit, ICMD,
IDIBAPS and Ciberehd Hospital Clinic, Barcelona, Spain
Increased resistance to portal blood flow is the primary factor in the pathophysiology
of portal hypertension. In cirrhosis, increased intrahepatic vascular resistance is the
consequence of the distortion of the liver vascular architecture caused by fibrosis,
scarring and nodule formation. In addition, the active contraction, in response to
several agonists, of different contractile cell types in the liver promotes a further
increase in the intrahepatic resistance a component that may represent up to 40% of
the increased intrahepatic vascular resistance in cirrhosis. An increased activity of
several endogenous vasoconstrictors, such as endothelin, norepinephrine, angiotensin II, vasopressin and more recently leukotrienes and thromboxane A2, together
with an augmented vasoconstrictive response of the hepatic vascular bed to some of
them have been implicated in the pathogenesis of the dynamic component of the
raised intrahepatic resistance of the cirrhotic liver.
Besides the increased production and hyperresponse to vasoconstrictors the
intrahepatic vascular bed of cirrhotic livers present with an insufficient release of
vasodilators together with an insufficient response to them, which further contributes
to increase intrahepatic vascular tone. A reduced bioavailability of NO within cirrhotic
livers has been shown to play a major role in these disturbances. This occurs despite
a normal expression of endothelial NO synthase (eNOS) mRNA and normal levels of
eNOS protein and is attributed both to a posttranslational regulation of the enzyme
and to increased NO scavenging by superoxide. The potential role of other
endogenous vasodilators such as carbon monoxide and H2S has been less
extensively evaluated.
In normal conditions, the endothelium of the hepatic vascular bed is able to generate
vasodilator stimuli in response to increases in blood volume, blood pressure or
vasoconstrictor agents in an attempt to attenuate the consequent increase in portal
pressure. In several pathological conditions there is impairment in this endotheliumdependent vasodilatation that has been named endothelial dysfunction. The hepatic
vascular bed of cirrhotic livers exhibits endothelial dysfunction that has been shown
to be, at least in part, the consequence of the imbalance between vasoconstrictor
and vasodilator forces within the cirrhotic liver.
59
Remodeling portal hypertension: Preprimary prophylaxis
Guadalupe Garcia-Tsao, M.D.
Professor of Medicine, Yale University, VA-CT Healthcare System, New Haven, CT,
USA
Portal hypertension is the main complication of cirrhosis, regardless of its etiology.
Gastroesophageal varices are a direct consequence of portal hypertension and
develop after a minimal portal pressure gradient of 10–12 mmHg is reached, the socalled “clinically significant” portal hypertension [1].
Variceal hemorrhage is the main clinical consequence of gastroesophageal varices
and one responsible for the high morbidity and mortality in cirrhosis. Randomized
controlled trials have demonstrated that non-selective β-adrenergic blockers are
effective in preventing first variceal hemorrhage and, together with nitrates, they are
effective in preventing recurrent variceal hemorrhage [2].
Nonselective β-adrenergic blockers (propranolol, nadolol) reduce portal pressure
through a reduction in portal venous inflow. This in turn is achieved in two ways: by
normalizing the increased cardiac output (β-1 adrenergic blockade) and by
normalizing the increased splanchnic blood flow (β-2 adrenergic blockade). This last
effect is the result of an unopposed α-adrenergic-mediated constriction of the
splanchnic vasculature.
Pharmacological reduction in portal pressure (as determined by the hepatic venous
pressure gradient or HVPG) in patients with cirrhosis has been shown to significantly
decrease first and recurrent variceal hemorrhage. Patients in whom the HVPG
decreases below 12 mmHg are essentially protected from bleeding and those in
whom the HVPG decreases by more than 20% from baseline have a significantly
lower rate of recurrent hemorrhage [3; 4]. Importantly this reduction in HVPG has
also been associated with a reduction in the incidence of other complications of
portal hypertension such as ascites, spontaneous bacterial peritonitis and
encephalopathy and with a decreased mortality [3–6].
The above data is derived from studies of primary or secondary prophylaxis of
variceal hemorrhage, that is, patients who have varices (usually large-sized) but have
never bled (primary prophylaxis) and patients who have varices and have bled
(secondary prophylaxis). In a murine model of chronic portal hypertension, the
reduction in portal pressure and portal venous inflow induced by beta-adrenergic
blockers was to prevent the development of portosystemic collaterals [7].
By decreasing portal pressure at earlier stages, it may be possible to prevent or
ameliorate not only the development of gastroesophageal varices but also the
detrimental hepatic and systemic metabolic effects due to portal systemic shunting
and the development of ascites, thereby improving the survival of patients with
cirrhosis and portal hypertension. Therefore, a large multicenter randomized placebocontrolled trial was performed with the objectives of evaluating the efficacy of a nonselective beta-adrenergic blocker, timolol, in preventing the development of
gastroesophageal varices and to assess the usefulness of baseline and sequential
60
measurements of HVPG in predicting the development of varices and other
complications of cirrhosis [8].
In this trial 213 patients with cirrhosis without varices but with portal hypertension
(HVPG of at least 6 mmHg) were randomly assigned, 108 to the timolol group and
105 to the placebo group. Every year, an upper endoscopy, abdominal ultrasound
and HVPG measurements were performed. The primary end points were
development of varices or variceal hemorrhage. In a median follow-up time of
54.9 months, 84 (39%) patients developed a primary endpoint, 42 in the timolol group
(35 small varices, 4 large varices, 2 variceal hemorrhage, 1 bleeding from gastropathy) and 42 in the placebo group (35 small varices, 4 large varices,
3 variceal hemorrhage) (NS). No differences in development of ascites, encephalopathy, liver transplants or death were found between groups. More patients in the
timolol
group
developed
serious
adverse
events
(18%
vs.
6%,
p = 0.006).
On Cox multivariate regression analysis two parameters were independently
predictive of a primary endpoint: HVPG > 10 mmHg (p = 0.005) and AST (p = 0.007).
Furthermore, primary endpoints developed at a significantly lower rate in patients
who had a reduction in HVPG >10% at 1 year (19/69 or 28%) compared to those in
whom the reduction was ≤ 10% (44/85 or 52%) (p-0.003). Notably, 53% patients
randomized to timolol, had a reduction in HVPG >10% while this reduction occurred
in only 38% patients in the placebo group (p = 0.04).
A follow-up study in this patient cohort aimed at identifying predictors of clinical
decompensation (defined as the development of ascites, variceal hemorrhage or
hepatic encephalopathy) showed that, in a median follow-up time of 51.1 months,
62/213 patients (29%) developed decompensation: 46 (21.6%) ascites, 17 (8%)
encephalopathy and 6 (3%) variceal hemorrhage. On multivariable analysis,
3 predictors of decompensation were identified: HVPG, MELD score and albumin.
Diagnostic capacity of HVPG was greater than for MELD or Child-Pugh score. An
HVPG of 10 mmHg distinguished two populations with a different rate of incidence of
decompensation with a negative predictive value of 90% [9].
In conclusion, nonselective β-adrenergic blockers are not effective in preventing the
development of varices and are associated with a significant rate of adverse events.
This therapy is therefore not recommended in compensated cirrhotic patients without
varices at large. However, even in this very compensated patient population (stage 1
per reference [10], we have confirmed the predictive value (both for the development
of varices and for the development of clinical decompensation) of baseline HVPG
levels and of reductions in HVPG > 10%, a therapeutic goal that could be achieved
through the use of β-blockers or other drugs being developed for the treatment of
portal hypertension.
61
References and recommended reading:
1. D'Amico G, Garcia-Tsao G, Cales P, Escorsell A, Nevens F, Cestari R, Caletti G,
and Zoli M. Diagnosis of portal hypertension: how and when. In: DeFranchis R.
Portal Hypertension III. Proceedings of the Third Baveno International
Consensus Workshop on Definitions, Methodology and Therapeutic Strategies.
Oxford: Blackwell Science, 2001: 36–64.
2. D'Amico G, Pagliaro L, Bosch J. Pharmacological treatment of portal
hypertension: an evidence-based approach. Semin Liver Dis 1999, 19: 475–505.
3. D’Amico G, Garcia-Pagan JC, Luca A, Bosch J. HVPG reduction and prevention
of variceal bleeding in cirrhosis. A systematic review. Gastroenterology 2006,
131: 1624.
4. Bosch J and Garcia-Pagan JC. Prevention of variceal rebleeding. Lancet 2003,
361: 952–954.
5. Turnes J, Garcia-Pagan JC, Abraldes JG, Hernandez-Guerra M, Dell'era A,
Bosch J. Pharmacological reduction of portal pressure and long-term risk of first
variceal bleeding in patients with cirrhosis. Am J Gastroenterol 2006, 101:
506–512.
6. Abraldes JG, Tarantino I, Turnes J, Garcia-Pagan JC, Rodes J, Bosch J.
Hemodynamic response to pharmacological treatment of portal hypertension and
long-term prognosis of cirrhosis. Hepatology 2003, 37: 902–908.
7. Sarin SK, Groszmann RJ, Mosca PG, Rojkind M, Stadecker MJ, Reuben A,
Bhatnagar R, Dayal D. Propranolol ameliorates the development of portalsystemic shunting in a chronic murine schistosomiasis model of portal
hypertension. J Clin Invest 1991, 87: 1032–1036.
8. Groszmann RJ, Garcia-Tsao G, Bosch J, Grace ND, Burroughs AK, Planas R,
Escorsell A, Garcia-Pagan JC, Patch D, Matloff DS, Gao H, Makuch RW, for the
Portal Hypertension Collaborative Group. Beta-blockers to prevent
gastroesophageal varices in patients with cirrhosis. N Engl J Med 2005, 353:
2254–2261.
9. Ripoll C, Groszmann R, Garcia-Tsao G, Grace N, Burroughs AK, Planas R,
Escorsell A, Garcia-Pagan JC, Makuch RW, Patch D, Matloff DS, Bosch J, and
the Portal Hypertension Collaborative Group. Hepatic venous pressure gradient
(HVPG) predicts clinical decompensation in patients with compensated cirrhosis.
Hepatology 2006, 44 Suppl 1: 203A.
10. D'Amico G, Garcia-Tsao G, Pagliaro L. Natural history and prognostic indicators
of survival in cirrhosis. A systematic review of 118 studies. J Hepatol 2006, 44:
217–231.
62
When and how to scope in portal hypertension
Prof. Roberto de Franchis
University of Milan, Italy
Gastroesophageal varices eventually develop in all cirrhotic patients, and once they
have developed, they tend to increase in size and to bleed. Mortality for bleeding
varices is about 20% within 6 weeks from the bleeding episode. Therefore, identifying
patients at risk of bleeding, in order to start prophylactic treatment is crucial. Noninvasive means for identifying patients with varices have given disappointing results;
as a consequence, the current recommended policy is to screen all cirrhotic patients
by upper GI endoscopy at the time of the diagnosis of cirrhosis. By doing so, three
different patient populations will be identified: a majority of patients without varices, a
smaller proportion with small varices and a minority with medium-sized to large
varices. The latter patients will undergo prophylactic treatment to prevent bleeding
(usually with beta-blockers or endoscopic band ligation), while those with no or small
varices will enter endoscopic surveillance programs.
Knowledge of the rate of development and growth of esophageal varices would have
important implications, because it would help optimising the intervals for follow-up
endoscopy. Concerning the incidence of new varices, the published figures range
between 5 and 10% per year. As far as deciding the timing of follow-up endoscopy is
concerned, we must first decide which level of risk (i.e. what proportion of patients
bleeding before starting prophylactic treatment) we are willing to accept. If we set this
level at 10%, then patients with no varices at baseline can be re-endoscoped at
3-years intervals. In fact, the rate of development of large varices and the rate of
bleeding in these patients are both below the 10% threshold. For patients with small
varices at baseline, the picture is much more complicated, since the reported
incidences range between 6% and 70% at two years for the development of large
varices, and between 4% at 16 months and 24% at two years for bleeding. Clearly,
more information is needed on this point. For the time being, it appears reasonable to
re-endoscope these patients at intervals of 1–2 years, adopting the shorter interval
for patients with alcoholic cirrhosis, with more severe impairment of liver function and
with endoscopic risk signs. Screening programs are somewhat hampered by patients'
aversion to undergo conventional endoscopy, which is perceived as unpleasant.
Recent data suggest that the new esophageal videocapsule endoscope might be a
valid alternative to endoscopy, primarily in patients unable or unwilling to undergo
conventional upper GI endoscopy.
63
Prevention of first variceal bleeding
Paul Calès
Department of Hepato-Gastroenterology, University Hospital and HIFIH Laboratory,
UPRES EA 3859, IFR 132, University, Angers, France
Prophylaxis of variceal bleeding consists of the prevention of varices development
(pre-primary prophylaxis), the prevention of a first variceal bleeding episode (primary
prophylaxis), or the prevention of rebleeding (secondary prophylaxis).
Endoscopic screening for the presence of esophageal varices should be done in all
patients at the diagnosis of cirrhosis. Endoscopic follow-up should then relate to the
initial variceal size. Endoscopy should be repeated every 3 years in patients without
varices and every 2 years in those with small varices. In case of large varices,
primary prophylaxis by nonselective β-adrenergic blockers (propranolol, nadolol)
should be begun and endoscopic follow-up is not necessary. Band ligation is also
useful in preventing variceal bleeding in patients with medium or large esophageal
varices, however the long-term benefit of band ligation requires further research and
is not currently proposed in primary prophylaxis unless there are contra-indications to
or side effects of nonselective β-adrenergic blockers.
Nonselective β-adrenergic blockers are effective in reducing the risk of first bleeding
from esophageal varices in patients with medium or large varices. The chronic
administration consists of a singly daily oral intake of long acting propranolol at a
dose of 80 or 160 mg, depending on the available dosage in each country. Another
empirical approach consists of b.i.d. administration and a titration of the oral dose
according to patient tolerance and treatment objectives. In all cases, doses should be
adjusted to obtain a 20–25% reduction in heart rate or a heart rate of less than
55 beats/min. After acute or chronic administration, the hemodynamic effects of
propranolol last a few days in cirrhotic patients; then, there is a risk of withdrawal
syndrome with variceal bleeding due to adrenergic rebound phenomenon during the
first years of treatment. Thus, patients (and physicians) have to be advised for a
stringent adherence to treatment (therapeutic education). β-blockade therapy is
maintained indefinitely as late withdrawal can be deleterious on survival despite the
lack of an increased bleeding risk. In patients with intolerance or contraindications to
β-adrenergic blockers, band ligation is recommended. Nitrates (isosorbide
mononitrate) are ineffective in preventing variceal bleeding if used alone and their
use in primary prophylaxis is not recommended. As current prevention reduces the
incidence of first bleeding of about 50%, other approaches have to be encouraged:
pre-primary prevention, new therapies with either hemodynamic objective or fibrosis
target, treatment of cause of chronic liver disease.
64
Session VI
Complications of cirrhosis
65
Management of acute variceal bleeding
Prof. Dr. A.K. Burroughs
Royal Free Hospital School of Medicine, Department of Medicine, London, UK
Therapy not specifically related to bleeding, in both cirrhotic and non-cirrhotic
patients with variceal haemorrhage, remains a cardinal point in management.
Effective resuscitation, attention and protection of the airway, particularly with severe
bleeding and disturbed conscious level, especially during endoscopy are initial
priorities. A normal central venous pressure should be maintained taking into account
the severity of ascites if present, in order to reduce the likelihood of renal failure, and
diminish the risk of infection.
Over transfusion in animal models provokes continued bleeding due to a portal
pressure increase but in humans the balance should be towards establishing an
effective circulating volume.
The most important therapeutic manoeuvre in terms of increasing survival is the use
of prophylactic antibiotics1, oral quinolones (and recently cephalosporins which have
been shown to be superior in one study2). The relative risk of mortality is reduced to
0.39 (95% CI 0.32 to 0.48), which is a greater reduction than that seen with specific
vasoactive drugs. Prophylactic antibiotics also reduce the incidence of early
rebleeding, supporting the hypothesis of infection as a trigger for bleeding3.
Data on vasoactive drugs which reduce portal pressure favour the use of terlipressin,
as mortality is reduced4;5, and it may have an added role in maintaining renal
function. Starting vasoactive therapy before diagnostic endoscopy is supported by
trials and prolonged therapy up to 5 days has been used, as this is the period of
greatest risk of early rebleeding6–8.
Therapeutic endoscopic therapy remains a mainstay of therapy. In randomized
studies, sclerotherapy and banding ligation result in similar survival, with only a small
advantage of ligation in terms of control of bleeding – relative risk reduction of 3.3%8.
Choice of therapy depends on operator experience, and control of the airway with
respect to severity of bleeding and the need to perform a double intubation
(diagnostic and then therapeutic endoscopy with the banding apparatus attached). In
the best conducted trial there was no difference in the control of bleeding in grade C
cirrhotics and those with active variceal bleeding9.
There is no accepted definition regarding failure of drug and endoscopic therapy6, but
in practice most units will only use two sessions of emergency therapeutic
endoscopy8. Failure is most likely in cirrhotics with severe liver dysfunction6, in whom
it has been shown that portal pressure is higher10. The latter particularly if hepatic
venous pressure gradient ≥ 20 mmHg is associated with increased mortality, and
more early rebleeding10.
Tissue adhesives, bucrylate and fibrin glue have been used as rescue therapy but
rarely as primary therapy in patients most at risk of failing sclerotherapy or ligation.
There is room for randomized studies versus tissue adhesives in this sub-group.
Alternative rescue therapy, if available, is the transjugular intrahepatic stent shunt
67
(TIPS), which is very effective in stopping bleeding, and often avoids the need to
keep a patient intubated11. It is also most useful for bleeding fundal varices
(subcardial varices usually respond to treating oesophageal ones)11. These also
respond well to injection with tissue adhesives, and poorly to sclerotherapy or
banding. Randomized studies of TIPS versus adhesives would be ethical and should
be carried out- particularly as no randomized studies exist of TIPS as rescue therapy.
Recombinant human thrombin has been used in small studies. Recombinant factor
VIIα has not been shown to have significant therapeutic benefit12 but a further
randomized study in decompensated cirrhotics is being evaluated.
Mortality from variceal bleeding has fallen in recent decades13 most likely due to
better specific therapies and better care of the acutely ill cirrhotic, particularly the
prevention of infection1.
Reference List:
1. Soares-Weiser K, Brezis M, Tur-Kaspa R et al. Antibiotic prophylaxis of bacterial
infections in cirrhotic inpatients: a meta-analysis of randomized controlled trials.
Scand. J. Gastroenterol 2003; 38: 193–200.
2. Fernandez J, Ruiz del AL, Gomez C et al. Norfloxacin vs ceftriaxone in the
prophylaxis of infections in patients with advanced cirrhosis and hemorrhage.
Gastroenterology 2006; 131: 1049–1056.
3. Goulis J, Armonis A, Patch D et al. Bacterial infection is independently
associated with failure to control bleeding in cirrhotic patients with gastrointestinal
hemorrhage. Hepatology 1998; 27: 1207–1212.
4. Gotzsche PC. Somatostatin analogues for acute bleeding oesophageal varices.
Cochrane Database Syst Rev 2002: CD000193.
5. Ioannou G, Doust J, Rockey DC. Terlipressin for acute esophageal variceal
hemorrhage. Cochrane Database Syst Rev 2003: CD002147.
6. Ben-Ari Z, Cardin F, McCormick AP, Wannamethee G, Burroughs AK.
A predictive model for failure to control bleeding during acute variceal
haemorrhage. J Hepatol 1999; 31: 443–450.
7. D'Amico G, Pietrosi G, Tarantino I, Pagliaro L. Emergency sclerotherapy versus
vasoactive drugs for variceal bleeding in cirrhosis: a Cochrane meta-analysis.
8. Triantos CK, Goulis J, Patch D et al. An evaluation of emergency sclerotherapy
of varices in randomized trials: looking the needle in the eye. Endoscopy 2006;
38: 797–807.
68
9. Villanueva C, Piqueras M, Aracil C et al. A randomized controlled trial comparing
ligation and sclerotherapy as emergency endoscopic treatment added to
somatostatin in acute variceal bleeding. J Hepatol 2006; 45: 560–567.
10. Moitinho E, Escorsell A, Bandi JC et al. Prognostic value of early measurements
of portal pressure in acute variceal bleeding. Gastroenterology 1999; 117:
626–631.
11. Chau TN, Patch D, Chan YW et al. "Salvage" transjugular intrahepatic
portosystemic shunts: gastric fundal compared with esophageal variceal
bleeding. Gastroenterology 1998; 114: 981–987.
12. Bosch J, Thabut D, Bendtsen F et al. Recombinant factor VIIa for upper
gastrointestinal bleeding in patients with cirrhosis: a randomized, double-blind
trial. Gastroenterology 2004; 127: 1123–1130.
13. McCormick PA, O'Keefe C. Improving prognosis following a first variceal
haemorrhage over four decades. Gut 2001; 49: 682–685.
69
Prevention of variceal rebleeding
Didier Lebrec
INSERM and Service d'Hepatologie, Hopital Beaujon, Clichy, France
In patients with cirrhosis and portal hypertension, the rebleeding rate is well known
with the analysis of control groups in clinical trials. At 2 years, the rebleeding rate is
approximately 70% and the mortality 30%. In this summary, the Baveno consensus
statements are partly reported.
The secondary prophylaxis should start as soon as possible after the episode of
bleeding when the hemodynamic condition of the patient is control.
In patients with cirrhosis who have not received primary prophylaxis, non selective
beta-adrenergic antagonist (propranolol or nadolol), band ligation or both should be
used to prevent recurrent bleeding. The combination of beta-blockers and band
ligation is probably the best treatment particularly in Child-Pugh C patients. More
trials are, however, needed. The assessment of hemodynamic response to
vasoactive drugs by the hepatic venous pressure gradient measurement may provide
information on the risk of rebleeding but more investigations are needed.
In patients with cirrhosis who received beta-blockers for primary prevention and
bleed, band ligation should be performed or added to beta-blockers. Beta-blockers
should be continued in case of discontinuation and when the dose of beta-blockers
was too low.
In patients who have contraindications or intolerance to beta-blockers, band ligation
is the best treatment for prevention of rebleeding. N-butyl-cyanoacrylate may be used
in particular in patients who bleed from gastric varices
In patients who fail endoscopic and pharmacological treatment for the prevention of
rebleeding, TIPS or surgical shunts are effective for Child-Pugh A–B patients and
should be used. In non-surgical candidates, TIPS is the only option. In Child-Pugh
B–C patients, liver transplantation provides good long-term outcomes and should be
considered.
70
From sodium retention to therapy for refractory ascites: The
role of new drugs
Florence Wong
University of Toronto, Toronto, Canada
Ascites is a common complication of liver cirrhosis, affecting 10% of patients with
cirrhosis at any one time. The appearance of ascites heralds the onset of
decompensation and adversely affects survival, with a 50% two-year survival,
worsening significantly to 20–50% at 1 year when the ascites becomes refractory to
medical therapy. Since sodium retention is the hallmark of liver cirrhosis, and it plays
a pivotal role in the pathogenesis of ascites formation in cirrhosis, traditional
treatment of ascites has evolved around dietary sodium restriction, and diuretic
therapy which are natriuretic. However, as the cirrhotic process progresses, patients
will gradually become resistant to diuretics, with a gradually decreasing natriuretic
response, until they eventually become refractory to diuretic therapy. Alternative
treatments such as repeat large volume paracentesis or the insertion of transjugular
intrahepatic portosystemic stent shunt (TIPS) are associated with increasing costs
and their unique complications, therefore, other treatment options for the
management of ascites are now being developed.
Albumin is an “old” drug that has recently been proposed as an adjunct therapy in the
management of ascites in cirrhosis. In one randomized controlled trial assessing the
effects of albumin in addition to standard diuretic therapy in cirrhotic patients with
ascites, weekly infusions of 25 grams of albumin was shown to produce significantly
better diuretic response, shorter hospital stays, lower probability of re-accumulation
of ascites, and lower likelihood of readmission to hospital. Survival, however, was not
affected. The rationale for using albumin was to improve the filling of the effective
arterial blood volume in these patients, thereby improving renal perfusion. Despite
the fact that this makes physiological sense, the practice was not shown to be cost
effective. Hyponatremia is a common side effect complicating diuretic therapy in the
management of cirrhotic ascites. Hyponatremia is usually the result of over-activity of
vasopressin, in response to a reduction in the effective arterial blood volume. The
presence of hyponatremia prevents further diuretic use. The volume-expanding
property of albumin has also been shown to successfully correct hyponatremia and
this may permit the re-introduction of diuretic treatment.
A novel class of agents has recently been development to specifically correct the
hyponatremia in cirrhosis. Vasopressin (V2) receptor antagonists are aquaretics,
which mobilize free water, thereby correcting the hyponatremia. Indeed in cirrhotic
patients with ascites and hyponatremia, lixivaptan, a V2 receptor antagonist, has
veen shown to improve serum sodium without significant side effects. However, in
patients with ascites but without hyponatremia, aquaretics have also been to be
effective as adjunct therapy to diuretics in the management of ascites. In a
multicenter, double-blind, randomized controlled study, three fixed doses of
satavaptan (5 mg, 12.5 mg, 25 mg once daily), a V2 receptor antagonist, versus
placebo were administered to 148 patients with cirrhosis and ascites, in addition to
daily doses of standard diuretics consisting of spironolactone 100 mg and furosemide
20–25 mg per day for 14 days. Satavaptan was able to significantly increase the daily
urinary volume in a dose dependent manner, associated with a significant reduction
in abdominal girth and body weight, demonstrating potential clinical benefit of the
71
addition of a vasopressin V2 receptor antagonist to standard diuretic therapy in the
management of ascites in patients with cirrhosis without hyponatremia.
In another multi-center single-blind randomized controlled study, 151 patients with
liver cirrhosis and recurrent ascites requiring frequent large volume paracentesis
were given the same doses of satavaptan with 100 mg of spironolactone. The effects
of satavaptan on ascites control were compared to those of placebo over a 12-week
period. In those patients receiving satavaptan with spironolactone, the time to the first
paracentesis was delayed. In addition, the patients receiving Satavaptan had
significantly less number of paracentesis during the 12-week study period as well as
less ascites collected per week. Urine volume increased and urine osmolarity dosedependently decreased, suggesting that an increase of free water clearance alone
without natriuresis is also helpful in the control of ascites.
Thus the treatment of ascites will be a little easier in the near future when the role of
albumin will be better defined. The availability of aquaretic agents is likely to make
ascites patients less dependent on paracentesis, and overall improve the
management of these patients.
References:
1) Gerbes AL, Gülberg V, Gines P, et al. Therapy of hyponatremia in cirrhosis with a
vasopressin receptor antagonist: a randomized double-blind multicenter trial.
2) Wong F, Blei AT, Blendis LM & Thuluvath PJ. A vasopressin receptor antagonist
(VPA-985) improves serum sodium concentration in patients with hyponatremia:
a multicenter, randomized, placebo-controlled trial. Hepatology 2003; 37:
182–191.
3) Gerbes AL. The patient with refractory ascites. Best Pract Res Clin Gastroenterol
2007; 21: 551–560.
4) Wong F. The role of albumin in the management of chronic liver disease. Nat Clin
Pract Gastroenterol Hepatol 2007; 4: 43–51.
5) Laffi G, Gentilini P, Romanelli RG, La Villa G. Is the use of albumin of value in the
treatment of ascites in cirrhosis? The case in favour. Dig Liver Dis 2003; 35:
660–663.
6) Wong F, Gines P, Watson HR, Kujundzic M, Angeli P, Horsmans Y, for the SPA
investigators. Effects of a selective vasopressin V2 receptor antagonist,
satavaptan (SR121463B), on recurrence of ascites after large volume
paracentesis. [Abstract] Hepatology 2007; 44 (Suppl 1): 445A.
7) Gines P, Wong F, Watson HR, Ruiz del Arbol LR, Bilic A, Dobru D. Effects of a
selective vasopressin V2 receptor antagonist, satavaptan (SR121463B), in
patients with cirrhosis and ascites without hyponatremia. [Abstract] Hepatology
2007; 44 (Suppl 1): 256A.
72
The role of TIPS in the treatment of refractory ascites
Martin Rössle
Internist, Freiburg, Germany
Refractory ascites is one of the most serious complications of liver cirrhosis
compromising the quality of life and survival markedly. It may be associated with an
increased risk of further complications, such as spontaneous bacterial peritonitis and
hepatorenal syndrome. Accordingly, an effective treatment should be able to reduce
the recurrence of tense ascites and the rate of complications, as well as to improve
survival.
So far the standard therapy for patients with refractory ascites is large-volume
paracentesis that rapidly relieves abdominal distension, dyspnea, and reduces the
time of hospitalisation without improving survival. In more than 80% of cases,
however, it cannot prevent the re-accumulation of ascites because it does not correct
the mechanisms causing ascites formation. In addition, it does not sufficiently control
accompanying complications such as spontaneous bacterial peritonitis, hepatic
hydrothorax, or large umbilical hernia. With respect to the renal function which is
often impaired in patients with refractory ascites, it may even have a negative effect
and increases the risk of the hepatorenal syndrome. Conversely, transjugular
intrahepatic portosystemic shunt (TIPS) relieves portal hypertension which is one of
the main pathogenetic mechanisms of ascites formation and, therefore, it should be
able to reduce the recurrence of ascites and the incidence of complications related to
portal hypertension. Indeed, many uncontrolled studies reported elimination or
reduction of ascites after TIPS implantation in most cases. However, randomized
controlled trials (RCTs) comparing uncovered TIPS with large-volume paracentesis
showed that the greater efficacy of TIPS is counterbalanced by an increased rate of
hepatic encephalopathy (HE), whereas the results on survival were not uniform.
Three meta-analyses, performed on the aggregated data of those RCTs, confirmed
that TIPS significantly improves ascites but increases the risk of HE without a
significant improvement in survival (1–3).
In contrast to the 3 previous meta-analyses a recent meta-analysis (4) compared the
effects of TIPS and large-volume paracentesis by pooling individual data from the
RCTs (5–8). One RCT by Lebrec et al (9) was excluded because no data were
submitted. The study population consisted of 305 patients: 149 allocated to TIPS and
156 to paracentesis. Tense ascites recurred in 42% of TIPS patients and 89% of
paracentesis patients (P < 0.0001). Sixty-five patients in the TIPS group and 78 in the
paracentesis group died. The actuarial probability of transplant-free survival was
significantly better in the TIPS group (P = 0.035). Cox regression analysis performed
in a subgroup of 235 patients (114 TIPS and 121 paracentesis) showed that age,
serum bilirubin, plasma sodium and treatment allocation were independently
associated with survival. A subgroup analyses to evaluate any possible difference in
the risk for mortality according to different MELD scores showed that mortality was
lower in the patients allocated to TIPS compared to patients allocated to paracentesis
in low (= 10), intermediate (= 15) and high (= 19) MELD scores.
The average number of episodes of encephalopathy was higher in TIPS patients
(1.13 ± 1.93 vs. 0.63 ± 1.18, P = 0.006), although the cumulative probability of
developing the first episode of encephalopathy was similar between the two groups
(P = 0.19). Ascites associated complications such as spontaneous bacterial
73
peritonitis (2% vs. 3%) or hepatorenal syndrome (4.6% vs. 12.7%) were lower in the
TIPS arm.
This meta-analysis based on individual patient data shows that TIPS significantly
improves transplant-free survival of cirrhotic patients with refractory ascites. This
advantage does not disappear with higher MELD scores. Thus, patients with a poor
liver function must not be excluded from TIPS implantation. Certainly, liver
transplantation should be considered in any patient with refractory ascites. This
option is, however, limited to a minority due to advanced age, ongoing alcohol abuse,
poor performance, hepatocellular carcinoma or infections. Thus, in the RCTs
reported, only 10–20% of patients with refractory ascites were transplanted. On the
other hand, in patients awaiting transplantation, TIPS is not of disadvantage. In
contrary, many of these patients may be taken from the list or transplantation can be
delayed (10).
In conclusion, TIPS may improve survival in patients with refractory ascites of any
MELD score. TIPS may be of particular advantage in the presence of ascites
associated complications, such as recurrent spontaneous bacterial peritonitis and
hepatorenal syndrome. Liver transplantation is the treatment of choice in patients
with decompensated liver function with or without prior TIPS treatment.
References:
1. Albillos A, Banares R, Gonzalez M, Catalina MV, Molinero LM. A meta-analysis
of transjugular intrahepatic portosystemic shunt versus paracentesis for
refractory ascites. J Hepatol 2005; 43: 990–996.
2. D’Amico G, Luca A, Morabito A, Miraglia R, D’Amico M. Uncovered transjugular
intrahepatic portosystemic shunt for refractory ascites: a meta-analysis.
3. Saab S, Nieto JM, Ly D, Runyon BA. TIPS versus paracentesis for cirrhotic
patients with refractory ascites. Cochrane Database Syst Rev 2004; 3:
CD004889.
4. Salerno F, Cammà C, Enea M, Rössle M, Wong F. Transjugular intrahepatic
portosystemic shunt for refractory ascites: A Meta-Analysis of individual patient
data. Gastroenterology 2007, accepted for publication.
5. Rössle M, Ochs A, Gulberg V, Siegerstetter V, Holl J, Deibert P, Olschewski M,
Reiser M, Gerbes AL. A comparison of paracentesis and transjugular intrahepatic
portosystemic shunting in patients with ascites. N Engl J Med 2000; 342:
1701–1707.
6. Sanyal AJ, Genning C, Reddy KR, Wong F, Kowdley KV, Benner K,
Mc Cashland T. The North American Study for the treatment of refractory ascites.
74
7. Salerno F, Merli M, Riggio O, Cazzaniga M, Valeriano V, Pozzi M, Nicolini A,
Salvatori F, and GIST. Randomized controlled study of TIPS versus paracentesis
plus albumin in cirrhosis with severe ascites. Hepatology 2004; 40: 629–635.
8. Ginès P, Uriz J, Calahorra B, Garcia-Tsao G, Kamath PS, Del Arbol LR,
Planas R, Bosch J, Arroyo V, Rodes J. Transjugular intrahepatic portosystemic
shunting versus paracentesis plus albumin for refractory ascites in cirrhosis.
9. Lebrec D, Giuily N, Hadengue A, Vilgrain V, Moreau R, Poynard T, Gadano A,
Lbenhamou JP, Erlinger S. Transjugular intrahepatic portosystemic shunt:
comparison with paracentesis in patients with cirrhosis and refractory ascites: a
randomized trial. J Hepatol 1996; 25: 135–144.
10. Ochs A, Haag K, Rössle M. TIPS for ascites – reply. N Engl J Med 1995; 333:
879 (Letter).
75
Spontaneous bacterial peritonitis: A disease of the gut?
Therapeutic implications
Andres Cárdenas and Pere Ginès
GI and Liver Unit, Institut de Malalties Digestives i Metabòliques, Hospital Clínic,
Barcelona, Spain
Spontaneous bacterial peritonitis (SBP) is a common and severe complication of
cirrhotic patients with ascites characterized by spontaneous infection of ascitic fluid
(defined as a polymorphonuclear leukocyte count greater than 250 per cubic
millimeter) which occurs in the absence of any intra-abdominal source of infection.
