Core I Objectives - Three-Dimensional Orthopaedic Animations

Transcription

Cells, Tissues, Organs Obejctives
08-15-03
The Human Genome and Medicine of the Future
CS&F B1
Dr. Anderson
1. Describe the general characteristics of the human genome.
40k+ genes. Sequenced x2.5 years ago
90% introns
2. Discuss current and potential future applications of DNA sequence information to
medical problems.
CF
Page 1 of 29
08-18-03
Protein Structure & Function (Review)
CS&F B2
Dr. Levy
1. Define pH, acid, base, and buffer.
pH = -log[H+]
Acid: A proton donor. Examples: hydrochloric acid, phosphoric acid, acetic acid, carbonic acid,
ammonium chloride
Base: A proton acceptor. Examples: sodium hydroxide, ammonia, sodium acetate, sodium carbonate.
Buffer = A buffer is a mixture of a weak acid and its conjugate base.
Such a mixture tends to resist changes in pH when either acids or bases are added.
The buffering tendency is greatest at pH values near the pKa of the weak acid (pKa = logKa).
2. Explain the difference between strong and weak acids.
Strong acid: An acid which dissociates fully, so that the concentration of protons released is equal to the
concentration of acid added to water.
Weak acid: An acid which dissociates partially. The concentration of protons released will be a function
of the dis sociation constant of the acid. Most important acids in biological systems are weak acids,
i.e. amino acids, carbonic acid, citric acid, H2PO43. Define the Ka and pKa for a weak acid.
Acid dissociation constant. i.e. the equilibrium constant for the breaking apart of a weak acid into its
hydrogen and conjugate base in a water solution.
HA <======> H+ + A Ka = [H+ ][A -]/[HA]
pKa = -logKa
4. Write and explain the Henderson-Hasselbalch equation and draw a typical titration curve for a weak
acid.
•
A mathematical relationship, derived from the equilibrium equation for
of a weak acid, that describes the behavior of buffers.
•
Relates the pH to the concentrations of the weak acid [HA]
and its conjugate base [A -] and to the pKa of the weak acid.
dissociation
pH = pKa + log[conjugate base]/[weak acid] or pH = p Ka + log[A -]/[HA]
5. Describe the general properties of amino acids.
Amino acids are the building blocks of proteins.
COO|
+NH3 - C - H
|
R
•
The alpha carboxyl group of amino acids has a pKa of about 2-3 and, thus, is ionized at neutral pH.
(It exists as the conjugate base.)
Page 2 of 29
CS&F B2 Protein Structure & Function (Review) Page 2
•
The alpha amino group has a pKa of about 9-10 and, thus, is protonated at neutral pH. (It exists in
the acid form.)
• At pH values around neutrality an amino acid has a negative charge on its carboxyl group and a
positive charge on its amino group and is in the so-called zwitterion form. See the titration curve
of an amino acid (Leucine).
• The R groups of certain amino acids can also ionize and will contribute to the overall charge of the
amino acid in a pH-dependant way. See the titration curve of an amino acid with an ionizable R
group (Glutamic Acid).
• The isoelectric point, or pI, is the pH at which the net charge of a molecule is zero.
