Dr: Mohamed I Kotb El

Transcription

Clinical Biochemistry --------------------------- Biochemistry Department ------------------------- Helwan University
Lectures Notes
On
CLINICAL CHEMISTRY OF KIDNEY
By
Dr: Mohamed I Kotb El-Sayed
For
4th Year – Pharmacy Students
(1st Semester – 2015-2016)
Biochemistry and Molecular Biology Department
Faculty of Pharmacy
Helwan University
Cairo
EGYPT
For Contact
Email: [email protected]
[email protected]
https://web.facebook.com/drmohamed.k.elsayed
https://scholar.google.com.eg/citations?user=47ZPvuAAAAAJ&hl=ar&oi=ao
Kidney Function Tests ----------------------------------------------------------------------- Dr.: Mohamed I Kotb El-Sayed
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1. INTRODUCTION
1.1. Definition:
 Kidney function tests are a collective term for a variety of individual tests and procedures
that can be done to evaluate how well the kidneys are functioning.
 A doctor who orders kidney function tests and uses the results to assess the functioning of
the kidneys is called a nephrologist.
1.2. Physiology of the kidney:
1.2.1. Functions of the kidney:
1. To maintain the constancy of the extra-cellular fluid by:

Excreting dietary surpluses and metabolic end-products e.g. NPN; urea, creatinine,
urate, H+.

Retaining necessary substances, either by not letting them be filtered (e.g. proteins)
or by reabsorbing them in the tubules (e.g. glucose, amino-acids, HCO3-).
2. Regulatory functions: Mechanisms of differential Na+, H2O, CO2 reabsorption and
secretion: this operates under complexed controlling system.
3. To act as an endocrine gland;

Erythropoietin.

Renin.

One-alpha-hydroxylation of Vitamin D (to make 1:25 di-hydroxycholecalciferol,
calcitriol).
 For these functions the kidneys use ¼ of the total cardiac output, and produce 180 L/day
(or 125 mL/min) of glomerular filtrate.
 Each kidney has about 1 million nephrons. Aspects of sodium, potassium, and acid-base
pathology will be touched on
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1.2.2. Glomerular Function:
 The glomerulus has three layers separating the blood in the glomerular capillaries from the
glomerular lumen: capillary endothelial, basement membrane, and visceral epithelium.
The basement membrane is the primary barrier.
 An overall negative charge due to abundant sialic acid groups in the glomerular layers
helps prevent large anions, such as most proteins, from crossing.
 The hydrostatic pressure across the membrane which carries the ultra-filtrate is only
about 1 kPa, and if blood pressure falls only moderately, the oncotic pressure due to the
plasma proteins is sufficient to cause filtration to slow or even stop. This explains the
oliguria in shocked patients.
 N.B: polyuria = more than normal urine; oliguria = less than normal; anuria = no urine.
1.2.3. Tubular Function:
1. Proximal tubule:

Designed to reabsorb only what you need - 80% of sodium and water - high capacity
active sodium uptake, with chloride following passively.