The isolation of aerobic gram-negative bacteria (GNB), mainly Escherichia coli, in
most cases suggests that the gastrointestinal tract is the most common source of
infection. The pathogenesis of SBP involves passage of bacteria from the intestinal
lumen to the systemic circulation through translocation of bacteria to mesenteric
lymph nodes, bacteremia secondary to the impairment of the reticuloendothelial
system phagocytic activity, and infection due to poor opsonization and defective
bactericidal activity of ascitic fluid. Bacterial translocation, the process by which
enteric bacteria migrate across the intestinal mucosa and enter mesenteric nodes,
seems to play a key role in the pathogenesis of SBP. The enteric microflora has
more anaerobic bacteria than aerobic bacteria and unlike GNB, anaerobes in general
do not translocate across the intestinal mucosa and play a protective role against the
colonization and overgrowth of pathogenic GNB. Structural abnormalities of the
intestinal wall may also contribute to an increased potential for bacterial translocation
in cirrhosis. The hemodynamic abnormalities that occur in cirrhosis have unfavorable
effects on intestinal motility, increased permeability and the microflora. The
overactive sympathetic nervous system causes slow transit and favors bacterial
stasis and overgrowth. Additionally, edema of the submucosa disrupts gut integrity
and the mucosal barrier. All these changes result in an imbalance of gut microflora
and overpopulation of enteric GNB, increased bacterial toxins that lead to mucosal
damage and diminished mucosal defense. Therefore, SBP seems to be a result of
increased overgrowth of bacteria in the presence of altered intestinal permeability
and an impaired host immune response. Blood seeding with GNB facilitates
contamination of the sterile ascites that then results in SBP. Third-generation
cephalosporins are the antibiotics of choice for patients with SBP and the resolution
of the infection is achieved in 80–90% of patients. Prophylaxis of recurrent SBP is
based on the continuous administration of norfloxacin, however GNB resistant to
quinolones are a limiting factor for long term use. In consequence, there is a need for
non-antibiotic-based strategies to prevent intestinal bacterial overgrowth, bacterial
translocation and SBP in patients with cirrhosis.
76
Hepatorenal syndrome in cirrhosis
A.L. Gerbes
Med. Klinik und Poliklinik II, Klinikum der LMU München-Großhadern, Munich,
Germany
¾
Hepatorenal syndrome (HRS) is a functional renal failure that mainly occurs in
patients with cirrhosis, ascites and liver failure. It is characterized by impaired
renal function, marked alterations of cardiovascular function and over-activity of
the endogenous vasoactive systems. HRS is diagnosed by excluding other
types of renal failure.
¾
Type-1 HRS is caused by acute and severe impairment of circulatory function
related to increased peripheral vasodilatation. Progression to severe renal
failure is rapid. Survival of patients is extremely short, and they often show rapid
deterioration of liver function and hepatic encephalopathy. Type-1 HRS may
occur after a precipitating event, usually spontaneous bacterial peritonitis (SBP).
Type-1 HRS can be prevented in patients with SBP by albumin infusion.
¾
Type-2 HRS is the result of spontaneous progression of the circulatory
dysfunction in cirrhosis, which is mainly determined by increased splanchnic
vasodilatation. It is characterized by moderate, slowly evolving renal failure and
is frequently associated with refractory ascites.
¾
Patients who develop type-1 HRS can improve by administration of
vasoconstrictors (terlipressin, midodrine or norepinephrine) plus albumin, or by
transjugular intrahepatic portosystemic shunt (TIPS). However, liver
transplantation is the only treatment that provides long-term survival.
¾
Although vasoconstrictors plus albumin and TIPS can reverse type-2 HRS,
randomized controlled trials are required in order to determine whether their use
is suitable.
References:
1.
Arroyo V, Gines P, Gerbes A, et al. Definition and diagnostic criteria of
refractory ascites and hepatorenal syndrome in cirrhosis. Hepatology 1996; 23:
164–176.
2.
Gülberg V, Bilzer M, Gerbes AL. Long-term therapy and retreatment of
hepatorenal syndrome type 1 with ornipressin and dopamine. Hepatology 1999;
30: 870–875.
3.
Salerno F, Gerbes AL, Gines P, Wong F, Arroyo V. Definition, diagnosis,
prevention and treatment of hepatorenal syndrome in cirrhosis. A consensus
workshop of the international ascites club, Gut 2007, in press.
77
Infections and hepatic encephalopathy
Dieter Häussinger
Department of Internal Medicine, Heinrich Heine University Düsseldorf, Germany
Hepatic encephalopathy is frequent neuropsychiatric complication of liver cirrhosis,
which is known to exacerbate during infections. One example is spontaneous
bacterial peritonitis in cirrhotic patients with ascites, which frequently becomes
symptomatic through precipitation or worsening of hepatic encephalopathy (HE) only.
HE is nowadays seen as the clinical manifestation of a low grade cerebral edema
(1, 2), which induces an oxidative stress response in brain compartments leading to
covalent protein modifications and mRNA oxidation. As a result, astrocyte function
changes with impact on glioneuronal communication, neurotransmitter and receptor
processing with impact on synaptic plasticity. These alter oscillatory networks in the
brain, which may explain cognitive and fine-motoric dysfunction in HE (3).
Inflammatory cytokines, which are increasingly produced during infections, augment
these pathogenetic processes and precipitation of HE episodes. This is supported by
findings demonstrating a close relationship between circulating levels of tumor
necrosis factor-α (TNFα) and HE severity in cirrhotic patients (Odeh et al. Liver Int.
2004; 24:110-116). Several mechanisms have been identified by which
inflammmatory cytokines augment HE. Among these an important role is played by
(a) a cytokine-induced tyrosine nitration of hepatic glutamine synthetase (GS), which
inactivates the enzyme (4). As a consequence hepatic ammonia detoxication is
inhibitied due to an impaired ammonia-scavenging function of perivenous GScontaining hepatocytes. (b) TNFα induces in vitro astrocyte swelling, which is a
hallmark of HE. (c) Inflammatory cytokines, such as TNFα and interferon-α induce an
oxidative stress response and protein tyrosine nitration of astroglial proteins
synergistic with benzodiazepines and ammonia, i.e. other well-known precipitants of
HE in cirrhotics (5). (c) Preliminary studies suggest that TNFα triggers cerebral
mRNS oxidation. (d) Inflammatory cytokine augment cerebral NO production via
nNOS, which may contribute to cerebral hyperemia.
References:
1. Häussinger, D., Laubenberger, J., vom Dahl, S., Ernst, T., Bayer, S., Langer, M.,
Gerok, W. & Hennig, J. (1994) Proton magnetic resonance spectroscopic studies
on human brain myo-inositol in hypoosmolarity and hepatic encephalopathy.
Gastroenterology 107, 1475–1480.
2. Häussinger, D., Kircheis, G., Fischer, R. & vom Dahl, S. (2000) Hepatic
encephalopathy in chronic liver disease: a clinical manifestation of astrocyte
swelling and chronic low grade cerebral edema? J. Hepatol. 32, 1035–1038.
3. Timmermann, L., Gross, J., Butz, M., Kircheis, G., Häussinger, D. & Schnitzler,
A. (2003) Mini-Asterixis in hepatic encephalopathy is associated with pathological
thalamo- motorcortical coupling. Neurology 61, 689–692.
78
4. Görg, B., Wettstein, M., Schliess, F. & Häussinger, D. (2005) Lipopolysaccharideinduced tyrosine nitration and inactivation of hepatic glutamine synthetase.
Hepatology 41, 1065–1073.
5. Görg, B., Bidmon, H.J., Foster, N., Goehrlich, R., Schliess, F. & Häussinger, D.
(2006) Inflammatory cytokines induce protein-tyrosine nitration in rat astrocytes.
Arch. Biochem. Biophys. 449, 104–114.
79
Liver and lung – Treatment of hepatopulmonary diseases
Peter Schenk, M.D.
Associate Professor of Internal Medicine, Department of Internal Medicine III,
Medical University Vienna, Austria
Both vascular complications of liver disease, hepatopulmonary syndrome (HPS) and
portopulmonary hypertension commonly present with dyspnoea but have opposite
pathophysiological concepts.
Although HPS has been described in acute liver disease, such as acute hepatitis,
acute liver failure and hypoxic hepatitis, it is mainly relevant as a complication of liver
cirrhosis, where the reported prevalence is 4–32%. HPS is defined as arterial
deoxygenation induced by intrapulmonary vasodilation associated with hepatic
disease. Prognosis of patients with cirrhosis and HPS is poor, HPS is an independent
prognostic parameter for mortality and patients with HPS should be prioritized on the
waiting list for orthotopic liver transplantation (OLT). OLT is the only established
effective therapy, it improves HPS in > 80% of the patients and many centers
consider HPS as an indication for OLT. After OLT, HPS usually resolves within
12 months, increased postoperative mortality exists in patients with severe
hypoxaemia (PaO2 < 50 mm Hg) and high intrapulmonary shunt (> 40%).
HPS patients with severe hypoxaemia should receive long-term oxygen therapy. In
selected patients, embolisation of direct intrapulmonary arterio-venous
communications improved arterial oxygenation. Portal hypertension plays a central
role in the pathogenesis of HPS. Therefore reduction of portal pressure by
transjugular intrahepatic portosystemic shunt (TIPS) may improve HPS. However, a
series of 7 patients did not show any change in pulmonary gas exchange.
Pulmonary vasodilation is assumed to result from excessive production of
vasodilators, particularly nitric oxide (NO). Inhibition of cyclic GMP by methylene blue
transiently improved the hyperdynamic state, pulmonary vasodilation and arterial
oxygenation in 8 patients with moderate to severe HPS. Inhibition of NO by NG-nitroL-arginine methyl ester (L-NAME) improved gas exchange in 1 patient, however, a
recent study found that inhaled L-NAME decreased the hyperdynamic state and
intrapulmonary vasodilation but did not change arterial deoxygenation. In animal
studies, inhibition of bacterial translocation with norfloxacin prevented macrophage
accumulation in pulmonary vasculature thus decreasing TNF-α and NO production
which reduced HPS development. A human study with inhibition of bacterial
translocation by antibiotics is currently performed. In addition, inhibition of TNF-αinduced macrophagic NO production by pentoxifylline prevented HPS development in
rats. Garlic powder improved oxygenation in a case report and an open trial.
Portopulmonary hypertension has a reported prevalence of 2–8.5% in patients with
cirrhosis. Postoperative OLT mortality is higher with mean pulmonary artery pressure
> 35 mm Hg. Therapeutic experience exists with i.v. prostacyclin, p.o. bosentan and
sildenafil. Betablockers should be withdrawn, if possible.
Correspondence:
Peter Schenk, M.D., Associate Professor of Internal Medicine, Department of Internal
Medicine III, Medical University Vienna, Allgemeines Krankenhaus, Währinger Gürtel
18–20, A-1090 Vienna, Austria, Tel.: 0043-1-40400-4767 or 4502, Fax: 0043-140400-4797, E-mail: [email protected]
80
Allocation in liver transplantation: Is MELD the solution? North
American perspective
Douglas M. Heuman, M.D.
Virginia Commonwealth University, McGuire VA Medical Center, Richmond, VA, USA
Since 2/27/02, deceased donor livers for adult transplantation in the U.S. have been
allocated on a “sickest first” basis, using the Model for End Stage Liver Disease
(MELD) to assign priority. MELD was originally derived by analyzing variables
associated with 90-day survival after TIPS placement in a group of cirrhotic patients
with variceal bleeding or refractory ascites. For use in the transplant setting, it was
modified arbitrarily in several ways: eliminating points for etiology of liver disease,
assigning minimal values of 1 to bilirubin, INR and creatinine, capping creatinine at a
maximum value of 4, and assigning a creatinine value of 4 to patients receiving renal
dialysis. The resulting index empirically was found to predict short term survival in
several large cirrhotic populations. Early claims that MELD was superior to the more
traditional Child-Turcotte-Pugh (CTP) score were inflated by biases in patient
selection, but MELD is at least as accurate as the CTP score for predicting short term
survival, and has the advantage of being based on easily obtained, verifiable
laboratory data.
The five years since adoption of MELD-based organ allocation have seen only
minimal reduction in adult U.S. waiting list mortality (less than 2%, 2000–01
compared to 2005–06), despite a 33% increase in number of adult liver transplants
during the same period. Even this minimal improvement in mortality may be an
artifact resulting from listing practices: with the elimination of waiting time as a
criterion, cirrhotics with MELD scores less than 10–14, who might previously have
been listed, are now routinely deferred, and any deaths among these “pre-list”
cirrhotics no longer register as waiting list mortality. On the positive side, median time
on the waiting list prior to transplantation has decreased (e. g., for cirrhotics aged
50–64, median wait went from 987 days in 1999–2000 to 416 days in 2003–04).
The practice of using MELD as a minimal listing criterion to identify patients who
could benefit from transplantation has become widespread, but has little objective
justification. Though a relationship of MELD to mortality can be demonstrated
throughout its range, the performance of MELD at the low end of the range is poor. In
patients with MELD scores ≤ 20, we found that hyponatremia and uncontrolled
ascites were independent predictors of six-month pre-transplant survival, whereas
MELD was not. In this low-MELD population, CTP score was a better predictor of
short term survival than MELD. MELD is also less accurate than CTP score in
identifying patients at risk for death in the intermediate term (between 6 and
24 months). For intermediate term survival, we found that hemoglobin, albumin, INR
and bilirubin were important independent predictors of survival or transplantation, but
creatinine was not. These findings suggest that indicators other than MELD remain
useful in guiding listing decisions.
The impact of MELD-based organ allocation on post transplant outcomes has
probably been negative. Transplanting sicker patients leads to longer post transplant
hospitalization, increased complications and increased costs. Very high MELD
scores are associated with increased post transplant mortality. The precise effect of
“sickest first” organ allocation on post transplant survival, morbidity and cost is
difficult to pin down accurately because of a number of concurrent developments,
81
including: increasing average age at transplantation, an increasing proportion of
patients infected with hepatitis C, transplantation of many patients under MELD
exceptions unrelated to cirrhosis severity (e. g. hepatocellular carcinoma), evolution
of living donor transplantation, reduced proportion of patients undergoing
retransplantation, and growing use of extended criteria donor livers.
MELD is widely regarded as an objective measure of liver disease severity, but each
of the components of MELD can be altered by factors unrelated to severity of
cirrhosis. Intrinsic renal disease, transient biliary ductal obstruction, and warfarin
therapy each can increase MELD score to 20 or more in the absence of any chronic
liver disease. Because it is logarithmic, MELD is particularly sensitive to changes at
the low end of the scale, and fluctuations of 3 to 5 points can result when transient
dehydration raises creatinine or when fasting increases the bilirubin level in patients
with Gilbert’s syndrome. Substantial variation in INR from laboratory to laboratory can
occur because of differences in reagents used. Physicians have learned to
manipulate these factors to increase transplant priority for their patients. It is possible
to circumvent some of these concerns; for example, a surrogate MELD score
excluding the INR can give a reasonable approximation of true risk in patients treated
with warfarin. However, the organ allocation system remains blind to these nuances,
and patients with artifactual elevations of MELD continue to receive enhanced priority
for transplantation.
Although MELD has always been described as a work in progress, it has not been
modified since its adoption, and attempts to improve MELD face considerable
resistance. The relative weighting factors for INR, bilirubin, and creatinine have never
been reassessed, despite the fact that the original coefficients were based on a
relatively small, selected population and probably are not optimal. There is an
unnecessary element of “smoke and mirrors” associated with MELD’s arithmetic
complexity. Use of base 10 rather than natural logarithms and elimination of the
constant component (both simple linear transformations) would make the index more
intuitively accessible. It may be that logarithms can be eliminated altogether; we
found that an index calculated by taking the sum of creatinine, INR and 10% x
bilirubin was as accurate as the MELD score for predicting short term mortality. Most
important, there is growing evidence that MELD’s accuracy can be improved by
incorporating additional independent predictors of short term mortality. Numerous
studies from the U.S., Europe and South America have found that low serum sodium
is a predictor of short term cirrhotic mortality independent of MELD. We found that
uncontrolled ascites, documented objectively by imaging or paracentesis, also was
predictive of 180 day survival independent of MELD, and data of Kamath et al.
suggest that hepatic encephalopathy likewise has independent prognostic
significance. The current system of organ allocation permits routine MELD
exceptions for some uncommon clinical complications of cirrhosis (e. g. hepatopulmonary syndrome), but a consensus panel in 2006 rejected routine MELD
exceptions for hyponatremia, ascites and encephalopathy.
In conclusion, MELD has proven to be a good but imperfect predictor of short term
survival in cirrhosis. It is a poor minimal listing criterion. MELD based organ allocation
has eliminated much of the subjectivity from the process. The waiting list now
contains a smaller number of sicker patients. However pretransplant mortality
probably has not improved and post transplant outcomes are probably worse. A
comprehensive, evidence based revision of MELD is warranted, but may not be
politically feasible.
82
European perspective
A.O. Rahmel
Eurotransplant International Foundation, Leiden, The Netherlands
Eurotransplant International was founded in 1967 and is responsible for organ
allocation in seven European countries (Austria, Belgium, Croatia, Germany,
Luxemburg, Slovenia and The Netherlands). The liver allocation rules were changed
several times since the first liver was allocated by Eurotransplant in 1979. In
December 2006, a MELD-based liver allocation system was introduced in the
Eurotransplant countries. This latest change had become necessary, because the
discriminative power of the previous allocation system was no longer considered to
be sufficient. The old allocation system was based on 3 different urgency classes for
patients with chronic liver disease: “T2” included all patients in urgent need of
transplantation, “T3” all patients with chronic liver disease developing complications,
“T4” covered stable patients. In addition a separate urgency class was existing for
patients with acute liver failure without prior chronic liver disease (high urgency status
– ”HU”). Available donor livers were primarily allocated to “HU”-patients, thereafter to
the different elective patient groups. Within each of the elective urgency classes
allocation was based on waiting time.
Between 01.01.2000 and 31.12.2006 the number of patients on the waiting list
increased from about 600 to more than 2300. At the same time the number of
available donor organs did only rise from 1132 to 1427. In spite of the above
mentioned efforts to limit the access to the highest elective urgency class, the
number of patients with a “T2”-status continuously increased over the years, so that
finally about two thirds of all donor livers were allocated to this group of patients. As a
result allocation was more and more driven by waiting time. The increase in the
number of patients waiting for an organ transplant on the one hand and the lack of
discriminative power concerning urgency for transplantation of the allocation system
on the other hand leads to a major increase of the waiting list mortality. In 2006
346 patients died while waiting for a suitable liver transplant.
As a consequence in December 2006 a MELD-based allocation was introduced for
elective allocation of after extensive discussion within the transplant communities of
the different ET-countries. The well accepted “HU”-system with mandatory
international exchange of donor livers stayed in place.
The MELD-Score is calculated based on three objective lab values: INR, bilirubin and
serum creatinine:
MELD Score = 10 {0.957 ln(creatinine) + 0.378 ln(total bilirubin) + 1.12 ln(INR) +
0.643}
The calculate MELD-scores (lab MELD) are limited to 40 as a maximum and reflect
the probability to die within 3 months without liver transplantation and thus the
urgency of a transplantation. Within each urgency class organs are allocated based
on expected ischemic time (by taking into account the distance between donor and
recipient center) and were necessary waiting time as a tie-breaker.
83
For patients with liver diseases, where the MELD-score does not reflect the urgency
of the transplantation, a list of so called “standard exceptions” (SE) is defined. The
list of diseases includes hepatocellular carcinoma, polycystic liver disease, primary
hyperoxaluria type I, small for size syndrome after liver transplantation, cystic
fibrosis, hepatopulmonary syndrome, FAP, cholangiocarcinoma and other specific
diseases. They receive a MELD-score that matches the urgency of the underlying
disease to allow a fair allocation of donor livers also to this group of patients
(matchMELD).
In addition in case of a patient with a liver disease where neither the MELD-score
reflects the urgency of a transplantation nor a standard exception is defined, a so
called “non-standard exception” (NSE) can be applied for. The individual details of
the medical history and the current situation of these patients are eluded to an
auditor group that decides about granting an accurate matchMELD-score.
During the first six months after the introduction of the new MELD-based allocation
system 833 liver transplantations took place. 713 livers (85%) were allocated to
elective patients, 120 (15%) to high urgency patients. Acute liver failure was the most
common reason for high urgency transplantation. The most common underlying
diseases for an elective liver transplant were liver cirrhosis after viral infections,
mainly hepatitis B and C (34%), hepatocellular carcinoma (20%) and alcoholic liver
disease (20%).
72% of the livers allocated to elective patients were allocated based on the
calculated MELD-score (labMELD), 22% to patients with a SE, 2% to patients with
NSE and 4 to children with a pediatric MELD.
During the first 6 months the average labMELD of those patients receiving their
transplant based on their labMELD-score decreased from 28.6 to 24.4. Although it is
too early to judge about the effectiveness of the new allocation system it is of note
that the transition from the old to the new allocation system went without problems,
was well received and lead during the first 6 months to a reduction of the mortality on
the waiting list.
84
Emerging future therapies for portal hypertension
Jaime Bosch, M.D.
Professor of Medicine, Hepatic Hemodynamic Laboratory, Liver Unit, Hospital ClínicIDIBAPS, University of Barcelona Centro de Investigación Biomédica de
Enfermedades Hepáticas y Digestivas (CIBEREHD), Barcelona, Spain
New methods for the development and evaluation of new treatments
HVPG is a strong prognostic indicator in chronic liver disease and portal
hypertension. Moreover, changes in HVPG during therapy for portal hypertension
closely correlate with clinical outcomes and survival. There is an increasing
awareness that HVPG may be the best surrogate end-point in the evaluation of new
therapies for portal hypertension and its complications, as well as to assess
progression of alcoholic, viral, metabolic (and perhaps cholestatic) cirrhosis. The use
of HVPG as a surrogate of clinical events in initial clinical trials will greatly facilitate
research in treatments testing new paradigms.
However, to make its use reliable worldwide HVPG measurements need to be
standardized following internationally agreed guidelines.
The old paradigm: Use of vasoactive drugs
Until now, the medical treatment of portal hypertension has been based on the use of
vasoactive drugs that could decrease splanchnic blood flow and portal (and variceal)
pressure. These include terlipressin, somatostatin (and analogs) and non-selective
beta-blockers, alone or in association with a low dose of a vasodilator (to enhance
the reduction in portal pressure and minimize systemic side-effects).
New paradigms
Emerging treatments for portal hypertension are testing new paradigms, for which
objective evidence has been accumulating in recent years. These include:
a) Modification of the structural abnormalities increasing hepatic vascular
resistance, by means of specific treatments for the underlying liver disease, of
drugs promoting sinusoidal remodelling, or by cell transplantation.
b) Correction of the dynamic component of increased hepatic resistance, by means
of posttranslational upregulation of eNOS and SH2 and inhibition of COX-1 and
TXA2.
c) Targeting collateral formation and hyperkinetic circulation by
angiogenesis interfering with VEGF and PDGF signalling pathways.
inhibiting
These new paradigms are currently being tested both in animal models and in
preliminary clinical trials.
85
List of Speakers, Moderators and Scientific Organizers
Prof. Dr. D.H. Adams
Queen Elizabeth Hospital
Institute of Clinical Sciences
Liver Research Laboratories
Birmingham B15 2TH
Great Britain
Prof. Dr. M. Beaugrand
Hôpital Jean Verdier
Service d'Hépato-Gastro-Entérologie
Ave. du 14 Juillet
F-93140 Bondy
France
Prof. Dr. J. Bosch
Hospital Clinico y Provincial
Universidad de Barcelona
Villarroel 170
E-08036 Barcelona
Spain
D.A. Brenner, M.D.
Professor of Medicine
UCSD School of Medicine
University of California, San Diego
1318A Biomedical Sciences Building
9500 Gilmore Drive
La Jolla, CA 92093-0602
USA
Prof. Dr. A.K. Burroughs
Royal Free Hospital
School of Medicine
Department of Medicine
Pond Street
London NW3 2QG
Great Britain
Prof. Dr. P. Calès
Centre Hospitalier
Département de Hépatologie
F-49033 Angers
France
Dr. M. Fernández
Hepatic Hemodynamic Laboratory
Liver Unit
Villarroel 170
E-08036 Barcelona
Spain
Prof. Dr. R. de Franchis
Università di Milano
IRCCS Ospedale Maggiore Pol.
Dipartimento di Medicina Interna
Servizio di Gastroenterologia
Via Pace, 9
I-20122 Milano
Italy
S.L. Friedman, M.D.
Mount Sinai School of Medicine
Room 11-70F
1425 Madison Ave.
New York, NY 10029
USA
Dr. J.C. Garcίa-Pagán
Hepatic Hemodynamic Laboratory
Villarroel 170
E-08036 Barcelona
Spain
G. Garcia-Tsao, M.D.
Yale University
School of Medicine
Section of Digestive Diseases
333 Cedar Street
New Haven, CT 06520
USA
87
Prof. Dr. A.L. Gerbes
Innere Medizin II
Klinikum der Universität
München-Großhadern
Marchioninistr. 15
D-81377 München
Germany
Prof. Dr. P. Ginès
Unidad de Hepatologia
Villarroel 170
E-08036 Barcelona
Spain
Prof. Dr. A.M. Gressner
Universitätsklinikum Aachen
Klinische Chemie/Zentrallabor
Pauwelsstr. 30
D-52074 Aachen
Germany
Prof. Dr. D. Häussinger
Gastroenterologie/Hepatologie
Universitätsklinikum Düsseldorf
Moorenstr. 5
D-40225 Düsseldorf
Germany
PD Dr. J. Heller
Medizinische Klinik I
Universitätsklinikum Bonn
Sigmund-Freud-Str. 25
D-53127 Bonn
Germany
PD Dr. C. Hellerbrand
Klinik für Innere Medizin I
Klinikum der
Universität Regensburg
D-93042 Regensburg
Germany
88
D.M. Heuman, M.D.
Virginia Commonwealth University
McGuire VA Medical Center
Hepatology/Liver Transplantation
P.O. Box 980341
Richmond, VA 23298
USA
H.W. Jaeschke, Ph.D.
Professor of Toxicology
University of Kansas
Medical Center
Pharmacology & Toxicology, MS1018
3901 Rainbow Blvd.
Kansas City, KS 66160
USA
Dr. C. Kordes
Gastroenterologie/Hepatologie
Universitätsklinikum Düsseldorf
Moorenstr. 5
D-40225 Düsseldorf
Germany
Prof. Dr. F. Lammert
D-53127 Bonn
Germany
Prof. Dr. D. Lebrec
Hôpital Beaujon
Inserm U 481
100, Bd. Général Leclerc
F-92118 Clichy
France
Dr. A. Moschetta
Ospedale Consorziale
Policlinico
Department of Internal and
Public Medicine
I-70124 Bari
Italy
Prof. Dr. M. Pinzani
Università di Firenze
Ospedale Careggi
Fisiopatologia Epatica
Clinica Medica II
Viale G.B. Morgagni 85
I-50134 Firenze
Italy
V. Shah, M.D.
Assistant Professor of Medicine
Mayo Clinic
GI Research Unit
Alfred 2-435
200 First Street SW
Rochester, MN 55905
USA
Dr. A. Rahmel
Eurotransplant
Internationa Foundation
P.O. Box 2304
NL-2301 CH Leiden
The Netherlands
Prof. Dr. U. Spengler
D-53127 Bonn
Germany
Prof. Dr. Dr. h.c. G. Ramadori
Gastroenterologie
Universitätskliniken Göttingen
Robert-Koch-Str. 40
D-37075 Göttingen
Germany
Dr. D. Thabut
Hôpital Pitié Salpêtrière
Hepato-Gastroenterology
AP-HP Groupe
47–83, boulevard de l'Hôpital
F-75013 Paris
France
Prof. Dr. T. Roskams
Catholic University of Leuven
University Hospital St. Rafael
Department of Pathology
Minderbroederstraat 12
B-3000 Leuven
Belgium
Prof. Dr. M. Rössle
Internist
Bertoldstr. 48
D-79098 Freiburg
Germany
Prof. Dr. T. Sauerbruch
D-53127 Bonn
Germany
Dr. P. Schenk
Medizinische Universität Wien
Innere Medizin IV, 13H3
Währinger Gürtel 18–20
A-1090 Wien
Austria
Prof. Dr. C. Trautwein
Medizinische Klinik III
Pauwelsstr. 30
D-52074 Aachen
Germany
PD Dr. H.E. Wasmuth
Medizinische Klinik III
Pauwelsstr. 30
D-52074 Aachen
Germany
PD Dr. R. Weiskirchen
Klinische Chemie/Pathobiochemie
Pauwelsstr. 30
D-52074 Aachen
Germany
PD Dr. R. Wiest
Klinik für Innere Medizin I
Klinikum der Universität Regensburg
D-93042 Regensburg
Germany
89
Prof. Dr. F. Wong
University of Toronto
Toronto General Hospital
9EN/220
200 Elizabeth Street
Toronto, ON M5G 2C4
Canada
90
POSTER ABSTRACTS
Poster Numbers 1 – 98
Author Index to Poster Abstracts
1
Hepatic cirrhosis in the elderly in a Hispano-American
population
E. Aravena, F. Contesse, R. Muñoz, H. Iturriaga
Chilean Japanese of Digestive Diseases, Santiago, Chile
Introduction: In recent years a progressive increase in the number of elder
inpatients with cirrhosis has been observed. The aims of our study are to compare
the etiologies of cirrhosis in the elderly (> 60 years) with those observed in younger
population to verify if there has been an epidemiologic change in cirrhosis.
Methods: Patients with cirrhosis who had their first decompensation between 1998
and 2001 were selected from a database of hospital discharges. We analyzed age,
etiology and Child-Pugh classification. Etiologic study was based on clinical data
including obesity, diabetes and dyslipidemia, HBsAg, antiVHC, protein
electrophoresis, antibodies and iron parameters. Patients with incomplete study were
excluded (17 out of 225).
Results: The age of the 208 patients ranged from 18 to 83 years (mean ± SD, 57.6 ±
12.2) with a similar gender proportion between males (51%) and females (49%). The
most common etiology was alcohol (56%), followed by cryptogenic (15%), hepatitis C
virus (9%), autoimmune (9%), non-alcoholic steatohepatitis (5%), primary biliary
cirrhosis (4%) and hepatitis B virus (2%). Patients were classified in Child-Pugh A
(30%), B (38%) and C (32%). In patients more than 60 years of age (47%) as well as
in the youngest age group (53%) the most frequent etiology was alcoholic, followed
by cryptogenic and hepatitis C virus. In the group of elderly patients women were
slightly predominant (52%), where the cryptogenic and hepatitis C virus etiology was
more prominent.
Conclusion: Compared with previous studies in our country, the proportion of elder
patients and women with cirrhosis has increased significantly, and alcohol is a less
frequent etiology. There are no significant differences in etiology and cirrhosis
severity between elder and younger patients. Further studies will be necessary for
validation of this epidemiologic change.
2
Liver affection in different types of glycogen storage disease in
children
Dublina E., Uvarova E., Strokova T., Potapov A.
Scientific Centre of Children’s Health, Moscow, Russia
Introduction: Glycogen storage disease (GSD) is a group of relatively frequent
metabolic disorders with liver involvement. The aim of this work was to follow up liver
affection in different types of GSD.
Methods: 97 children with GSD (type I – 18, type III – 30, types VI and IX – 49) were
followed up during 5–10 years. They underwent clinical examination, blood
biochemistry, USS, liver histology and histochemistry.
Results: GSD type I mostly presented in neonates with hepatomegaly, cramps,
severe hypoglycaemia, hyperlacticacidaemia, dyslipidaemia, hyperuricaemia, while
the signs of liver cytolysis and cholestasis were mild. Liver histology indicated
hepatocytes’ distension with glycogen, prominent lipid vacuoles, mild fibrosis,
glucose-6-phosphatase deficiency. Long-term observation in conditions of dietary
treatment showed hepatomegaly decrease, however cirrhosis developed in
2 (11.1%) children by 8–9-year age.
GSD type III usually manifested later in infancy with hepatosplenomegaly, growth
retardation, hypoglycaemia, hypercholesterolaemia, elevated transaminases and
cholestasis markers, increased creatinekinase. Liver histology was distinguished by
more expressed fibrosis, less steatosis, amylo-1,6-glucosidase deficiency. 7 children
had isolated liver affection with benign disease course. 23 had progressive
myopathy, gradual liver cytolysis and cholestasis, which resulted in cirrhosis in
7 (22.3%) cases. Liver adenomas were observed in 3 children.
GSD types VI, IX presented on the third year of life with protuberant abdomen,
hepatomegaly, failure to thrive. Hypoglycaemia, hyperlipidaemia, hyperketosis,
transaminase elevation were mild if presented. These types mainly had benign
course with hepatomegaly reduction, although 2 (4%) cases of cirrhosis were
registered in puberty.
Discussion/Conclusion: Liver affection in GSD and its prognosis depend on type of
disease.
3
Wilson disease in childhood: Results of a single center
experience
M. Georgieva1, D. Konstantinova-Kanazireva1, I. Krasnaliev2, A. Atanassova3,
S. Tomova4, A. Savov5, V. Tzaneva1
Dept. of Pediatrics & Genetics1 , Dept. of Pathology2, Clinic of Hepatogastroenerol3,
University Hospital “St. Marine”, Instit. of Economics4, Varna, National Genetic
Laboraory5, Sofia, Bulgaria
Introduction: Copper serves as an integral component of a variety of key enzymes.
High concentration of copper is responsible for toxicity. Wilson disease (WD) is
associated with a clinically significant accumulation of copper and exhibits an
autosomal recessive inheritance.
Aim is to evaluate the clinical findings, diagnosis (+ genetic analysis) & treatment of
WD in children & adolescents.
Methods: In 1987–2006 the diagnosis was proved in 26 patients (14 girls & 12 boys,
3–18 years, median age of 10.9 years) by clinical, laboratory (+ specific), genetic
(22 pts.), MRI (3 pts.), liver biopsy (6 pts.).
Results: Twenty of these children were with hepatic dysfunction, one – with mixed
neurologic and hepatic involvement, four – with neurological form, one – without any
symptoms. Hepatic symptoms were: fatigue (18/20), abdominal pain (17/20),
jaundice (12/20), oedema (8/20), ascites (17/20), hepatomegaly (20/20),
splenomegaly (10/20), osteoporosis (2/20); elevated aminotransferases (20/20),
decreased: prothrombine time (18/20) & serum albumin (15/20) & cholesterol (6/20).
Neurologic manifestations were presented in 4 adolescents: asterixis (4/4), dysarthria
(3/4), dysphagia (2/4), rigidity (1/4), choreic movement (1/4) without changes in
hepatic function. One boy was with hepatic complains + severe neurological
manifestations – rigidity, dysphagia, dysarthria. Fifteen of pts. had Kayser-Fleischer
corneal ring. Specific examinations showed: decreased ceruloplasmin (19/26),
decreased serum copper (25/26), increased urinary excretion of copper for 24h.
(26/26). Test with 1.0 D penicillamine provocated the elevation of urinary excretion of
copper from 10–20 times. In 20/22 pts. mutation analysis showed: 15 – homozygotes
H1069Q, 1 – homozygote 3400delC/3007G>A, 4 – compound heterozygotes.
Ultrasound imaging showed portal hypertension in 16/26 pts., ascites in 10/26,
hepatomegaly in 21/26. MRI showed changes in the basal ganglia in 3 of the pts.
with neurologic symptoms. Liver biopsy showed signs of chronic liver damage – large
drops steatosis, vacuolar degeneration of the hepatocytes, disrupt architectonics,
regenerative nodules of hepatocytes, surrounded by fibrous connective tissue,
bridges between portal tracts, lymphocytes, and proliferation of bile ducts. The
treatment was with D penicillamine, zinc preparations, in 4 pts. – infusions with RBC,
platelets, humanalbumin, plasma, antibiotics, Ursofalk®, transmetil, hyper bar
oxygenation, hemoperfusion dialysis. Five of the children developed hepatorenal
syndrome & died with the picture of severe brain and pulmonal edema. Disease
progression under treatment was more likely for neuropsychiatric than for hepatic
symptoms. Side effects of treatment occurred in 69.23% of patients.
Discussion/Conclusion: In conclusion the diagnosis of WD is difficult because of
the variable clinical manifestation & laboratory tests. In children & adolescents with
unclear hepatic and/or neurologic symptoms copper metabolism & molecular genetic
analysis should be examined.