6. Name the 20 common amino acids and be able to group them into categories based on their R groups.
Here is list where amino acids are grouped according to the characteristics of the side chains:
* Aliphatic - alanine, glycine, isoleucine, leucine, proline, valine
* Aromatic - phenylalanine, tryptophan, tyrosine
* Acidic - aspartic acid, glutamic acid
* Basic - arginine, histidine, lysine
* Hydroxylic - serine, threonine
* Sulphur-containing - cysteine, methionine
* Amidic (containing amide group) - aspara gine, glutamine
Amino Acid
---------alanine
arginine
3- & 1- letter codes
-------------------ala
A
arg
R
Side Chain
----------CH3
+
-CH2-CH2 -CH2-NH-C=NH2
|
NH2
asparagine
asn
N
aspartic acid
asp
D
-CH2-CO-NH2
_
-CH2-COO
cysteine
cys
C
-CH2-SH
glutamine
gln
Q
-CH2-CH2 -CO-NH2
glutamic acid
glu
E
-CH2-CH2 -COOH
glycine
gly
G
-H
histidine
his
H
-CH2-C = CH
|
|
+N=C-NH
H
isoleucine
ile
I
-CH-CH2-CH3
|
CH3
leucine
lysine
leu
lys
L
K
-CH2-CH-CH3
|
CH3
+
-CH2-CH2 -CH2-NH3
methionine
met
M
Amino Acid
3- & 1- letter codes
----------------------------phenylalanine
phe
F
-CH2-CH2 -S-CH3
Side Chain
----------CH2-Ph
proline
pro
P
-CH2-CH2 -CH2(the terminus is bonded
to the
main chain amine
nitrogen to
form a ring)
serine
ser
S
-CH2-OH
threonine
thr
T
-CH-CH3
|
OH
tryptophan
trp
W
-CH2-C = CH
|
|
Ph -NH
(the phenyl group is
bonded at carbons 1 and
2 ie 'ortho' positions)
tyrosine
tyr
Y
-CH2-Ph-OH
(the phenyl group is
bonded at carbons 1 and
4 ie 'para' positions)
valine
val
V
-CH2-CH3
|
CH3
7. Draw the peptide bond and describe its properties.
Proteins are made up of long chains of amino acids
linked together by peptide bonds.
The peptide bond has partial double-bond character.
This restricts rotation about the bond and influences
the possible conformations of proteins.
Page 3 of 29
08-18-03
Protein Structure & Function (Lecture)
CS&F B2
Dr. Levy
Lecture Objectives
1. General properties of proteins: functions, size, shape, charge, solubility
fxs: catalysis, regulators, transporters, contractile elements, defense, (most) structural; collagen most
abundant (90%)
size: 6000 – 4x107 microns
shape: fibrous – long, globular – globular
charge: @ isoelectric point, sum of all charges of protein = 0. changes with pH
Solubility: some aqueous, water soluble, lipids not. Solubility of protein depends on its AA
composition & pH of sol’n
2. Levels of protein structural organization:
primary: AA sequence
secondary: Stx from folding polypeptide chain into repeating pattern stabilized by H bonds
e.g. α-helix, β-sheet
tertiary: three dimensional stx of protein stabilized by H bonds, ionic bonds, hydrophobic
interactions. Restricted by steric hindrance, bond rotations, electrostatic interxns
quaternary structure: non-covalent association of protein subunits
protein complexes: e.g. fatty acid synthetase, viral coat protein
CLINICAL: sickle -cell anemia results from gluàval. Val apolar, so HgB forms into rod
3. Protein folding: mediated by chaperones (helper molecules block non-productive folding)
denaturation: unfold protein, e.g. ribonuclease, insulin
4. Structure-function relationships (CFTR, hemoglobin)
CFTR (cystic fibrosis transmembrane conductance regulator): stx allows chloride ions to cross.
Hemoglobin: structure allows it to carry oxygen at multiple sites
Page 4 of 29
08-19-03 & 08-20-03
Enzymes
CS&F B3 B4
Dr. Levy
Describe the following:
1. General properties of enzymes
ID: A protein molecule produced by living organisms that catalyses chemical reactions of other
substances without itself being destroyed or altered upon completion of the reactions.
globular proteins
coenzyme: composed of protein & nonproteinous moiety e.g. metal ion
1 subunit or multiple
multisubunit: subunits may do same thing; some regulate, some carry out activity
consecutive reactions, enzymes aggregate to increase efficiency of synthesis of fatty acid
2. Interaction of enzyme with substrate
binding site
non-covalent forces drive site bonding
ionic interactions
H-bond
Hydrophobic interactions
Dynamic interactions: substrate touch protein, induce conformations change so enzyme and
protein can bind.
3. Enzyme catalysis; Michaelis-Menten equation
enzymes decrease the energy required to start the reaction
∆G = ∆G0 + 2.3RT log Keq
∆G0 = - 2.3RT log Keq; where Keq = [P]/[S]
∆G = potential to do work
∆G < 0, eXergonic, reaction releases energy in the form of heat, light, etc., S à P
∆G > 0, eNDergonic, reaction consumes energy, S ß P
Michaelis -Menten
v = Vmax [S] where Km = [S] at 1/2 Vmax
Km + [S]
Km = A kinetic parameter used to
characterize an enzyme, defined as the
concentration of substrate that
permits half maximal rate
of reaction.