A low plasma chloride can limit recovery of sodium – important later for understanding
how this can cause or perpetuate alkalosis after prolonged vomiting.
 K+: 95% absorbed usually, but diet dependent.
 Phosphate: active reabsorption which is inhibited by PTH.
 HCO3- : mostly absorbed.
 Glucose and amino acid absorption is normally nearly complete.
 Also secretes: organic acids, urate, and drugs.
 The Fanconi syndrome is loss (inherited or acquired) of proximal tubular functions and
is characterised (logically from the above) by glycosuria, amino aciduria, and
phosphaturia.
o May also have acidosis and polyuria.
o Classic cause is cystinosis, an inherited disease of lysosomal membrane
transport, but it can also be acquired.
2. Loop of Henle:
 Only found in birds and mammals.
 Its counter-current multiplier effect is the basis for creating either dilute or concentrated
urine.
 Key features are an active NaCl pump in the thick ascending limb, and water
impermeability of the whole of the ascending limb.
 Fluid emerges hypotonic at the end of the loop of Henle, at about ½ the osmolality of body
fluids (~ 120-150 mosmoles/L, as compared with 250-300 mosmoles/L in plasma).
 Further salt may be removed in the collecting ducts under conditions of water diuresis,
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causing further dilution.
3. Distal Convoluted Tubule (DCT):
 Little change in volume or concentration.
 Secretion of Aldosterone (see renin-angiotensin system) causes sodium to be exchanged
for K+ and/or H+.
 Conn's syndrome, Addison's disease, and RTA type I all affect DCT function.
4. Collecting Ducts:
 Anti-Diuretic Hormone (ADH, Vasopressin, Pitressin), synthesized in the hypothalamus
and released from the posterior pituitary in response to an increase in extracellular
osmolality, increases water permeability of tubular cells (and urea permeability in the
lower medullary part).
 ADH linked pathology exerts its effect here and consists of;
4.1- Diabetes Insipidus:
 Pituitary form, where no ADH is synthesized due to damage to the pituitary.
 Nephrogenic form, where renal tubular cells do not respond to normal levels of ADH.
 Both forms give rise to polyuria with dilute urine.
4.2- Syndrome of inappropriate secretion of ADH (SIADH): low output of inappropriately
concentrated urine in the presence of hypervolaemia and dilutional hyponatraemia.
2. KIDNEY FUNCTIONS TESTS
2.1. Introduction:
 There are a number of urine tests that can be used to assess kidney function.
 A simple, inexpensive screening test a routine urinalysis is often the first test conducted if
kidney problems are suspected. A small, randomly collected urine sample is examined
physically for things like color, odor, appearance, and concentration (specific gravity);
chemically, for substances such a protein, glucose, and pH (acidity/alkalinity); and
microscopically for the presence of cellular elements (red blood cells [RBCs], white blood
cells [WBCs], and epithelial cells), bacteria, crystals, and casts (structures formed by the
deposit of protein, cells, and other substances in the kidneys’s tubules).
 If results indicate a possibility of disease or impaired kidney function, one or more of the
following additional tests is usually performed to pinpoint the cause and the level of decline
in kidney function.
 Glomerular function tests depend on examination of substances which depend on
glomerular function for their elimination (creatinine, urea, and eGFR).
 The determination of non-protein nitrogenous substances (NPN) in the blood has
traditionally been used to monitor renal function.
 The majority of these compounds arise from the catabolism of proteins and nucleic acids.
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2.2. Non-protein nitrogenous substances (NPN):
 The most important NPN compounds used in evaluation of kidney functions are urea,
creatinine, creatine uric acid, and ammonia.
 As clearance levels decrease, blood levels of creatinine, urea, and uric acid increase.
2.2.1. Blood Urea Nitrogen (BUN): Glomerular function test:
1. Introduction
o It is formed in the liver from protein catabolism by releasing free amino groups (NH2).
o
o
o
o
o
Then it is transported to blood and then excreted in urine by kidneys
It represent about more than 75% of excreted NPN compounds.
Urea is the major excretory product of protein metabolism.
It is a useful measure of decreased filtration (considered as glomerular filtration test).
Since historic assays for urea were based on measurement of nitrogen, the term
blood urea nitrogen (BUN) has been used to refer to urea determination. Urea
nitrogen (urea N) is a more appropriate term.
o Its levels are affected by high protein intake, catabolic states, post-surgery and
trauma, and gastro-intestinal hemorrhage, all of which cause increased urea
production from protein.
Clinically Significant Non-protein Nitrogen Compounds
2. Physiology of Urea:
 Protein metabolism produces amino acids that can be oxidized to produce energy or
stored as fat and glycogen.
 These processes release nitrogen, which is converted to urea and excreted as a waste
product.
 Following synthesis in the liver, urea is carried in the blood to the kidney, where it is
readily filtered from the plasma by the glomerulus.
 Most of the urea in the glomerular filtrate is excreted in the urine, although some urea is
reabsorbed by passive diffusion during passage of the filtrate through the renal tubules.
 Under conditions of normal flow and normal renal function, about 40% of the filtered
urea is reabsorbed; when the flow rate is decreased, the amount passively reabsorbed
5
increases.
 The amount reabsorbed depends on urine flow rate and extent of hydration.
 Small quantities of urea (≤10% of the total) are excreted through the gastrointestinal tract
and skin.
o Small of urea portion is diffused to intestine to be cleaved by bacterial urease
enzyme to CO2 and NH3. NH3 is partially lost in stool and partially reabsorbed to
blood.
 The concentration of urea in the plasma is determined by renal function and perfusion,
the protein content of the diet, and the rate of protein catabolism.
 The BUN test measures the amount of nitrogen contained in the urea. High BUN levels
can indicate kidney dysfunction, but because BUN is also affected by protein intake and
liver function, the test is usually done together with a blood creatinine, a more specific
indicator of kidney function.
 The serum concentration of urea nitrogen is influenced by factors not connected with
renal function or urine excretion as it is affected strongly by the degree of protein
catabolism. A marked change in dietary protein consumption will be reflected in BUN
values.
 The injection or ingestion of steroids produces a rise in BUN as do stressful situations
that cause the adrenal gland to secrete additional cortisol. For these reasons, the
measurement of serum creatinine is a better indicator of kidney status than is that of
BUN although in many cases, they go up and down simultaneously.
3. Clinical Application of Urea:
 Measurement of urea is used
o
o
o
o
o
to evaluate renal function,
to assess hydration status,
to determine nitrogen balance,
to aid in the diagnosis of renal disease, and
to verify adequacy of dialysis.
 In renal failure ↑ urea →↑ NH3 Hyperammonemia. So that Neomycin and/or laculose is
given as prophylactic therapy in hepatic failure.
 Assays for urea were originally performed on a protein-free filtrate of whole blood and
based on measuring the amount of nitrogen in the sample.
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 Urea is not used to determine GFR due to:
o 40-70% of high diffusible urea moves passively out from the renal tubules to
plasma.
o This back diffusion depends on urine flow rate.
o Urea production is depending on several non renal factors e.g. Diet and hepatic
factors.
o So that urine urea is limited to be used to estimate urea production rate but
serum urea give useful clinical information in particular circumstance.
4. Pathophysiology of Urea:
 After considering the effects of high protein diet, administration of cortisol-like steroids
and stressful situations, increased BUN is seen in the prerenal, renal, and post-renal
factors.
 An elevated concentration of urea in the blood is called azotemia not uriceimia.
 Very high plasma urea concentration accompanied by renal failure is called uremia, or
the uremic syndrome.
 This condition is eventually fatal if not treated by dialysis or transplantation.
 Conditions causing increased plasma urea are classified according to cause into three
main categories: prerenal, renal, and post-renal.
o Prerenal azotemia is caused by reduced renal blood flow.
 It is increased the plasma urea levels without increased plasma creatinine.
 Less blood is delivered to the kidney; consequently, less urea is filtered.
 Causative factors include;

Decreased renal blood flow (renal ischemia) as in congestive heart
failure (↓COP).