4
Liver cirrhosis in cystic fibrosis
K. Kalinova, K. Todorova
University Hospital, Stara Zagora, Bulgaria
Introduction: Liver disease is the second cause of death in cystic fibrosis. The most
deleterious complication of liver disease is portal hypertension, which has an
estimated prevalence of up to 8%. Portal hypertension may manifest itself by
splenomegaly, hypersplenism, gastro-oesophageal bleeding and ascites. The aim of
our study was to determine the prevalence, risk factors and invasive management of
portal hypertension at our centre.
Methods: Forty patients with cystic fibrosis were followed up between 1995 and
2006. Nine patients (27%) had liver disease. All underwent clinical evaluation and
laboratory and imaging studies.
Results: All patients underwent clinical, ultrasonographic, and biochemical
examinations at entry and then every 6 months up to 3 years of follow-up. Patients
with cirrhosis underwent endoscopy every 12 months. In a sub-group of patients
without cirrhosis, who consented, liver biopsy was repeated at the end of the study.
Portal hypertension was diagnosed in 10 patients (25%), of whom six were male. The
mean age at diagnosis was 35 years (range, 7–57 years). All had severe mutations
of the cystic fibrosis transmembrane conductance regulator gene (the CFTR gene),
pancreatic insufficiency, or distal intestinal obstruction syndrome and variceal
bleeding. Six patients underwent sclerotherapy to control acute bleeding. Four
underwent portosystemic shunting. Two patients with severe lung and liver disease
underwent transjugular intrahepatic portosystemic shunting, which provided bleeding
control, but both died while waiting for liver transplantation. Treatment with
ursodeoxycholic acid (15–20 mg/kg days) regularly improves the biochemical
markers of liver disease. Treatment with ursodeoxycholic acid regularly improves the
biochemical markers of liver disease.
Discussion/Conclusion: Liver cirrhosis and portal hypertension is more common
complications in patients with cystic fibrosis. The unique genetic composition of our
population may explain this phenomenon. Risk factors include male gender,
pancreatic insufficiency, severe CFTR mutations, and meconium ileus equivalent.
Sclerotherapy is the main option to control oesophageal variceal bleeding, while
portosystemic shunts offer a prolonged alternative treatment for refractory bleeding.
A transjugular intrahepatic portosystemic shunt and liver transplantation may also be
effective, but further research is required in order to establish their role. Long-term
ursodeoxycholic acid therapy did not alter disease progression in primary biliary
cirrhosis patients despite a significant improvement in serum bilirubin and alkaline
phosphatase consistent with, and similar to, those seen in ursodeoxycholic acid
cohorts in randomized trials.
5
Niemann-Pick disease-type B
Konstantinova-Kanazireva D., M. Georgieva, V. Sinigerska*, V. Tzaneva,
L. Angelova
Dept. of Pediatrics & Medical Genetics, University Hospital “St. Marine”, Varna,
*National genetic laboratory, Sofia, Bulgaria
Introduction: Niemann-Pick disease (NPD) is a lipid storage disorder that results
from the deficiency of a lysosomal enzyme, acid sphingomyelinase. Till now
6 subtypes of NPD have been described, including type B, the second most
common.
The aim is to evaluate the clinical findings, diagnosis, mutation analysis & treatment
of NPD in 5 children.
Methods: For 5 years (2002–2007) the diagnosis NPD was proved in 5 patients
(4 boys and 1 girl) by clinical + laboratory findings and mutation analysis. All children
are of gypsy origin and unrelated.
Results: All children have been with normal appearance at birth. In four of them
clinical manifestation has started up to 3 years of age, in one – at 7 years with
enlargement of liver ± spleen. Hepatic symptoms were: fatigue (4/5), abdominal pain
(3/5), oedema (2/5), hepatomegaly (5/5), severe splenomegaly (5/5), osteoporosis
(1/5), elevated aminotransferases (4/5), pancytopenia (3/5), elevated cholesterol
(5/5). Neurologic manifestation was present in the patient with the latest onset; mild
mental retardation in 1/5. Growth retardation is found in all children. Cherry-red spot
(2/5). Myelogram with foamy cells (4/5). Specific lab test: decreased
sphingomyelinase activity (5/5). Acid sphingomyelinase mutation analysis – all
patients were found homozygotes for a specific mutation (W391G), characteristic for
patients of gypsy origin.
Discussion/Conclusion: NPD is difficult to diagnose because of the variable clinical
manifestation and rarity of the disease. In children with unclear hepatic and spleen
involvement NPD should be considered and specific lab tests and mutation analysis
should be performed.
6
Clinical features of pediatric liver cirrhosis resulting from
alpha1-antitrypsin deficiency
I. Lastauka, M. Halabarodzka
Belarusian Medical Academy of Post-Graduate Education, Minsk, Belarus
Introduction: Aim was to assess clinical features of pediatric liver cirrhosis (LC)
resulting from alpha1-antitrypsin deficiency (ATD).
Methods: 13 children with LC resulting from ATD were observed. Age at diagnostics:
in 9 children (69.2%) – from 2 weeks to 4 months, in 4 – 7–11 years old. Follow-up is
2.9 (1.3–8.0) years.
Diagnosis of ATD has been made by detecting of serum alpha1-antitripsin level
decreasing (0.62, 0.40–0.84 mg/l) and PiZZ genotype determined in all observed
patients. Diagnosis of LC was confirmed histologically in 61.5%.
Results: Cholestatic hepatitis (in 7 infants) and hepatosplenomegaly with normal
liver function tests (in 2) were presentations of ATD in infants up to 4 months of life.
LC manifestation was first presentation of ATD in children 7–11 years old. All
observed newborns with ATD will develop LC in 4.4 (3.0–5.9) years.
Clinical signs were evaluated in 4.7 (2.5-8.0) years after LC was diagnosed. Elevated
serum ALT level was detected in 10 children (76.9%), but it was not severe – 0.90
(0.77–1.33) mmol/l/h. Mild increasing of direct bilirubin, alkaline phosphatase and
gamma-glutamyltranspeptidase had 2 children (15.4%). All the children survived, all
had PELD score < 10, but 2 children had liver transplantation.
Discussion/Conclusion: Diagnosis of ATD has been made in children with
2 different clinical presentations: symptomatic liver disease (cholestatic hepatitis or
hepatosplenomegaly) in first months of life or advanced liver disease (LC) in older
children. LC is also natural outcome of neonatal ATD-associated hepatitis. Children
with LC resulting from ATD had relatively mild clinical signs and were characterized
by prolonged period of clinical compensation.
7
US scanning data in liver cirrhosis in infants of first 3 years old
T. Raeuneva, M. Halabarodzka
Minsk Children’s Infectious Clinic, Belarusian Medical Academy of Post-Graduate
Education, Minsk, Belarus
Introduction: Aim was to reveal US signs presented in liver cirrhosis (LC) in infants
of first 3 years old.
Methods: 79 children were observed. Diagnosis of LC was estimated during first
3 years of life in 30 (research group), 4–18 years – in 49 (control group).
Diagnosis of LC was based on complex of clinical data and was confirmed
histologically in 70.9% (22 and 34 children respectively by groups). Following US
features were evaluated: liver lobes enlargement, liver vascular pattern depletion,
portal vein dilatation, cavernous transformation of portal vein, arterialization of
hepatic blood flow, hepatic artery dilatation, increasing of spleen length, splenic vein
dilatation, absence of superior mesenteric vein reaction to breath phases, vein
anastomoses visualization, ascites, retrograde flow in portal vein.
Results: There were the features significantly more often presented in research
group: right liver lobe enlargement (86.0% vs. 55.2%, p = 0.0080), portal vein
dilatation (53.4% vs. 22.4%, p = 0.0103), arterialization of hepatic blood flow (53.4%
vs. 20.4%, p = 0.0055), retrograde flow in portal vein (30.0% vs. 6.2%, p = 0.0109).
There were the features significantly rarer presented: hepatic artery dilatation (3.3%
vs. 26.6%, p = 0.0205) and vein anastomoses visualization (6.6% vs. 38.7%,
p = 0.0041).
Discussion/Conclusion: Evidence of late US signs of portal hypertension
(arterialization of hepatic blood flow and retrograde flow in portal vein) with failure of
timely compensatory manifestations (vein anastomoses, hepatic artery dilatation)
development may lead to severe course and early decompensation of LC in infants
of first 3 years old compare to older children.
8
H63D heterozygotes with alcoholic liver disease are proned to
hypercholesterolemia: A single centre, prospective study
Joanna Raszeja-Wyszomirska*, Iwona Zawada*, Janina Suchy**,
Grzegorz Kurzawski**, Jan Lubiński**, Piotr Milkiewicz*
*Liver Unit, Division of Gastroenterology, **Department of Pathology and Genetics,
Pomeranian Medical School, Szczecin, Poland
Background: Heterozygocity for HFE mutation has been linked with predisposition to
alcoholic liver disease (ALD). As iron overload precipitates the progression of alcohol
induced liver damage in animal models, HFE mutations may play a role in the
development/progression of ALD in humans. HFE mutations were also linked with
progression of hepatic damage in patients with non-alcoholic fatty liver.
Aims: to (i) determine the prevalence of C282Y and H63D mutations in well defined
cohort of patients with ALD (ii) to analyze laboratory findings in patients with
particular HFE mutations
Methods: Blood fasting samples for biochemistry and genetic analysis were taken
from 154 consecutive patients with ALD admitted to the tertiary liver centre.
Mutations were detected by PCR-RFPL as previously described and study protocol
accepted by the local ethic committee.
Results: The distribution of HFE mutations in analyzed cohort was as follows:
1 (0.63%) homozygote and 8 (5%) heterozygotes for C282Y mutation; 8 (5%)
homozygotes and 40 (25%) heterozygotes for H63D mutations and one (0.6%)
compound heterozygote. Patients who were H63D heterozygotes when compared to
patients not having HFE mutations had significantly higher levels of total cholesterol
(235 ± 38 vs 170 ± 9, p = 0.01); LDL cholesterol (160 ± 34 vs. 105 ± 8, p = 0.009)
and AST (228 ± 58 vs. 138 ± 15; p = 0.03). No significant difference between these
groups was found when patients age, gender, bilirubin, ALT, ALP, Fe, TIBC or ferritin
levels were compared. C282Y heterozygotes had significantly higher serum Fe levels
than all the other groups.
Conclusions: (i) The prevalence of HFE mutations in patients with liver cirrhosis due
to ALD is not different from the prevalence of these mutations in general population.
(ii) Patients with ALD who are H63D heterozygotes have higher total and LDL
cholesterol levels. In view of the potential role of H63D heterozygocity in a
progression of NAFLD, this novel finding may be of help in elucidation of the
pathogenesis of liver damage in ALD.
9
Natural history of primary sclerosing cholangitis in patients
with inflammatory bowel disease: Our experience
Svorcan P1, Bojic D1, Protic M1, Bojic B1, Djorovic S2, Djordjevic J1, Jojic Nj1,
Kazic S1, Dapcevic B1
1
University Clinical Center Zvezdara-Gastroenterology Ward, Belgrade, Serbia
2
University Clinical Center Zvezdara-Radiology Ward, Belgrade, Serbia
Introduction: Primary sclerosing cholangitis (PSC) is a chronic cholestatic disease
commonly associated with inflammatory bowel disease (IBD).
Aims: To evaluate clinical course of PSC in PSC-IBD patients diagnosed and treated
in our referral center.
Methods: 411 IBD patients, admitted to our Center in 2000–2005 were evaluated:
262 (64%) had ulcerative colitis (UC) and 149 (36%) had Crohn’s disease (CD). The
diagnosis of PSC was based on cholestasis and endoscopic retrograde
cholangiography (ERCP) finding. All patients with PSC were treated with
ursodeoxycholic acid (Ursofalk®) in dose 15 mg/kg body weight daily.
Results: 16 (3.9%) IBD patients had PSC: 12 (75%) had UC and 4 (25%) CD.
Male:female ratio was not significantly different (58%/42%, respectively). 56% of all
PSC-IBD patients had no IBD symptoms. ERCP findings showed large duct PSC in
7 (43.7%) patients, small duct PSC in 6 (37.5%) and both large and small duct PSC
in 3 (18.8%) patients. No significant difference was found in liver function tests
regarding the type of PSC. Pancolitis was found in 83% of our patients and 81% had
mild/moderate disease. Median age at PSC diagnosis was 39 years (range 15–51).
PSC in 88% patients was diagnosed after IBD was established. Colectomy was
performed in 5/16 patients and an all of them PSC was diagnosed after colectomy.
Conclusion: PSC was diagnosed after IBD diagnosis in majority of patients. Large
duct PSC tended to be predominant. The course of IBD in our group PSC-IBD
patients was mild with clear predominance of pancolitis.
10
Copper and ceruloplasmin in sera of patients with alcoholic
liver cirrhosis
E. Uhlikova, V. Kupcova, M. Szantova, I. Vozar, L. Turecky
Medical School, Comenius University, Bratislava, Slovakia
Introduction: Copper is an essential trace element. The major fraction of copper
present in the blood plasma is integrated in the molecule of ceruloplasmin
Ceruloplasmin is synthesized in the liver and secreted into the plasma as an alpha-2glycoprotein. Although its precise physiological roles are still unknown, it exerts an
oxidative activity toward iron, low density lipoprotein and homocysteine, and is
reported to have roles related to angiogenesis, antioxidant defense and copper
transport. Changes in serum ceruloplasmin levels have been reported under many
conditions, among these also in patients with liver diseases.
The purpose of this study was to examine the effect of liver cirrhosis on plasma
levels of copper and ceruloplasmin and to compare the changes of apoceruloplasmin
with its enzymatic activity.
Methods: 48 patients suffering from alcoholic liver cirrhosis were enrolled in the
study. The serum levels of copper, immunoreactive ceruloplasmin and oxidase
activity of ceruloplasmin were determined.
Results: The copper level in patients with liver cirrhosis was moderately decreased
in comparison to healthy controls. Enzymatic activity of ceruloplasmin was also
decreased in liver cirrhosis in comparison with healthy controls. The difference in
ceruloplasmin apoprotein levels between patients with liver cirhosis and healthy
controls was not significant, but the specific activity of ceruloplasmin in patients was
significantly decreased in comparison to controls (0.64 vs. 0.82, P < 0.001).
Discussion/Conclusion: The decreased specific activity of ceruloplasmin in spite of
normal levels of apoceruloplasmin in patients with alcoholic liver cirrhosis suggests
some problems in copper metabolism in these patients.
The work was supported by VEGA grant 01/4268/07.
11
Hepatic stellate cells and liver regeneration in chronic viral
hepatitis C
Abdulkhakov S., Cheremina N., Gumerova A., Titova M., Kiassov A., Fatkheeva L.,
Smetannikova T., Andreeva D.
Kazan State Medical University, Kazan, Russia
There is still a question about sources of liver regeneration as well as hepatic stellate
cells as probable pretender for the role of stem cell.
The aim pf our study was to examine cellular reactions in liver in case of chronic viral
hepatitis C.
105 liver biopsies were stained immunohistochemically with antibodies to alphaSMA, CD 31, CD 34 and GSTpi.
The number of CD 34-positive cells in liver parenchyma correlated with histology
activity index (HAI) and increased from HAI 4–8 to biopsies with HAI 13–18. While
comparing biopsies with different fibrosis score we’ve got the same result: the more
advanced fibrosis was more CD 34-positive cells were found in liver parenchyma.
These CD 34-positive cells could be either the result of sinusoids’ capillarization or
haematopoietic stem cells getting to the liver. To find out the latter we stained
biopsies with antibodies to CD 31 which is known as the marker of endothelial cells in
case of their capillarization. Number of CD 31-positive cells in areas of inflammation
grew both with increasing of HAI and fibrosis score while decreased in liver
sinusoids. These results prove that some CD 34-positive cells in liver parenchyma
could be haematopoietic stem cells while increase of their number in areas of
inflammation is mostly due to capillarizaion of sinusoids.
Dynamics of the number of alpha-SMA-positive and GSTpi-positive cells was the
same: their number increased with the growth of HAI or fibrosis grade in portal areas
while continuously goes down in liver sinusoids. Their common behaviour and
stellate shape of GSTpi-positive cells allow us to suppose that GSTpi could probably
be the marker of hepatic stellate cells in human liver which play key role in liver
regeneration.
12
TGF-β and ethanol: Two substances that exponentiate each
other leading to increased liver damage
K. Breitkopf, H. Gaitantzi, L. Ciuclan, P. Godoy, S. Dooley
Dept. of Medicine II, Molecular Alcohol Research in Gastroenterology, Faculty of
Medicine at Mannheim, University of Heidelberg, Germany
Introduction: TGF-β levels are high in fibrotic livers and its' signaling is induced in
liver cells of patients with alcoholic organ damage. We analyzed if TGF-β is directly
involved in the adverse actions of alcohol and if ethanol and TGF-β act independently
or via connected pathways in hepatocytes.
Methods: Ethanol and/or TGF-β target genes were identified in hepatocytes using
DNA microarrays, by RT- or real-time PCR and by Western blot analyses. Cell death
was evaluated with LDH assay, apoptosis and necrosis by a quantitative kit and
Caspase-Western blots.
Results: Ethanol and TGF-β, both display a dose-dependent cytotoxic effect on
hepatocytes shown by release of lactate dehydrogenase (LDH). Treatment with
ethanol and TGF-β together significantly increased cell death compared to each
treatment alone. Furthermore this combination led to clearly detectable apoptosis
which was under our experimental conditions only very low for TGF-β alone and
absent after ethanol treatment. The pro-fibrogenic mediator CTGF is induced by
TGF-β but not by ethanol and was superinduced by both together. We further
investigated expressions of other known TGF-β and/or Ethanol target genes and
found similar superinductions by both substances together. We are currently
investigating the underlying signal transduction pathway(s) that regulate these
effects.
Discussion/Conclusion: While ethanol alone mainly leads to necrosis of
hepatocytes and TGF-β alone causes only mild apoptosis both together lead to
stronger apoptosis and superinduction of a diverse set of target genes including profibrogenic ones like CTGF. These findings imply that there exists intracellular crosstalk between Ethanol and TGF-β induced pathways most likely contributing to
enhanced organ damage.
13
Cholestasis but not steatosis potentiates liver injury by
activated Kupffer cells: Evidence for a role for cxyclooxygenase-2
M. Bystron, C.J. Steib, J.M. Härtl, F. Roggel, M. Bilzer, A.L.Gerbes
Medizinische Klinik und Poliklinik II, Klinikum Großhadern, Ludwig-MaximiliansUniversität München, Munich, Germany
Introduction: A growing body of evidence shows a central role for endotoxin in
development of chronic liver disease. Relation between hepatotoxic effects of KCactivation and liver-disaease is unclear. We therefore compared KC-activation in
normal to steatotic and cholestatic livers.
Methods: Cholestasis was induced by bile-duct-ligation (BDL). Livers were nonrecirculating perfused with Krebs-Henseleit-buffer 2 weeks after BDL or shamoperation (each n = 5). To investigate effects of steatosis, livers of 14 weeks old male
Zucker-rats were perfused (n = 5). KC were activated after 40 minutes by infusion of
zymosan A (150 µg/ml and 75 µg/ml) for 6 minutes. In a subgroup of BDL-animals,
the cyclooxygenase-2 inhibitor Dup 697 (1 µM) was infused. LDH-efflux was
measured, liver tissue was analysed by HE- and ED2-staining.
Results (mean x ± SD, LDH-efflux in mU/min x g liver): Activation of KC by zymosan
resulted in a significant increase (p < 0.05) of LDH-efflux from 3 ± 1 to 60 ± 14 in
sham-operated animals. LDH-efflux reached only 7 ± 3 in obese Zucker-rats. Fibrotic
livers from BDL rats responded with a significant higher LDH-efflux (637 ± 215) upon
KC-activation. Even half the Zymosan concentration increased LDH-efflux to 260 ±
97. The different effects of KC-activation in the groups were correlated with KCdensity: 454 ± 16 ED2+KC/10 HPF in normal liver tissue, 320 ± 40 ED2+KC/10 HPF in
fatty livers and 792 ± 60 ED2+KC/10 HPF in cholestatic livers. Cyclooxygenase-2
inhibition significantly lowered LDH efflux in cholestatic livers after KC-activation to
160 ± 64.
Conclusions: Cholestasis, but not steatosis, potentiates liver injury by activated KC.
Aggravation of liver injury by cholestasis is related to an increase in KC density and
dependent on cyclooxygenase-2.
14
Expression of C-kit in liver after partial hepatectomy in rats
Gazizov IM, Kaligin MS, Andreeva DI, Pevnev GO, Burganova GR, Gumerova AA,
Kiassov AP
Kazan State Medical University, Kazan, Russian Federation
Introduction: Unless there are a lot of articles in the field of liver regeneration,
identification of liver stem cell is still controversial. One of the recent stem cell
markers is C-kit-receptor to stem cell factor. In our research we studied activation of
stem cell compartment in liver by detection of cells expressing C-kit after partial
hepatectomy in rats.
Methods: We performed partial hepatectomy in rat by method described by Higgins
and Anderson. Histological sections of liver after 1, 2, 3, 5, 7 posthepatectomy days
were studied immunohistochemically with antibodies against C-kit.
Results: Expression of C-kit started in hepatocytes after 1 day and persisted until 8th
posthepatectomy day. It had 3 patterns of expression: on one cell, on group of cells
and on layer of hepatocytes surrounding central vein. Weak expression of C-kit on
sinusoidal cells was detectable after 1 day and it increased gradually until 6th day.
Our previous results showed activation and increase of perisinusoidal cells after
partial hepatectomy. After 5 days in liver parenchyma we also found rare cells with
intermediate phenotype between hepatocytes and sinusoidal cells. These cells
resembled small hepatocytes with high distribution of C-kit which was comparable
with sinusoidal cells.
Discussion/Conclusion: Our results suggest that in liver after partial hepatectomy
characteristics of stem cells are possessed by hepatocytes and some sinusoidal
cells.
15
Perisinusoidal cells after partial hepatectomy in rats
Gazizov IM, Kaligin MS, Andreeva DI, Pevnev GO, Burganova GR, Gumerova AA,
Kiassov AP
Kazan Medical State University, Kazan, Russian Federation
Introduction: Perisinusoidal cells are type of nonparenchymal liver cells and can be
in activated and nonactivated condition. The purpose of our investigation was
studying of phenotypical changes of perisinusoidal cells after partial hepatectomy in
rats.
Methods: We performed partial hepatectomy in rat by method described by Higgins
and Anderson. Histological sections of liver after 1, 2, 3, 5, 7 posthepatectomy days
were studied immunohistochemically with antibodies against desmin (marker of
perisinusoidal cells), α-SMA (α-smooth muscle actin – marker of myofibroblasts).
Results: Desmin expression showed that after 1st posthepatectomy day number of
perisinusoidal cells increases 6.4 times and their maximum is achieved after 3 days
(12 times more than in control). That increase in perisinusoidal cells number after 1st
posthepatectomy day can not be explained just by proliferation, because it happens
mostly after 3rd day. The only possible explanation is starting expression of desmin in
previously desmin-negative perisinusoidal cells and it can be accepted as their
activation. Our study of α-SMA expression did not identify any myofibroblast in
regenerating liver, so activation of perisinusoidal cells after partial hepatectomy is not
total and there is no fibrosis in liver.
Discussion/Conclusion: We suggest that activation and increase of perisinusoidal
cells after partial hepatectomy is needed for supply of hepatocytes regeneration and
for that process they have all possibilities: they synthesize Hepatocyte Growth
Factor, Stem Cell Factor, remodel intercellular matrix which is needed in early stages
of hepatocytes regeneration. At last, if perisinusoidal cells are regional stem cells –
their progeny can genrerate hepatocytes.
16
Hepatic overexpression of RhoA- and Rho-kinase and elevated
Rho-kinase activity in livers from rats with CCl4-induced
micronodular cirrhosis
Hennenberg M, Trebicka J, Eckhardt A, Sauerbruch T, Heller J
Department of Internal Medicine I, University of Bonn, Germany
Introduction: In cirrhosis, increased resistance of the intrahepatic vasculature and
intrahepatic hyperresponsiveness to vasoconstrictors contributes to portal
hypertension. We have previously shown that upregulation of RhoA and Rho-kinase
and a subsequently increased Rho-kinase activity critically participate in the
development of increased intrahepatic vascular resistance in rats with secondary
biliary cirrhosis after bile-duct occlusion (Gut 2006; 55 [9]). Moreover, Rho-kinase
activity is increased in livers from patients with alcohol-induced cirrhosis (Gut 2006;
55 [9]). In contractile cells, the small monomeric GTPase is activated by
vasoconstrictors through G-protein coupled receptors. RhoA in turn activates Rhokinase, which mediates vasocontraction via inhibition of myosin-light-chain
phosphatase. Here, we investigated the expression of RhoA and Rho-kinase, as well
as Rho-kinase activity in livers of rats with CCl4-induced micronodular cirrhosis.
Methods: Micronodular cirrhosis in male rats was induced by chronic application of
CCl4 (by inhalation). Livers from cirrhotic rats and non-cirrhotic controls were shockfrozen and homogenized. Expressions of RhoA and Rho-kinase were studied by
Western-blot analysis of liver homogenates. Rho-kinase activity in liver homogenates
was assessed as phosphorylation of the Rho-kinase substrate, moesin (Thr-558), by
Western-blot analysis with a site- and phosphospecific antibody.
Results: Expression of RhoA and Rho-kinase are upregulated in livers from CCl4rats, compared to livers from non-cirrhotic rats. Levels of phospho-moesin (Thr-558)
are increased in homogenates of cirrhotic livers, compared to non-cirrhotic controls.
Discussion/Conclusion: Upregulation of RhoA and Rho-kinase expression, and
increased Rho-kinase activity and Rho-kinase-mediated contraction might contribute
to increased intrahepatic vascular resistance in cirrhosis of different etiologies.
17
The association of intestinal iNOS expression and oxidative
stress in the intestine in experimental cirrhosis
1
Mas MR, 2Savas B, 1Ocal R, 1Tasci I, 2Ensari A, 1Comert B, 1Yilmaz S, 1Isik AT,
3
Mas N, 2Erden E, 4Uzunalimoglu O
1
Gulhane School of Medicine, Department of Internal Medicine, Ankara, Turkey
2
Ankara University, Department of Pathology, Ankara, Turkey
3
Baskent University, Department of Anatomy, Ankara, Turkey
4
Turkish Hepatology Foundation, Ankara, Turkey
Introduction: In cirrhosis, increased portal pressure results in generation
proinflammatory cytokines, reactive oxygen species and nitric oxide leading to
alterations in intestinal barrier function. In this study, the association of inducible nitric
oxide synthase (iNOS) expression and oxidative stress in the intestinal wall as well
as the changes after administration of a selective iNOS inhibitor were examined in
cirrhotic rats.
Methods: Ninety male Sprague Dawley rats were injected subcutaneous (SC) three
times a week with CCl4 for 12 weeks. Five out of 33 surviving animals were tested for
the establishment of cirrhosis. The remaining 28 rats were divided into two groups.
Group 1 received SC salin and group 2 (S-methylisothiourea [SMT] (5 mg/kg/day) for
four weeks. Caecum tissue samples were tested for iNOS expression and oxidative
stress markers at the end of treatment.
Results: SMT treated group displayed significant improvements in inflammation
lamina propria (2.14 ± 0.36 vs. 2.86 ± 0.36, p < 0.002) and iNOS expression in
epithelium (2.07 ± 0.27 vs. 3.00 ± 0, p < 0.001). The degree of inflammation in lamina
propria and epithelium were in strong correlation with caecum tissue with
malondialdehyde (MDA) levels (r = 0.730, r = 0.785, respectively).
Discussion/Conclusion: Caecum iNOS expression and MDA levels are markedly
increased in cirrhotic rats. SMT treatment successfully decreased intestinal İNOS
expression. The results strengthen the role of altered iNOS activity in intestinal
barrier function in cirrhosis. The clinical effectiveness of iNOS inhibition in prevention
of bacterial translocation should be investigated.
18
Large-conductance calcium-activated potassium
modulate vascular tone in experimental cirrhosis
channels
Aina Rodríguez-Vilarrupla, Mariona Graupera, Vasilica Matei, Ramón Bataller,
Juan G Abraldes, Jaume Bosch, Joan-Carles García-Pagán
Unitat d’Hémodinàmica Hepática, Institut de Malalties Digestives, Hospital Clίnic,
Fundació IDIBAPS, Barcelona, Spain
Background: Portal hypertension is characterized by an increase in resistance
partially due to reversible functional alterations, including a deficient response of the
hepatic vascular bed to vasodilators and an exaggerated response to
vasoconstrictors. Large conductance calcium-activated potassium channels (BKCa)
regulate vascular tone in different vascular systems. Moreover, activated hepatic
stellate cells (HSC), a contractile cell-type that regulate hepatic blood flow in
cirrhosis, contain BKCa channels.
Aim: to evaluate the role of BKCa channels in the regulation of vascular tone in
control and cirrhotic rat livers.
Methods: Liver cirrhosis was induced by chronic carbon tetrachloride (CCl4)
administration. Changes in portal perfusion pressure (PP) in response to
methoxamine (Mtx), an α1-adrenergic agonist, were assessed in the presence of:
Iberiotoxin (Ibtx; a BKCa channel blocker), NS1619 (a BKCa channel opener), Ibtx plus
the nitric oxide (NO) synthase inhibitor, NG-nitro-L-arginine (L-NNA), or L-NNA alone.
BKCa mRNA expression was assessed in liver homogenates, and protein expression
was measured in HSC isolated from both control and cirrhotic livers
Results: In cirrhotic livers, Ibtx increased baseline PP and exacerbated Mtx-induced
increase in PP, whereas in control livers these effects were not found. Conversely,
NS1619 preincubation induced a mild decrease of the baseline PP and attenuated
the hyperresponse to Mtx of cirrhotic livers. Importantly, Ibtx had no effects after NO
inhibition by L-NNA. Cirrhotic livers showed an up-regulation of the gene encoding αsubunit of BKCa, and cultured HSCs had increased protein expression.
Conclusions: BKCa channels are overexpressed in cirrhotic livers and might
represent a compensatory mechanism modulating the increased hepatic vascular
tone of cirrhosis. Opening of these channels seems to be mainly mediated by NO.
19
Kupffer cell (KC) activation in fibrotic livers by β-glycans: The
COX-2 inhibitor Parecoxib reduces portal pressure (PP)
increase in vivo
Steib CJ, Bystron M, Härtl JM, Opelz C, Liss I, Göke B, Gerbes AL
Medizinische Klinik II, Klinikum Großhadern, Ludwig-Maximilians Universität
München, Munich, Germany
Background: We investigated whether COX-2 is involved in the generation of
thromboxane A2 in fibrotic livers and whether COX-2 inhibition attenuates PP
increase following KC activation in fibrotic livers in vivo.
Methods: Fibrosis was induced by bile duct ligation (BDL). 2 weeks after BDL or
sham operation livers were isolated perfused or PP, mean arterial pressure (MAP)
and heart rate were monitored in vivo. KC were activated in isolated livers by
Zymosan A (75 µg/ml) with or without different COX-inhibitors. A stock solution of
32 mg Zymosan A/ml was infused (100 µl/min) over 6 minutes in the in vivo setting
either with acute administration of vehicle or Parecoxib (5 mg/kg). Thromboxane B2
(TxB2) was measured in effluent perfusate by ELISA.
Results: (mean x ± SD, PP and MAP in mmHg, TxB2 in pg/min x g liver) KC
activation in the perfused liver resulted in a more significant increase of PP in BDL
(∆7.9 ± 1.2) than in sham operated animals (∆3.2 ± 0.4) paralleled by augmentation
of TxB2 efflux (sham: 1284 ± 382, BDL: 6769 ± 2825). COX-1 inhibition by SC 560
(0.4 µM) reduced significantly increase of PP in sham operated (∆1.5 ± 0.3) and in
BDL animals (∆4.7 ± 0.7) ongoing with reduced TxB2 efflux (sham: 120 ± 43, BDL:
738 ± 78). In contrast, COX-2 inhibition by Dup 697 (1.0 µM) reduced increase of PP
(∆4.9 ± 1.1) and TxB2 (2029 ± 329) in BDL, but not in sham operated rats (PP ∆3.1 ±
0.5, TxB2 1139 ± 269). KC activation in vivo also increased PP in sham operated
(from 6.1 ± 1.3 to 12.2 ± 2.7) and in BDL animals (from 11.4 ± 1.1 to 19.5 ± 2.8)
following vehicle administration. Interestingly, the COX-2 inhibitor Parecoxib reduced
increase of PP about 50%. Following KC activation MAP decreased in sham
operated animals from 107 ± 12 to 52 ± 10, in BDL animals from 113 ± 14 to 58 ± 9
and by Parecoxib treatment from 115 ± 12 to 73 ± 10, while heart rate was stable.
Conclusion: COX-2 inhibition attenuates the thromboxane A2 generation in fibrotic
livers, but not in normal livers. Therefore administration of Parecoxib, a COX-2
inhibitor, improves portal hypertension in vivo following KC activation in portal
hypertensive fibrotic livers.
20
Stem cell properties of hepatic stellate cells
Gumerova A.A., Kiassov A.P., Abdulkhakov S.R., Kaligin M.S., Gazizov I.M.,
Andreeva D.I.
Kazan Medical Unversity, Kazan, Russian Federation
The existing theory of liver development is considered that hepatocytes develop from
duodenal epithelial cells. But it was shown that hepatocytes can develop from
hematopoietic and mesenchymal stem cell. We immunohistochemically studied the
peculiarities of expression of wide range of phenotypic markers as well as the
markers of different cell types’ differentiation and stem cell markers in human (after
medical abortions) and rat prenatal development and liver regeneration.
We found out that cells of ductal plate of human emryons don’t express hepatocyte
specific antigen and have different from hepatocytes phenotype. Bile duct formation
during development and regeneration occurs due to migration and proliferation of
cholangiocytes, but not because of hepatoblasts to cholangiocytes differentiation. So
phenotype and differentiation peculiarities of hepatocytes and cholangiocytes are
quite different and they can have different sources of development.
At the 4–7 weeks of gestation human mesenchymal cells in the place of liver laying
are desmin+/cytokeratin 18+/cytokeratin 19+. Moreover, in desmin+ mesenchymal
cells antiapoptotic protein Bcl-2 is present and it is typical for the progenitor cells
compartment. Desmin is present in hepatoblasts. Analogous results were obtained in
rats. Desmin is marker of Hepatic Stellate Cells (HSC), so these cells can be source
of hepatocytes development.
Our experiments on organotipic embryonal rat liver culturing and on pure adult rat
HSC population culturing confirmed our hypothesis: in vitro HSC cells acquire the
phenotype of hepatoblasts/hepatocytes – so we‘re the first who described
mesenchymo-epithelial transfdifferentiaton of HSC. So we suppose that HSC can be
regional liver stem cells.
21
Effects of pentoxifylline on established liver fibrosis and
hepatic stellate cells after bile duct ligation in rat
1
Mas MR, 1Ocal R, 2Bilgin G, 3Karslioglu Y, 1Comert B, 1OzsariI L, 1Tasci I, 4Mas N,
1
Isik AT, 5Uzunalimoglu O
1
Gulhane School of Medicine, Department of Internal Medicine, Etlik 06018 Ankara,
Turkey
2
Gulhane School of Medicine, Department of Surgery, Etlik 06018 Ankara, Turkey
3
Gulhane School of Medicine, Department of Pathology, Etlik 06018 Ankara, Turkey
4
5
Introduction: Liver fibrosis is characterized by overproduction and accumulation of
extracellular matrix proteins in the sinusoids. Modulation of hepatic stellate cell (HSC)
transdifferentiation may be an effective target to prevent liver fibrosis. Pentoxifylline,
a xanthine derivative fosfodiesterase inhibitor, was reported to prevent fibrous tissue
production. However, this effect has been questioned by some authors. The present
study searches its effects on established liver fibrosis and HSCs in an animal model.