Page 5 of 29
CS&F B3,B4 Enzymes page2
4. Enzyme inhibition
Inhibitor: any substrate that decreases the Vmax or changes the Km
Competitive Inhibitor: compete with substrate at active site
No ∆ Vmax, increase Km (need more S to react at ½ Vmax)
Noncompetitive Inhibitor: bind to non-active site
May induce conformational ∆
Rxn will never reach Vmax, there is less enzyme available. No ∆ Km. Enzyme is not inhibited.
5. Mechanism of enzyme reactions
a. Specificity (vast spectrum)
Ultimate specificity: only 1 3-dimensional stx will fit
Need to Not be specific, e.g. digestive enzymes, peptidases
b. Functional Groups
What Amino Acids are involved in binding/catalytic process
c. Catalytic Efficiency
i. Proximity and Orientation
Enz binds S exactly so [ ] locally high, lower PE barrier
ii. Covalent Intermediates
E + S à ES à E + P
ES = covalent intermediate
iii. General Acid-Base Catalysis
many rxns require acid/base. In active site, AA side chains act as acids or bases in localized
volume of active site. Effective pH in site very potent. Doesn’t affect outside
iv. Distortion of Substrate
make bond more easily broken by distorting the bond
Page 6 of 29
CS&F B3 B4 Enzymes
6. Regulation of enzyme activity
a. Protein synthesis and degredation
b. Activation of enzyme precursors
response to environment. E.g. digestive enzymes. Don’t want active if nothing around to
digest cuz they’ll start eating the stomach
c. Isozymes (Isoenzymes)
Variants of enzymes that catalyse the same reaction. Different tissues often have different
isoenzymes. The sequence differences generally confer different enzyme kinetic parameters
that can sometimes be interpreted as fine tuning to the specific requirements of the cell types
in which a particular isoenzyme is found.
e.g. fetus, Hgb chains change during development post birth
e.g. cancer cells, some revert to more basic state
e.g. hexokinase (other cells), glucokinase (liver)
d. Enzyme derivitization
e.g. Phosphorylation: phosphate switch. Add a phosphate and make the enz more active
e. Alteration of enzyme subunit interaction
e.g. adenylate cyclase: cAMP change enzyme and break off active catalytic subunits
f. Allosteric Enzymes
A regulatory enzyme whose activity is modified by the noncovalent binding of a particular
metabolite at a site (the allosteric site) other than the active site.
(+) cooperativity: one enzyme subunit turned on, next will be easier
(_ ) cooperativity: one enzyme subunit turned on, next will be more difficult
Clinical Correlations:
a. Tissue specific enzymes as markers of pathology
e.g. lactate dehydrogenase levels increase immediately following a heart attack
b. Isoenzyme Patterns
more H in heart, more M in liver.
c. Immobilized enzymes as reagants
DM, urine sample, color development proportional [glucose]
same for cholesterol
d. Blood clotting treat immediately with streptokinase (a plasminogen activator)
Page 7 of 29
08-21-03
DNA Replication
CS&F B5
Dr. Reddy
Learning objectives prior to class (from self-study review of nucleotides and nucleic acids)
1. State the names of the major nucleotides found in DNA and RNA as well as how they base pair.
Nucleosides (Base + Sugar)
• Adenosine, Ado, A Deoxyadenosine dAdo dA (Structures)
• Guanosine, Guo, G Deoxyguanosine dGuo dG
• Cytidine, Cyd, C Deoxycytidine dCyd dC
• Uridine, Urd, U Deoxyuridine dUrd dU
• Thymidine, Thd, T Deoxythymidine dThd dT
Base Pair:
T=A
G≡ C
Nucleotides (general form, N = any nucleoside)
NMP, NDP, NTP = nucleoside mono-, di-, or tri-phophate.
Above nomenclature implies P’s are 5’ unless specified otherwise.
Examples: AMP, ADP, ATP (adenosine triphosphate).
dNMP, dNDP, dNTP = deoxynucleoside mono-, di-, or tri-phosphate.
d implies that the deoxy position is 2’ unless specified otherwise.
Examples: dGMP, dTDP, dCTP.