Shock,

Bloody meal as in GI hemorrhage,

Mild dehydration,

Cortisone therapy,

High protein diet,

Increased protein catabolism as in starvation, and
 Significant decrease in blood volume.
 The amount of protein metabolism also induces prerenal changes in blood
urea concentration. A high-protein diet or increased protein catabolism,
such as occurs in stress, fever, major illness, corticosteroid therapy, and
gastrointestinal hemorrhage, may increase urea concentration.
o Renal azotemia: Decreased renal function causes an increase in plasma urea
concentration as a result of compromised urea excretion.
7

Renal causes of elevated urea include acute and chronic renal failure,
acute and chronic glomerular nephritis, tubular necrosis, and other
intrinsic renal disease.
o Post-renal azotemia can be due to obstruction of urine flow anywhere in the
urinary tract by renal calculi, tumors of the bladder or prostate, or severe
infection.
 It is increased plasma urea levels with increased plasma creatinine as in:
obstruction of urine out flow through ureters, bladder or urethra as in
renal stones, prostatism or urinary tract tumours
 The major causes of decreased plasma urea concentration include low protein intake
and severe liver disease.
o Plasma urea concentration is decreased during late pregnancy (when the fetus is
growing rapidly and utilizing maternal amino acids) and infancy as a result of
increased protein synthesis.
o In starvation, and in patients whose diet is grossly deficient in protein
o The conditions affecting plasma urea concentration are summarized in;
Causes of abnormal plasma Urea concentration
 Differentiation of the cause of abnormal urea concentration is aided by calculation of
the urea nitrogen/creatinine ratio, which is
o Normally 10-20.
o Prerenal conditions tend to elevate plasma urea, whereas plasma creatinine remains
normal, causing a high urea N/creatinine ratio.
o A high urea N/creatinine ratio with an elevated creatinine is usually seen in post-renal
conditions.
o A low urea N/creatinine ratio is observed in conditions associated with decreased urea
production, such as low protein intake, acute tubular necrosis, and severe liver disease.
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 NB: - Serum urea Nitrogen mg/dL = Serum urea mg/dL X 0.467.
5. Urea clearance:
 The urea clearance test requires a blood sample to measure the amount of urea in the
bloodstream and two urine specimens, collected one hour apart, to determine the amount
of urea that is filtered, or cleared, by the kidneys into the urine.
 It is plasma volume which is cleaned from urea per minute.
U V
= ml / minute.
P
o U = Urine urea (mg/dl).
o Urea Clearance =
o P = Plasma or serum urea (mg/dL).
o V = Urine flow rate (ml/minute).
 Normal values of urea clearance:
o If urine flow rate ≥ 2 mL / minute → urea clearance = 60 - 95 ml/minute.
o If urine flow rate < 2 mL / minute → urea clearance = 40 - 65 ml/minute.
 Urea clearance % (Van Slyk percent):
o If urine flows rate ≥ 2 mL / minute: Urea Clearance %
Urea clearance
100 .
75
o If urine flow rate < 2 mL/
minute:
Urea Clearance %
=
=
Urea clearance
100 .
54
 Clinical significance of urea clearance %:
o If urea clearance % about 50 – 20% → Mild renal failure.
o If urea clearance % about 20% → sever renal failure.
o If urea clearance % about 5 % → Coma and death.
2.2.2. Creatine and Creatinine: Glomerular function test:
1. Introduction
 Creatinine is formed from creatine and creatine phosphate in muscle and is excreted into the
plasma at a constant rate related to muscle mass.
 Plasma creatinine is inversely related to glomerular filtration rate (GFR) and, although an
imperfect measure, it is commonly used to assess glomeular filtration function.
2. Physiology of Creatine and Creatinine:
 Creatine is synthesized primarily in the liver from arginine, glycine, and methionine.
 It is then transported to other tissues, such as muscle, where it is converted to creatine
phosphate, which serves as a high-energy source.
 About 1-2% of the total muscle creatine pool is converted daily to creatinine through the
non-enzymatic loss of water.
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 The structures and relationship of these compounds are shown in Figure 1.
Figure 1- Interconversion of creatine, creatine phosphate, and creatinine.
 Creatinine is released into the circulation at a relatively constant rate that has been shown to
be proportional to an individual’s muscle mass and is not affected by diet, age, sex, or
exercise.
 With normal kidney function, then, the amount of creatinine in the blood remains relatively
constant and normal.
3. Clinical Application of Creatine and Creatinine:
 Plasma creatinine concentration is a function of relative muscle mass, the rate of creatine
turnover, and renal function.
 Measurement of creatinine concentration is used to determine sufficiency of kidney function
and the severity of kidney damage and to monitor the progression of kidney disease.
 Creatinine is filtered freely unchanged at the glomerulus and eliminated without significant
reabsorption or secretion in the tubules (not altogether true but works in practice).
 The amount of creatinine in the bloodstream is reasonably stable, although the protein
content of the diet does influence the plasma concentration.
 For this reason, and because creatinine is affected very little by liver function, an elevated
blood creatinine level is a more sensitive indicator of impaired kidney function than the
BUN.
 The serum creatinine is a better indicator of renal function than either that of BUN or uric
acid because it is not affected by diet, exercise, or hormones
 Therefore the glomerulus filtration rate (GFR) was most often assessed by determining the
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urinary creatinine clearance.
4. Creatinine Clearance (CrCl):
 Because of the constancy of endogenous production, urinary creatinine excretion has been
used as a measure of the completeness of 24-hour urine collections in a given individual,
although the uncertainty associated with this practice may exceed that introduced by the use
of urine volume and collection time for standardization.
 Urinary constituents may be expressed as a ratio to creatinine quantity rather than as mass
excreted per day.
Creatinine clearance:
 It is a measure of the amount of creatinine eliminated from the blood by the kidneys, and
GFR are used to gauge renal function.
o Owing to creatinine not being significantly reabsorbed or secreted by the renal
tubules, CrCl provides a measure of the glomerular filtration rate (GFR). It is
calculated as follows;
CrCl = (Urine Creatinine conc. x volume) / (Plasma Creatinine conc.)
Creatinine clearance 
U V 1.73

 ml/ minute/m 2
P
A

U = Urine creatinine (mg/dL).