Methods: Forty female Spraque Dawley rats underwent bile duct ligation. Four out of
36 survived animals were killed after four weeks and establishment of liver fibrosis
was confirmed. The remaining 32 rats were randomized into two groups. Group I
(n = 16) was treated with oral pentoxifylline (50 mg/kg/day) for four weeks. Group II
(n = 16) did not receive any treatment. After the animals were killed, liver fibrosis,
liver tissue hydroxyproline, ductal proliferation and the number of activated stellate
cells were determined.
Results: Tissue hydroxyproline content and the degree ductal proliferation were the
same in the two groups [1.71 ± 0.22 vs. 1.88 ± 0.09, p > 0.05, and 3 (2.5–4) vs.
4 (4–4), p > 0.05, respectively]. Serum ALT levels and semiquantitative fibrosis were
lower in group I (365.6 ± 33.2 vs. 406.7 ± 57.7, p < 0.05, and 44.5 ± 16.7 vs. 67.2 ±
11.8, p < 0.003). alfa-SMA (+) HSC counts were also lower in the pentoxifylline
treated group (15.56 ± 4.22 vs. 29.56 ± 2.56, p < 0.001).
Discussion/Conclusion: Pentoxifylline improved histological parameters of rat liver
fibrosis due to bile duct ligation. It showed no significant effect of tissue
hydroxyproline content. Histological improvement was associated with the amount of
activated HSCs.
22
CD95 ligand induces proliferation and CD95-tyrosine nitration
in quiescent rat hepatic stellate cells
Roland Reinehr, Annika Sommerfeld, Stephan Becker and Dieter Häussinger
Clinic for Gastroenterology, Hepatology and Infectiology, University Hospital
Düsseldorf, Moorenstr. 5, D-40225 Düsseldorf, Germany
Introduction: Quiescent hepatic stellate cells (HSC) are fairly resistant to CD95
ligand (CD95L)-induced apoptotic cell death, while apoptosis upon CD95L-treatment
has been observed in activated HSC/myofibroblasts when cycloheximide (CHX) was
co-administered (Cariers et al., Cell Physiol Biochem 2002). Recently, quiescent
HSC have been reported to express CD133 and to represent a hepatic progenitor
cell compartment (Kordes et al., BBRC 2007).
Methods: Rat hepatic stellate cells were either cultivated for 1–2 days (quiescent
HSC) or 10–14 days (activated HSC) on plastic culture dishes. Activation of Src,
EGFR, Erks and JNKs were measured by W. blot using respective phospho-specific
antibodies. CD95 was immunoprecipitated and detected for CD95/EGFR-association,
CD95-tyrosine phosphorylation, CD95-tyrosine nitration and DISC-formation using
W. blot technique. HSC proliferation and apoptosis were analyzed by BrdUincorporation measurements and TUNEL-staining, respectively.
Results: In quiescent HSC, CD95L leads to an activation of c-Src, EGFR and Erks
which was sensitive to inhibitors of Src-kinases (PP-2), sheddases (GM6001) and to
EGF-neutralizing antibodies, while no JNK-activation occurred. Furthermore, an
increased bromodesoxyuridine incorporation was observed in quiescent HSC after
CD95L-addition, while no pro-apoptotic CD95-activation became detectable with
respect to CD95/EGFR-association, CD95-tyrosine phosphorylation, DISC-formation,
TUNEL-staining and PARP-cleavage. In addition, in quiescent HSC CD95L induced
CD95-tyrosine nitration in a time- and dose-dependent manner, which was previously
shown to inhibit CD95-tyrosine phosphorylation and thereby exhibits anti-apoptotic
properties. In contrast, in activated HSC no CD95-tyrosine nitration upon CD95L
administration was observed, whereas there was still activation of the EGFR. When
CD95L and CHX were co-administered, a sustained JNK-activation occurred which
allowed for CD95/EGFR-association, subsequent CD95-tyrosine phosphorylation,
DISC formation and apoptotic cell death.
Discussion/Conclusion: In quiescent HSC, CD95L is a mitogen through a liganddependent activation of the EGFR which requires Src-kinase- and sheddase-activity.
Simultaneously, apoptotic signaling is prevented by CD95L-induced CD95-tyrosine
nitration, which may contribute to the recently described progenitor cell properties of
quiescent HSC and which is lost after differentiation to activated HSC/myofibroblasts.
23
TGF-β signaling in hepatocytes promotes EMT and liver
fibrogenesis, whereas hepatocyte-specific expression of TGF-β
antagonist Smad7 suffices to protect from liver damage and
fibrogenesis
Honglei Weng1, Loredana Ciuclan1, Patricio Godoy1, Stephan Kanzler2,
Rainer Heuchel3, Elke Ueberham4, Rolf Gebhardt4, Jafar Hamzavi1 Peter Mertens
and Katja Breitkopf1, Steven Dooley1
1
Dept. of Medicine II, Molecular Alcohol Research in Gastroenterology, University
Hospital at Mannheim, University of Heidelberg, Germany, 21. Medical Clinic, Univ.
Hospital Mainz, Germany, 3Ludwig Institute for Cancer Research, Uppsala, Sweden,
4
Institute for Biochemistry, University of Leipzig, Germany
Chronic liver intoxication results in overrepresentation of TGF-β in the liver, which
induces activation of hepatic stellate cells, deposition of extracellular matrix and
hepatocyte (HC) apoptosis. Since TGF-β signal transduction and its biological
responses are highly complex, we endeavored a Systems Biology approach to
understand the diverse observations for TGF-β and Smads in liver cells. Apoptosis
assays revealed that TGF-β dependent apoptosis is only an inferior function of this
cytokine on hepatocytes.
Using a top down approach by Affymetrix array gene expression profiling in mouse
HC stimulated with 5 ng/ml TGF-β1, we identified target genes critically involved in
epithelial mesenchymal transition (EMT; snail, E-cadherin, fibronectin) and
fibrogenesis (type I collagen, TIMP-1, CTGF). According to this, TGF-β enhanced
morphology changes of HCs in culture from typical honeycomb to fibroblastic shape,
indicating profibrogenic activity of TGF-β on hepatocytes. To confirm this finding in
vivo, we established a mouse model, expressing TGF-β antagonist Smad7 under the
control of the HC specific C-reactive protein promoter. CCl4 treatment of animals
triggered to overexpress Smad7 in HCs by LPS lack activated TGF-β signaling,
accumulate less collagen and display improved values of damage and fibrosis as
compared to controls.
The data indicate that HCs undergo TGF-β dependent EMT like phenotypic changes
in culture and produce profibrogenic proteins. Furthermore, blocking TGF-β signaling
specifically in this cell type is sufficient to abrogate fibrogenesis.
24
The role of ADH and ALDH gene polymorphism in alcohol liver
cirrhosis
Halina Cichoż-Lach, Krzysztof Celiński, Jadwiga Partycka, Maria Słomka
Department of Gastroenterology, Medical University of Lublin, Poland
Alcohol abuse poses a serious medical problem, social and economic. Alcohol
disease of the liver is estimated to occur in 77% individuals who abuse alcohol
consumption. Genetic polymorphism of enzymes involved in alcohol metabolism
plays a relevant role in etiopathogenesis of alcoholism and alcohol liver disease.
The aim of the present study was to find in the Polish population the ADH2, ADH3
and ALDH2 genotypes, which are likely to be responsible for higher susceptibility to
alcohol liver cirrhosis. The ADH2, ADH3 and ALDH2 genotype and alleles
frequencies were examined in 202 patients: 77 with alcoholic liver cirrhosis, 64
abusing alcohol but without damage to gastrointestinal organs –“healthy drinkers”
and 61 nondrinkers as a control group. Genotyping of the ADH2, ADH3 and ALDH2
was performed using polymerase chain reaction -restriction fragment length
polymorphism methods on white cell DNA.
The genotype ADH3*1/ADH3*1 and allele ADH3*1 were found to be significantly
more frequent in alcohol abusers compared to non-drinkers. Frequency of ADH3*1
allele in alcohol liver cirrhosis group was 62.8%, and ADH3*1/ADH3*1 genotype was
observed in 45.4% and was significantly higher than in the control group. The
differences between of the group of patients who abuse alcohol were not statistically
significant. In the group of nondrinkers ADH2*2 and ADH3*2 alleles were more
frequent in comparison to the alcohol liver cirrhosis patients and alcohol addicts. All
examined patients were ALDH2*1/ALDH2*1 homozygotic.
Conclusion: Our studies suggest that in the Polish population examined ADH3*1
allele and ADH3*1/ADH3*1 genotype favor developing alcoholism and alcohol liver
cirrhosis. However ADH2*2 allele is likely to protect against them. Genetic
polymorphism of ALDH2 shows no correlation with alcohol addiction or alcohol
cirrhosis.
25
Effect of copy number repeats of the CCL3L1 gene one
susceptibility to HCV infection and fibro-inflammatory
response
Frank Grünhage1, Olav Gressner1, Hilde Keppeler1, H.E. Wasmuth2, M. Odenthal3,
U. Drebber3, H.P. Dienes3, C. Hellerbrand4, Tilman Sauerbruch1, Frank Lammert1
1
Department of Internal Medicine I University Hospital Bonn, Rheinische FriedrichWilhelms-Universität Bonn, Germany
2
Department of Medicine III, University Hospital Aachen, Aachen University, Aachen,
Germany
3
Institute of Pathology, University of Cologne, Cologne, Germany
4
Department of Internal Medicine I, University Regensburg, Regensburg, Germany
Introduction: Variation in the copy numbers of the CCL3L1 gene has recently been
associated with susceptibility to HIV infection (Gonzales et al.; Science 307:
1434–1440). CCL3L1 is a potent ligand for CCR5 which in turn plays a central role in
anti-viral lymphocyte activity. Variation in the CCR5 gene has already been
associated with differences in treatment response in HCV therapy. Hence, variation
in the gene dose of CCL3L1 by variation in the copy number repeats may also
contribute to differences the inflammatory response in HCV and general susceptibility
to the infection. Thus, we compared (1) the copy number distribution between
patients with HCV infection and controls and (2) tested for association with
inflammatory grade and fibrosis stage in HCV patients.
Patients and methods: Patients with chronic HCV infection and uninfected controls
were included in this study. All participants gave written informed consent. Copy
number repeats of the CCL3L1 gene were determined using a real time, fluorescent
dye labelled PCR method analogue to the publication of Gonzales et al. (Science
2005; 307 [5714]: 1434–1440 ). In short, the genomic sequence of the CCL3L1 gene
is amplified with gene specific primers and a fluorescent dye labelled probe using a
real time PCR based system. Melting curves are compared in relation to a house
keeping gene (beta-globulin), and a probe known to have exactly two copies of the
CCL3L1 gene. Copy number repeats are then calculated analogue to the methods of
Gonzales et al. and compared between the different groups using non-parametric
tests (Man-Whitney).
Results: We included 255 HCV patients. Men were over-represented in the HCV
cohort (70.2%). Overall copy number distribution among patients and controls was in
line with previously published data for Non-Africans (mean 2.7 copies). Copy number
frequency distribution differed significantly between HCV patients and controls
(p = 0.036). However, no difference in copy number frequency distribution was
detected between patients with different grades/stages of inflammation / fibrosis.
Conclusion: Non-infected controls carried slightly more copy numbers than HCV
patients. Thus, susceptibility to HCV infection may be influenced by variation in copy
numbers of the CCL3L1 gene. However, inflammation grade and fibrosis stage in
HCV is unswayed by copy numbers in this gene.
26
Genome wide scans for fibrosis susceptibility genes in
experimental crosses of inbred mouse strains
Rabea Hall, Sonja Hillebrandt, Katrin Hochrath, Frank Grünhage, Tilman Sauerbruch
and Frank Lammert
Department of Internal Medicine I, University Hospital Bonn, University of Bonn,
Bonn, Germany
Introduction: Recently we identified quantitative trait loci (QTLs) that determine
susceptibility to liver fibrosis in experimental crosses between fibrosis-susceptible
and resistant inbred mouse strains (Hillebrandt et al. 2002, 2005). For these
analyses, we modelled the effects of single QTLs only. Our aim now was to assess
additive and interacting effects of multiple QTLs for liver fibrosis.
Methods: We intercrossed F1 hybrids of the susceptible strain BALB/cJ and the
resistant strain FVB/NJ to obtain 294 F2 progeny. To induce liver fibrosis, we
challenged the mice twice weekly with CCl4 (1 mg/kg i.p. for 6 wks). For phenotypic
characterization, we determined histological stages of liver fibrosis and measured
hepatic collagen contents. We performed one QTL genome scans with 141
microsatellite markers, followed by simultaneous genome scans of pair wise
interactions, as implemented in R/qtl. LOD thresholds were determined by
permutation tests.
Results: The F2 progeny show a normal distribution of collagen concentrations,
consistent with polygenic inheritance of fibrosis susceptibility (mean fibrosis stage
1.8 ± 0.02; mean collagen content 322 ± 122 µg/g). In addition to our previously
described fibrogenic QTLs (Hfib1 and Hfib2) on chromosomes 2 and 15, one QTL
genome scans show peak linkage for collagen accumulation on chromosomes 1, 2,
and 8. In the two QTL scans, we identified additive gene-gene interactions of the loci
on chromosomes 2 and 8. Analyzing gene-gender interaction, we observed
differences between male and female progeny.
Discussion/Conclusion: This genome analysis indicates gene-gene and genegender interactions during liver fibrogenesis and provides a basic experimental
framework to model networks of genes during chronic liver injury.
27
Immunogenetic factors in the development of HCV-related liver
cirrhosis
A.O. Romanov, T.V. Belyaeva, E.V. Esaulenko
Pavlov State Medical University, Saint-Petersburg, Russia
Introduction: Persistent hepatitis C virus infection ranks as the leading cause of
cirrhosis in the world. Progression to cirrhosis in HCV infected patients could be a
genetically complex and its pathogenesis is likely to involve an interaction between
multiple alleles located on different locuses. In the present study, joint effect of the
functional polymorphisms in the RNASEL, MBL, CTLA-4, MCP-1, TGF-beta1 genes
on the development HCV-related liver cirrhosis has been investigated.
Methods: Polymorphisms +1385G/A RNASEL, +230G/A MBL, -2518A/G MCP-1,
+49A/G CTLA-4, +29T/C TGF-beta1 were analyzed by PCR-RFLP techniques in
76 patients with decompensated liver cirrhosis due to chronic HCV infection and
88 patients with chronic hepatitis C. The control group comprised of 122 healthy,
unrelated blood donors.
Results: The rate of composite wild-type genotype +1385 GG RNASEL/+230 GG
MBL/-2518 AA MCP-1/+49 AA CTLA-4/+29 TT TGF-beta1 was 1% (1) in the HCV
infected patients with cirrhosis, significantly lower than in the patients without
cirrhosis 12% (11), while that of healthy controls was between the two groups 4% (5).
In HCV infected patients, those with cirrhosis more frequently had composite variant
genotypes +1385 GA(AA) RNASEL/+230 GA(AA) MBL/-2518 AG(GG) MCP-1/+49
AG(GG) CTLA-4/+29 TC(CC) TGF-beta1 when compared with those bearing
composite wild-type genotype (5 [6%] versus 0 [0%]; p = 0.0009). There were no
differences in frequencies of these genotypes between HCV patients with or without
cirrhosis and healthy controls (p = 0.10 and p = 0.13, respectively).
Discussion/Conclusion: These results indicate that inheritance of the composite
variant genotypes, which appear to affect the immunoregulatory proteins production,
is thus a potential predictor of genetic susceptibility to the development of HCVrelated liver cirrhosis that may prove useful in determining which patients are at
elevated risk for developing of decompensated cirrhosis due to chronic HCV infection
to estimate the need and urgency of therapy.
28
Hypersecretory haplotype of interleukin-1
increases the risk of alcoholic liver cirrhosis
gene
cluster
Sperl J1, Petrasek J1, Reznakova S1, Hubacek JA1, Stickel F2, Trunecka P1, Berg T3,
Spicak J1, Jirsa M1
1
Institute for Clinical and Experimental Medicine, Prague, the Czech Republic
2
Department of Clinical Pharmacology, University of Berne, Switzerland
3
Department of Medicine, Charité - Campus Virchow, Humboldt University Berlin,
Germany
Introduction: Interleukin 1-beta (IL-1β) is an important pro-inflammatory cytokine.
The objective of the study was to test the hypothesis that the IL-1B promoter
polymorphism -31C/T known to increase the expression of IL-1β, and the pentaallelic 86-bp tandem repeat in intron 2 of IL-1 receptor antagonist (IL-1RN) known to
increase the production of IL-1β are associated with susceptibility to alcoholic liver
cirrhosis (ALC).
Methods: Allele frequencies for both mentioned polymorphisms were determined in
198 heavy drinkers with clinically proven alcoholic cirrhosis and 246 non-abstaining
age and sex matched healthy controls. The principle positive finding was further
validated in an independent cohort of 173 patients with ALC.
Results: The haplotype [IL-1RN*2/*2; IL-1B -31T+] was associated with an increased
risk of ALC in both the primary (odds ratio 4.9, 95% CI 1.4–17.8, P = 0.016) and the
second confirmatory cohort (odds ratio 8.0, 95% CI 2.2–28.4, P = 0.001). The
population attributable fraction that estimates the proportion of ALC in the study
population that is attributable to the haplotype was 4.9% (95% CI 1.3–8.6) in the pilot
cohort and 7.5% (95% CI 3.1–12.0) in the confirmatory cohort.
Conclusion: Our observation is in line with the previous studies indicating that
synergistic action of the IL-1B -31T and IL-1RN*2 polymorphisms are responsible for
IL-1β high secretory phenotype. Such phenotype promotes inflammation and fibrosis
and explains the molecular background of the increased risk of ALC in homozygotes
for the IL-1RN*2 allele that are at least heterozygous for the IL-1B -31T allele.
29
Beta-glucosidase 2 (GBA2) deficiency leads to zonal glucosylceramide accumulation in the liver
Yildiz Y.¹, Matern H.², Weber S.¹, Matern S.², Russell D.W.³, Sauerbruch T.¹,
Lammert F.¹
¹Department of Internal Medicine I, University of Bonn, Germany ²Department of
Internal Medicine III, University of Aachen, Germany, ³Department of Molecular
Genetics, University of Texas Southwestern Medical Center at Dallas, TX, USA
Background and aims: The enzyme beta-glucosidase 2 (GBA2) has been purified
and characterized from human liver microsomes. GBA2 is highly conserved and
ubiquitously present, and was known to hydrolyze bile acid 3-O-glucosides. To gain
insight into the pathobiological functions of this enzyme and its substrates, we
recently generated Gba2 knockout mice (J Clin Invest 2006; 116: 2985–94).
Methods: Vector construction for Gba2 -/- generation, ES cell transformation,
screening and blastocyst injections were carried out according to standard protocols.
GBA2 protein expression was determined by immunoblot and immunofluorescence
analyses. Glycolipids were extracted from different tissues, and characterized by
thin-layer chromatography and mass spectrometry. Liver sections of Gba2 -/knockout mice and wild-type controls were characterized immunohistochemically,
and tunel assays were performed to quantify apoptosis in the liver.
Results: Gba2 -/- mice have a normal life span with no profound defect in bile acid
metabolism, since bile acid pool sizes and secretion rates do not differ. In contrast,
Gba2 -/- mice display impaired male fertility with globozoospermia, and accumulate
glucosylceramide in different tissues. In liver, glucosylceramide accumulation
predominates in the perivenous zone (zone 3). Consistent with zonal heterogeneity,
highest GBA2 expression is also observed around the central veins. In addition, we
observe significantly increased apoptosis in GBA2 deficient livers.
Conclusions: We conclude that GBA2 is a critical glucosidase that clevaes
glucosylceramide as endogenous compound in various tissues including liver. We
speculate that mutations of the orthologous GBA2 gene might contribute to
alterations of cell cycle, modulate cell signaling, and modify the phenotypes of
glucosylcerebrosid storage disorders such as Gaucher disease in humans.
30
Impact of peginterferon α-2b and ribavirin combined therapy on
serum gastrin and progastrin concentrations in Helicobacter
pylori (HP) infected patients with chronic hepatitis C (CHC)
W. Bielanski1, W.W. Mazur2; Z. Gonciarz3; M. Gonciarz3; S.J. Konturek1;
M. Magiera2; J.F. Rehfeld4
1
Department of Physiology, Jagiellonian University Medical College , Cracow, Poland
2
Department of Infectious Diseases, Silesian University Medical School, Chorzów,
Poland
3
Department of Internal Medicine, District Hospital No 5, Sosnowiec, Poland
4
Department of Clinical Chemistry, Rigshospitalet, Copenhagen, Denmark
Introduction: Leptin, seems to be also involved in hepatic fibrogenesis in pts with
CHC. It was reported that leptin serum levels decreased during antiviral therapy
(AVT) based on Peg-IFN and ribavirin. The aim of our study was to assess serum
gastrin and progastrin levels changes during and after AVT in pts with CHC in
relation to HP infection
Methods: 40 pts with CHC, elevated ALT and positive HCV-RNA (25M/15F, age 46 ±
13,4 yr., genotype 1 – 87,5%) were treated according to genotype for 24–48 weeks
with standard PegIFN α-2b plus ribavirin. Gastrin and progastrin levels were
determined at the baseline (BS), the end of treatment (ET) and the end of 6-months
follow-up (FU). HP status was assessed by capsulated 13C-UBT.
Results: The mean gastrin levels were significantly higher in Group I (HP-positive) in
comparison to Group II (HP-negative) at ET (18.6 pg/ml vs. 15.8 pg/ml; p = 0.032)
and FU (21.2 pg/ml vs. 13.8 pg/ml; p = 0.001). Progastrin serum levels decreased
significantly during AVT (BS: 480 pg/ml vs. ET: 422 pg/ml; p = 0,000) but returned to
the base levels after stopping therapy. The progastrin values did not differ between
Group I and II at BS and ET, but at FU the progastrin levels were significantly higher
in Group I vs. II (655 pg/ml vs. 417.5 pg/ml; p = 0.012).
Discussion/Conclusion: Combined IFN plus ribavirin therapy in pts with CHC
induces significant changes on serum gastrin and progastrin levels. The hormonal
outcomes after the same AVT in Hp infected CHC pts are different
31
Researches on the immunologic-inflammatory answer at the
patients chronically infected with the hepatitis virus C
Deac M1, Boitan M1, Mihaila R1, Fraticiu A2, Rezi EC2, Zaharie AV2, Olteanu A2,
Dascalu D2, Tiurean L2, Porr PJ1, Cipaian C2, Vulcu D1, Petrascu O1, Beca C2,
Boicean A1, Chicea L1, Orga-Dumitriu D1, Bidlo C2, Catana A2, Mihaila R3
1
Lucian Blaga University Sibiu, Faculty of Medicine, Romania
2
Departmental Clinical Hospital Sibiu, Sibiu, Romania
3
Public Health Authority Sibiu, Sibiu, Romania
Introduction: At the patients with chronic hepatitis C, there were observed values of
IL-6 which were significantly higher, compared with healthy people. Even more, it is
now discussing if basal levels of IL-8 could constitute a predictive factor for the
answer to the therapy with Interferon. We proposed ourselves to compare the serum
level of two markers of inflammation – CRP and IL-8 and to study their possible
correlations with the biochemical liver tests at the patients chronically infected with
the C virus.
Methods: We have studied a group formed by 33 patients which were hospitalized in
the medical departments of the County Clinical Hospital Sibiu, during 12.02–
26.03.2007. The group was divided into two: group A with 13 patients chronically
infected with the C virus, and group B with 20 patients without any liver or
inflammatory disease (control group). At both groups we have studied: age, BMI,
biochemical liver tests, cholesterol, triglycerides, IL-8, CRP. We have compared the
medium values at the two groups and we have analyzed the existence of any
correlation between the markers of inflammation and the biochemical liver tests. The
results were statistically analyzed, using the t Student test and the Pearson index of
correlation.
Results: The average age of the studied group was 42.12 ± 14.57 years. The gender
repartition was: 75.75% women and 24.25% men. The TGO level in the C virus
group was, in average, of 63.46 U/l compared with 16.05 U/l in the control group
(p < 0.001); the TGP level, was, in average, 73.23 U/l in the C virus group, compared
with 14.65 U/l in the control group (p < 0.001). The medium value of CRP in the
group A was 9.38 mg/l, compared with the group B, were it was only 3.89 mg/l, but
the difference was not statistically significant. The medium value of IL-8 was 170.25
pg/ml at the patients with the C virus, compared with only 12.46 pg/ml in the control
group (p < 0.01). There is an important correlation between the level of transaminasis
and CRP, at the patients with the C virus (r = 0.65, between TGO and CRP,
respectively r = 0.759 between TGP and CRP). Also, in the control group, there is a
tight correlation between the level of TGO and CRP (r = 0.504). In this group, there
isn’t such a tight correlation between TGP and CRP (r = 0.204). The IL-8 level is
correlating very well with the TGO level in the control group (r =0.542), but not in the
C virus group. The level of TGP does not correlates itself with the level of IL-8, in
none of the two groups.
Discussion/Conclusion: Compared with the control group, the serum level of IL-8 is
significantly higher at the patients chronically infected with the C virus, suggesting its
implication in the inflammatory process, but does not correlates itself with the liver
cytolysis. CRP does not vary significantly in the two groups, but is well correlating
with liver cytolysis, especially at those chronically infected with hepatitis virus C.
32
Detection of SEN virus in chronic hepatitis C virus infected
patients genotype 4 and sequencing in Egypt
Samia A. Girgis and Ahmed Ali Monis*
Clinical Pathology Department and *Internal Medicine Department, Faculty of
Medicine, Ain Shams University, Cairo, Egypt
Introduction: A novel virus, SEN virus (SENV), was recently discovered. Eight
different genotypes SENV (A–H) have been described, two of them SENV-D and H
have been assumed to be responsible for post-transfusion hepatitis in humans. The
pathogenic role of SENV in liver diseases is still controversial. Among patients with
acute and chronic hepatitis C virus (HCV) infection, 24–40% is coinfected with SENV.
The effect of SENV on the course of HCV infection and response to interferon (IFN)
treatment, as well as the effect of IFN on the clearance of SENV, are unknown.
Aim: The present study aims were to detect SENV in healthy blood donors and in
chronic HCV infected patients, to identify the influence of SENV coinfection on
clinical, virological criteria and response to combination therapy, IFN and ribavirin.
Methods: SENV-D and H genotypes were detected in serum samples from
50 patients with chronic hepatitis C virus (HCV) and from 60 control subjects of
healthy blood donors by polymerase chain reaction (PCR) followed by sequencing to
identify specificity. HCV virus antibodies, reverse transcription PCR (RT-PCR) and
genotyping were detected for diagnosis. Follow up was carried over by detection of
HCV viral load for all patients under interferon (IFN) and ribavirin therapy.
Results: Forty (80%) of 50 chronic HCV patients were positive for SENV DNA which
was highly statistically different from the 60 blood donors who were all negative for
SENV (p < 0.001). By sequencing, the specificity of genotype-specific PCR was
confirmed. There was a highly significant association between SENV and HCV
infection (p < 0.001). Among the patient group; 26 (52%) were infected with both
types D and H, 13 (26%) with SENV-H and one (2%) with SENV-D alone. The most
prevalent genotype in the patient group was SENV-H (39/50) (78%) followed by
SENV-D (27/50) (54%). All patients had HCV genotype 4. A highly statistically
significant difference was found between the patient and control groups as regards
history of blood transfusion, surgical operation, clinical examination, laboratory
investigations including transaminases (ALT and AST), HCV-RNA and SENV-DNA
(p < 0.001). While no statistical significance was found between the HCV infection
alone (10/50) and coinfected patients (p > 0.05). Hepatosplenomegaly, anaemia and
higher bilirubin levels were more detected among coinfected patients with HCV and
SENV (p < 0.05). HCV Response rate to combination therapy with interferon and
ribavirin did not differ significantly between SENV positive 23/40 (57.5%) and
negative 4/10 (40%) patients (p > 0.05). In addition, HCV response occurred in
19 (70%) of the 26 patients with mixed SENV-D- and -H, 4 (31%) in the 13 SENV-H
cases and 4 (40%) of the 10 only HCV infected cases.
Discussion/Conclusion: SENV infection is frequently detected in patients with
chronic HCV infection and is not present in healthy blood donors. The most prevalent
genotype is SENV-H followed by SENV-D (mainly mixed SENV-H and D). There was
a highly significant association between SENV and HCV infection and with history of
blood transfusion and surgical operation. SENV coinfection does not influence the
clinical course of hepatitis C virus or the response rate of HCV to the combination
therapy. More studies with larger number of patients infected by this novel virus are
required before it is concluded that SENV cannot cause liver disease.
33
Clearance of HCV improves insulin resistance and early insulin
secretion of pancreas in chronic hepatitis C patients
Ryoichi NARITA, Masaaki HIURA, Shintaro ABE, Akinari TABARU, Makoto OTSUKI
Third Department of Internal Medicine, Kitakyushu, Japan
Background/Aims: Hepatitis C virus (HCV) infection is linked to greater insulin
resistance and glucose intolerance. Although HCV itself is a candidate for
development of insulin resistance and glucose intolerance, the effects of antiviral
treatment on impaired glucose metabolism and insulin secretion remain unclear. This
study aimed to examine the effects of clearance of HCV on insulin resistance,
pancreas β-cell function by oral glucose loading.
Methods: We analyzed 10 biopsy-proven patients with chronic HCV infection (CHC).
Patients received peginterferon α-2b and ribavirin for 6 or 12 months. Patients were
classified into two groups at 6 months after the conclusion of antiviral therapy
according to the antiviral therapy: sustained responders (N = 6) and nonresponders
(N = 4). Insulin resistance and insulin secretion were determined in response to oral
loading of 75 g glucose.
Results: In nonresponders, there were no significant changes in HOMA-R, HOMA-β
and insulinogenic index values after antiviral therapy. On the other hand, in sustained
responders, HOMA-R significantly decreased to 1.5 ± 0.5 from 2.7 ± 2.1 (P < 0.05).
Similarly, HOMA-β decreased to 71 ± 16 from 165 ± 58. Insulinogenic index
significantly increased to 1.4 ± 0.7 from 0.6 ± 0.3 (P < 0.05).
Conclusion: We showed that clearance of HCV improves insulin resistance and
pancreas β-cell function.
34
Relationship of serum interleukin-6 and alpha tumor necrosis
factor levels with the metabolic syndrome in NASH patients
Rezi EC1, Boitan M2, Mihăilă R2, Fraticiu A1, Zaharie AV1, Olteanu A1, Dascalu D1,
Tiurean L1, Deac M2, Porr PJ2, Cipaian C1, Vulcu D2, Petrascu O2, Beca C1,
Boicean A2, Chicea L2, Orga-Dumitriu D2, Bidlo C1, Crangaciu V1, Catana A1,
Mihaila R3
1
Departmental Clinical Hospital Sibiu, Romania
2
Lucian Blaga University Sibiu, Faculty of Medicine, Sibiu, Romania
3
Public Health Authority Sibiu, Romania
Introduction: The objective of this study was to study associations of circulating
interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-alpha) levels with
anthropometric and abdominal fat distribution in patients with non-alcoholic
steatohepatitis (NASH) compared with a control group.
Methods: We have studied a group of patients which were hospitalized between
February 15–March 31, 2007 in the Medical Departments of the County Clinical
Hospital from Sibiu, Romania, to whom we have performed several blood samples.
We have analyzed: the BMI, abdominal waist line, liver enzymes, circulating
interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-alpha) levels. The results
were statistically analyzed using the t Student test and the Pearson index of
correlation.
Results: We have studied 36 patients (16 were diagnosed with NASH and
20 patients formed the control group). There is an important correlation between the
abdominal waist line and the level of IL-6 (r = 0.4834) and between the abdominal
waist line and the level of TNF-alpha (r = 0.554), at the patients with NASH. In the
control group we have find the same correlations, but not so tight (r = 0.2120
between the abdominal waist line and the level of IL6 and r = 0.2321 between the
abdominal waist line and the level of TNF-alpha). There is an acceptable association
between the BMI and the level of IL-6 (r = 0.3340) and between the BMI and the
TNF-alpha level (r = 0.3904), at the patients with NASH. This association is not so
evident in the control group. The glycemic level is correlating with IL-6 (r = 0.535) and
TNF-alpha level (r = 0.629) in patients with NASH. This correlation is less important
in the control group. There is also, an acceptable association between the
triglycerides level and IL-6 and TNF-alpha in the NASH group (r = 0.297,
respectively, r = 0.237). This correlation is not so relevant in the control group.
Discussion/Conclusion: The BMI and the abdominal waist line are correlating with
the IL-6 and TNF-alpha levels in patients with NASH. IL-6 and TNF-alpha are well
correlated with the glycemic level, in patients with NASH. Also, there is a correlation
between the triglycerides level and IL-6 and TNF-alpha, at patients with NASH.
These correlations are not so significant, statistically speaking in the control group.
These findings suggest that the pro-inflammatory cytokines are involved in the
metabolic syndrome’s pathogenic mechanisms.
35
Correlation between development of liver fibrosis and
intrahepatic bile ducts’ state in chronic viral hepatitis and PBC
L. Fatkheeva, A. Goumerova, A. Kiassov, F. Sozinov, M. Fatkheeva
Kazan State Medical University, Department of Normal Human Anatomy, Kazan,
Russian Federation
Both in chronic viral hepatitis and PBC development of liver fibrosis starts from
periportal areas; this fact allows to suppose there are common ways of fibrogenesis
in these diseases.
PBC is primary destruction of cholangiocytes so the reason for development of
predominantly portal fibrosis is clear. In hepatitis hepatocytes are primarily injured,
and cholangiocytes’ damage is secondary. As in PBC and hepatitis primary injured
cells are different and fibrosing starts from periportal areas in both cases, the aim of
our study was to find if there is any correlation between fibrosis development and
state of intrahepatic bile ducts in case of these diseases.
Liver biopsies (123 in number) from patients with chronic viral hepatitis B and C and
PBC were observed. Intrahepatic bile ducts were studied using immunohistochemical
staining with antibodies to cholangiolar cytokeratins 7 and 19.
Ductopenia index was evaluated as bile ducts’ number to portal tracts’ number ratio
(0.9–2 – normal ratio, 0.6–0.8 – moderate ductopenia, ≤ 0.5 – expressed
ductopenia).
Our results showed that in chronic hepatitis B in case of normal bile ducts liver
fibrosis of different stages was observed in 45.5% of cases, however, fibrosis wasn’t
observed in biopsies with ductopenia. In chronic hepatitis C and normal bile ducts
fibrosis was found in 40%, in cases with expressed ductopenia – in 58.8% of
biopsies. In PBC bile ducts’ injury was observed on all cases; in cases with
expressed ductopenia fibrosis of different grades was found in 62.5% of patients. So
bile duct injury directly influences development of liver fibrosis in patients with PBC
and chronic hepatitis C while in case of chronic hepatitis B such correlation was not
found.
36
Serum fibronectin as marker of liver regenerative process
Smaranda Laura Gotia1, Smaranda Rodica Gotia1, Doina Verdes2, Lelia Susan3
1
Department of Physiology, 2Cell and Molecular Biology, 34th Clinic of Internal
Medicine, University of Medicine and Pharmacy “Victor Babes” Timisoara, Romania
Introduction: The hepatocytes are the main site of fibronectin synthesis that is
involved in regenerative and tissues repair processes and in non-specific immune
response mechanisms, as phagocytosis.
Methods: Were investigated in vitro phagocytic capacity using the nitroblue
tetrasolium test (NBT%) and the latex phagocytic index (LPI%) in 20 patients with
reactive chronic hepatitis (RCH), in 12 patients with liver cirrhosis, and in 13 normal
subjects, as controls. The serum fibronectin was determinated by radial immune
diffusion.