2. Explain the differences between a nucleotide, a nucleoside, and a purine or pyrimidine base and
recognize typical structures for each.
nucleotide
<biochemistry> Phosphate esters of nucleosides. The metabolic precursors of nucleic acids are monoesters
with phosphate on carbon 5 of the pentose (known as 5' to distinguish sugar from base numbering).
nucleosides
nucleotide
Page 8 of 29
CS&F B5 DNA Replication Page 2
nucleosides
Purine or pyrimidine bases attached to a ribose or deoxyribose.
purine
<biochemistry, molecular biology> A heterocyclic compound with a fused pyrimidine/imidazole ring.
Planar and aromatic in character. The parent compound for the purine bases of nucleic acids.
Pyrimidine
<biochemistry> A family of 6-membered heterocyclic compounds occurring in nature in a wide variety of
forms. They are planar and aromatic in character and include several nucleic acid constituents (cytosine,
thymine, and uracil) and form the basic structure of the barbiturates.
3. List the biological roles of nucleotides and nucleic acids.
I. Importance of Nucleic Acids and Nucleotides
A. Key roles of nucleic acids and nucleotides in biochemistry
1. Nucleic acids (composed of nucleotide building blocks) are essential for the storage, transfer, and
expression of genetic information.
a. DNA -- genetic information storage and replication
b. RNA -- roles in expression of genetic information as protein (transcription and translation)
2. Nucleotides are essential components of many cofactors and high energy metabolites (ATP, UDPG,
CoA, NAD, etc.).
3. Some nucleotides are metabolic regulators (i.e. cAMP).
B. Some relationships of nucleotides and nucleic acids to medical problems
1. Hereditary diseases -- Due to mutations in DNA passed from parent to child.
2. Somatic mutations (cancer) -- Due to mutations in DNA which occur in cells of the body and
alter the growth properties of those cells.
3. Abnormalities of nucleic acid and nucleotide metabolism -- Such as excessive degradation of
purines, leading to gout.
4. Targets for clinically useful drugs -- Used to treat cancer, microbial infections, etc.
5. Genetic diagnosis / gene therapy -- Nucleic acids are the basic material for these approaches.
Page 9 of 29
4. Describe the distinguishing structural characteristics of DNA and the three major types of RNA.
General characteristics of the polynucleotides formed from nucleotide building blocks
1. The nucleotide units are linked by 3',5'-phosphodiester bonds.
2. Negative charges are present at physiological pHs due to ionized phosphates in the sugarphosphate backbone
3. Nucleic acid chains have polarity due to the structure of the sugars (3' à 5' or 5' à 3'
orientations)
4. Ribose (RNA) or deoxyribose (DNA) sugars determine the nature of the nucleic acid.
5. Four major bases are typically found in each kind of nucleic acid (A, G, C, T in DNA and
A, G, C, U in RNA).
6. Nucleic acids may be single or double stranded (the two strands are antiparallel when
double stranded).
7. Complementary base pairing rules: A pairs with T or U (2 H-bonds) ;
G pairs with C (3 H-bonds)
8. Additional images of a T-A base pair and a G-C base pair
Structural features of the B form of DNA (the major form in cells) (Image 1 | Image 2)
a. H-bonding and base stacking -- forces which stabilize the double helix.
b. Inner base pairs and outer sugar-phosphate backbone (negatively charged).
c. Antiparallel nature of the two strands.
d. Right-handed helix: 10 base pairs per turn, 3.4 A per base pair, ~20 A diameter
e. Major and minor grooves allow proteins to interact with the base pairs of the structure.
f. Very high molecular weight.
g. May be linear (human nuclear chromosomes) or circular (bacterial chromosome, human
mitochondrial genome).
RNA (ribonucleic acid)
1. General characteristics and major functional types
a. Differences from DNA -- ribose instead of deoxyribose, generally single stranded instead of
double stranded, U instead of T.
b. Major types and functions
i. Ribosomal RNA (rRNA) -- part of ribosome, the workbench for protein synthesis.
ii. Transfer RNA (tRNA) -- the adapter molecule, connecting amino acid to anticodon.
iii. Messenger RNA (mRNA) -- the template for protein synthesis.