P = Plasma or serum creatinine (mg/dL).
V = Urine flow rate (mL/minute).
A = Body surface area (m2).
1.73/A = factor normalize clearance for average body surface area
because creatinine execration is proportion to muscle mass.
o Note that the units are a flow rate, ml/min. It can be difficult to measure well
since the timed urine collection (should be over 24 hrs) is in practice notoriously
unreliable and requires good patient and staff cooperation.
o However, CrCl changes more linearly in proportion to renal mass loss than does
plasma creatinine or urea (Cr Cl decreases, the latter increase) so is the best
measure of progress in chronic renal failure.
o It is of no value in acute renal failure since it needs a steady state situation to
obtain a meaningful result.
o Normal CrCl. is about 120 ml/min.
o Creatinine clearance may be used as indicator for GFR because:
 Creatinine is endogenously produced.
 Creatinine is released into body fluid at constant rate.
 Its plasma level maintained within narrow limits.
 Its plasma level not affected by dietary factors.
11
 Creatinine clearance is usually reported in units of mL/minute and can be corrected for body
surface area.
 For a 24-hour urine collection, normal results are 90 mL/min–139 mL/min for adult males
younger than 40, and 80–125 mL/min for adult females younger than 40. For people over
40, values decrease by 6.5 mL/min for each decade of life.
 Creatinine clearance overestimates GFR because a small amount of creatinine is reabsorbed
by the renal tubules and up to 10% of urine creatinine is secreted by the tubules. However,
CrCl provides a reasonable approximation of GFR.
 This relationship between plasma creatinine and GFR and the observation that creatinine
concentrations are relatively constant should make the analyte a good endogenous filtration
marker.
5. Reference Intervals
 Reference intervals vary with assay type, age, and gender. Creatinine concentration
decreases with age beginning in the fifth decade of life.
 Results expressed in conventional units of milligrams per deciliter can be converted to
international units using the molecular mass of creatinine (113 g/mol).
Creatinine
6. Pathophysiology:
1. Creatinine
High plasma creatinine:
1- Non renal causes of increased plasma creatinine include:
 High protein (meat) intake → temporary increase of plasma creatinine.
 Exercise → transient increase of plasma creatinine after vigorous exercises.
 Analytical over estimation: some analytical methods are not specific for creatinine;
they measure the endogenous and exogenous interfering substances e.g. plasma
acetoacetate and pyruvate.
 Drugs e.g. salicylates and cimetedine which reduce tubular secretion of creatinine →
elevating plasma creatinine level.
2- Renal causes of increased plasma creatinine include:
 Diseases in which there is impaired renal perfusion e.g. reduced COP and in case of
renal artery stenosis.
 Diseases with loss of nephrotic functions e.g. acute and chronic glomerulo-nephritis.
12
 Diseases with increased pressure on the tubular side of nephrones e.g. urinary tract
obstruction due to prostatic enlargement.
 Elevated creatinine concentration is associated with abnormal renal function,
especially as it relates to glomerular function.
 Plasma concentration of creatinine is inversely proportional to clearance of
creatinine.
 Therefore, when plasma creatinine concentration is elevated, GFR is decreased,
indicating renal damage or when urine elimination is obstructed.
 Plasma creatinine is a relatively insensitive marker and may not be measurably
increased until renal function has deteriorated more than 50%.
Low plasma creatinine:
 Creatinine production is determined by the size of creatine pool hence a smaller muscle
mass leads to daily lower creatinine production.
 Physiologically pregnancy is accompanied with decreased plasma creatinine level.
o Also, females and children show low plasma creatinine levels when compared
with adult men.
 Pathologically low plasma level of creatinine is found in wasting diseases, starvation,
and in patients treated with corticosteroids due to their protein catabolic effect.
o
Low serum creatinine values have been associated with muscular dystrophy.
2. Creatine
 In muscle disease such as muscular dystrophy, poliomyelitis, hyperthyroidism, and
trauma, both plasma creatine and urinary creatinine are often elevated. Plasma creatinine
concentrations usually are normal in these patients.
 Measurement of creatine kinase is used typically for the diagnosis of muscle disease
because analytic methods for creatine are not readily available in most clinical
laboratories.
 Plasma creatine concentration is not elevated in renal disease.
2.2.3. Estimated GFR (eGFR): Glomerular function estimation:
 Measured creatinine concentration used in combination with other variables in one of
several empirically determined equations provides a better assessment of renal disease,
in part because the equations estimate GFR, not creatinine clearance .
 Laboratories are making increasing use of estimated GFR derived from creatinine
estimations, more especially because of the problems with measuring creatinine clearance.
 There are a number of equations that have been introduced. These make use of the patient’s
weight and creatinine level to estimate the GFR. The most popular equation in use is the
MDRD (Modification of Diet in Renal disease) equation (The abbreviated Modification of
Diet in Renal Disease (MDRD) equation is advocated by the National Kidney Foundation).
13
 This is only used in adults (>18 years) estimated GFR (eGFR) should be calculated using
the 4-variables (i.e. serum creatinine concentration, age, gender and ethnic origin)- and
makes the assumption that all filtered creatinine is excreted.
 The MDRD equation is most useful when serum creatinine results are produced in an assay
that has been calibrated to be traceable to an isotope dilution mass spectrometry (IDMS)
method.
 When serum creatinine is measured using an IDMS traceable method, the MDRD equation
for estimated glomerular filtration rate (eGFR) is:
GFR (mL/min/1.73 m2) = 175 x [serum creatinine (µmol/L) x 0.011312]-1.154 x [age]-0.203 x [1.212 if
black] x [0.742 if female]
 Where serum (plasma) creatinine concentration in mg/dL and age is in years.
 Results are normalized to a standard body surface area (1.73 m2). The equation is valid for
adults older than 18 years and younger than 70 years of age.
 The equation was developed using data from non-hospitalized patients known to have
chronic kidney disease and is reasonably accurate for this population. Effectiveness of the
equation in other groups is being investigated.
 Clinical laboratories have been strongly encouraged to report eGFR when serum creatinine
is ordered as a means to increase detection of kidney disease and improve patient care.32
2.2.4. Uric Acid:
1. Introduction:

Uric acid is a metabolite of nucleic acids, purines, and nucleoproteins catabolism and is
the end product of protein (purine) metabolism in man.