Results: The LPI was significant lower (p < 0.001) in RCH patients (45.4 ± 1.25%)
and LC patients (47.166 ± 1.659%), than in controls (56.92 ± 1.22%). The NBT test
presented lower values in RCH (6.48 ± 2.35%) and LC patients (4.25 ± 0.686%), than
in controls (9.25 ± 2.21%). The serum fibronectin levels were significant increased in
RCH patients (401.5 ± 35.55 microg/ml) than in controls (265.47 ± 19.12 microg/ml).
Discussion/Conclusion: The increase of fibronectin levels is correlated with the liver
regenerative processes that occur in RCH patients. In LC patients were normal levels
of fibronectin because the fibrosis is prevalent in these patients. We emphasize the
decrease of phagocytic capacity in investigated RCH and LC patients, which in
association with the diminished non-specific immune response ascertains the
increase of infection susceptibility in these patients.
37
Is serum hyaluronic acid a marker of liver fibrosis in acute and
chronic liver diseases?
J. Keiss, V. Sondore, N. Sevastjanova, B. Rozentale, L. Viksna1
State agency “Infectology Center of Latvia”, Riga, Latvia
1
Chair of Traditional infectology, tuberculosis and AIDS, Riga Stradins University,
Riga, Latvia
Introduction: Liver biopsy is not always possible or reproducible, it cannot be
performed frequently and is a costly invasive procedure with a certain, although low,
risk of serious complications. Histological examination of the liver does not provide
information about the dynamics of hepatic fibrogenesis, liver biopsy provides only
static information about fibrotic process. Non-invasive markers that reflect
fibroproliferative activity in the liver and the treatment response would be preferable.
Serum hyaluronic acid (HA) has been identified now as a potential marker of fibrosis
evolution.
The aim of study was to evaluate the importance of serum HA measurements for
assessment of fibrosis in acute and chronic liver diseases.
Patients and methods: 16 patients with acute hepatitis B, 9 – with acute hepatitis C,
22 – with acute alcoholic hepatitis and 33 patients with chronic hepatitis C were
included in this study. Diagnosis was based on contemporary immunochemical
hepatitis markers assays, clinical, biochemical, epidemiological and morphological
findings. Serum HA was measured by ELISA (Corgenix Inc., Colorado, USA)
according to description of manufacturer. Liver biopsies were scored mainly
according to Ishak K. et al. (1995).
Results: Very high serum concentration of HA was found in patients with acute
alcoholic hepatitis (1015.50 ± 58.83 ng/ml). Serum HA level was significantly higher
in acute hepatitis B (228.13 ± 51.71 ng/ml) than in acute hepatitis C (58.33 ± 27.22
ng/ml, p < 0.001). Patients with acute hepatitis C showed no significant difference in
levels of HA in chronic hepatitis C (64.30 ± 14.78 ng/ml, p < 0.2). Performed
histological investigations of liver biopsies from 30 patients with chronic hepatitis C
illustrated the heterogeneity of hepatic fibrosis degree. Serum HA levels didn’t
significantly correlate with the degree of hepatic fibrosis found in these patients.
HA increases when collagen synthesis is accelerated and may be a marker of
hepatic fibrosis progression. In chronic hepatitis C, serial HA levels have been
advocated as a means to monitor disease progression and to limit the need for
follow-up liver biopsy.
Conclusion: As fibrogenic process is dynamical, serum hyaluronic acid levels may
reflect fibrogenesis rather than developed and formed fibrosis. Circulating HA
measurement has been proposed for operative monitoring of fibrotic lesion dynamics
in acute and chronic liver damages.
38
APRI in chronic hepatitis C patients with AST levels equal or
below 1.5 of upper limit of normal. To do or not to do?
O. Kosseva, E. Pophristova, K. Antonov, D. Adjarov, Z. Krastev
University Hospital “St. Ivan Rilski” Sofia, Bulgaria
Background: The AST/platelet ratio (APRI) is a simple biochemical index for hepatic
fibrosis. When first proposed by Wai et al (Hepatology 2003), the cut offs were < 0.5
for F0–F1; > 1.5 for F2–F4 and > 2 for cirrhosis.
Aim: To evaluate the diagnostic utility of APRI and proposed from the authors cut
offs in patients with chronic hepatitis C and AST levels ≤ 1.5 of the upper limit of
normal (ULN).
Methods: During 3 years period 124 CHC patients underwent liver biopsy and
66 (53%) of them had AST ≤ 1.5 ULN. Liver fibrosis was staged according to the
Metavir system.
Results: The median age and AST levels were respectively 33 years and 43 IU/l.
34 patients (51%) had F0–F1 and 32 patients (49%) F2–F4. 10 patients were
cirrhotic. APRI did not show diagnostic values to discriminate F0–F1 patients from
F2–F4 CHC patients with ALT ≤ 1.5 ULN (AUROC 0.591 p > 0.05). For discriminating
non cirrhotic versus cirrhotic patients AUROC of APRI was 0.800 but the proposed
from Wai cut offs could not correctly identified any from cirrhotic patients and 3 of
them incorrectly would be diagnosed as F0–F1.
Discussion/Conclusion: We do not recommend APRI as non-invasive test in CHC
patients with AST ≤ 1.5 ULN.
39
Comparison of three non-invasive methods for the diagnosis of
liver fibrosis in chronic hepatitis B
O. Kosseva, E. Pophristova, E. Kaneti, S. Bocheva, D. Adjarov, Z. Krastev
University Hospital “St. Ivan Rilski” Sofia, Bulgaria
Introduction: Non-invasive markers of liver fibrosis have recently been developed as
an alternative to liver biopsy. Our aim was to compare diagnostic performance of
3 tests: hyaluronic acid (HA), APRI and SOS FS to predict significant fibrosis (F2–F4)
and cirrhosis.
Methods: Metavir staging of liver specimens and APRI, HA and SOS FS were
evaluated in 61 consecutive patients with chronic HBV infections. SOS FS (SOfia
Simplified Fibrosis Score) represents the difference between levels of alpha2macroglobulin and haptoglobin and was previous validated by us.
Results: The median age was 41 years. 45 patients (74%) had F2–F4 fibrosis at liver
biopsy and 10 (16%) of them had cirrhosis. The AUROCs of APRI, SOS FS and HA
for predicting significant fibrosis were, respectively: 0.80; 0.76 and 0.67. AUROCs for
predicting cirrhosis were: 0.85 for HA and 0.80 for SOS FS. Surprisingly, APRI did
not shown diagnostic values to discriminate cirrhotic from non cirrhotic patients
(AUROC 0.62, p > 0.05).
Discussion/Conclusion: The diagnostic performance of the HA and APRI were
different according to fibrotic target. SOS FS gave reliable information on all
diagnostic targets.
40
Assessment of cholestasis and fibrogenesis in patients with
primary biliary cirrhosis
V. Kupčová, L. Turecký, M. Vaľková, M. Szántová, E.Uhlíková, S Žigraiová
Med. School, Comenius University Bratislava, Slovakia
Background and aims: Progressive liver tissue fibrosis always follows liver
diseases, which are characterized by chronic inflammatory bile duct damage
(i.e. primary biliary cirrhosis). The precise relationship between cholestasis and liver
tissue fibrosis is still poorly defined. Fibrogenesis is currently viewed as a dynamic
process strictly related to the extent and duration of parenchymal injury. Primary
biliary cirrhosis (PBC) is a chronic, progressive biliary autoimmune disease of the
liver, exhibiting immunologic abnormalities and leading to inflammatory destruction of
small- and medium-sized bile ducts. In the advanced stage, the biliary inflammation
has progressed to fibrosis leading finally to cirrhosis and liver failure. Aim of the study
was to investigate the levels of plasma total bile acids (BA), which are accepted as
sensitive cholestatic marker, and liver fibrogenesis markers - hyaluronic acid, (HA) in
patients with PBC and to compare the levels of BA with levels of HA.
Patients and methods: The concentrations of BA and HA were determined in a
group of patients with PBC (diagnosis was histological verified). The level of HA was
estimated with “Chugai“ sandwich HA binding protein assay.
Results: The levels of plasma BA were significantly increased in patients with PBC in
comparison to controls. When patients were divided according to their clinical status,
the patients with progressive more advanced stage had higher levels of plasma BA
than patients with earlier stage of disease. The levels of HA were also significantly
higher in patients with progressive disease than in patients with earlier stage of
disease. The regression analysis showed statistically significant correlation between
the levels of BA and HA (r = 0.72).
Conclusions: The results of our study showed that cholestasis could be one of the
factors, which are involved in the pathogenesis of fibrosis in patients with PBC.
According to the results of our study we can conclude that the estimation of HA could
be helpful in the diagnosis and monitoring of the stage of liver fibrosis also in patients
with PBC.
This study was supported by VEGA grant 1/4277/07
41
Serum gammaglutamyltranspeptidase, steatosis and fibrosis in
chronic virus C hepatitis
Bogdan Mihai (1), Catalina Mihai (2), Cristina Cijevschi (2), Cristina Lăcătuşu (1),
Petrut Gogalniceanu (3)
1. University Hospital „Sf. Spiridon”, Iasi, Romania
2. Institute of Gastroenterology and Hepatology, Iasi, Romania
3. University College of London, UK
Background: Steatosis has emerged as a histological finding of importance to the
progression of hepatitis C virus – associated liver disease. Serum gammaglutamyltranspeptidase (GGT) activity is a predictor of poor response to antiviral treatment in
chronic hepatitis C.
Aim: to study the correlation between GGT, steatosis and fibrosis in chronic hepatitis
C.
Material: We studied 80 patients with chronic hepatitis C, with no history of alcohol
intake, with normal body mass index, normal values of glycemia, cholesterol and
tryglycerides, admitted in Institute of Gastroenterology and Hepatology Iasi in 2006.
We performed histological exam in all these patients. Liver histology was classified
according to Knodell and hepatic steatosis according to Brunt. In every patient we
determined conventional hepatic tests and viral load. Viral genotyping was not
performed, as hepatitis C genotype 1b is present in over 95% of cases diagnosed in
Romania.
Results: 55 patients (68.75%) had GGT > 1 the upper limit of normal (group A) and
25 had normal GGT (group B). There were no significantly differences in hepatic
tests and viral load among the 2 groups. Hepatic steatosis score was significantly
higher in group A than in group B. Severe fibrosis (F3–F4) was present in 15 patients
in group A (27.27%) and in 4 patients group B (16%) (p < 0.01).
Conclusion: A substantial proportion of patients with chronic hepatic C that are not
alcohol consumers have GGT activity above the upper limit of normal. GGT could be
a marker of steatosis and severe fibrosis in chronic hepatitis C.
42
Alpha-2-macroglobulin in sera of patients with chronic
hepatopathies and effect of liver transplantation on level of this
marker
L. Turecky, V. Kupcova, M. Szantova, E. Uhlikova
Introduction: The term fibrosis is used to define the excessive accumulation of
connective tissue in parenchymal organs. Liver biopsy is the “gold standard” for
assessing liver fibrosis. Unfortunately, liver biopsy is invasive and prone to
complications. The use of biochemical parameters as fibrosis markers could
substantially reduce the number of biopsies.
The aim of our study was to investigate the levels of alpha-2-macroglobulin (AMG),
potential fibromarker, in patients with chronic hepatopathies. We compare the levels
of this marker also in patients before and after liver transplantation.
Methods: 52 blood samples from patients with fatty liver, 64 blood samples from
patients with liver cirrhosis and 60 blood samples from healthy controls were assayed
for AMG. AMG was estimated immunochemically.
Results: The levels of AMG were significantly higher in patients with liver cirrhosis
than in patients with liver steatosis (3459 vs. 2006 mg/l). When patients were divided
according to the histological findings, the levels of AMG were significantly higher in
patients with severe fibrosis than in group with minimal and mild fibrosis.The follow
up of patients after liver transplantation (before transplantation and 3, 7, 12 months
after transplantation) showed significant decrease of AMG levels (3584 mg/l –
2244 mg/l – 2240 mg/l – 2108 mg/l).
Discussion/Conclusion: The results of our study suggest that the estimation of
AMG could be helpful in monitoring of liver fibrosis in patients with chronic
hepatopathies.
The study was supported by VEGA grant 1/4268/07
43
Is IGF-1 response to growth hormone administration in
cirrhosis reflect more hepatocellular dysfunction rather than
portal hypertension and malnutrition?
1,6
Assy N, 2Pruzansky Y, 3,6Hochberg Z, 4,6Gaitini D, 5Shen Orr Z, and 2,6Baruch Y
Liver Unit, Ziv Medical Center, Zefat, Israel; 2Liver Unit, 3Department of Pediatrics,
4
Ultrasound Unit, 5Laboratory of Endocrinology, Rambam Hospital, Haifa, Israel,
6
Bruce Rappaport Faculty of Medicine, Technion, Haifa, Israel
1
Background: IGF-1 response after GH administration shows good correlation with
severity and short term survival in cirrhosis.
Aim: Correlation of IGF-1 with MELD and CP score and to determine impact of portal
hypertension (HTN) and malnutrition on IGF-1 response.
Methods: 53 patients (56 ± 2 yrs) with post viral and cryptogenic cirrhosis and
normal renal function were enrolled. IGF-1 levels were measured before and
24 hours after single sc GH (0.06 mg/kg/body weight) injection by RIA.
Results: IGF-1 levels increased from 16.7 ± 6.8 (nmol/L) to 25.4 ± 9.1 in Child A
(+52%), from 13.3 ± 4.6 to 17.1 ± 7.3 in Child B (+29%), and from 10.9 ± 3.8 to 12.6
± 4.3 in Child C (+16%) as compared to healthy controls (from 17.3 ± 6.3 to 40.3 ±
12.1, 133%, p < 0.001). 37% of patients (n = 20) had blunted IGF-1 response (< 20%
change from baseline) (Child A 2/24; 8%, Child B 7/15; 47%, Child C 11/15; 73%). By
univariate analysis stimulated, IGF-1 correlates with albumin r = 0.5, congestive
index r = 0.4, and MAMC r = 0.25. By multivariate analysis, only the CP and MELD
score (OR 5.7, CI 3.1–10.5, P < 0.0001 and OR 4.5, CI 1.5–5, P < 0.001)
differentiate accurately between blunted or responsive IGF-1 and not portal
hypertension (OR 0.9) or malnutrition (OR 1.35). Stimulated IGF-1 had better
discriminant power than baseline IGF-1 levels (78% vs. 59%, p < 0.01) and the
addition of stimulated IGF-1 to CP score increased the diagnostic accuracy from 84%
to 91%.
Conclusion: The lower IGF-1 response in advanced cirrhosis may reflect more
hepatocellular dysfunction rather than portal hypertension or malnutrition
44
Investigation of isoenzymes of gamma-glutamyltransferase and
bile acids in chronic liver diseases
V. Kupčová, L. Turecký, E. Uhlíková, M. Szántová, A. Kršáková
Medical School Comenius University, Bratislava, Slovakia
Background and aims: Gamma-glutamyltransferase (GMT) is a membrane-bound
enzyme, found in canalicular segment of the cytoplasmic membrane of hepatocytes
and in membranes of smooth endoplasmic reticulum. Serum GMT is not a
homogeneous protein, but is found in several isoforms, differing mainly in the
structure of glycosidic part of the molecule. The particular isoforms are dimeres
consisting of two monomeres, which originate from one propeptide modified by
proteolysis. We have investigated patients with histological confirmed primary biliary
cirrhosis (PBC), chronic hepatitis (CHH) and primary sclerosing cholangitis (PSC).
Patients and methods: The levels of total serum bile acids (BA), total GMT and
GMT complexed with low-density lipoprotein plus very low-density lipoprotein (LDL +
VLDL) have been evaluated in serum from healthy subjects and patients with PBC,
PSC and CHH. Serum GMT activity complexed with LDL + VLDL (iGMT) was
estimated according to Sachetti et al. (1988) using the commercial kit from
Boehringer.
Results: In patients with PBC and PSC were values of iGMT and BA markedly
higher than in patients with CHH - there was a statistically significant difference
between PBC and CHH groups. There was also a statistically significant difference
between the healthy subjects and CHH group and between CHH and PSC groups. In
the group of patients with PBC, we have found a statistically significant correlation of
iGMT with bilirubin (r = 0.50), IgM (r = 0.71) and IgA (r = 0.50). In patients with CHH
we have found a statistically significant correlation between iGMT and bilirubin
(r = 0.81) too.
Conclusion: The results of our work have confirmed the possibility to use the
examination of GMT activity bound to LDL + VLDL in differentiation of some chronic
hepatopathies (PBC, CHH and PSC). This test could be a useful addition to other
methods for discriminating PBC and PSC from CHH. We confirmed a significant
benefit of treatment with ursodeoxycholic acid in patients with PBC and PSC.
Measurement of BA can be useful in the evaluation and follow up of chronic liver
disease as a parameter of severity of the disease and can be used in addition to
other tests in hepatology.
45
Investigation of serum cytokines in chronic viral hepatitis after
IFN-α treatment
V. Kupčová, L. Turecký, Z. Zelinková, M. Vaľková, E. Šeligová
Background and aims: Several studies have suggested that a specific cytokine
profile is related to chronic evolution of viral infection. The aim of the study was
assessment of the effect of the IFN-α treatment on serum IL-10 levels in patients with
Chronic hepatitis B (VHB) and C (VHC) and to investigate the effect of therapy with
IFN-α on serum hyaluronic acid (HA) concentration as a potential biochemical marker
of liver fibrosis. The relationship of the IL-10 pattern to short virological response to
the treatment was also studied.
Methods: Serum IL-10 was measured by a commercially available kit Quantikine®
(R&D Systems). Blood samples for serum IL-10 analysis were obtained before the
treatment, in weeks 4 to 8 of the treatment and in weeks 20 to 28 of the treatment.
Hyaluronic acid (HA) concentrations (hyaluronic acid “Chugai“) before and after
treatment with IFN-alpha (48 week) in patients were assayed.
Results: Serum HA level before the treatment correlated with the extent of liver
fibrosis (r = 0.87, p < 0.001). We observed statistically significant decrease in HA in
all patients (good responders and non-responders as well), after the finishing of the
treatment by IFN alpha. All patients with VHB and good responders in the VHC group
had significantly higher pre-treatment IL-10 levels, when compared to controls.
During the treatment, a constant decrease in IL-10 was observed in VHB good
responders subgroup, reaching the significant difference only in month 6. In VHC
patients in the good responders subgroup a significant decrease in IL-10 levels was
observed in month 1, while an increase was observed in non-responders subgroup.
Conclusions: Serum HA measurement is a good and clinically useful non-invasive
marker of liver fibrosis. It could be therefore used for monitoring of the stage of
fibrosis as a measurement of response to antifibrotic therapy. Serum IL-10
assessment might be used as a response-predicting marker in management of
patients with chronic viral hepatitis treated with IFN-α.
46
Study of the relationship between chronic hepatitis C infection,
non-alcoholic fatty liver disease (NAFLD) and insulin
resistance
*Mashaal NM, *Abdelmoety AA, **Shaat EA, *El Sheikh RAO
*Department of Internal Medicine and hepatology, **Department of Biochemistry,
Alexandria Medical University, Egypt
Background: Hepatitis C infection is considered the leading cause of chronic liver
disease in Egypt. Hepatic steatosis (NAFLD) has been reported in 30–70% of
patients with HCV. Insulin resistance (IR) emerges as a very important host factor in
patients with chronic hepatitis C (CHC).
Aim of the work: Assess the relationship between chronic hepatitis C infection,
NAFLD and insulin resistance in Egyptian patients.
Material: 20 patients with chronic HCV infection and hepatic steatosis were proven
by liver biopsy were included in the study. Other 20 healthy adults were enrolled as a
control group.
Methods: Liver function tests, HCV antibodies by third generation ELISA, HCV-RNA
by PCR, fasting serum glucose, fasting serum insulin and serum ferritin. Insulin
resistance was evaluated using the homeostasis model assessment (HOMA-IR)
method. Also, ultrasound examination of the abdomen.
Results: Insulin resistance (IR) by (HOMA) index was significantly higher in patients
than in control group (mean ± SD 8.36 ± 6.61 and 2.17 ± 0.08, respectively). Fasting
Insulin level was significantly higher in chronic HCV patients than in control group
(P = 0.000). Serum ferritin level was significantly higher in patients (124.04 ± 123.18
ng/ml) than in control group (65.22 ± 46.27 ng/ml). HOMA-IR positively correlated
with staging (P = 0.034, r = 0.477) and with grading (P = 0.022, r = 0.509) in HCV
infected patients.
Conclusion: HCV infected patients have high degree of IR. IR facilitates fibrogenesis
in chronic HCV patients. High serum ferritin may be implicated in (IR).
47
The study of correlation between IL-6 and TNF alpha and the
biochemical liver tests at the patients chronically infected with
the hepatitis virus C
Mihaila R1, Boitan M1, Fraticiu A2, Rezi EC2, Zaharie AV2, Olteanu A2, Dascalu D2,
Tiurean L2, Deac M1, Porr PJ1, Cipaian C2, Vulcu D1, Petrascu O1, Beca C2,
Boicean A1, Chicea L1, Orga-Dumitriu D1, Bidlo C2, Catana A2, Mihaila R3
1
2
3
Introduction: The releasing of high levels of pro-inflammatory cytokines determines
the augmentation of the EPO synthesis. At its turn, EPO determines the inhibition of
some pro-inflammatory cytokines, as IL-6, alpha TNF, cytokines which are involved
in the chronic hepatopaties pathogenesis. That is why we proposed ourselves to
study if there is any correlation between the above mentioned cytokines and EPO, or
other liver biochemical tests at the patients chronically infected with the C virus.
Methods: We have studied a group formed by 35 patients which were hospitalized in
the medical departments of the Clinical County Hospital Sibiu, during 15.02–
31.03.2007. The group was divided in two: group A – with 12 patients, which were
chronically infected with the C hepatitis virus and group B – with 23 patients without
any liver or inflammatory disease (control group). At both groups we have studied:
age, BMI, the biochemical tests of the liver, cholesterol, triglycerides, IL-6,
TNF alpha, EPO. We have compared the average values of the parameters from the
two groups and we have statistically analyzed the existence of any correlation
between the cytokines and EPO or the liver biochemical tests. The results were
statistically analyzed using the t Student test and the Pearson index of correlation (r).
Results: The medium age of the examined patients was 39.67 ± 14.73 years. The
gender repartition was: 77.14% women and 22.86% men. There is an important
direct linear correlation between IL-6 and TNF alpha, in group A (r = 0.6215), and in
group B, as well (r = 0.7716). At the patients with C virus, we have found higher
values of the EPO in group A, comparing with group B, but the differences are not
statistically significant. There is an acceptable association between EPO and IL-6, in
group B (r = 0.266), and in group A as well (r = 0.163). Despite this, at those with C
virus, the correlation between EPO and TNF alpha is stronger than in the control
group (r =0.483, versus r =0.333). At the patients from group A, there is very tight
linear direct correlation between the level of TGO and IL-6 (r = 0.826), respectively
between TGP and IL-6 (r = 0.886). This correlation does not exists in group B
(r = 0.054 for TGO, respectively r = 0.07 for TGP). Similarly results were obtained by
correlating the transaminasis with the level of TNF alpha. Correlating the level of
EPO with the markers of the liver dysfunction, at the patients from group A, there
could be observed an inverse linear correlations between the level of the protrombine
index and EPO (r = -0.605) and between the level of seric albumin and EPO
(r = -0.319).
Discussion/Conclusion: The data we have obtained are in accordance with the
data from the literature, and suggest the implication of the immune answer and of the
cytokines in the pathogenesis of the chronic infection with the C virus. EPO is directly
correlating with the pro-inflammatory cytokines in an acceptable way, and indirectly,
more strongly, with the biochemical syndrome of the hepatocytar failure.
48
The study of correlation between IL-6, TNF alpha and
erythropoietin with the biochemical liver tests at the patients
with non-alcoholic fatty liver disease
Mihaila R1, Boitan M2, Mihăila R2, Rezi EC3, Fraticiu A3, Zaharie AV3, Olteanu A3,
Dascalu D3, Tiurean L3, Deac M2, Porr PJ2, Cipaian C3, Vulcu D2, Petrascu O2,
Beca C3, Boicean A2, Chicea L2, Orga-Dumitriu D2, Bidlo C3, Catana A3
1
2
3
Introduction: IL-6 and TNF alpha are markers of inflammation, involved in the
pathogenesis of the chronic liver diseases. It was suggested that the level of
endogenous erythropoietin (EPO) can increase together with the level of IL-6. We
proposed ourselves to study if there is any correlation between IL-6, TNF-alpha and
the biochemical liver tests, and if the EPO level is correlating with the markers of
inflammation, or with the biochemical liver tests at the patients with non-alcoholic
fatty liver disease (NAFDL).
Methods: We considered a group formed by 54 patients which were hospitalized in
the medical departments of the Departmental Clinical Hospital Sibiu, during 15.02–
31.02.2007. The group was divided into two: group A – with 31 patients with NAFDL
(among them, 16 had NASH – A1, and 15 had only liver steatosis – A2) and group B
– with 23 subjects without any liver or inflammatory disease (control group). At both
groups we have studied: age, BMI, biochemical liver tests, cholesterol, triglycerides,
IL-6, TNF-alpha, EPO. We have compared the medium values at the two groups and
we have analyzed the existence of any correlation between the cytokines and EPO
and the biochemical liver tests. The results were statistically analyzed, using the
t Student test and the Pearson index of correlation.
Results: The medium age of the studied group was 35.68 ± 14.36 years. The gender
repartition was: 72% women and 28% men. The average value of TNF-alpha in
group A was 7.568 pg/MI, and in group B - 5.226 pg/MI (p < 0.001). Also, in group A,
it can be observed that IL-6 has higher values than in group B (p = 0.0036). There
are no statistically significant differences between the two groups regarding the
medium level of EPO. In group A1 there is a direct linear correlation between the
levels of IL-6 and TGO (r = 0.6774). This association is not so obvious regarding the
TGP level (r = 0.1731). Also, it can be observed an acceptable association between
the TNF-alpha level and TGO (r = 0.3843). There is an inverse linear correlation
between the level of endogenous EPO and TGP (r = -0.606). It can be observed that
there is an acceptable association between the EPO level and the TNF-alpha level
(r = 0.3357). In group B there are no correlations between the level of transaminases
and the level of IL-6 and TNF-alpha. There is an acceptable association between the
level of endogenous EPO and the level of IL-6 (r = 0.266). This correlation does not
exists in group A (r = -0.0241).
Discussion/Conclusion: The levels of IL-6 and TNF-alpha correlate mainly with the
level of TGO at the patients with NAFDL. The level of EPO increases together with
the increasing of IL-6 level, at the patients from the control group, which is in
accordance with some data from the literature. At the patients with NAFDL there is
an inverse correlation between EPO and TGP, which means that EPO levels
decrease in the liver cytolysis.
49
Serum apoptotic cytokeratin 18 neoepitope in acute and
chronic liver damage
V. Sondore, J. Keiss, N. Sevastjanova, L. Viksna1, B. Rozentale
State agency “Infectology Center of Latvia”, Riga, Latvia
1
Chair of Traditional Infectology, Tuberculosis and AIDS, Riga Stradins University,
Riga, Latvia
Introduction: Apoptosis is a common property of multicellular organisms to destroy
cells that represent a threat to the integrity of the organism. The apoptotic process
appears to be a host defense mechanism against different pathological stimuli.
During apoptosis the sequential activation of caspases (they all are proteases that
cleave proteins at aspartic acid residue) creates an expanding cascade of proteolytic
activity which leads to digestion of structural proteins in cytoplasm and generates e.g.
apoptotic cytokeratin 18 (CK-18) neoepitopes.
The aim of present study was to detect the levels of caspase-generated CK-18
neoepitope in acute and chronic liver damages.
Patients and methods: In 9 patients with acute hepatitis B, 23 – with acute alcoholic
hepatitis and in 22 patients with chronic hepatitis C, all treated in the Infectology
Center of Latvia, the quantitative detection and estimation of CK-18 neoepitope in
blood serum were performed. Non-invasive method for caspase-generated CK-18
fragments quatifying in serum (M30-Apptosense® ELISA kit, PEVIVA) was used.
Diagnosis was based on contemporary immunochemical hepatitis markers assays
and clinical, biochemical, morphological and epidemiological findings. Results were
expressed as means ± SEM; for two group comparison the unpaired Student’s group
t-test was used; P ≥ 95% was considered significant.
Results: Performed studies yielded interesting data: very high serum level of CK-18
neoepitope was found in patients with acute hepatitis B (1349.0 ± 119.7 U/L), that is
higher than in patients with alcoholic hepatitis (1003.9 ± 104.3 U/L; 98% > P > 95%).
CK-18 neoepitope concentration in chronic hepatitis C was significantly lower (258.2
± 27.5 U/L; 99% < P < 99.9%). About 1/3 of chronic hepatitis C patients with normal
aminotransferase activity had elevated serum CK-18 neoepitope if compared with
normal values of this apoptosis indicator.
Conclusions: 1) Apoptotic CK-18 neoepitope level is more sensitive measure of liver
damage than conventional surrogate marker ALT and therefore should be
recommendable as a compulsory diagnostic test in cases of liver diseases occured
with normal serum aminotransferases. 2) If in alcoholic hepatitis apoptosis may be a
principal cause of cell death, then in acute hepatitis B cell death would be associated
with both cell apoptosis and cell necrosis. 3) Monitoring of serum apoptotic CK-18
neoepitope concentration might provide a reliable tool to estimate the efficacy of
therapy of liver diseases and might open challenging approaches to therapeutic
strategies for different liver diseases.
50
Study of the correlation between nitric oxide profile, degree of
liver injury and structural vascular changes of the gastric
mucosa in chronic hepatitis C virus related liver diseases
*Taher MY, *Hassan El Saed, **Baddour N, *Abdelmoety AA, *Zeid A
*Department of Hepatology, **Department of Pathology, Alexandria Faculty of
Medicine, Alexandria, Egypt
Background: The mechanisms underlying liver injury and fibrogenesis in chronic
hepatitis C virus (HCV) infection are still poorly understood. Together with other
factors, nitric oxide (NO) plays an important role in HCV associated hepatic
dysfunction and in the pathogenesis of portal hypertension.
Aim of the work: This study was designed to correlated serum nitrite and nitrate
levels with the degree of liver injury and gastric mucosal changes in HCV patients.
Subjects and methods: 80 HCV infected patients were classified equally into
4 groups; chronic hepatitis C, Child A, B and C cirrhotic groups. 20 healthy subjects
were allocated as a control group. For all patients, serum nitrite and nitrate levels,
HCV RNA and liver test profile were evaluated and for chronic hepatitis and Child A
cirrhotic patients, liver biopsies were obtained for grading, staging and expression of
INF-γ and pentosidine. Gastric mucosal biopsies were taken from all patients and
evaluated for the degree of portal hypertensive gastropathy (PHG) and expression of
vascular endothelial growth (VEGF).
Results: Serum NO profile was significantly higher in all HCV infected patients than
healthy subjects and was correlating with the severity of Child-Pugh classification.
Examination of liver biopsies revealed no significant correlation between grading or
staging and serum NO level and a significant correlation between INF-γ expression
and both of serum NO and viral load. Also, hepatic pentosidine expression was
correlating with staging and fibrosis. In cirrhotic patients, the enhanced production of
NO was significantly correlated with the severity of ascites and the grade of
esophageal various. Also both of serum NO and gastric VEGF were over expressed
and correlating with the degree of PHG.
Conclusion: In HCV infected patients, serum NO was significantly correlating with
the severity of chronic liver disease. Our study supports the role of direct viral
cytopathic effect in HCV patients because of the significant correlation of viral load
with both of serum NO and hepatic INF-γ expression. Pentosidine might be
considered a marker of oxidative stress and fibrosis in chronic HCV liver disease.
Both NO and VEGF seem to play a role in the development and severity of PHG.
51
Serum selenium and carnitine concentrations of cirrhotic
children
Nuray Uslu, İnci Nur Saltık-Temizel, Hülya Demir, Figen Gürakan, Hasan Özen,
Aysel Yüce
Hacettepe University, Faculty of Medicine, Department of Pediatrics, Section of
Gastroenterology, Hepatology and Nutrition, Ankara, Turkey
Introduction: Selenium is an essential trace element for humans and functions as a
component of antioxidant enzymes like glutathione peroxidases. Decreased plasma
selenium concentration was reported especially in adults with cirrhosis. Carnitine is
an important biofactor which acts in mitochondrial energy production, heart and
smooth muscle function. Liver plays an important role in carnitine metabolism, but
plasma carnitine concentration in cirrhotic patients was found to be variable. So we
wanted to investigate the serum selenium and carnitine concentration in cirrhotic
children.
Methods: A total of 48 cirrhotic children were enrolled. The serum selenium
concentration of 38 patients was determined by spectrofluometric method (normal >
50 µg/L) while the serum carnitine concentration of 43 patients was determined by
“tandem mass spectroscopy” (normal range 10–60 µmol/mL). The results of the
patients were compared with those of age and gender matched healthy children.
Results: The mean age of the patients was 10.6 ± 4.3 and 5.4 ± 4.3 years for
selenium and carnitine group respectively. The mean serum selenium concentration
of cirrhotic children was 42.4 ± 8.2 µg/L and of the control’s was 64.4 ± 16.9 µg/L
(p < 0.05). The mean serum carnitine concentration was found as 28.6 ± 12.9
µmol/mL and of the control’s was 28.5 ± 9.3 µmol/ml (p > 0.05).
Discussion/Conclusion: The serum selenium concentration in cirrhotic children was
significantly lower than the control’s. Supportive selenium administration may be
beneficial in cirrhotic children. There was no difference between serum carnitine
concentrations of patients and control group.
52
On the optimization of the 13C-alpha-ketoisocaproic acid breath
test – An in-depth study speaks for an extended breath
sampling period
K Jonderko1, A Kasicka-Jonderko1, M Kamińska1, M Bielecka2,
B Błońska-Fajfrowska1
Dept. of Basic Biomedical Science1, Student Scientific Association Circle at the Dept.
of Basic Biomedical Science2, Medical University of Silesia, Sosnowiec, Poland
Introduction: From among 13CO2 breath tests designed for a functional examination
of the liver, the alpha-ketoisocaproic acid (13C-KICA) breath test was proposed as a
tool enabling the evaluation of xenobiotics-induced impairment of the human liver
mitochondrial function, as well as for the differentiation between non-alcoholic and
alcohol-induced liver steatosis. In the current study we searched for a quantitative
parameter which would offer the best reproducibility of a standard 13C-KICA breath
test.
Methods: Fourteen healthy volunteers (7F, 7M, aged 23.6 ± 0.5 years) were
examined. On three separate days they drank 200 ml orange juice containing
1 mg/kg body mass 13C-KICA and 20 mg/kg L-leucine. Samples of expiratory air for
13
CO2 measurement with the use of the isotope ratio mass spectrometry were
collected every 5 min during the first hour, and then at 70, 80, 90, 105, 120, 150, 180,
240, 300, 360, 540, 720 and 1440 min. Short-term reproducibility (STR) was
assessed on the basis of paired examinations taken 2–4 days (median 3 days) apart,
whereas paired examinations separated by 20–27 days (median: 23 days) served for
the medium-term reproducibility (MTR) assessment. The order of intervals between
the consecutive sessions was randomized. Coefficients of variation for paired
examinations (CVp) were calculated for three quantitative descriptors of the test: Dmax
– the maximum momentary 13C recovery, Tmax – the time to reach the Dmax, and AUC
– the cumulative 13C recovery.
Results: Tmax had an unsatisfactory reproducibility (CVp 19.99% STR, 24.75% MTR),
Dmax displayed a good reproducibility yet probably insufficient from the point of view
of diagnostic applications (CVp 11.24% STR, 13.94% MTR). The reproducibility of the
cumulative 13C recovery relative to the administered substrate dose depended on the
time span considered for calculation of the AUC. Accordingly, as from AUC0–80 the
CVp was lower than 10%, and a further improvement of reproducibility was found with
AUC0–90, AUC0–105, and AUC0–120, whereas inclusion of the data from beyond the
second hour did not result in any more refinement of the reproducibility of the AUC.