In most mammals, uric acid is oxidized in the liver to allantoin.
 Although it is filtered by the glomerulus and secreted by the distal tubules into the urine,
most uric acid is reabsorbed in the proximal tubules and reused.
 Uric acid is relatively insoluble in plasma and, at high concentrations, can be deposited in
the joints and tissue, causing painful inflammation.
2. Physiology of Uric Acid:
 Purines, such as adenosine and guanine from the breakdown of ingested nucleic acids or
from tissue destruction, are converted into uric acid, primarily in the liver.
 Uric acid is transported in the plasma from the liver to the kidney, where it is filtered by the
glomerulus.
 Reabsorption of 98% to 100% of the uric acid from the glomerular filtrate occurs in the
proximal tubules.
 Small amounts of uric acid are secreted by the distal tubules into the urine.
 Renal excretion accounts for about 70% of uric acid elimination; the remainder passes into
the gastrointestinal tract and is degraded by bacterial enzymes.
14
 Nearly all of the uric acid in plasma is present as monosodium urate. At the pH of plasma
(pH<7), urate is relatively insoluble; at concentrations greater than 6.8 mg/dL, the plasma is
saturated. As a result, urate crystals may form and precipitate in the tissues.
 In acidic urine (pH<5.75), uric acid is the predominant species and uric acid crystals may
form.
3. Clinical Application of Uric Acid:
 Uric acid is measured to assess inherited disorders of purine metabolism,
o to confirm diagnosis and monitor treatment of gout,
o to assist in the diagnosis of renal calculi,
o to prevent uric acid nephropathy during chemotherapeutic treatment, and
o to detect kidney dysfunction.

Increased levels of uric acid have been observed in renal failure, chronic lead poisoning,
polycythemia, some leukemias, and toxemia of pregnancy.
Reference Intervals of Uric Acid (Uricase Method)8
4. Pathophysiology of Uric Acid:
 Inherited disorders of purine metabolism are associated with significant increases in
physiological uric acid concentrations.
 Lesch-Nyhan syndrome is an X-linked genetic disorder (seen only in males) caused by the
complete deficiency of hypoxanthine guanine phosphoribosyltransferase (HGPRT, EC
2.4.2.8), an important enzyme in the biosynthesis of purines.
 Lack of this enzyme prevents the reutilization of purine bases in the nucleotide salvage
pathway and results in increased de novo synthesis of purine nucleotides and high plasma
and urine concentrations of uric acid.
 Neurologic symptoms, mental retardation, and self-mutilation characterize this extremely
rare disease.
 Mutations
in
the
first
enzyme
in
the
purine
synthesis
pathway,
phosphoribosylpyrophosphate synthetase (PRPP synthetase, EC 2.7.6.1), also cause elevated
uric acid concentration.
 Increased uric acid is found secondary to glycogen storage disease (deficiency of glucose-6phosphatase, EC 3.1.3.9) and fructose intolerance (deficiency of fructose-1-phosphate
aldolase, EC 2.1.2.13).
15
 Metabolites such as lactate and triglycerides are produced in excess and compete with urate
for renal excretion in these diseases.
 Elevated plasma uric acid concentration is found in gout, increased catabolism of nucleic
acids, and renal disease.
 Gout is a disease found primarily in men and usually is first diagnosed between 30 and 50
years of age.
 Affected individuals have pain and inflammation of the joints caused by precipitation of
sodium urates. In 25% to 30% of these patients, hyperuricemia is a result of
overproduction of uric acid, although hyperuricemia may be exacerbated by a purine-rich
diet, drugs, and alcohol.
o Plasma uric acid concentration in affected individuals is usually greater than 6.0
mg/dL. Patients with gout are very susceptible to the development of renal calculi,
although not all persons with high serum urate concentrations develop this
complication.
o In women, urate concentration rises after menopause. Postmenopausal women may
develop hyperuricemia and gout.
o In severe cases, deposits of crystalline uric acid and urates called tophi form in
tissue, causing deformities.
o Another common cause of elevated plasma uric acid concentration is increased
metabolism of cell nuclei, as occurs in patients on chemotherapy for such
proliferative diseases as leukemia, lymphoma, multiple myeloma, and polycythemia.
o Monitoring uric acid concentration in these patients is important to avoid
nephrotoxicity.
o Allopurinol, which inhibits xanthine oxidase (EC 1.1.3.22), an enzyme in the uric
acid synthesis pathway, is used as treatment.
 Chronic renal disease causes increased uric acid concentration because filtration and
secretion are impaired.
o However, uric acid is not useful as an indicator of renal function because many other
factors affect its plasma concentration.
 Patients with hemolytic or megaloblastic anemia may exhibit elevated uric acid
concentration. Hyperuricemia is a common feature of toxemia of pregnancy (preeclampsia)
and lactic acidosis, presumably as a result of competition for binding sites in the renal
tubules.
o Increased urate concentrations may be found following ingestion of a diet rich in
purines (liver, kidney, sweetbreads, shellfish) or as a result of increased tissue
catabolism due to inadequate dietary intake (starvation).
 Hypouricemia is less common than hyperuricemia and is usually secondary to severe liver
16
disease or defective tubular reabsorption, as in Fanconi syndrome.
o Hypouricemia can be caused by chemotherapy with 6-mercaptopurine or
azathioprine, inhibitors of de novo purine synthesis, and as a result of overtreatment
with allopurinol. The conditions affecting plasma urate concentrations are shown in
Table 5.
Table 5- Causes Of Abnormal Plasma Of Uric Acid
2.2.5. Ammonia:
1. Introduction:
 Ammonia is formed in the deamination of amino acids during protein metabolism.
 It is removed from the circulation and converted to urea in the liver.
 Free ammonia is toxic; however, ammonia is present in the plasma in low concentrations.
2. Physiology of Ammonia
 Ammonia (NH3) is produced in the catabolism of amino acids and by bacterial metabolism
in the lumen of the intestine.
 Some ammonia results from anaerobic metabolic reactions that occur in skeletal muscle
during exercise.
 Ammonia is consumed by the parenchymal cells of the liver in the production of urea.
 At normal physiologic pH, most ammonia in the blood exists as ammonium ion (NH4-).
17
 Figure 2 shows the pH-dependent equilibrium between NH3 and NH4-.
 Ammonia is excreted as ammonium ion by the kidney and acts to buffer urine.
Figure 2- Interconversion of ammonium ion and ammonia.
3. Clinical Applications of Ammonia:
 Clinical conditions in which blood ammonia concentration provides useful information are
hepatic failure, Reye’s syndrome, and inherited deficiencies of urea cycle enzymes. Severe
liver disease is the most common cause of disturbed ammonia metabolism. The monitoring
of blood ammonia may be used to determine prognosis, although correlation between the
extent of hepatic encephalopathy and plasma ammonia concentration is not always
consistent. Arterial ammonia concentration is a better indicator of the severity of disease.
 Reye’s syndrome, occurring most commonly in children, is a serious disease that can be
fatal. Frequently, the disease is preceded by a viral infection and the administration of
aspirin. Reye’s syndrome is an acute metabolic disorder of the liver, and autopsy findings
show severe fatty infiltration of that organ.
 Blood ammonia concentration can be correlated with both the severity of the disease and
prognosis. Survival reaches 100% if plasma NH3 concentration remains below five times
normal.
 Ammonia is of use in the diagnosis of inherited deficiency of urea cycle enzymes. Testing
should be considered for any neonate with unexplained nausea, vomiting, or neurological
deterioration associated with feeding.
 Assay of blood ammonia can be used to monitor hyperalimentation therapy and
measurement of urine ammonia can be used to confirm the ability of the kidneys to produce
ammonia.
4. Reference Interval of Ammonia:
 Values obtained vary somewhat with the method used.
 Higher concentrations are seen in newborns.
Ammonia
5. Pathophysiology of Ammonia:
 In severe liver disease in which there is significant collateral circulation or if parenchymal
liver cell function is severely impaired, ammonia is not removed from the circulation and
blood concentration increases.
 High concentrations of NH3 are neurotoxic and often associated with encephalopathy.
18
 Toxicity may be partly a result of increased extracellular glutamate concentration and
subsequent depletion of adenosine triphosphate (ATP) in the brain.
 Hyperammonemia is associated with inherited deficiency of enzymes of the urea cycle.
Measurement of plasma ammonia is important in the diagnosis and monitoring of these
inherited metabolic disorders.
2.3- Miscellaneous Tests:
2.3.1. Tubular Function Tests:
1. Urinary Na+ concentration. Normally low relative to serum concentration unless on a
dietary high salt intake. Urine sodium. A 24-hour urine sodium should be within 75–
200 mmol/day.
2. Concentration tests (usually after Pitressin), and Dilution tests, after a water load. Ratio
of osmolality (or urea) in urine relative to that in plasma is a simple practical measure.
3. Acidification tests, after administration of NH4Cl (→ NH3 + H+ + Cl-). Seldom done
except in differentiation of type I and II renal tubular acidosis (RTA, see later).
2.3.2. Plasma Protein: > 2.5 g/day indicates nephrotic syndrome;
o Bence-Jones Protein indicates myeloma (see protein lectures)
o ß2-microglobulin, small protein, filtered then absorbed by tubules, which is a sensitive
test of tubular function (but also ↑ in some malignancies and inflammatory
conditions).
o Modest proteinuria is associated with many types of pathology but a mild increase can
sometimes be normal (orthostatic, pregnancy).
1. Normal value of total plasma proteins:
o Total plasma proteins concentration about 6.3 – 8.3 g/dL.
2. Site of Protein synthesis:
o All types of plasma proteins are synthesized in the liver except γ-globulins that are
synthesized in plasma cells.
o Liver synthesizes about 12 gm albumin per day.
o Nearly all types of plasma proteins are synthesized in the pre-protein form then it is
subjected to modifications such as "phosphorylation & glycosylation to become
mature protein".
3. Types of plasma proteins:
o Plasma is reported to contain over than 100 individual proteins but mainly classified
into:



Albumins.
Globulins.
Fibrinogen.
19
Albumins:

Pre- albumin:
o It is present in very low concentration about 25 mg/dL.
o It helps in the transport of T3, T4 and Vitamin-A.

Albumin:
o It is the major type of plasma protein in a concentration of 3.4 – 4.7 g/dL. It
synthesized in the liver (about 12 g/day).
o It has low M. Wt about 68 KD acting about 60% of plasma proteins. 40% of
albumin is present in the plasma and the remaining 60% is present in extracellular space.
Mature human albumin consists of one polypeptide chain of 585 amino acid with
17 di-sulphide bonds.
o Albumin has a long plasma half- life (t ½) about 20 days. So that its plasma level
is normal in acute liver diseases but give clinical significance in chronic liver
diseases.
o Due to low M. Wt and high concentration albumin is responsible for about 7580 % of plasma osmotic (oncotic) pressure.
Globulins:
 Its normal plasma concentration is about 1.5 - 3 g/dL and of high M. wt about 90
– 1000 KD. It is classified into:
A. Alpha-I globulins: e.g. prothrombin and -1- anti-trypsin.
B. Alpha-II globulins: e.g. ceruloplasmin (Cu2+ transporter).
C. Beta globulins: e.g. transferrin (Fe2+ transporter).
D. Gamma globulins: Including all antibodies, which are IgG, IgM, IgD, IgA
and IgE.
 IgG: The major type (about 80%) of serum antibodies.
o It is responsible for second immune response.
o It is the only type which can cross the placental barrier.