Taking into account the numerical values of the CVp, AUC0–90 and AUC0–120 exhibited
excellent reproducibility – AUC0–90 CVp 8.18% STR, 8.98% MTR, AUC0–120 CVp
7.37% STR, 8.01% MTR. In no case did the medium-term reproducibility prove any
worse than the short-term one.
Conclusion: From among the three parameters considered, AUC computed on data
involving at least 90 min after administration of the substrate offers the level of
reproducibility sufficient for clinical applications of the 13C-KICA breath test.
53
Band ligation vs. miniloops in the endoscopic treatment of
esophageal varices – Comparative pilot study
Chr. Banciu, Lelia Susan, O. Chirileanu, Loredana Marian, I. Romosan
IV. Department of Internal Medicine, University of Medicine and Pharmacy Timisoara,
Romania
Introduction: Variceal hemorrhage is one of the most frequent causes of upper GI
bleeding in hepatic cirrhosis.
Objectives: This study evaluates rubber band ligation (RBL) vs. ligation with endominiloops (LEM) for the endoscopic treatment of: 1.Variceal bleeding; 2.Prophylactic
treatment of esophageal varices (EV).
Material and methods: 23 pts (14M, 9F) with Child B and C cirrhosis presenting
large EV were randomized in two groups, one group receiving RBL (12 pts), the
others LEM (11 pts). Devices used: Olympus loop application device HX-20U-1,
miniloops type MAJ-339 and Wilson Cook Saed six shooter device. For LEM
procedure we used a cap at the tip of the endoscope. Ligation of the EV started at
the GE junction. Mean follow-up: 9 months (6–12).
Results: Hemostasis was achieved in all pts of both groups, RBL & LEM,
respectively. During follow-up 2 pts of the LEM group rebled due to early miniloop
dislodgement, but they were successfully treated with endoloops and the bleeding
stopped; 4 pts of RBL group and 2 pts of the LEM one presented esophageal ulcer,
treated with PPIs. In both groups there was a significant reduction of variceal size.
Conclusions: 1. Both LEM & RBL are effective and safe methods to control variceal
bleeding; 2. RBL is better, faster and easier to use than LEM, the latter taking more
time for reloading; 3. Regarding the endoscopic view, LEM has a better one
compared to RBL; 4. Looking at the cost benefit ratio, both have about the same
price, but in our opinion RBL is overall easier to perform, less time-consuming, more
successful and efficient.
54
Upper digestive bleeding in liver cirrhosis
Cristina Cijevschi (1), Catalina Mihai (1), Vasile Drug (1), Petrut Gogalniceanu (2)
1. Institute of Gastroenterology and Hepatology Iasi, Romania
2. Universty College of London, UK
Background: Upper digestive bleeding (UDB) is one of the most frequent
complications of liver cirrhosis (LC), with a high mortality.
Aim: to appreciate the incidence of UDB in liver cirrhosis, and to find some risk
factors.
Material and method: We studied 840 patients with UDB, in which we performed
emergency upper digestive endoscopy in the first 24 hours, between 1st January
2006 and 1st January 2007.
Results: In this group we found 230 patients with LC (27.3%), 137 men and
93 women, aged between 28 and 83 years .The etiology of LC was: virus C
(45 cases), alcohol (50 cases), virus B (29 cases), both virus and alcohol – 80 cases
and unknown or rare causes – 26 cases. In 40 cases (17.39%) there was
nonsteroidal anti-inflammatory drugs (NSAID) intake previous the bleeding. In
150 patients (65.21%) the cause of bleeding was variceal efraction; and in the rest
80 patients portal gastropathy, erosive gastritis, duodenal ulcer.22 patients were in
the A Child class, 85 patients in B and 123 in C.
Conclusions: In 17.3% cases NSAID intake is a precipitant factor for UDB in
patients with LC; the emergency upper digestive endoscopy need to be a rule
because in 34.71% patients with LC the cause of bleeding is not variceal efraction;
the incidence of UDB is correlated with Child-Pugh class and not with the etiology.
55
Upper digestive haemorrhage in cirrhosis – Prevalence and
precipitating factors
Dana Damian, Mircea Grigorescu, Andrea Berecz, Monica Hogea, Mihaela Rusu
Third Medical Clinic, Cluj Napoca, Romania
Introduction: The aim of this study is to evaluate the causes and the precipitating
factors of the upper digestive bleedings in patients with or without hepatic cirrhosis,
who were admitted in our department during one year period.
Methods: We studied retrospectively 118 patients with upper digestive haemorrhage,
who underwent gastroscopy for diagnosis and treatment. Using the information from
the case history, we analysed the possible risk factors to precipitate the bleeding.
Results: Of a total of 4792 patients admitted in our department in 2005, 118 patients
(2.46%) were diagnosed with upper gastrointestinal bleeding. There were 75 males
and 43 females with a mean age of 57.3 ± 13.4 yrs. A group of 34 patients (28.81%)
were cirrhotics and the bleeding site could be localized: 28 cases of oesophageal
and/or gastric varices (82.35%) and 6 cases (17.65%) of other esogastroduodenal
lesions, as: duodenal ulcer (66.6%), erosive gastritis and oesophagitis (16.67%),
oesophageal cancer and duodenal ulcer (16.67%). The remaining 84 non-cirrhotic
patients (71.19%) presented with non-variceal bleeding. Precipitating factors for
variceal vs. non-variceal haemorrhage were: alcohol ingestion (50% vs. 34.44%,
p = 0.138), Helicobacter pylori infection (46.42% vs. 60%, p = 0.205), smoking
(14.28% vs. 27.77%, p = 0.147) and non-steroidal anti-inflammatory drugs (7.14% vs.
33.33%, p = 0.006), anticoagulant/antiaggregant drugs (0% vs. 16.67%, p = 0.046).
Discussion/Conclusion: In the whole study group the non-variceal haemorrhage
was more frequent (71.19%), but in patients with cirrhosis the variceal haemorrhage
was predominant.
For variceal haemorrhage, the most frequent precipitating factors are alcohol
consumption and Helicobacter pylori infection, and for the non-variceal ones,
Helicobacter pylori infection, alcohol ingestion and the use of anti-inflammatory
drugs.
56
Upper gastrointestinal bleeding in cirrhosis – Clinical aspects
and evolution
Dana Damian, Mircea Grigorescu, Andrea Berecz, Monica Hogea, Mihaela Rusu
Third Medical Clinic, Cluj Napoca, Romania
Introduction: The aim of this study is to evaluate the clinical aspects and evolution
of the upper gastrointestinal bleedings in patients with or without hepatic cirrhosis.
Methods: We studied retrospectively 118 patients admitted in our hospital in 2005 for
upper gastrointestinal hemorrhage, who performed endoscopy for diagnosis and
treatment, if necessary.
Results: We analysed 118 patients (75 males and 43 females, mean age 57.3 ± 13.4
yrs). 34 of all patients (28.81%) were cirrhotics and 28 presented oesophageal and/or
gastric variceal bleeding (82.35%). The rest of 84 non-cirrhotic patients (71.19%)
presented non-variceal bleeding. Regarding the Child-Pugh classification, the
distribution of patients was as follows: A class 4 patients (11.76%), B class
14 patients (41.18%), C class 16 patients (47.06%). The admission of patients with
vs. without cirrhosis was decided for haematemesis (11.76% vs. 19.05%), melena
(14.70% vs. 32.14%), haematemesis and melena (64.71% vs. 40.48%) and
hematochesia (2.94% vs. 1.19%), occult digestive haemorrhage (2.94% vs. 8.33%).
Anemia was present in both patients groups (85.29% vs. 69.04%, p = 0.069), but the
severe anemia was observed mainly in cirrhotic patients (29.41% vs. 16.67%,
p > 0.05) in parallel with the need for transfusions (70.58% vs. 27.38%, p < 0.001)
and the mean number of blood units transfused per patient (3.51, respectively 2.91
blood units). Re-bleeding rate was higher in the first patients group (32.35%) than in
the second (5.95%) and the mortality rate was also different (11.76% vs. 2.38%,
p > 0.05).
Discussion/Conclusion: The severe anemia, the re-bleeding and the mortality rates
are more frequent in patients with cirrhosis and upper gastrointestinal bleeding.
57
Major complications in liver cirrhosis
Catalina Mihai (1), Cristina Cijevschi (1), Bogdan Mihai (2), Vasile Drug (1),
P. Gogalniceanu (3)
1. Institute of Gastroenterology and Hepatology, Iasi, Romania
2. University Hospital “Sf. Spiridon”, Iasi, Romania
3. University College of London, UK
Background: Liver cirrhosis (LC) is a severe disease, with a lot of complications and
a poor prognostic. In the last years the management of LC complications improved
without a significantly decreasing of the mortality.
Aim: assessment of death causes in patients with LC.
Material: We have studied death causes in patients with LC admitted in the Institute
of Gastroenterology and Hepatology between 1st January 2006 and 1st January 2007.
We have tried to put into evidence a possible correlation between etiology,
complications, Child class, obesity, diabetes mellitus, the duration of the disease.
Results: We studied 730 patients; 28th deaths have been recorded (3.89%) (group
A). The death causes according to their frequency were as follows: hepatic coma – 8,
haemorrhage shock induced by upper digestive haemorrhage (UDB)-7, hepato-renal
syndrome (HRS) – 6, spontaneous bacterial peritonitis (SBP) – 5, and in two cases
were associated diseases. HRS and SBP were the complications with highest
mortality (80%, respectively 76%). UDB, although in comparison with other
complications was associated with a low mortality rate in itself, has been met as a
risk factor in 18 of the deaths. The death rate was correlated with a long history of the
disease, the male sex, Child class, type II diabetes mellitus, but was not correlated
with etiology.
Conclusions: Due to its complications LC is a high-risk mortality disease. SBP and
HRS are the complications with the highest death risk, but in almost all cases UDB is
the most important risk factor. Death risk is linked to the long history of the disease,
its complications, and the liver potential assessed by Child score.
58
Use of clonidine in cirrhotic patients with large diuretic
resistant ascitis – Prospective pilot study
Chr. Banciu, Lelia Susan, Violetta Vacariu, C. Dascau, Loredana Marian, I. Romosan
IV. Dept. of Internal Medicine, University of Medicine and Pharmacy Timisoara,
Romania
Introduction: Large refractory ascitis resistant to diuretics is frequently associated
with splanchnic arterial vasodilatation.
Objectives: The present prospective pilot study wants to evaluate the efficacy of oral
clonidine (alpha agonist agent) administration in the treatment of diuretic resistant
ascitis.
Material and methods: 9 pts (7 males, 2 females) with cirrhosis Child C class and
persistent refractory ascitis and repeated paracentesis, with serum creatinine levels
< 130 µmol/L and mean arterial pressure under 80 mmHg were randomized to
receive either diuretic treatment only (spironolactone 300 mg/day and furosemide
120 mg/day) or in association with clonidine (0.1 to 0.2 mg bid), respectively. Several
parameters were followed (liver tests, serum sodium concentration, urinary sodium
excretion, serum creatinine, PTT, ascitis formation – liter/week).
Results: After 4 weeks ascitis production and consequently, the need for
paracentesis decreased significantly in 4 of 5 pts treated with clonidine (80%),
compared with 1/4 pts receiving only diuretics (25%). Serum creatinine levels were
reduced and urinary sodium excretion increased. There was also observed an
increase of the mean arterial pressure. Ni important side effects of clonidine were
noted, only some mild ones (weakness, headache, dizziness).
Conclusions: Although this pilot study was carried out on a small number of pts and
cannot obtain statistical significance, it points out and suggests that clonidine in
association with diuretics reduces refractory ascitis significantly and consequently the
need for paracentesis, by improving the circulatory dysfunction in these pts. Thus,
clonidine would be efficient in the management of refractory diuretic-resistant ascitis.
59
Superiority of the serum-ascites albumin gradient over the
ascites total protein concentration in diseases with ascites
Colic N, Vrinic D, Colic A, Kazic S, Svorcan P, Necic D, Dapcevic B
Department of gastroenterology and hepatology, Zvezdara Clinical Center, Belgrade,
Serbia
Introduction: We compared serum-ascites albumin gradient (SAAG), the difference
between serum and ascitic fluid albumin concentrations, to the ascites total protein
concentration in patients with ascites and estimated diagnostic significance of SAAG
in differential diagnosis of ascites.
Methods: We studied 103 patients with ascites. 79 of them had portal hypertension
(56 patients with decompensated liver cirrhosis, 11 with heart failure, 4 with massive
liver metastases and 8 with mix ascites). The other 24 patients did not have portal
hypertension (15 with peritoneal carcinosis, 5 with pancreatic ascites, 3 with
peritoneal mesothelioma and 1 with TBC peritonitis). SAAG was determined in all
patients by a person who was unaware of diagnosis (blind study).
Results: SAAG values were greater than 1.1 g/dl in all patients who had ascites and
portal hypertension (average value: 2.2 g/dl ± 0.5 g/dl). SAAG values were less than
1.1 g/dl in the patients without portal hypertension (average value: 0.8 g/dl ± 0.5 g/dl).
Discussion/Conclusion: SAAG values above 1,1 g/dl point to portal hypertension
as the cause of ascites. SAAG values below 1,1 g/dl point to the other causes of
ascites. SAAG categorizes ascites better than ascites total protein concentration
(which classifies ascites into exudate and transudate), and has significant diagnostic
value in differential diagnosis of ascites.
60
The evolution of spontaneous bacterial peritonitis under
medical treatment in cirrhotic patients with ascites
Amelia Genunche-Dumitrescu, P. Mitrut, Daniela Badea, M. Badea
University of Medicine and Pharmacy, Clinical Hospital of Emergency, Craiova,
Romania
Aim: to estimate the incidence and the evolution of spontaneous bacterial peritonitis
(SBP) under treatment in cirrhotic patients with ascites.
Methods: We studied 188 patients with cirrhosis (25 cases Child-Pugh A, 43 cases
Child-Pugh B and 120 cases Child-Pugh C). The incidence of SBP was 18.62%
(35 cases, mean age 44.6 ± 8.4 years). The diagnosis was confirmed by clinical
picture and laboratory test (ascitic neutrophilic leukocytes count ≥ 250/mmc). Three
therapeutic schemes was used: Cefotaxime 4 g/day for 7 days then 2 g/day for 7day
(in 18 cases – A group), ciprofloxacinum 1000 mg/day 10 days (in 8 cases – B group)
and C group (9 cases) treated with 1 g amoxicillin and 0.2 g clavulanic acid every
6 hours for 12 days.
Results: SBP usually develops in patients with advanced liver cirrhosis (Child’s C
class in 27 cases) and alcohol intake. All SBP episodes were symptomatic: jaundice
(54.28%), abdominal pain (48.57%), fever (62.86%), tenderness and fatigue. In 23 of
the total cases the culture of the ascites fluid was positive: 16 cases with E. coli,
5 cases with Klebsiella, 2 cases with Enterococcus. In A group the evolution was
favourable in 15 cases, one patient died in the course of the primary SBP attack and
2 patients was presented recurrences of SBP within 15–20 days after therapy was
discontinued. In A group 3 patients was presented a new episode during 6 months.
The evolution was favourable in all B group patients and in 8 cases at C group
(one patient died because developed renal insufficency). To prevent recurrent SBP
we used quinolone or Normix 15 mg/kg/day.
The parameters that correlated with development SBP was: lower ascites protein
level (≤ 1 g/dl; r = 0.49, p < 0.05), total bilirubin (r = 0.37, p = 0.01), creatinine
(r = 0.30, p < 0.05), longer prothrombin time (INR ≥ 2.5; r = 0.19, p < 0.05).
Discussion/Conclusion: By monitoring the ascitic fluid PMN cell count is possible to
determine the efficacy and optimal duration of treatment in patients with SBP.
Antibiotic therapy was initiated imediaterly after the diagnosis paracentesis and
before culture results are available.
61
E. coli lipopolysaccharides antibodies (CLA) and some species
of chronic hepatitis and liver cirrhosis
B. Levitan, A. Umerova, G. Levitan
Medical Academy, Astrakhan, Russia
The aim of the investigation was to study the serum CLA concentration in chronic
hepatitis (CH) and liver cirrhosis (LC) and to determine the correlation between CLA
and some laboratory markers.
Methods: 37 CH and 60 LC patients and 50 healthy volunteers were included in the
investigation. CLA concentrations were assessed by the ELISA method.
Results: The average CLA level in CH & LC was twice increased in comparison with
the donors (24.0 ± 0.4 IU/ml cons 14.1 ± 0.4 IU/ml. The increased level of CLA
concentration was determined in about 90% of CH & LC patients. The CLA level was
significantly increased in viral CH & LC in comparison with the alcoholic etiology and
also correlated with circulation immune complexes (CIC) level (r = 0.463; p = 0.02)
that testify intercommunication with immunocomplex liver failure. The opposite
correlative dependence between CLA indications, albumin level, albumin/globulin
index and thymol test was determined that confirm the connection with violation of
protein-synthetic liver function and dysproteinemia. LC & CH patients with low and
normal CLA level had innocent clinical course (low clinical-biochemistry activity of LC
& CH; normal bilirubin and thymol test indications; cholestasia and icterus absence;
low level of cytolysis). High CLA level in LC & CH was associated with increased
cholestasia frequency and high clinical-biochemistry activity.
Conclusions: The increased CLA level indicates the E. coli participation in LC & CH
pathogenesis. That is additional reason to use prebiotics and probiotics in LC & CH
therapy.
62
Bacterial translocation in cirrhotic rats: Therapeutic role of
iNOS inhibitor S-methylisothiourea
1
Mas MR, 1Yilmaz S, 2Uzunalimoglu O, 3Ozyurt M, 1Tasci I, 1Isik AT, 4Ozcan A,
1
Comert B, 5Mas N, 6Akay C, 7Bozkaya H, 7Yurdaydin C
1
Gulhane School of Medicine, Department of Internal Medicine, Ankara, Turkey
2
3
Gulhane School of Medicine, Department of Microbiology, Istanbul, Turkey
4
Gulhane School of Medicine, Department of Pathology, Ankara, Turkey
5
6
Gulhane School of Medicine, Department of Toxicology, Ankara, Turkey
7
Ankara University, Institute for Hepatology, Ankara, Turkey
Introduction: Bacterial overgrowth and translocation (BT) play important roles in the
pathogenesis of spontaneous bacterial peritonitis. Nitric oxide, though beneficial in
physiological conditions, may facilitate translocation of bacteria from the intestine at
high concentrations. In this animal study, the effects of S-methylisothiourea (SMT),
an iNOS inhibitor, on BT in cirrhosis and the behavior of oxidative stress (OS)
markers were studied.
Methods: To induce cirrhosis, 90 Sprague-Dawley rats were injected with CCl4. The
surviving 28 animals at 12th week were divided into two groups. Group-1 received
intraperitoneal saline, and group 2 received SMT (5 mg/kg/day). All animals were
killed at day 30. Caecum, mesenteric lymph node (MLN), and liver samples were
collected for histological assessment, determination of BT and OS markers. 10 rats
served as controls.
Results: Degree of cirrhosis was similar in the two study groups. BT to MLNs and
peritoneum was not observed in any of the controls. However, it was evident in
group1 [10/14 (71%), p < 0.01 and 7/14 (50%), p < 0.05, respectively). BT to MLNs
and peritoneum decreased in the SMT treated cirrhotic group 2 compared to group 1
(2/14 vs.10/14, p < 0.01 and 1/14 vs. 7/14, p < 0.05, respectively). MDA levels
decreased, and GSHPx and SOD levels increased in caecum tissues in group 2
compared to group 1 (p < 0.01, for all). The decrease in the degree of OS and BT
correlated positively.
Discussion/Conclusion: This study confirms that BT is increased in cirrhotic rats.
Reduction in contamination of MLNs in rats treated with SMT suggests that NO might
have a role in BT possibly through increasing OS.
63
Enzymes of purine degradation in lymphocytes of patients with
liver cirrhosis
L. Turecky, V. Kupcova, V. Rendekova, M. Szantova, E. Uhlikova
Introduction: Adenosine deaminase (ADA) and purine nucleoside phosphorylase
(PNP) are enzymes which participate in the degradation of purines. Interest in the
purine degradative enzymes of lymphoid cells was stimulated by the discovery that
inherited deficiency of these enzymes was associated with immunodeficiency
syndromes. It was observed that activity of both enzymes especially that of ADA was
very close connected with T-lymphoid cells.
The present study was undertaken to determine the activities of ADA and PNP in
lymphocytes separated from peripheral blood of patients with alcoholic and
nonalcoholic liver cirrhosis.
Methods: Lymphocytes were isolated from blood of 31 patients with alcoholic (17)
and nonalcoholic (14) liver cirrhosis. Enzyme activities were determined by sensitive
and specific radiochromatographic methods.
Results: Activity of ADA was decreased in lymphocytes of patients with nonalcoholic
liver cirrhosis. The activity of PNP was significantly increased mainly in alcoholic liver
cirrhosis. Index PNP/ADA was increased in patients with liver cirrhosis in comparison
to healthy controls.
Discussion/Conclusion: The increase of PNP (higher activities reported in
helper/inducer T-cells) in alcoholic liver cirrhosis is in agreement with results which
described impaired suppressor function in patients with alcoholic liver disease.
Changes of ADA and PNP activities in lymphocytes of cirrhotic patients probably
reflects the changes of functional status of lymphocytes in these patients.
64
Hepatorenal syndrome is a major complication of end stage
liver cirrhosis and has a poor prognosis
B. Appenrodt, J. Zielinski, J. Heller, T. Sauerbruch, M. Schepke
Department of Internal Medicine I, University of Bonn, Bonn, Germany
Background and aim: Hepatorenal syndrome (HRS) is a frequent and major
complication of end-stage liver cirrhosis. HRS type I shows a very poor prognosis.
However, the predictive factors for improved renal function and survival are still
unknown in patients with HRS type I.
Methods: In the present study we analysed retrospectively all patients with HRS type
I (defined by the International Ascites Club) (1) seen at our department between
1993 and 2004. Patients received standard treatment for HRS type I, i.e.
vasoconstrictors (terlipressin or midodrine) together with albumin as well as TIPS if
suitable. Furthermore, pentoxifylline was administered in 33 (36%) patients.
Patients had to fulfill the entry criteria as defined by the international ascites club.
Patients were analysed with respect to the endpoints 1) improved renal function
(defined as drop of creatinine to less than 1.5 mg/dl) and 2) survival by using
univariate analysis (p < 0.1) and multivariate Cox model in order to test for
independence (p < 0.05).
Results: 91 Patients fulfilled the entry criteria (Child A/B/C 0/13/72, alcoholic
cirrhosis 66 [73%], posthepatic cirrhosis 10 [11%] and by other causes 25 [16]);
mean follow up was 105 months ± 19.5 months and mean survival was 2.7 months
(1.5;3.8) by using the Kaplan-Meier analysis (mean [95% CI]).
Improvement of renal function occures in 29 (26%) patients. It was significantly
associated with liver function (low Child-Pugh score, p = 0.0001) and administration
of pentoxifylline (p = 0.004). Survival correlated with a low Child-Pugh score
(p = 0.0001), improvement of renal function (p=0.001) and administration of albumin
(p = 0.004).
Summary:
Our analysis 1) confirms the bad prognosis of HRS type I (only one forth responded
to treatment) and 2) shows that liver function and response to treatment are the most
important predictors of survival.
Reference:
Arroyo V. et al.; Hepatology 1996, 23: 164–176.
65
Assessment of the renal hemodynamics in liver cirrhosis with
duplex-Doppler ultrasonography
Alex. Caraba, Lelia Şuşan, Alina Păcurari, I. Romoşan
The IVth Medical Clinic, University of Medicine and Pharmacy “Victor Babeş”
Timişoara, Romania
Aim: Kidney dysfunction represents a serious complication of liver cirrhosis. Kidney
involvement secondary to liver cirrhosis is produced by an important renal
vasoconstriction, finally generating hepatorenal syndrome. Resistive index (RI) of
interlobar renal arteries, determined by means of duplex – Doppler ultrasonography,
is used for the assessment of renal vasoconstriction.
Material and methods: RI was determined with ALOKA ProSound 4400
ultrasonographic device in 15 healthy subjects (group A), 15 patients with
compensated liver cirrhosis (group B) and 15 patients with decompensated liver
cirrhosis with ascites (group C). Previous renal pathology was absent; the levels of
blood urea nitrogen and serum creatinine were normal in all patients.
Results: The values of RI were: 0.63 ± 0.05 (group A), 0.67 ± 0.02 (group B), 0.71 ±
0.01 (group C), p < 0.001. RI increased with the severity of liver cirrhosis and the
appearance of ascites.
Conclusion: RI of the interlobar renal arteries is a parameter used for the
assessment of renal hemodynamics disturbances in patients with liver cirrhosis. By
means of this parameter, it is possible to select the cirrhotic patients with high risk for
the development of hepatorenal syndrome, in order to correct the precipitating
factors.
66
Predictive diagnosis of hepatorenal syndrome
Mitrut Paul, MD, Ph.D, Genunche Amelia, MD, Ph.D
UMF Craiova, Emergency District Hospital of Craiova – The IInd Medical Clinic –
Romania
Introduction: Renal disorders of cirrhotic patients represents one of the key points
involved in the unleash of the ascites and development of its complications.
Hepatorenal syndrome is a severe complication of the cirrhotic ascites with an
extrem unfavorable prognosis.
Aim of study: Our study intends to select specific predictive parameters for
assessing the risk of renal dysfunction development.
Material and methods: We studied a group of 496 patients with hepatic cirrhosis
with ascites followed over a 6 years period. Global statistics evaluations showed a
majority of male patients (70.5%) compared to the female patients (29.5%); mean
age was 59.07 ± 6.50 years). The research protocol contained a clinical, biological
and a complete imagistic evaluation of the liver and port system, the ascitic fluid
analysis and especially tests for the evaluation of the systemic hemodynamic
changes and of the renal function (Holter monitoring of medium blood pressure and
cardiac frequency, urea and creatinin quantification, seric and urinar ionogram,
creatinin clearance and water diuresis).
Results and discussions: During the study period of 6 years, a number of
51 patients, meaning 10.8%, developed hepatorenal syndrome. Statistically the
predictive parameters helpful for early diagnosis of the hepatorenal syndrome were:
creatinin clearance, water diuresis, protein’s level of the ascitic fluid, medium blood
pressure and urinary sodium. The evaluation of these parameters has to be done in
dynamic, following a good schedule.
Conclusion: The dynamic evaluation that followed good set protocols of the
parameters witch characterize the systemic hemodynamic and renal function is the
most efficient early diagnosis method concerning these symptoms. Hemodynamic
systemic parameters and those evaluating the renal function are the most efficient in
assessing the risk of renal complication within the cirrhotic patients and is the only
way to indicate full security for hepatic transplant.
67
Predictor factors in development of
syndrome in patients with liver cirrhosis
hepatopulmonary
Amelia Genunche-Dumitrescu, P. Mitrut, Daniela Badea, M. Badea
University of Medicine and Pharmacy, Clinical Hospital of Emergency, Craiova,
Romania
Introduction: Aim was to evidentiated the relationship between severity of liver
disease, arterial blood gas pressures, clinical features, liver transaminases and
intrapulmonary vasodilation in patients with liver cirrhosis.
Methods: A prospective analysis of 133 patients from Emergency Clinical Hospital
Craiova with cirrhosis was conducted between June 2005 and January 2006. We
excluded 45 patients because of encephalopathy (17 cases) or cardiopulmonary
disorders: chronic obstructive pulmonary disease (13 cases), cardiac disorders
(9 cases), pleural effusion (6 cases).
Arterial blood gas pressures, pulmonary function tests, chest radiography, serum
AST and ALT levels were determined in all patients. Ultrasonography was done for
evidentiated of ascites and contrast echocardiography was performed to assessment
intrapulmonary vasodilation.
A comparative study was performed on two groups of patients: A group composed of
65 normoxemic patients and B group consisting of 23 hypoxemic patients (20 mild to
moderate hypoxemia and 3 severe hypoxemia). Statistical analysis was performed
using the Wilcoxon and ANOVA tests, Pearson and Spearman rank correlation.
Results: In A group, the Child Pugh class repartition was: A – 11 cases, B – 24
cases and 30 cases in C stage. In B group most patients had stage C (16 cases).
The mean duration of liver disease was 4.4 ± 2.78 years in A group and 3.12 ± 1.23
years in B group. All the B group patients had cyanosis, clubbing fingers and spider
naevi (present in only 73.9% of cases). Prevalence of ascites was higher in
hypoxemic patients (86.86%), comparativ with normoxemic patients (73.84%). ALT
and AST level was more higher in hypoxemic group. There was statistically
significant difference between mean spirometric values in hypoxemic and
normoxemic patients (p = 0.001), but spirometric values was not correlated with
serum transaminase levels (r = 0.17, p > 0.05). In whole group (88 cases), a
statistically significant relationship was evidentiated between arterial blood gas
pressures, ascites and high levels of AST (r = 0.358, p = 0.001).
Discussion/Conclusion: Presence of ascites and relatively high levels of serum
transaminase appear to be predictors of hypoxemia in cirrhotic patients without
cardiopulmonary disorder or encephalopathy.
68
Pulmonary hypertension, a rare but dreaded complication of
liver cirrhosis
Monica Marin, Olimpia Chira, Sorin Tanu, V. Mercea
The 3rd Medical Clinic, Spitalul Clinic de Urgenta "Octavian Fodor", Cluj-Napoca,
Romania
Background: Patients with liver cirrhosis and portal hypertension may develop
severe pulmonary hypertension (PH). Cardiac ultrasound (US) criteria of PH are
systolic pulmonary artery pressure (sPAP) > 35 mm and right ventricular diameter
(RVD) > 30 mm. The purpose of our study was to evaluate the prevalence of PH, by
cardiac US in a population with severe liver cirrhosis.
Material and methods: We measured the RVD and sPAP by cardiac US in
40 patients with Child-Pugh class C liver cirrhosis. Patients with known pulmonary
pathology were excluded.
Results: The etiology of liver cirrhosis was alcoholic (62.5%), viral (25%), viral and
toxic (7.5%) and unknown (5%). Cardiac US measurements were abnormal in
2 patients. The first patient with viral cirrhosis and portal hypertension had a RVD =
38 mm and a sPAP = 80 mm Hg. These findings were incidental and confirmed by
cardiac catheterization. He was treated with prostacyclin analogs with PAP
normalization. The second patient with alcoholic cirrhosis and portal hypertension
presenting with dyspnea had a RVD = 40 mm and sPAP = 60 mm. Treatment with
calcium channel blockers and nitrates was ineffective and the patient died in
3 months of cardiorespiratory failure. Sonographic measurements in the remaining
38 patients were normal: mean sPAP = 23.4 mm Hg (range 19–32) and mean RVD =
26 mm (range 20–29).
Conclusion: PH is a rare complication of liver cirrhosis which can be revealed by
cardiac ultrasound. The prevalence of PH in our patients with Child-Pugh class C
was 5%. In the absence of treatment response, its consequences are severe with
right heart failure and death.
69
Impaired cardiovagal autonomic function in patients with
chronic hepatitis C: Is there an association with sensory nerve
function and cryoglobulinaemia?
Osztovits J1, Horvath T3, Visnyei Zs1, Csák T1, Bekő G1, Lakatos PL1, Lengyel G2,
Ibrányi E4, Toth T1, Abonyi M1, Fehér J2, Kempler P1, Kollai M2, Szalay F1
1 st
1 Dept. of Internal Medicine; 22nd Dept. of Internal Medicine; 3Institute of Human
Physiology and Clinical Experimental Research, Semmelweis University, 4Szt. László
Hospital, Budapest, Hungary
Background: Autonomic neuropathy in certain chronic liver diseases is associated
with higher stage of liver cirrhosis, increased cardiovascular mortality, and lower
quality of life. It is not known, howewer, how chronic hepatits C virus (HCV) infection
influences autonomic function, and whether autonomic function relates to sensory
function and cryoglobulinaemia in these patients.
Aims: 1. to assess cardiovagal autonomic function in HCV patients and healthy
controls; 2. to examine the relation of autonomic function to sensory nerve function
and cryoglobulinaemia in HCV patients.
Methods: Cardiovagal autonomic function was assessed in 41 HCV PCR positive,
treatment-naive patients and in 38 healthy, age-matched controls, by determining
baroreflex sensitivity (BRS) and heart rate variability (HRV) indices. R-R interval and
radial artery pressure was measured simultaneously. In 25 HCV patients, peripheral
sensory nerve function was characterized by current perception thresholds, and
mixed cryoglobulin was detected.
Results: BRS and HRV time- and frequency-domain indices were decreased in HCV
positive patients as compared to healthy controls (7.3 ± 3.9 vs. 10.0 ± 5.9 ms/mmHg
for BRS, 30.8 ± 13.2 vs. 39.0 ± 18.0 ms for a HRV index [mean ± SEM]; p < 0.05). In
HCV group, patients with sensory dysfunction had lower autonomic function indices
as compared to patients with normal sensory function (p < 0.05). HCV patients with
and without cryoglobulinaemia had no difference in autonomic function.
Conclusion: We found decreased cardiovagal autonomic function in HCV infected
patients. The lower autonomic function correlated with sensory dysfunction, but was
independent of cryoglobulinaemia. Our results highlight the importance of
cryoglobulin-independent mechanisms of autonomic nerve injury in patients with
chronic hepatitis C.
70
The etiological spectrum of portal hypertension in pediatric
population – Evolution and treatment (5 years retrospective
study)
A. Constantinescu1, C. Gheorghe2, L. Gheorghe2, C. Nicolae3, I. Lupescu3,
V. Herlea4, A. Constantinescu5, I. Popescu6, C. Arion1
1
Fundeni Clinical Institute, Department of Pediatrics, Bucharest, Romania
2
Fundeni Clinical Institute, Department of Gastroenterology and Hepatology,
Bucharest, Romania
3
Fundeni Clinical Institute, Department of Radiology, Bucharest, Romania
4
Fundeni Clinical Institute, Department of Pathology, Bucharest, Romania
5
University of Medicine and Farmacology Carol Davila, Bucharest, Romania
6
Fundeni Clinical Institute, Department of General Surgery and Liver Transplantation,
Bucharest, Romania
Background: Portal hypertension (PHT) is common in children and extrahepatic
portal venous obstruction or liver cirrhoses constitute the majority of cases in our
clinic.
Aim: The etiology of portal hypertension (PHT) at children with implications in
evolution and differentiated therapeutical approach of these patients.
Methods: Between February 2001–May 2006, 152 children with PHT were included
in the study.
Patients with PHT present with gastrointestinal bleeding, hypersplenism,
splenomegaly, ascites and growth retardation.
The diagnosis is usually made by abdominal ultrasound, upper gastrointestinal
endoscopy, liver histology.
Treatment for PHT include primary prophylaxis for variceal bleeding and prevention
of repeat bleeding using drugs like beta-blockers, endoscopic sclerotherapy and
endoscopic band ligation of varices.
The children were followed endoscopically every 3 weeks and band ligation
undertaken till the eradication of esophageal varices (EV) and then every 3 months
for surprising the recurrence of EV.
Results: The characteristic of the patients were: F:M ratio = 1:1; age between
3 months and 17 years; 26 patients underwent splenectomy in average with
1.5 years ago.
In 24% of cases the etiology of PHT is extrahepatic obstruction (vena porta
thrombosis, portal cavernoma and hepatic fibrosis) and 76% are intrahepatic.
24 patients had endoscopic variceal band ligation (EVL), 5 patients had endoscopic
sclerotherapy.
116 patients with intrahepatic PHT were listed for liver transplantation.
Conclusion: Extrahepatic PHT is the most frequent cause of upper gastrointestinal
bleeding in children.
Endoscopic variceal ligation is highly effective in obliterating esophageal varices in
children.
71
Prehepatic portal hypertension in patients with
thrombosis: Clinical and laboratory characteristics
portal
E. Kitsenko*, E. Lukina, E. Sysoeva, E. Voronkova, E. Lubivy*, G. Sukhanova,
S. Vasiljev
National Research Center for Haematology, Russia
*National Research Center for Surgery, Moscow, Russia
Background: The prehepatic portal hypertension due to portal thrombosis was
believed to be a rare condition (about 10–20% of all cases of portal hypertension).
However, during the last 3 years we observed more than 60 patients with portal
thrombosis who developed PPH.
The aim of the investigation: to study clinical and laboratory signs of PPH in patients
with portal thrombosis.
Materials and methods: We studied 60 patients (mediana of age 43 years) with
portal thrombosis confirmed by Doppler sonography. The period from the first
manifestation of portal hypertension (splenomegaly, varicose dilatation of esophageal
veins) to examination in our Center varied from 4 to 480 months (mediana =
72 months). 24 patients had blood picture compatible with diagnosis of chronic
myeloproliferative disorders (MPD). Other patients had PPH due to hereditary or
acquired thrombophilia. Basic laboratory and clinical examinations were made. Also
we studied laboratory parameters concerning the hemostatic status and
thrombophilia markers.
Results: The main clinical signs were the following: splenomegaly (100%),
hepatomegaly (61%), varices (95%), bleedings from esophageal and gastric varices
(58%). The majority of patients had normal albumin, bilirubin and transaminase
levels. Cholestatic markers were increased in more than half of the patients (alkaline
phosphatase – in 56% of patients, γGT – in 52% of patients). Regarding the
hemostatic status, we observed the decrease of prothrombin level (mediana = 77)
and increase of von Willebrand factor (mediana = 195). Blood and plasma viscosity
did not differ from those in the control group. Index density of erythrocytes was
elevated in 59% of patients, rate of erythrocyte aggregation was increased in 74% of
patients.
Conclusion: Cholestatic markers and microhemorrheological disturbances
(increased erythrocyte aggregation and low deformability of erythrocytes) are the
distinctive clinical and laboratory features of PPH in patients with portal thrombosis.
We found no significant differences in these parameters in patients with portal
thrombosis associated with MPD and without MPD.
Correspondence:
Dr. Evgeny Kitsenko, Municipal Clinical Hospital № 20, 15, Lenskaja, 129327,
Moscow, Russia, E-mail: [email protected]
72
Myeloproliferative disorders in patients with prehepatic portal
hypertension
E. Sysoeva, E. Lukina, E. Semenova, E. Voronkova, N. Khoroshko, G. Frank
National Research Center for Haematology, Moscow, Russia
Background: The most frequent cause of prehepatic portal hypertension in adults is
thrombosis of portal vein or its trunks. Chronic myeloproliferative disorders (MPD)
were considered to be the main cause of thrombotic complications in adult patients.
Aim: to study the frequency of myeloproliferative disorders in patients with prehepatic
portal hypertension.
Materials and methods: 54 patients (21 males, 33 females, Median age – 43 years)
with portal thrombosis confirmed by Doppler sonography were included into this
study. The period from the first manifestation of portal hypertension (splenomegaly,
varicose dilatation of esophageal veins) to examination in our Center varied from 4 to
480 months (Median 60 months). All patients had bone marrow examination and
were screened for polymorphism of 10 genes of hemocoagulation system.
Results: The bone marrow morphology of myeloproliferative disorders was revealed
in 46% patients. The platelet level increase more 400 x 109/l was found in 35%
patients, 11% patients with MPD had normal platelets counts. The other patients had
normal pattern of bone marrow, normal blood picture or cytopenias. Combination of
MPD and polymorphisms of genes hemocoagulation system was found in 23 (43%)
pts.
Conclusions: All patients with thrombosis should be screened for chronic
myeloproliferative disorders. In case of MPD cytoreductive therapy with hydroxyurea
and anticoagulant/antiaggregant therapy should be prescribed.
Correspondence:
Dr. E. Sysoeva
National Research Center for Haematology
Novozykovsky pr. 4a
125167, Moscow, Russia
Tel/Fax 7(495) 612-4332
E-mail:[email protected]
73
Minimal hepatic encephalopathy: The role of neuropsychological assessment in the diagnosis
Janković G, Djurić Z, Mijač D, Stefanova E
Clinic for gastroenterology and hepatology and Institute of neurology, Clinical centre
of Serbia
BACKGROUND: The detection of neurological impairment by neuropsychological
tests is of crucial importance for the diagnosis of minimal hepatic encephalopathy
(minHE). There is no defined battery of tests which can be recommended as
standard diagnostic tool.
AIM: The aim of the study was to assess the value of several neuropsychological
tests in the diagnosis of minHE.
MATERIAL AND METHODS: We examined 35 patients with non-alcoholic liver
cirrhosis without clinically manifested hepatic encephalopathy, and 20 non-cirrhotic
gastroenterological patients using a battery of neuropsychological tests.
RESULTS: The results of test (units) in patients vs. controls (X ± SD), were: Trail
making test-Reitan-A (s): 74.0 ± 50.8 vs. 42.3 ± 15.6, P = 0.009; Trail making testReitan-B (s): 218.4 ± 133.6 vs. 103.6 ± 42.7, P = 0.000; Digit symbol test (N/90s):
31.6 ± 13.8 vs. 38.0 ± 9.5, P = 0.04; Hooper visual organization test (N/30s): 18.1 ±
6.7 vs. 21.5 ± 3.6, P = 0.022; The complex figure test Rey-Osterreith (N): 31.8 ± 7.7
vs. 35.5 ± 0.7, P = 0.008; Boston naming test (N/90s): 41.4 ± 10.9 vs. 46.6 ± 7.5,
P = 0.04. Results of other tests: Mini mental state examination-Folstein, The complex
figure test Rey-Osterreith 1 and 2, Benton judgment of line orientation test - phonetic
and category, Ray auditive learning test 1 and 2; Digit span - verbal stimulus; Purdue
pegboard test-left hand, right hand and l.+ r. hand, Finger tapping test-left hand and
right hand, were not significantly different between two groups.
CONCLUSION: The data suggests that Trail making test-Reitan-A and -B, Digit
symbol test, Hooper visual organization test, the complex figure test Rey-Osterreith
and Boston naming test have the potential to be used for the detection of minHE.
74
Efficacy of flumazenil administered in small repeated
intravenous doses in patients with hepatic encephalopathy
Jasna Jovic*, Nebojsa Jovic**, Tanja Pecelj-Brocic*, Branka Roganovic*
*Clinic for gastroenterology and hepatology, Military Medical Academy, Belgrade,
Serbia
**Clinic for Neurology and Psychiatry for Children and Youth, Medical School,
Belgrade, Serbia
Etiopathology of hepatic encephalopathy (HE) is unclear. Endogenous
benzodiazepines (Bz) may play a role in pathogenesis of HE, trough GABA-Bzreceptor-complex. Acting as imidazo-Bz with competitive antagonistic and partial
inverse agonistic properties on GABAA-Bz-receptor-complex, flumazenil is potentially
effective in patients with HE.
The aim of this open uncontrolled study was to evaluate efficacy and safety of
repeated small doses of flumazenil in patients with HE.
Eleven (2 female and 9 male) patients aged 24 to 74 years with proven hepatic
cirrhosis were enrolled. Etiological analysis disclosed alcohol in 9 patients, and HCV
infection and PSC + HCV each in one patient. They had 26 episodes of HE with
Portal Systemic Encephalopathy (PSE) grade 1 to 4 and PSE score between 8–16
points. Severity of cirrhosis was Child Pugh C in 22 (85%) and B in 4 (15%)
episodes. One patient had 10 episodes of HE with PSE of grade II and III during
18 months of follow-up. Precipitating factors were identified in 11 episodes (42%).
Flumazenil was administered in repeated i.v. boluses of 0.3–0.4 mg/8 hours. The
therapeutic response was evaluated after 1 mg (24 h) and 2 mg (48 h) of flumazenil,
with PSE score values (0–16 points).
Three patients were treated with lactulose and-L-ornitin-L-aspartate infusion without
clinical benefit in 6 episodes of HE. An improvement of PSE score (ranged of 2 to
12 points) after 24 and 48 hours of flumazenil administration was observed in
24 (92%) episodes. Two patients had no favorable therapeutic effect after 1 mg of
flumazenil. They died in hepatic coma within 3–5 days. Flumazenil was previously
beneficial in one of them (PSE score 16, 13 after 1 mg and 4 after 2 mg/48 h). Three
patients died, despite significant decrease of PSE score.
Both tolerability and safety of flumazenil were satisfactory.
Flumazenil in repeated small i.v. doses seems effective and safe therapy for HE in
some patients. Our favorable results perhaps are due to prevalence of patients with
liver cirrhosis of alcohol etiology.
75
Prognostic significance of bacterial infection as precipating
factors of chronic hepatic encephalopathy in cirrhosis
S. Naumovski-Mihalic, M. Katicic, V. Colic-Cvrlje, B. Papa, T. Filipec-Kanizaj,
M. Ticak, B. Sabaric, B. Skurla
Intensive Care Unit, Department of Medicine, University Hospital MERKUR, Medical
School, Zagreb, Croatia
Background: Hepatic encephalopathy is a well-recognized clinical complication of
chronic liver disease. About 30% of patients with cirrhosis die in hepatic coma. The
current study was conducted to clarify the mechanisms of level of deficiency in liver
cirrhosis and its involvement in hepatic encephalopathy via ammonia metabolism.
The aim of this study was to determine the incidence of bacterial infections in
cirrhotic patients with encephalopathy.
Methods and patients: In the study were included 235 patients (M/F 155/80) aged
24–86 yo, suffered from liver cirrhosis. They were divided into three groups according
to: the Child’s classification for cirrhotic severity (A – good, B – fair, C – poor) the
aetiology of liver cirrhosis and to the presence or absence of: ascites, splenomegaly,
esophageal varices, bilirubin level and known risk factor for hepatic encephalopathy.
All patients had upper gastrointestinal endoscopy. In all patients were measured level
of C-reactive protein, blood ammonia and other biochemical parameters. Blood, urine
and ascitic fluid cultures were systematicaly performed 1, 4 and 7 days after
admission.
Results: 189 patients (80.40%) had alcoholic cirrhosis (AC) and 46 patients
(19.60%) had non-alcoholic cirrhosis (NAC). In patients with AC 121 patients (64%)
had ascites and hepatic encephalopathy and in group with NAC 16 (32.60%) patients
(P < 0.001). In group with AC from 121 patients, 108 (89.20%) had levels of CRP
> 15 and in group NAC from 16 patients, 5 (31.20%) had levels of CRP > 15
(P < 0.001). Eighty-two bacterial infection were documented in 45 patients with AC
(68%) within 7 days of admission. In group with NAC were documented only
16 bacterial infection in 9 patients (22%), P < 0.001.
The serum CRP levels were inversely correlated with blood ammonia and with
development of hepatic encephalopathy.
Conclusion: In our study incidence of bacterial infection were significantly higher in
patients of alcoholic liver disease with hepatic encephalopathy and Childs C.
Incidence of bacterial infections were found significantly increasing with the severity
of hepatic encephalopathy.
76
The role of Helicobacter pylori infection for development of
portal encephalopathy
Daniela Neagoe, G. Ianosi, A. Saftoiu, C. Vere, Aurelia Racanel, Diana Tudorascu
University of Medicine and Pharmacy, Craiova, Romania
Aim of study: We want to evaluate the possible role of Helicobacter pylori (HP)
infection as a risk factor for development of portal encephalopathy. The pathogenesis
of Helicobacter pylori infection in human liver is not very clear.
Patients and method: We included in this study 92 patients (36 females), age
ranging 25–70 years with liver cirrhosis. 26% had Child A cirrhosis, 36% Child B
cirrhosis and 38% Child C. HP infection was diagnosed through the morphological
study of gastric mucosa biopsy.
Results: HP infection has been identified in 58.7% of cases. The proportion of
infected patients was significantly higher compared to non-infected cases (41.3%,
p = 0.001).
Portal encephalopathy was observed in 79.6% patients HP-positive and 73.7%
patients HP-negative. The level of portal encephalopathy in HP-positive group did not
differ significant statistically from HP-negative group. There was no significant
difference of portal encephalopathy forms in HP-positive versus HP-negative patients
with cirrhosis. Higher level of ammonia in HP-infected patients has been established
compared with non-infected patients.
Conclusion: HP infection can not be considered a determinate factor for
development of portal encephalopathy in patients with liver cirrhosis.
77
Hepatic encephalopathy does not correlate with depression
and quality of life in patients with liver cirrhosis: A single
centre, prospective study
Ewa Wunsch1, Iwona Zawada1, Mariusz Kaczmarczyk2, Ewa Gębala1
Piotr Milkiewicz1
1
Liver Unit, Division of Gastroenterology, Pomeranian Medical School, Szczecin,
Poland
2
Department of Laboratory Diagnostic and Molecular Medicine, Pomeranian Medical
School, Szczecin, Poland
Introduction: Hepatic encephalopathy (HE) comprises a range of neuropsychiatric
symptoms including personality abnormalities, altered consciousness and
neuromuscular dysfunction observed in patients with liver cirrhosis. It usually
correlates with the stage of liver failure but it may also occur in relatively stable
patients after well known precipitating factors.
Aim: to analyze the relationship between (HE) and selected parameters of life’s
quality, depression and clinical parameters in well defined cohort of patients with liver
cirrhosis.
Methods: Forty two, consecutive patients with liver cirrhosis, referred to the tertiary
liver centre were included. HE was assessed with psychometric tests: NTC-A, NTC-B
and Trail Making Test. Degree of depression was analyzed with McSad score and
quality of life with a modified NIDDK (National Institute of Diabetes, Digestive and
Kidney Disease) scale. MELD score was used for the clinical assessment of included
patients. Twenty healthy, age and gender matched subjects were used as controls in
psychometric tests. In statistical analysis with Spearman non-parametric correlationcoefficient test was performed and p values less than 0.05 considered significant.
Results: Thirty six (87.2%) of patients showed features of HE. No statistically
significant correlation between HE and depression (p = 0.124762); various aspects of
NIDDK score including psychological status (p = 0.197879); social status
(p = 0.325869), measures of disease (p = 0.303429) were seen. No correlation was
also observed between HE and MELD score (p = 0.930405). The only statistically
significant and negative correlation was seen between results of psychometric tests
and patients’ age (p = 0.000371).
Discussion/Conclusion: In our cohort of patients with liver cirrhosis hepatic
encephalopathy measured with commonly used psychometric tests did not show
correlatation with the degree of depression and parameters of life’s quality. It may
suggest either the significant effect of HE on the perception of life’s quality in these
patients or perhaps a disputable value of psychometric tests in this clinical context.
Correlation between younger age and superior performance in psychometric tests
may suggest better preservation of psychometric function in younger subjects with
liver cirrhosis.
78
UDCA in the treatment of primary biliary cirrhosis
Krasimira Kalinova
University Hospital, Stara Zagora, Bulgaria
Introduction: Ursodeoxycholic acid (UDCA) improves liver biochemistry in primary
biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC). Since UDCA acts
partly by reducing the intestinal absorption of hydrophobic endogenous bile salts and
is poorly absorbed from the intestine, a multiple dose regimen has been advocated.
Single dose treatment, on the other hand, may improve compliance.
Aim: The effects of a single or multiple dose regimens on liver enzymes and serum
and biliary bile salts composition were evaluated.
Methods: Twenty patients (12 PSC, 8 PBC), most with early stage disease, received
UDCA (15–20 mg kg-1 day-1) in a single dose at bed time (n = 13) or in three divided
gifts with meals (n = 14) over 3 months. Five patients had both treatment regimens in
random order with a 1-month wash-out period in between.
Results: Liver biochemistry equally improved in both groups. Biliary enrichment
(%UDCA of total bile salts, mean ± SEM) was 40.1 ± 2.4 in the single dose group vs
40.8 ± 2.8 in the multiple dose group (p = NS) and was positively correlated with
biochemical improvement (AP: r = 0.47, p = 0.02; GGT: r = 0.58, p = 0.002; ASAT:
r = 0.67, p = 0.002; ALAT: r = 0.52, p = 0.01). Biochemical improvement was not
correlated with the concentration or %UDCA in serum. Patients participating in the
cross-over design had comparable biochemical response and biliary %UDCA during
both regimens. In UDCA-treated patients, fall of serum aspartate aminotransferase,
alanine aminotransferase, alkaline phosphatase and gamma-glutamyltranspeptidase
activities started within 4 weeks after start of the trial and continued throughout the
trial period. Serum bilirubin concentration showed no significant change at the end of
the study in either of UDCA- and placebo-treated group of patients. There was no
significant difference between these two groups with respect to the frequency of
improvement of pruritus. In UDCA-treated patients, serum bile acid composition
changed markedly, though its concentration showed no significant change.
Discussion/Conclusion: Single and multiple doses UDCA have similar effects on
liver biochemistry and biliary enrichment in cholestatic liver disease. Biochemical
improvement appears to be related to biliary (but not serum) enrichment with UDCA.
79
The somatostatin analogue octreotide inhibits angiogenesis
and VEGF expression in the early, but not in advanced, stages
of portal hypertension in rats. Role of somatostatin receptor
subtype 2 downregulation during the evolution of the portal
hypertensive syndrome
Marc Mejias, Ester Garcia-Pras, Jaime Bosch and Mercedes Fernandez
Hepatic Hemodynamic Laboratory, Liver Unit, IDIBAPS, CIBEREHD, Hospital Clinic,
University of Barcelona (Spain)
Angiogenesis, the development of new blood vessels, is an important determinant of
the pathophysiology of portal hypertension, contributing to the formation of
portosystemic collateral vessels and the hyperdynamic splanchnic circulation
associated to this syndrome. Somatostatin and its analogues, like octreotide, have
been shown to be powerful inhibitors of experimental angiogenesis in vitro and in
vivo. The purpose of our study was to determine the effects of octreotide on
splanchnic neovascularization, expression of angiogenesis mediators, portosystemic
collateralization and hemodynamics in portal hypertensive rats. The expression of the
somatostatin receptor subtype 2 (SST2), which mediates the antiangiogenic effects
of octreotide, was also assessed in mesentery from sham-operated rats and 4 and 7
days after induction of portal hypertension.
Rats with portal hypertension induced by partial portal vein-ligation (PPVL) were
treated with octreotide or vehicle during 4 or 7 days, starting immediately after PPVL.
Splanchnic neovascularization (CD31 expression) as well as VEGF and SST2
expressions were determined by western blotting. Formation of portosystemic
collaterals was measured by radioactive microspheres. Hemodynamic studies were
performed by flowmetry.
Octreotide treatment during 4 days significantly decreased splanchnic
neovascularization and VEGF expression in portal hypertensive rats. Portal pressure
and heart rate were also significantly reduced by the 4-days octreotide treatment,
whereas portosystemic collateralization and splanchnic blood flow and resistance
were not significantly modified. After one week of daily octreotide injection, starting
on the day of PPVL, portal hypertensive rats escaped from octreotide therapy with
regard to inhibition of angiogenesis. The only significant effect observed after 7-days
octreotide treatment was a reduction in portal pressure. The mechanism underlying
the escape phenomenon could be related to the finding that expression of the SST2
receptor decreased progressively during the evolution of portal hypertension.
In conclusion, octreotide may be an effective therapy in early stages of portal
hypertension by reducing splanchnic neovascularization, VEGF expression and
portal pressure, but not in late stages most likely due to SSTR2 downregulation
during the progression of portal hypertension in rats.
80
Comparison of two statins efficacy on a reversibility of
experimental rat liver fibrosis
Lukivskaya O.1, Popov Y.2, Zaks J.2, Schuppan D.2, Buko V.1
1
Department of Experimental Hepatology, Institute of Biochemistry, Grodno, Belarus;
2
Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, USA
Background/Aims: Previous data indicate that statins may have antifibrotic
properties independently of its lipid-lowering efficacy. Currently, nothing is known on
efficiacy of these drugs to reverse liver fibrosis. The aim of this study was to assess
and compare the effects of two statins, fluvastatin (FLU) and simvastatin (SIM) in rat
model of thioacetamide (TAA)-induced liver fibrosis reversal.
Materials and methods: Advanced liver fibrosis was induced by TAA treatment
(200 mg/kg, i.p.) twice a week for 12 w. Resolution of fibrosis was assessed after
2 months of TAA withdrawal. During this period, fibrotic rats were daily administered
with FLU (10 mg/kg) and SIM (5 and 10 mg/kg) by oral gavage. The severity of liver
fibrosis and degree of its reversal was assessed by morphometric evaluation of liver
slides stained with Azan-Mallory, hydroxyproline (Hyp) determination and mRNA the
steady state levels of procollagen I, MMP-13 and TIMP-1 as assessed by QRT-PCR
using the TaqMan technique.
Results: The TAA treatment resulted in advanced fibrosis/cirrhosis with complete
fibrous septa formation and dramatic increase in liver hydroxyproline (Hyp) content,
procollagen I, TIMP-1 and MMP-13 mRNA expression. These signs were significaltly
impaired in rats with TAA withdrawal. Only treatment with high dose SIM (10 mg/kg)
significantly decreased the square of liver connective tissue stained by Azan-Mallory
compared to the vehicle-treated group, whereas no significant reversion was
observed in rats treated with low dose SIM (5 mg/kg) and FLU (10 mg/kg). The
tested statins did not changed total and relative HYP content in the liver.
Interestingly, both doses of SIM increased hepatic levels of interstitial collagenase
(MMP-13) mRNA expression, while FLU did not.
Conclusion: Our study suggests that statins with similar lipid-lowering properties
may have a different antifibrotic activity. Simvastatin, but not Fluvastatin
demonstrated a moderate antifibrotic effect in a model of TAA-induced liver fibrosis
reversal. This is likely due to a difference in MMP-13 induction.
81
Effect’s evaluation of the treatment with carvedilol versus
propranolol
and
isosorbide-5-mononitrate
on
hepatic
haemodynamic parameters in patients with liver cirrhosis
Tcaciuc Eugen, Matcovschi Serghei, Tcaciuc Angela, Nacu Natalia
State Medical and Pharmaceutical University “Nicolae Testemitanu”, Chair of Internal
Medicine Nr. 5, Chisinau, Moldova
Introduction: The effect of the treatment with β-blockers and nitrates for primary
prevention of variceal bleeding in patients with liver cirrhosis is well known.
The aim of the study was to evaluate the effect of carvedilol administration (β-blocker
with vasodilatation effect) versus treatment with propranolol and isosorbide-5mononitrate on hepatic haemodynamic parameters in cirrhotic patients.
Methods: The study included 65 patients with liver cirrhosis of viral B or C etiology
with different cirrhotic stage. We evaluated the portal system and hepatic artery
before and after 1 month of treatment using duplex Doppler ultrasonography.
35 patients (gr. 1) received propranolol (30–120 mg/daily) plus isosorbide-5mononitrate (20–40 mg/daily) and 31 patients (gr. 2) received only carvedilol (6.25–
12.5 mg/daily).
Results: The treatment with propranolol plus isosorbide-5-mononitrate by one hand,
and carvedilol administration by other hand, improve hepatic haemodynamic
parameters in cirrhotic patients. The diameter of portal vein significantly decreased in
both groups versus initial data (16.3 ± 0.8% vs. 15.8 ± 0.7%, p < 0.05). Portal blood
flow velocity significantly increased after 1 month of treatment in both groups (15.3 ±
0.9% vs. 14.9 ± 0.6%, p < 0.05) and hepatic artery resistance index the same
significantly decreased in both groups (13.3 ± 0.5% vs. 14.2 ± 0.6%, p < 0.05). We
didn’t find the statistical difference between carvedilol administration and treatment
with propranolol plus isosorbide-5-mononitrate.
Discussion/Conclusion: Our study indicates that the carvedilol may be
administrated for primary prevention of variceal bleeding, especially in patients with
liver cirrhosis who have intolerance to nitrates.
82
The efficacy of the treatment with Losartan and Ursofalk®
versus only Losartan on hepatic haemodynamic parameters in
patients with compensated liver cirrhosis
Tcaciuc Angela, Matcovschi Serghei, Tcaciuc Eugen, Nacu Natalia
State Medical and Pharmaceutical University “Nicolae Testemitanu”, Chair of Internal
Medicine Nr. 5, Chisinau, Moldova
Introduction: The efficacy of low-dose Losartan’s administration on hepatic
haemodynamic parameters in patients with liver cirrhosis is well known. Also, a lot of
authors mention, that the ursodeoxycholic acid improves the hepatic haemodynamic
in cirrhotic patients.
The aim of our study was to evaluate the effect of the treatment with Losartan and
Ursofalk® versus only Losartan on hepatic haemodynamic parameters in patients
with compensated liver cirrhosis.
Methods: We studied 52 patients with liver cirrhosis of viral C etiology in stage A
(according to Child-Pugh score): 28 patients (gr. 1) received Losartan (6.125–12.5
mg/daily) plus Ursofalk® (750 mg/daily) and 24 patients (gr. 2) received only
Losartan. We evaluated the portal system and hepatic artery before and after 6
months of treatment using duplex Doppler ultrasonography.
Results: The diameter of the portal vein the same decreased in both groups (12.5 ±
0.5% vs. 10.8 ± 0.4%, p > 0.05). Portal blood flow velocity significantly increased in
gr. 1 vs. gr. 2 (14.9 ± 0.6% vs 9.3 ± 0.4%, p < 0.05) and hepatic artery resistance
index significantly decreased in gr. 1 vs gr. 2 (16.3 ± 0.7% vs. 11.2 ± 0.5%, p < 0.05).
Discussion/Conclusion: Our study indicates that the administration of Ursofalk in
patients with compensated liver cirrhosis improve the hepatic haemodynamic,
probably, because they contribute to decrease the intrahepatic resistance in cirrhotic
patients.
83
Atorvastatin inhibits fibrogenesis by inhibiting Ras and RhoA
activation
Trebicka J1, Neef M2, Shir K1, Hennenberg M1, Lammert F1, Reichen J2,
Sauerbruch T1, Heller J1
1
Internal Medicine, University of Bonn, Bonn, Germany
2
Institute for Clinical Pharmacology, University of Berne, Berne, Switzerland
Background: Hepatic stellate cells (HSC) play an essential role in fibrogenesis.
Different stimuli induce transdifferentiation of HSC to myofibroblasts and synthesis of
extracellular matrix. For this process Ras- and RhoA-signaling is important
(J. Hepatol. 2000: 762). HMG CoA-reductase-inhibitors (statins) inhibit the synthesis
of lipid anchors which are necessary for membrane association and subsequent
activation of Ras and RhoA. Therefore, we investigated the effects of atorvastatin on
liver fibrogenesis and on Ras and RhoA activation in activated HSC and in bile duct
ligated rats (BDL).
Methods: BDL-rats were treated with atorvastatin (15 mg/kg/d; 7 d) at different times
(directly or 1 to 5 weeks) after BDL. Untreated BDL and sham-operated rats were
used as controls. Hepatic collagen content was determined by hydroxyproline
measurement and morphometry of Sirius-red staining. mRNA levels of TGF-β were
analyzed by RT-PCR in liver homogenates. The MMP-2 activity was analyzed by
zymography. For analysis of membrane-association of Ras and RhoA, in-vitro
activated HSC were separated into a cytosolic and membrane-fraction by
ultracentrifugation with western-blot-analysis in each fraction.
Results: Immediate treatment with Atorvastatin and one week after bile duct ligation
reduced significantly the collagen production in the liver. Later treatment (2 to
5 weeks after BDL) did not change collagen content of the liver. MMP-2-activity and
TGF-β-mRNA remained unchanged when treatment was started early, but were
significantly reduced by late treatment. Membrane-associated Ras and RhoA in
activated HSC were significantly reduced by atorvastatin incubation.
Discussion: Atorvastatin treatment during early stages of fibrosis reduces collagen
formation and subsequent liver fibrosis in BDL rats. The underlying mechanism might
be the inhibition of hepatic stellate cell activation through the inhibition of Ras and
RhoA activation. After establishment of fibrosis we found indirect hints for HSC
deactivation (reduced MMP-2, TGF-β), but no fibrolysis in our model.
84
Urotensin II receptor antagonist palosuran lowers portal
pressure by activation of mesenterial RhoA/Rho-kinase
signalling in liver cirrhosis
Trebicka J, Eckhardt A, Hennenberg M, Biecker E, Lammert F, Sauerbruch T,
Heller J
Internal Medicine, University of Bonn, Bonn, Germany
Background: Splanchnic vasodilation in liver cirrhosis is responsible for portal
hypertension. Urotensin II (UII) a vasodilator of mesenteric vessels is increased in
liver cirrhosis. Recently we have shown that the UII-receptor antagonist, palosuran,
causes splanchnic vasoconstriction and decreases portal hypertension in liver
cirrhosis (Hepatol. 2006; 44: 202A). Here, we investigated the effect of palosuran on
the vasoactive signalling in mesenteric arteries (SMA).
Methods: Bile duct liagted rats (BDL) were treated with palosuran (30 mg/kg/d; 3 d).
In SMA of sham-operated and cirrhotic BDL-rats UII plasma levels and nitrite/nitrate
(NOx) production were investigated via ELISA and Griess-reaction. In SMA of these
rats RhoA, Rho-kinase and eNOS mRNA levels and protein expression were
measured via rt-PCR and Western blot analysis. The activity of Rho-kinase and NOeffector, protein kinase G (PKG), were investigated by the phophorylation of their
substrates, respectively moesin at Thr-558 and VASP at Ser-239 (Westernblot
analysis with phosphospecific antibodies).
Results: UII-plasma levels were higher in BDL- than in sham-operated rats. In SMA
of BDL-rats Rho-kinase expression and activity was decreased when compared to
vessels of sham-operated rats. eNOS-mRNA level, protein expression, NOx
concentration und PKG-activity was increased in SMA of BDL-rats when compared to
those of sham-operated rats. Palosuran increased the mRNA levels of RhoA, Rhokinase and eNOS, as well as the Rho-kinase-activity, while the NOx-concentration
and PKG-activity were decreased by palosuran in BDL-rats.
Discussion: Palosuran causes splanchnic vasoconstriction via increased activity of
RhoA/Rho-kinase signalling and decreased activity of NO/PKG signalling in
mesenteric arteries of rats with secondary biliary cirrhosis. This vasoconstriction in
the splanchnic vasculature led to decreased portal venous inflow and reduced portal
pressure. Palosuran might represent a therapeutic option in liver cirrhosis with portal
hypertension.
85
Antioxidant taurine treatment in experimental liver fibrosis
1
Uzunalimoglu O, 2Mas MR, 3Vural SA, 4Deveci S, 5Akay C, 1Isik AT, 4Bozdayi M
1
2
Gulhane School of Medicine, Department of Internal Medicine, Etlik 06018 Ankara,
Turkey
3
Department of Pathology, Veterinarian Faculty, Ankara University. Ankara, Turkey,
4
Gulhane School of Medicine, Department of Pathology, Etlik 06018 Ankara, Turkey
5
Gulhane School of Medicine, Department of Toxicology, Ankara, Turkey
Introduction: Liver fibrosis, a healing response to injury, results from the
overproduction and diminished degradation of matrix proteins in the sinusoids. The
main source of excessive collagen synthesis in the liver is the hepatic stellate cells
(HSCs). Oxidative stress (OS) has an important role in fibrogenesis. HSC apoptosis
has been proposed to be involved in spontaneous recovery (SR) of liver fibrosis. We
studied the effects of a potent antioxidant taurine on experimental liver fibrosis.
Methods: Thirty male Sprague-Dawley rats were injected SC with CC14 for
12 weeks to induce liver fibrosis. After induction of fibrosis the rats were divided into
three groups. Group I (n = 15) was left for SR and Group II was treated with taurine
1200 mg/kg/day. The rats were killed after four weeks of treatment, and liver tissue
OS parameters, histopathological fibrosis scores, alpha-SMA (+) HSC counts and
apoptotic HSCs and hepatocytes, and were determined.
Results: Taurine treatment improved fibrosis. Tissue SOD, MDA, and GSHPx levels
improved significantly only in Group II (p < 0.01, for all). Fibrosis scores reduced
significantly in Group II Group I (p < 0.05). alpha-SMA (+) cell counts also reduced in
Group II when compared to Group I (p < 0.001). Hepatocyte apoptosis in the injured
liver increased after taurine treatment (p < 0.03). Activated HSC apoptosis increased
significantly in Groups II compared to Group I (p < 0.002).
Discussion/Conclusion: Taurine improved experimental hepatic fibrosis through
induction of activated HSC apoptosis and thereby accelerating SR capacity of the
liver, which may be another potential treatment approach.
86
Nutritional state in alcoholic liver cirrhosis
Jadwiga Daniluk1, Ewa Krzyżanowska2, Agnieszka Zwolak2
1
Clinic and Department of Gastroenterology, University Medical School,
Jaczewskiego 8, 20-950 Lublin, Poland
2
Chair and Department of Internal Nursing, University Medical School, Jaczewskiego
8, 20-950 Lublin, Poland
Background and aims: The aim of the study was to access nutritional state of
patients in different stages of alcoholic liver cirrhosis (Child A, B and C).
Methods: The study population consisted of 100 patients with alcoholic liver cirrhosis
and control group of 25. Nutritional status was assessed by anthropometry: body
mass index (BMI), triceps skin fold thickness (TST), mid-arm muscle circumferences
(MAMC) and laboratory parameters: albumins, total blood protein and total
lymphocyte count.
Results: Compared to controls, all groups of cirrhotic patients showed higher
malnutrition with greater impairment in advanced stages of cirrhosis. Energy
malnutrition, defined as BMI, TST and MAMC below the 50% of standard values, was
found in 15%, 32% and 19% of the study population, respectively. Malnutrition,
defined as TST was present in 25% patients of Child A, 31.25% of B and 43.75% of
C (11.53 ± 4.64 mm vs. 10.33 ± 4.33 mm vs. 9.09 ± 4.20 m, respectively). Energy
malnutrition, defined as MAMC was found in 12.5% of patients in Child A, 12.12%, B
and 31.43% Child C. Protein malnutrition (low albumin, and total protein
concentrations) were much more frequent (47% and 20%) than energy malnutrition.
Serum proteins correlated with the degree of liver function impairment (Child A 4.45 ±
0.85, Child B 4.42 ± 0.68, Child C 3.77 ± 0.47). Multivariate analysis showed
significant statistical differences between examined groups of patients which were
associated with the Child-Pugh score in the level of albumins (p < 0.0001), MAMC
(p = 0.003) and TST (p < 0.05).
Conclusions: The study showed that protein-energy malnutrition is widely prevalent
in cirrhotics, with greater impairment in more advances stages of cirrhosis. The
greatest disorders of nutritional status were observed in patients of Child C cirrhosis.
87
Affective alterations: Determinant factor of sleep disorders in
cirrhotic patients
Mauricio González, Jaime Poniachick, Miguel Cumsille*, Ana Madrid, Javier Brahm
Centro de Gastroenterología, Hospital Clínico José Joaquín Aguirre, Universidad de
Chile
*Departamento de Bioestadística, Facultad de Medicina, Universidad de Chile
Introduction: Cirrhotic patients frecuently suffer sleep disorders, but the pathogenic
mecanisms are not totally well-known. The aims of this study were to determine the
prevalence of insomnia in these patients, and to correlate it with affective alterations,
severity and cirrhosis’ etiology.
Methods: Thirty four cirrhotics without current alcoholic habit nor concomitant
pathologies and thirty healthy fellows, paired by age, sex, educational and
socioeconomic status, were interviewed. To each group, questionnaires to determine
sleep disorders (Pittsburg Sleep Quality Index) and affective dysfunctions (Hamilton
scales) were applied. We also looked for subclinic encephalopathy with
neurosicological tests (Number Connection Test and Digit Symbol Test). Patients’
clinical characteristics were analyzed considering severity (Child Pugh’s
classification) and etiology.