IgA: Present in the body secretions as milk and saliva.
IgM: Responsible for primary immune response.
IgE: Responsible for immunity against parasites.
IgD: Present in B-lymphocyte surface acting as specific antigen receptors.
Clinical Significance of Proteins:
 Variations in plasma proteins concentration occur due to one or more of the following
changes:
A. Change in the rate of synthesis.
B. Chang in the rate of excretion.
20
C. Change in the volume of distribution.
Hypoalbuminaemia or hypoproteinaemia:
Due to;
o Decrease in the rate of protein synthesis as in liver diseases, malnutrition and
malabsorption (diarrhoea).
o Increase in the rate of protein excretion (↑protein loss) as in: kidney disease e.g.
nephrotic syndrome, severe burns, haemorrhage, fever, cancer and preeclampsia.
o Increase the volume of distribution as in: over hydration, hypoxia and septicemia.
Hypo γ-globulinaemia:
o Either congenital or acquired leading to immune deficiency and recurrent infections.
Hyperalbuminaemia:
o Mainly due to decreased volume of distribution as in: dehydration.
Hyper γ-globulinaemia:
o Due to increase the rate of synthesis of γ-globulins as in: cancer.
Albumin / Globulin Ratio (A/G Ratio):
 Normal value of A/G ratio: is about 1.6 – 2.5.
Clinical Significance of A/G ratio:
Decreased A/G ratio: as in
o Chronic liver diseases due to decrease in the rate of albumin synthesis.
o Kidney diseases due to loss of albumin in urine due to its low M.wt. and high
concentration.
Increased A/G ratio: as in
o Hypo γ-globulinaemia as in immune deficiency.
o After IV albumin infusion.
2.3.3. Urine protein test:
 Healthy kidneys filter all proteins from the bloodstream and then reabsorb them, allowing no
protein, or only slight amounts of protein, into the urine.
 The persistent presence of significant amounts of protein in the urine, then, is an important
indicator of kidney disease.
 A positive screening test for protein (included in a routine urinalysis) on a random urine
sample is usually followed up with a test on a 24-hour urine sample that more precisely
measures the quantity of protein.
 Urine protein, a 24-hour urine collection should contain no more than 150 mg/day of
protein.
2.3.4. Urine osmolality (tubular function test):
 Urine osmolality is a measurement of the number of dissolved particles in urine. It is a more
precise measurement than specific gravity for evaluating the ability of the kidneys to
21
concentrate or dilute the urine.
 The test may be done on a urine sample collected first thing in the morning, on multiple
timed samples, or on a cumulative sample collected over a 24-hour period. The patient will
typically be prescribed a high-protein diet for several days before the test and be asked to
drink no fluids the night before the test.
 Concentration tests (usually after Pitressin), and Dilution tests, after a water load. Ratio of
osmolality (or urea) in urine relative to that in plasma is a simple practical measure.
 A 24-hour urine osmolality should average 300–900 mosm/kg.
 A random urine osmolality should average 500–800 mOsm/kg.
 If fluid intake is decreased, the kidneys excrete less water and the urine becomes more
concentrated.
o With restricted fluid intake (concentration testing), osmolality should be greater than
800 mOsm/kg of water.
 Kidneys that are functioning normally will excrete more water into the urine as fluid intake
is increased, diluting the urine.
o With increased fluid intake (dilution testing), osmolality should be less than 100
mOsm/kg in at least one of the specimens collected.
3. MISCELLANEOUS TOPICS
Polyuria:
 Water diuresis (urinary osmolality <200);
o Compulsive water drinking.
o Diabetes insipidus - neurogenic or nephrogenic.
 Osmotic diuresis (urinary osmolality + 300);
o Caused by the presence of incompletely reabsorbed solutes in the tubular lumen:
 Na+ - dietary, iatrogenic, diuretics, salt-losing nephritis.
 Urea – CRF, recovery phase of acute tubular necrosis or post-renal failure.
 Glucose (diabetes mellitus).

Mannitol and some other therapeutic agents.
Renal Stones (Nephrolithiasis):
Causes:
1. A high concentration of a substance in the urine due to;
 low urine volume

high excretion rate
2. pH changes;
 Alkaline urine predisposes to Ca deposition (e.g. infection).
 Acid urine predisposes to uric acid deposition.
22
3. Stagnation, usually due to obstruction.
Types of stones (or calculi);
1. Calcium - oxalate (+ phosphate)
2. Phosphate.
3. Uric acid - in about 10% of gouty cases. May be associated with low urinary pH due to
inadequate buffer production.
4. Rare forms;
o Cystinuria, a transport defect of dibasic amino acids and cysteine.
o Xanthine: xanthine oxidase deficiency.
o 2,8 dihydroxyadenine: Adenine Phosphoribosyl Transferase (APRT - a purine
salvage enzyme) deficiency cystine:
 Calculi are only partly mineral; up to 60% may consist of protein, the rest being varying
proportions of calcium, magnesium, ammonium, phosphate, etc.
Treatment;
 Fluids ++ to keep urine dilute (in all cases).
 Potassium citrate and thiazide diuretics may help prevent Ca2+ stone formation - urinary
citrate helps solubilize calcium.
 Alkalinisation may help prevent uric acid (but not 2, 8 dihydroxyadenine) stone formation.
Renal acidosis:

Tubular function test:
There are 2 components to H+ excretion by the kidney: (i) reabsorption of filtered
bicarbonate in the proximal tubule, and (ii) H+ secretion in the distal tubule.
1- Uraemic Acidosis;
 It is seen in acute or chronic renal failure.
 It is decreased H+ excretion due to both glomerular and tubular failure.
 It is increased anion gap due to retention of phosphate, sulphate and other anions.
 Acidosis develops only when the GFR falls below about 20 ml/min. (plasma creatinine >350
μM). It associated with hyperkalemia.
2. Renal Tubular Acidosis (RTA);
 A group of disorders characterized by tubular dysfunction, with normal or perhaps slightly
decreased glomerular function.
 The picture is that of a normal anion gap (i.e. hyperchloraemic) metabolic acidosis, in the
presence of a normal or near-normal plasma creatinine.
 The urine pH is often inappropriately high in the face of the systemic acidosis (but NOT
always - see below).
Type 1 (distal) RTA:
 Due to inability of distal nephron to excrete H+. The urine pH is inappropriately high (pH
> 5.5), but does not contain significant bicarbonate.
23
 Associated with hypokalemia, nephrocalcinosis and rickets. Hypokalemia occurs because
there is an increase in K+/Na+ exchange compensating for the defect in H+/Na+ exchange
in the distal tubule.
 There are genetic and acquired forms (e.g. heavy metal toxicity).
Type 2 (proximal) RTA:
 Due to defective proximal bicarbonate reabsorption.
 The renal threshold for bicarbonate is decreased (normal value 24 mmol/l). Whenever the
plasma bicarbonate level exceeds the (lowered) renal threshold (e.g. 16 mmol/l), the urine
contains large amounts of bicarbonate and the urine pH is inappropriately high (called
renal bicarbonate wasting).
 However, if the plasma bicarbonate drops to the level of renal threshold, then all filtered
bicarbonate can be reabsorbed, and the urine pH can be appropriately acidic (< 5.5) since
distal tubular H+ excretion is normal.
 Associated with hypokalemia, due to the increased delivery of Na+ to the distal tubule
where Na+/K+ exchange occurs.
 Proximal RTA may be isolated or may be associated with other proximal tubular defects:
glycosuria, phosphaturia and amino aciduria - the Fanconi syndrome. Can be genetic (e.g.
with cystinosis) or acquired (e.g. toxins).
 (Type 3 RTA is a mixed form of types 1 and 2 - not recognized nowadays as a specific
entity.)
 The acidosis due to mineralo-corticoid deficiency or to renal resistance to mineralocorticoid action is sometimes called Type 4 RTA. Associated with hyperkalemia.
Mineralo-corticoid resistance may be a specific genetic entity (pseudohypoaldosteronism,
see below) or may result from generalized tubular damage.
With my best wishes
Dr: Mohamed I Kotb El-Sayed
Email: [email protected]
[email protected]
https://web.facebook.com/drmohamed.k.elsayed
https://scholar.google.com.eg/citations?user=47ZPvuAAAAAJ&hl=ar&oi=ao
WhatsApp (00967736318958)
24
REFERENCES

Assessment of kidney function: Serum creatinine, BUN, and GFR. Retrieved Spring 2008 from
http://www.uptodate.com.

Coomes MW. Amino acid metabolism. In Devlin TM, ed. Textbook of biochemistry with clinical
correlations. 6th ed. Hoboken, N.J.: Wiley-Liss, 2006:743.

Cory JG. Purine and pyrimidine nucleotide metabolism. In Devlin TM, ed. Textbook of biochemistry
with clinical correlations. 6th ed. Hoboken, N.J.: Wiley-Liss, 2006:789.

Garde AH, Hansen AM, Kristiansen J, Knudsen LE. Comparison of uncertainties related to
standardization of urine samples with volume and creatinine concentration. Ann Occup Hyg 2004;
48:171.

Lamb E, Newman DJ, Price CP. Kidney function tests. In Burtis CA, Ashwood ER, Bruns DE, eds.
Tietz Textbook of clinical chemistry and molecular diagnostics. 4th ed. Philadelphia, Pa.: WB
Saunders, 2005:797.

Levey AS, Coresh J, Balk E, et al. National Kidney Foundation practice guidelines for chronic
kidney disease: Evaluation, classification, and stratification. Ann Intern Med 2003;139:137.

Levey AS, Coresh J, Greene T, et al. Expressing the Modification of Diet in Renal Disease Study
Equation for estimating glomerular filtration rate with standardized serum creatinine values. Clin
Chem 2007;53:766.

Matthews DE. Proteins and amino acids. In Shils ME, ed. Modern nutrition in health and disease.
10th ed. Philadelphia, Pa.: Lippincott Williams & Wilkins, 2006:23.

Myers GL, Miller WG, Coresh J, et al. Recommendations for improving serum creatinine
measurement: A report from the Laboratory Working Group of the National Kidney Disease
Education Program. Clin Chem 2006;52:5

National Kidney Disease Education Program. Retrieved Spring 2008 from www.nkdep.nih.gov.

Oh MS. Evaluation of renal function, water, electrolytes and acidbase balance. In Pincus MR,
Lifshitz MS, eds. Henry’s clinical diagnosis and management by laboratory methods. 21st ed.
Philadelphia, Pa.: WB Saunders, 2006:147.

Pincus MR, Tierno P, Dufour DR. Evaluation of liver function. In Pincus MR, Lifshitz MS, eds.
Henry’s clinical diagnosis and management by laboratory methods. 21st ed. Philadelphia, Pa.: WB
Saunders, 2006:263.

Rinaldo P, Hahn S, Matern D. Inborn errors of amino acid, organic acid, and fatty acid metabolism.
In Burtis CA, Ashwood ER, Bruns DE, eds. Tietz textbook of clinical chemistry and molecular
diagnostics. 4th ed. Philadelphia, Pa.: WB Saunders, 2005:2207.

Surveys 2007: C-B chemistry/therapeutic drug monitoring and U-B urine chemistry. Northfield, Ill.:
College of American Pathologists, 2007.

VITROS AMON Slides. Instructions for use. VITROS Chemistry Products. Rochester, N.Y.: OrthoClinical Diagnostics, 2002 (Publication no. MP2-90).

Wu AHB. Tietz clinical guide to laboratory tests. 4th ed. St. Louis, Mo.: WB Saunders, 2006.

Young DS. Effects of preanalytical variables on clinical laboratory tests. 3rd ed. Washington, D.C.:
American Association for Clinical Chemistry Press, 2007.
25

Dr: Mohamed I Kotb El

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