Results: Insomnia was present in 52.9% of cirrhotic patients and 26.6% of controls
(odds ratio = 3.09; p = 0.035). Cirrhotic patient’s probability of depression was 0.32
and anxiety was 0.38, that increased to 0.61 (OR = 13.6; p = 0.000) and 0.6
(OR = 11.6; p = 0.001) respectively, in presence of insomnia. Cirrhosis severity and
etiology didn’t correlate with insomnia (p > 0.05).
Discussion/Conclusion: This study shows a high prevalence of insomnia in cirrhotic
patients. The affective alterations are pathogenic factors of cirrhotic insomnia. The
severity and the etiology of liver damage don’t determine presence of insomnia.
88
Impaired health-related quality of life in Romanian patients with
hepatic cirrhosis
Cristina Pojoga, D.L. Dumitraşcu
3rd Medical Department, University of Medicine and Pharmacy “Iuliu Haţieganu”,
Cluj-Napoca, Romania
Introduction: Cirrhosis is an invalidating condition leading to the impairment of
quality of life. We looked for the impact of hepatic cirrhosis on health-related quality
of life. We also investigated relationship between the level of transaminases and the
scores of the quality of life questionnaire.
Methods: We investigated 37 patients with hepatic cirrhosis (10 viral, 27 alcoholic,
5 B + C, 25 males, 12 females). The control group consisted of 35 healthy persons
(17 males, 18 females). Both groups completed the SF-36 Health Survey.
Results: Significant differences between the two groups for every domain of quality
of life (physical functioning, role disability: physical, general health, vitality, social
functioning, role disability: emotional, mental health) were recorded. We did not find
any significant correlation between the level of transaminases and every item of the
health-related quality of life questionnaire.
Discussion/Conclusion: Patients with hepatic cirrhosis have an impaired quality of
life as estimated by SF-36 Health Survey.
89
Fatigue, depression, and functional disability in patients with
liver cirrhosis
G. Stefanescu, C. Stefanescu
Institute of Gastroenterology and Hepatology, University Hospital of Psychiatry, Iasi,
Romania
BACKGROUND: Previous studies reported an impairment of both the physical and
mental dimensions of quality of life in patients with cirrhosis. Very few data are
available on the psychological impact of the disease and its relation to liver function.
The aim of study is to examine the extent to which depressive symptoms correlate
with severity of fatigue and functional disability in patients with liver cirrhosis.
METHODS: 82 patients with liver cirrhosis were evaluated using two instruments: the
clinical psychiatric interview and Hamilton Depression Scale (HAMD).
Clinical and laboratory data were collected using standardised forms.
The prevalence of fatigue and its association with clinical manifestations
dermatological, rheumatological, neurological and nephrological manifestations; and
with comorbidities (diabetes; arterial hypertension, etc.) were assessed. Then, using
multivariate analysis, we identified demographic, biochemical, immunological and
virological factors associated with the presence of fatigue.
RESULTS: The HAMD scores were indicative of a depressed mood in 42 patients
(56.1% of patients), in relation to the presence and severity of clinical symptoms.
Depressed and nondepressed patients did not differ with regard to demographics.
Depressive symptoms was highly correlated with severity of fatigue and Child-Pugh
score (a comprehensive measure of disease severity) while measures comorbid
medical illness were not. Severity of depressive symptoms was associated with
functional disability and somatization.
Fatigue was present in 52.43% of patients (95% confidence interval 50–55). In
17.07% of patients (95% confidence interval 15–19) fatigue was severe, impairing
activity.
In univariate and multivariate analyses, fatigue, in comparison with the absence of
fatigue, was associated with female gender, age over 50 years, purpura and severity
of cirrhosis. There was no significant association between fatigue and the following
characteristics: etiology of cirrhosis, alcohol consumption, abnormal thyroid function.
CONCLUSIONS: Patients with cirrhosis have signs of psychological distress and
depression, as assessed by HAMD and clinical psychiatric interview, in relation to the
severity of liver disease. Disability and fatigue are closely related to depression.
90
Total paracentesis with albumin infusion in patients with
ascites before radiofrequency ablation (RFA) of hepatocellular
carcinoma (HCC)
J. Genov, N. Grigorov, R. Mitova, B. Golemanov
Clinic of Gastroenterology, University Hospital “Queen Joanna”
Sofia, Bulgaria
Aim: To assess the feasibility of total paracentesis (TP) in patients with cirrhosis as a
part of conditioning for percutaneous RFA of HCC.
Methods: 63 patients with 78 HCC (size 5.03 ± 3.05 cm) were treated with
ultrasound-guided monopolar RFA and 21 patients with 29 НСС (size 5.91 ± 3.22) –
with multipolar RFA. In the first group 24 patients were Child B and C, in the second
– 10 patients were Child B. Eleven patients of group I and 5 patients of group II had
ascites despite diuretic treatment. In these cases TP with simultaneous intravenous
albumin infusion 8 g/l ascitic fluid was performed on the day before RFA.
Results: None of the patients developed complication of TP. Complete destruction of
all lesions was achieved in 59% of the patients with Child B in group I and in 60% in
group II, without significant difference in the effectiveness of RFA between patients
without ascites vs. patients treated with TP in the two groups. Also there was not
difference in the effectiveness of RFA between patients with Child A vs. Child B in
the two groups, in the rate of major complications of RFA and rate of local
recurrences during the follow-up between patients without ascites and patients
treated with TP.
Conclusion: TP is effective and safe option in patients with cirrhosis and HCC
before RFA.
91
S-100 and GFAP-positive cells in human liver cirrhosis and in
human hepatocellular cancer
1
Gulubova M, 2Vlaykova T, 2Velev V
1
Dept. of General and Clinical Pathology, 2Dept. of Chemistry and Biochemistry,
Medical Faculty, Trakia University, Stara Zagora, 11 Armeiska Str, Stara Zagora,
6000, Bulgaria, E-mail: [email protected]
Introduction: Intrahepatic innervation plays an important role in the regulation of
hepatic microcirculation, biliary function, glycogen and lipid metabolism. We have
already shown that liver sinusoids contained substance P-, VIP-, serotonin-,
somatostatin-, and GFAP-positive peptidergic nerve fibres. The aim of the study is to
describe S-100-positive and GFAP-positive nerve structures in the liver in cirrhosis.
Methods: We investigated immunohistochemically 12 patients with cirrhosis and
12 with primary hepatocellular cancer (HCC), as 5 of them had cirrhosis. The
biopsies with cirrhosis were surgically resected for the purpose of diagnosis.
Results: In cirrhosis S-100-positive nerve fibres were located in the enlarged portal
tracts and septa as a large nerve bundle or as single nerve fibres in the connective
tissue and around bile ducts. In the sinusoids S-100-positive nerve fibres were
located in the space of Disse as long fibres attached to hepatocytes. GFAP-positive
nerve bundles and nerve fibres were observed in enlarged portal tract. Single GFAPpositive nerve fibres were detected in the septa. In the lobule there could be detected
GFAP-positive nerve fibres and cells. In HCC S-100-positive nerve fibres and GFAPpositive nerve fibres were distributed mainly in the capsule of the tumour. In the
tumour tissue in some tumours we could observed single S-100- and GFAP- positive
nerve In cirrhosis S-100-positive nerve fibres were located in the enlarged portal
tracts and septa as a large nerve bundle or as single nerve fibres in the connective
tissue and around bile ducts. In the sinusoids S-100-positive nerve fibres were
located in the space of Disse as long fibres attached to hepatocytes. GFAP-positive
nerve bundles and nerve fibres were observed in enlarged portal tract. Single GFAPpositive nerve fibres were detected in the septa. In the lobule there could be detected
GFAP-positive nerve fibres and cells. In HCC S-100-positive nerve fibres and GFAPpositive nerve fibres were distributed mainly in the capsule of the tumour. In the
tumour tissue in some tumours we could observed single S-100- and GFAP-positive
nerve fibres.
Discussion/Conclusion: In conclusion we may say that nerve fibres although
reduced in number appear in septa and in liver lobules in cirrhosis and in the tumour
stroma of HCC.
92
Frequency of hepatocellular carcinoma in chronic viral
infections
Dr Daniela Svarc
Department for infectious disaeses of liver and biliary tract, Clinic for infectious
diseases, Clinical center Vojvodina, Novia Sad/Vojvodina, Serbia
Introduction: Hepatitis B and hepatitis C chronic viral infections are two major risk
factors for hepatocarcinogenesis. Patients diagnosed with hepatocellular carcinoma
(HCC) in the majority of cases have positive viral markers. HBAg is positive in around
36.5% of the patients, antiHCV in around 26% of the patients and as much as 30.6%
have both HBsAg and antiHCV positive.
Aim: To determine the frequency of HCC in patients with chronic viral hepatitis and
also to asses the stage of the carcinogenesis when the diagnosis is established.
Method: A retrospective study reviewing medical histories of 606 patients with
chronic hepatitis B (216) and hepatitis C (390) during a 4 year period (2003–2006).
All patients had regular 3 month period check-ups which included ultrasound and
basic laboratory findings.
Results: In total 35 (5.8%) patients have developed HCC in the 4 year period
(17 HBsAg+, 18 antiHCV+). On average HCC was diagnosed in HBsAg+ in 1.97%
patients per year and in antiHCV+ in 1.1% of the patients per year with a steady rise
from 2003 to 2006. At the time of diagnosis 34.3% of the patients already had
thrombosis of portal/mesenteric veins and 37% had multiple focal leasions. Less then
half (42.8%) had elevated alfa fetoprotein levels.
Conclusion: Although the yearly incidence of HCC in patients with chronic viral
hepatitis is not very high they are too often recognised in late stages of the disease.
93
Expression of the reactive oxygen species-detoxifying
enzymes, GST-pi and Cu/Zn-SOD in cirrhosis and
hepatocellular carcinoma
1
Vlaykova T, 2Gulubova M
Dept. of Chemistry and Biochemistry, 2Dept. of General and Clinical Pathology,
Medical Faculty, Trakia University, Stara Zagora, 11 Armeiska Str, Stara Zagora,
6000, Bulgaria, E-mail: [email protected]
1
Introduction: Both liver cirrhosis and hepatocellular carcinoma (HCC) are
multifactorial processes and there is increasing evidence that the free radical
damage of hepatocytes is a possible mechanism involved in the pathogenesis of
those diseases. In this respect we aimed to study the changes of expression of two
enzymes implicated in cellular defense against excessive reactive oxygen species,
Cu/Zn-SOD (SOD1) and GST-pi in liver cirrhosis and HCC.
Methods: The expression of the enzymes was studied immunohistochemically in
routinely processed formalin-fixed paraffin-embedded biopsies obtained from
12 patients with liver cirrhosis and 12 patients with HCC (7 male and 5 female
patients aged between 25 and 82 years). The biopsies with cirrhosis were surgically
resected for the purpose of diagnosis.
Results: Cytoplasmic GST-pi immune deposits were observed in all of the biopsies
both from active and inactive cirrhosis, as in active cirrhosis the GST-pi expression
was more frequently stronger than in inactive. Very strong immune signal (+++) was
observed in the hepatocytes in areas with balloon degeneration. The epithelium of
small bile ducts was notably stained (+++) with anti-GST-pi antibody. The SOD1
immune signal was distributed to hapatocytes mainly in the patterned of cytoplasmic
diffusion and the level of this signal was markedly weaker compared to GST-pi.
In HCC we observed GST-pi expression with varying pattern of staining. There were
samples with weak immune signal (+, 17%), with strong (++, 33%) and very strong
signal (+++, 50%). The level of GST-pi expression in tumor cells was significantly
higher than that in adjacent “normal” liver. Analogously to the liver cirrhosis, the
expression of SOD1 in HCC was weak, especially comparing to the peritumoral liver
tissues
Discussion/Conclusion: The finding suggest that the changes in the expression of
ROS-detoxifying enzymes are possibly implicated in the pathogenesis of liver
cirrhosis and HCC via compromising the response to increased free radical
production, produced by the liver injuring factors, such as alcohol, toxins, viruses and
different hepatic carcinogens.
Key words: cirrhosis, hepatocellular carcinoma, GST-pi, SOD1, immunohistochemistry
This work is done with the financial support of research project VU-L-05, funded by
the Ministry of Education and Science, Bulgaria
94
Orthotopic liver transplantation for chronic hepatitis B
Fraňková S, Trunečka P, Šperl J, Adamec M, Špičák J, Vítko Š
Transplantcentre, Institute of Clinical and Experimental Medicine, Prague, Czech
Republic
Introduction: Successful outcome of orthotopic liver transplantation (OLT) in patient
with chronic hepatitis B is influenced by efficious hepatitis B recurrence prophylaxis in
the liver graft. It has been proved that the recurrence rate of HBV infection in patients
without HBV recurrence prophylaxis exceeds 80% and frequently leads to graft loss.
Aim: The study is aimed on evaluation of the current experience with liver
transplantation for chronic hepatitis B in IKEM.
Patients and methods: Liver transplantation for HBV infection has been performed
between 1999 and 2007 in 20 patients of mean age 51 ± 10 years (1 patient with
fulminant liver failure, 4 with hepatocellular carcinoma and 15 with end stage liver
disease). One of the patients had undergone kidney transplant and 1 patient lung
transplant before. Prior the OLT, 7 patients were treated with lamivudine, 3 with
adefovir and 10 patients were not receiving any antiviral treatment. At the time of
OLT and in posttransplantation period, prophylaxis with hyperimmune
immunoglobulin (HBIG) was administrated together with lamivudine (17 patients) or
adefovir (3 patients). The length of post-OLT follow-up was 0–8 years (mean
32 months).
Results: In our cohort, recurrent HBV graft infection appeared in 4 (20%) patients
after 4. 7. 9. and 12. month posttransplant, respectively, who were given combined
prophylaxis. Three of these patients were on lamivudine, one on adefovir, the HBIG
prophylaxis was stopped after HBs antigen positivity was revealed. Patients with
lamivudine prophylaxis were switched on adefovir, lamivudine was added to the
patient with adefovir prophylaxis. No graft was lost in these patients, in one patient
with high necroinflammatory activity, serological elimination of HBs antigen was
achieved after one year of adefovir therapy.
Three deaths occurred, 2 due to infectious complication, one due to recurrence of
hepatocellular carcinoma.
Conclusion: The efficacy of combined HBV recurrence prophylaxis is high, however,
recurrence can occur even when sufficient level of HBIG in combination with either
lamivudine or adefovir is kept. Although HBV recurrence prophylaxis considerably
increases the treatment costs, patients undegoing OLT for HBV constitute nowadays
a group with high survival rate.
95
Plasma value of endothelin-1 versus renal and liver function in
patients after orthotopic liver transplantation in long-term
follow up
Gottfriedová H, Schück O, Karasová L, Śtollová M, Malý J, Skibová J, Trunečka P,
Špičák J
Institute for Clinical and Experimental Medicine, Prague, Czech Republic
Introduction: Reduced renal and liver function after orthotopic liver transplantation
(OLT) is a significant high-risk factor of morbidity and mortality. The role of endotelin1 (ET-1) level in regulating renal and liver function after OLT remains uncertain. We
analyzed relationship between plasma value of endotelin-1 and level of renal and
liver function in patients after OLT in long-term follow up.
Methods: In 61 patients (31 M, 30 F, 51.4 ± 10.7 years old) in different time after
OLT (23.7 ± 20.8 [1–86] months) were examined renal function (serum creatinine
[Scr], creatinine clearence [Ccr], inulin clearence [Cin], and serum cystatine [Scyst]),
ET-1, blood pressure, number of hypotensive drugs, TGF-β, serum bilirubin (Sbil),
AST, ALT, protrombin time, serum cholesterol and triglycerides (Schol, Stg) and dose
of immunosuppressive agents.
ET-1 was also examined in 2 control group: 10 healthy volunteers and 10 patients
with end stage liver disease (ESLD) waiting for OLT.
Results: 1. in patients after OLT ET-1 values are higher in comparison with patients
with ESLD (p = 0.003**, p = 0.0001***, p = 0.0014**, p = 0.028*), values remain
elevated in long-term follow up (5 years) but there is a trend of decline. ET-1 values
in patients with ESLD are higher in comparison with healthy volunteers
(p = 0.0025**). 2. between ET-1 and level of renal function in patients after OLT no
correlation was found. 3. between ET-1 and blood pressure, number of hypotensive
drugs, dose of immunosuppressive agents, , Schol, Stg and TGF-β correlation was
not found. 4. correlation was found between ET-1 and liver function – Sbil (r = 0.328,
p < 0.05*) and ALT (r = 0.27, p < 0.05*).
Discussion/Conclusion: Plasma value of ET-1 in patients after OLT in long-term
follow up is elevated but there is a trend of decline. We did not show the influence of
ET-1 on the decline of renal function after OLT. Results suggest that there is a
relationship between ET-1 and liver function in patients after OLT.
96
Absence of teratogenicity of sirolimus used during early
pregnancy in a liver transplant recipient
Jankowska I1, Oldakowska-Jedynak U2, Jabiry-Zieniewicz Z3, Cyganek A3,
Pawlowska J1, Teisseyre M1, Kalicinski P4, Pączek L2, Socha J1
1
Department of Gastroenterology, Hepatology and Immunology, The Children’s
Memorial Health Institute, Warsaw, 2Department of Immunology, Transplantology
and Internal Diseases University Medical School, Warsaw, 3Department of Obstetrics
and Gynecology, University Medical School, Warsaw, 4Department of Pediatric
Surgery and Organ Transplantation, The Children’s Memorial Health Institute,
Warsaw, Poland
Introduction: Sirolimus (Sir) was introduced for renal transplantation in 1999, and
recently has been used by an increasing number of liver transplant centres. There is
little information on the possible effect of Sir on the human fetus.
Methods: The aim of our study was present the case of a successful delivery in a
21–year old woman after liver transplantation (LTx) treated with Sir during early
pregnancy.
Results: LTx was performed due to liver cirrhosis in the course of biliary atresia at
the age of 18 years. One year after Ltx, Sir was introduced due to renal dysfunction
(GFR /1.75 m2 – 46 ml/min). Tacrolimus (Tac) dose was reduced by half to maintain
the level of 4–8 ng/ml. The regimens of Sir, steroids and Tac were well tolerated by
the patient, the liver function was good and renal function improved significantly
(5 months later GFR /1.75 m2 – 75 ml/min). Three years after LTx the patient
developed amenorrhoea and ultrasonography revealed a 6-week gestation. Sir was
immediately discontinued but prednisone and Tac were maintained. The course of
pregnancy was uneventful. Due to fetal intrauterine threatening asphyxia the
pregnancy was finished by caesarean section (39th gestational week). The patient
was delivered of a healthy, weighting 2950 g, Apgar 10, female infants. Postoperative
course was uncomplicated. There was no congenital malformation or unusual
infections. Postnatal follow-up showed normal physical development.
Discussion/Conclusion: The course of pregnancy in our patient treated in the first
6 weeks Sirolimus was uneventful; there was no evidence of teratogenicity Sir in the
infant.
97
Recurrence of nonalcoholic steatohepatitis in transplanted
liver in a 13-year-old boy
Jankowska I1, Piotr Socha1, Pawłowska J1, Teisseyre M1, Gliwicz D1, Kaliciński P2,
Cielecka-Kuszyk J3, Socha J1
1
Department of Gastroenterology, Hepatology and Immunology, 2Department of
Pediatric Surgery and Organ Transplantation, 3Department of Pathology, The
Children’s Memorial Health Institute, Warsaw, Poland
Introduction: Nonalcoholic steatohepatitis (NASH) is a medical term used to
describe liver biopsy findings of fatty change and lobular hepatitis in the absence of
excess alcohol intake or underlying disease. The risk factors for NASH include
obesity, hyperlipidemia and obesity-related diabetes. Although fibrosis is common in
pediatric NASH, there are only a few report of NASH-related cirrhosis, which maybe
an indication for liver transplantation (LTx). Little information is available regarding
outcome of the liver allograft in patient with NASH related liver cirrhosis undergoing
LTx.
Methods: The aim of our study was to report on recurrent NASH in the transplanted
liver in a boy after hypothalamic tumor operation.
Results: The boy developed NASH and liver cirrhosis after surgery for hypothalamic
tumor that seemed to be related to severe obesity (BMI – 32.3, > 97% centile for
age). Six months after LTx transaminases activity increased, fatty infiltration and
hepatocellular degeneration in transplanted liver were found and recurrence of NASH
in the graft was diagnosed. As weight reduction after dietary counseling could not be
obtained and insulin resistance (fasted insulin = 8.2 µLU/ml, HOMA-IR = 2.2) was
diagnosed. Metformin therapy was started. Transaminases activity decreased (AlAT
from 322 U/l to 153 U/l, AspAt from 223 U/l to 83 U/l) within 6 months. Metformin
therapy has been continued for 2 years with a good effect. Follow-up period is now
5 years.
Discussion/Conclusion:
1. NASH may reccur after LTx if the obesity related risk factors are not diminished.
2. Metformin therapy should be considered in NASH patients non responsive to
dietary therapy.
98
Activation of latent TGF-β inside the hepatocyte leads to target
gene expression via autocrine, intracellular signalling –
Relevance for the initiation of acute and chronic liver injury
Gressner OA, Lahme B, Siluschek M, Rehbein K, Weiskirchen R, Gressner AM
Institute of Clinical Chemistry and Pathobiochemistry, University-Hospital of the
RWTH Aachen, Aachen, Germany
Introduction: TGF-β is regarded as the profibrogenic master cytokine. We aimed at
analyzing the regulatory link between TGF-β target gene- (i. . CTGF-) induction and
intracellular demasking of latent TGF-β in hepatocytes (PC) of healthy and injured
livers.
Methods: Primary rat PC were cultured in entirely serum- (i. . TGF-β-) free
conditions. Analyses performed included immunoblotting, reporter-assays, metabolic
labelling and immunocytochemistry (with/without the addition of specific
activators/inhibitors of TGF-β signalling) as well as Immunohistochemistry of healthy
rat livers and those subjected to bile duct-ligation (BDL), intraperitoneal D-GalN or
CCl4-injection.
Results: Immunocytochemistry of primary PC revealed a significant increase of
intracellular TGF-β within the first 24 h of culture under entirely TGF-β free
conditions. This correlated with spontaneous Smad2-phosphorylation and a strong,
time-dependent expression of CTGF. CTGF expression was abrogated by inhibition
of the intracellular kinase domain of the TGF-β-R1/Alk5-receptor. This phenomenon
was not detectable following broad spectrum inhibition of extracellular TGF-βsignalling.
TGF-β demasking was confirmed by immunohistochemistry, where an increase of
intracellular TGF-β and CTGF was observed in BDL, D-GalN or CCl4-treated rats
while healthy livers remained TGF-β and CTGF negative. In injured livers, TGF-β and
CTGF positive PC mostly showed a patch-like, periportal distribution.
Conclusion: We claim that intracellular TGF-β is demasked in PC during conditions
of cellular stress which enables it to trigger specific TGF-β dependent signalling
pathways within the PC that lead to target gene (i. e. CTGF) expression. This might
play an important role in the pathogenesis of apoptosis during early liver injury which
finally leads to fibrosis.
Author Index to Poster Abstracts
(Name - Poster Number)
Abdel Moety, A.A.
Abdulchakov, S.
Abe, S.
Abonyi, M.
Abraldes, J.G.
Adamec, M.
Adjarov, D.G.
Akay, C.
Andreeva, D.I.
Angelova, L.
Antonov, K.
Appenrodt, B.
Aravena, E.
Arion, C.
Assy, N.
Atanassova, A.
46, 50
11, 20
33
69
18
94
38, 39
62, 85
11, 14, 15, 20
5
38
64
1
70
43
3
Baddour, N.M.
Badea, D.
Badea, M.
Banciu, C.
Baruch, Y.
Bataller-Sifre, R.
Beca, C.B.
Becker, S.
Bekö, G.
Belyaeva, T.V.
Berecz, A.
Berg, T.
Bidlo, C.
Biecker, E.
Bielanski, W.
Bielecka, M.
Bilgin, G.
Bilzer, M.
Blonska-Fajfrowska, B.
Bocheva, S.
Boicean, A.B.
Boitan, M.
Bojic, B.
Bojic, D.
Bosch, J.
Bozdayi, M.
Bozkaya, H.
Brahm, J.
Breitkopf, K.
Buko, V.U.
50
60, 67
60, 67
53, 58
43
18
31, 34, 47, 48
22
69
27
55, 56
28
31, 34, 47, 48
84
30
52
21
13
52
39
31, 34, 47, 48
31, 34, 47, 48
9
9
18, 79
85
62
87
12, 23
80
Burganova, G.R.
Bystron, M.
14, 15
13, 19
Caraba, A.
Catana, A.
Celinski, K.
Cheremina, N.A.
Chicea, L.
Chira, O.
Chirileanu, O.
Cichoz-Lach, H.
Cielecka-Kuszyk, J.
Cijevschi, C.
Cipaian, C.C.
Ciuclan, L.
Colic, A.
Colic, N.
Colic-Cvrlje, V.
Comert, B.
Constantinescu, A.
Contesse, F.
Crangaciu, V.
Csák, T.
Cumsille, M.
Cyganek, A
65
31, 34, 47, 48
24
11
31, 34, 47, 48
68
53
24
97
41, 54, 57
31, 34, 47, 48
12, 23
59
59
75
17, 21, 62
70
1
34
69
87
96
Damian, D.
Daniluk, J.
Dapcevic, B.
Dascalu, D.
Dascau, C.
Deac, M.
Demir, H.
Deveci, S.
Dienes, H.P.
Djordjevic, J.
Djorovic, S.
Djuric, Z.
Dooley, S.
Drebber, U.
Drug, V.
Dublina, E.S.
Dumitrascu, D.L.
55, 56
86
9, 59
31, 34, 47, 48
58
31, 34, 47, 48
51
85
25
9
9
73
12, 23
25
54, 57
2
88
Eckhardt, A.
El Saed, H.
El Sheikh, R.A.O
16, 84
50
46
Ensari, A.
Erden, E.
Esaulenko, E.V.
Fatkheeva, L.
Fatkheeva, M.
Feher, J.
Fernandez, M.
Filipec-Kanizaj, T.
Frank, G.A.
Frankova, S.
Fraticiu, A.
17
17
27
11, 35
35
69
79
75
72
94
31, 34, 47, 48
Gaitantzi, H.
12
Gaitini, D.
43
Garcia-Pagán, J.-C.
18
Garcia-Pras, E.
79
Gazizov, I.M.
14, 15, 20
Gebala, E.
77
Gebhardt, R.
23
Genov, J.
90
Genunche-Dumitrescu, A.
60, 66, 67
Georgieva, M.
5
Georgieva-Shakola, M.
3
Gerbes, A.L.
13, 19
Gheorghe, C.
70
Gheorghe, L.
70
Girgis, S.A.
32
Gliwicz, D.
97
Godoy, P.
12, 23
Gogalniceanu, P.
41, 54, 57
Göke, B.
19
Golemanov, B.
90
Gonciarz, M.
30
Gonciarz, Z.
30
González, M.
87
Gotia, L.S.
36
Gotia, S.R.
36
Gottfriedova, H.
95
Goumerova, A. A.
35
Graupera, M.
18
Gressner, A.M.
98
Gressner, O.
25, 98
Grigorescu, M.
55, 56
Grigorov, N.
90
Grünhage, F.
25, 26
Gulubova, M.V.
91, 93
Gumerova, A.A.
11, 14, 15, 20
Gürakan, F.
51
Halabarodzka, M.
Hall, R.
Hamzavi, J.
Härtl, J.M.
Häussinger, D.
Heller, J.
Hellerbrand, C.
Hennenberg, M.
Herlea, V.
Heuchel, R.
Hillebrandt, S.
Hiura, M.
Hochberg, Z.
Hochrath, K.
Hogea, M.
Horvath, T.
Hubacek, J.A.
6, 7
26
23
13, 19
22
16, 64, 83, 84
25
16, 83, 84
70
23
26
33
43
26
55, 56
69
28
Ianosi, G.
Ibranyi, E.
Isik, A.T.
Iturriaga, H.
76
69
17, 21, 62, 85
1
Jabiry-Zieniewicz, Z.
Jankovic, G.
Jankowska, I.
Jirsa, M.
Jojic, N.
Jonderko, K.
Jovic, J.J.
Jovic, N.J.
Kaczmarczyk, M.
Kalicinski, P.
Kaligin, M.S.
Kalinova, K.
Kaminska, M.
Kaneti, E.
Kanzler, S.
Karasová, L.
Karslioglu, Y.
Kasicka-Jonderko, A.
Katicic, M.
Kazic, S.
Keiss, J.
Kempler, P.
Keppeler, H.
Khoroshko, N.
Kiassov, A.P.
Kitsenko, E.
96
73
96, 97
28
9
52
74
74
77
96, 97
14, 15, 20
4, 78
52
39
23
95
21
52
75
9, 59
37, 49
69
25
72
11, 14, 15, 20,
35
71
Kollai, M.
KonstantinovaKanazireva, D.
Konturek, S.J.
Kosseva, O.
Krasnaliev, I.
Krastev, Z.
Krsáková, A.
Krzyzanowska, E.
Kupcová, V.
Kurzawski, G.
Lacatusu, C.
Lahme, B.
Lakatos, P.L.
Lammert, F.
Lastauka, I.
Lengyel, G.
Levitan, B.N.
Levitan, G.
Liss, I.
Lubinski, J.
Lubivy, E.
Lukina, E.
Lukivskaja, O.
Lupescu, I.
Madrid, A.M.
Magiera, M.
Maly, J.
Marian, L.
Marin, M.
Mas, M.R.
Mas, N.
Mashaal, N.M.
Matcovschi, S.
Matei, V.
Matern, H.
Matern, S.
Mazur, W.W.
Mejias, M.
Mercea, V.
Mertens, P.
Mihai, B.
Mihai, C.
Mihaila, R.
Mijac, D.
Milkiewicz, P.
69
3, 5
30
38, 39
3
38, 39
44
86
10, 40, 42, 44,
45, 63
8
41
98
69
25, 26, 29, 83,
84
6
69
61
61
19
8
71
71, 72
80
70
87
30
95
53, 58
68
17, 21, 62, 85
17, 21, 62
46
81, 82
18
29
29
30
79
68
23
41, 57
41, 54, 57
31, 31, 34, 34,
47, 47, 48, 48
73
8, 77
Mitova, R.
Mitrut, P.
Monis, A.
Muñoz, R.
Nacu, N.
Narita, R.
Naumovski-Mihalic, S.
Neagoe, D.
Necic, D.
Neef, M.
Nicolae, C.
90
60, 66, 67
32
1
81, 82
33
75
76
59
83
70
Ocal, R.
17, 21
Odenthal, M.
25
Oldakowska-Jedynak, U.
96
Olteanu, A.
31, 34, 47, 48
Opelz, C.
19
Orga-Dumitriu, D.
31, 34, 47, 48
Osztovits, J.
69
Otsuki, M.
33
Ozcan, A.
62
Özen, H.
51
Ozsari, L.
21
Ozyurt, M.
62
Pacurari, A.
Paczek, L.
Papa, B.
Partycka, J.
Pawlowska, J.
Pecelj-Brocic, T.
Petrascu, O.
Petrasek, J.
Pevnev, G.O.
Pojoga, C.
Poniachick, J.
Popescu, I.
Pophristova, E.
Popov, Y.
Porr, P.-J.
Potapov, A.S.
Protic, M.
Pruzansky, Y.
Racanel, F.
Raeuneva, T.
Raszeja-Wyszomirska, J.
Rehbein, K.
Rehfeld, J.F.
Reichen, J.
65
96
75
24
96, 97
74
31, 34, 47, 48
28
14, 15
88
87
70
38, 39
80
31, 34, 47, 48
2
9
43
76
7
8
98
30
83
Reinehr, R.
Rendekova, V.
Rezi, C.E.
Reznakova, S.
Rodriguez-Vilarrupla, A.
Roganovic, B.
Roggel, F.
Romanov, A.O.
Romosan, I.
Rozentale, B.
Russell, D.W.
Rusu, M.
22
63
31, 34, 47, 48
28
18
74
13
27
53, 58, 65
37, 49
29
55, 56
Sabaric, B.
Saftoiu, A.
Saltik-Temizel, I.N.
Sauerbruch, T.
75
76
51
16, 25, 26, 29,
64, 83, 84
17
3
64
95
80
45
72
37, 49
46
43
98
5
95
75
24
11
96, 97
97
22
37, 49
35
28, 94
28, 94, 95
89
89
73
13, 19
28
95
2
8
71
36, 53, 58, 65
Savas, B.
Savov, A.
Schepke, M.
Schück, O.
Schuppan, D.
Seligová, E.
Semenova, E.
Sevastjanova, N.
Shaat, E.A.
Shen Orr, Z.
Siluschek, M.
Sinigerska, V.
Skibová, J.
Skurla, B.
Slomka, M.
Smetannikova, T.
Socha, J.
Socha, P.
Sommerfeld, A.
Sondore, V.
Sozinov, A.S.
Sperl, J.
Spicák, J.
Stefanescu, C.
Stefanescu, G.
Stefanova, E.
Steib, C.J.
Stickel, F.
Stollová, M.
Strokova, T.V.
Suchy, J.
Sukhanova, G.
Susan, L.M.
Svarc, D.
Svorcan, P.
Sysojeva, E.P.
Szalay, F.
Szántová, M.
92
9, 59
71, 72
69
10, 40, 42, 44,
63
Tabaru, A.
Taher, M.Y.
Tanu, S.
Tasci, I.
Tcaciuc, A.
Tcaciuc, E.
Teisseyre, M.
Ticak, M.
Titova, M.A.
Tiurean, L.
Todorova, K.
Tomova, S.
Toth, T.
Trebicka, J.
Trunecka, P.
Tudorascu, D.
Turecky, L.
33
50
68
17, 21, 62
81, 82
81, 82
96, 97
75
11
31, 34, 47, 48
4
3
69
16, 83, 84
28, 94, 95
76
10, 40, 42, 44,
45, 63
3, 5
Tzaneva, V.
Ueberham, E.
Uhliková, E.
Uhlíková, E.
Umerova, A.
Uslu, N.
Uvarova, E.
Uzunalimoglu, Ö.
23
10, 40, 42, 44,
63
40
61
51
2
17, 21, 62, 85
Vacariu, V.
Valková, M.
Vasiljev, S.
Velev, V.
Verdes, D.
Vere, C.
Viksna, L.
Visnyei, Z.
Vitko, S.
Vlaykova, T.
Voronkova, E.
Vozár, I.
Vrinic, D.
Vulcu, D.V.
Vural, S.A.
58
40, 45
71
91
36
76
37, 49
69
94
91, 93
71, 72
10
59
31, 34, 47, 48
85
Wasmuth, H.
Weber, S.
Weiskirchen, R.
Weng, H.
Wunsch, E.
Yildiz, Y.
Yilmaz, S.
Yüce, A.
Yurdaydin, C.
Zaharie, A.V.
Zaks, J.
Zawada, I.
Zeid, A.
Zelinkova, Z.
Zielinski, J.
Zigraiová, S.Z.
Zwolak, A.Z.
25
29
98
23
77
29
17, 62
51
62
31, 34, 47, 48
80
8, 77
50
45
64
40
86
Falk Foundation
Dr. Falk Pharma
Where medicine
and pharmaceuticals meet –
a tried and trusted link
We help
by providing innovative medicines for bowel and liver diseases.
We promote knowledge
from 1967 onwards, there have been more than 200 international
Falk Symposia and Workshops with over 100,000 participants from
more than 100 countries
1 million doctors and patients have attended over 11,000
Falk educational meetings during the last 30 years
our Literature and Patient Information Service –
including on-line ordering through our website.
www.falkfoundation.de
www.drfalkpharma.de
Leinenweberstr. 5 79108 Freiburg Germany Tel +49 (0) 761/1514-0 Fax +49 (0) 761/1514-321 E-Mail [email protected]

Liver Cirrhosis - Dr. Falk Pharma GmbH

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