2. Ribosomal RNA
a. Four species in eukaryotic ribosomes: 5S, 7(or 5.8)S, 18S, 28S (S is the sedimentation
coefficient)
b. Contain some methylated bases
c. Form a two-subunit complex with specific proteins to make up the ribosome (site of protein
synthesis)
3. Transfer RNA (Image 1 | Image 2 | Image 3 | Image 4 | Image 5)
a. Small size (4S)
b. Adapter function in protein synthesis
c. Multiple species (carry different amino acids)
d. Many modified bases
e. Common amino acid attachment site -- CCA at 3' end
Page 10 of 29
f. Anticodon to recognize triplet codons in mRNA
g. Cloverleaf structure
4. Messenger RNA
a. Greatest variations in size and sequence among the RNAs
b. Code for the sequence of amino acids in proteins
c. Contain some methylated bases and sugars
d. In eukaryotes most have 3' poly (A) tails
e. Have a novel 5' cap structure in eukaryotes:
m7G(5')ppp(5')N'(m)-----------------3'
f. Overall mRNA structure
Lecture Objectives
Compare and contrast the general characteristics and mechanism of DNA replication and the proteins
involved in prokaryotic and eukaryotic replication.
Replication
1) Separate strands @ origin of replication
DNA-A bind @ origin of replication
Eukaryotes have multiple origins
SSB (single stranded DNA binding proteins) – keep strands separate & protect DNA from nucleases
Helicases – unwind double helix
DNA Topoisomerases – remove supercoils
I – nick/reseal. Cut a single strand of DNA
II – bind to double strands, transient breaks in both strands
2) Replication Fork
3) Direction
read 3’ -> 5’. Write 5’-> 3’
-- Leading Strand
continuous. Copied in direction of replication fork
-- Lagging Strand
discontinuous. Copied in direction away from replication fork. Okazaki fragments.
4) RNA Primer
Primase, an RNA polymerase
Primosome. Binds to DNA, displaces SSB, moves along lagging strand, synthesize occ. RNA primers
5) Chain Elongation
DNA Polymerases
III – elongation, proofreading (exonuclease activity)
I – RNA primer excised & fill gap
6) DNA Ligase – link DNA chains made by PolyIII & PolyI
Differences between Eukaryotes/Prokaryotes
Eukaryotes:
Have multiple origins of replication
DNA Polymerases:
Pol α − Primase
PCNA - proliferating cell nuclear antigen
displaces Polα and interrupts leading
strand synthesis
Pol δ - elongate leading strand.
Exonuclease (proofread)
Pol ε - elongate lagging. Exonuclease
(proofread)
Pol β :: DNA Poly I, excise primers,
“repair”
Pol γ − replicates mitochondrial DNA
Page 11 of 29
08-22-03
Transcription & Control of Transcription
CS&F B6 & B7
Dr. Stallcup
1. Compare the basic transcription machinery, the basic structure of genes (including
promoters) and transcription units, and the basic mechanism of transcription in prokaryotes
and eukaryotes.
A. General mechanism
• RNA polymerase reads the template strand of DNA from 3' to 5'.
• The RNA transcript (complementary to the template DNA strand) is synthesized 5' to 3'.
• Substrates: ATP, GTP, CTP, UTP
• No primer required
• Local unwinding of the DNA double helix is required, aided by DNA topoisomerases
B. Structure of the gene or transcription unit
• Transcription start site is defined by promoter region. TATA box
• Transcription stop site is defined by terminator sequences.
C. Types of RNA % of cellular RNA number of species
rRNA 80% 3-4
tRNA 15% about 50
mRNA 5% 1000s
D. Basal transcription machinery
Prokaryotes
• 1 RNA polymerase composed of four subunits (two alpha, one beta, one beta-prime)
• sigma subunit recognizes promoter and directs the polymerase to that site
Eukaryotes
• RNA pol I (large rRNAs); RNA pol II (mRNAs); RNA pol III (tRNAs & small rRNA) (1,2,3:rmt)
• Different “basal transcription factors” help each polymerase recognize promoter and initiate
transcription.
Mitochondria
• Have their own RNA polymerase which transcribes mitochondrial DNA into rRNAs,
mRNAs, and tRNAs
2. Explain the structure of prokaryotic operons and various mechanisms for induction or
repression of their transcription.
• 3000 total genes, organized into clusters or operons of coordinately regulated genes.
• Transcription of an operon produces a poly-cistronic mRNA, encoding multiple proteins.
• When genes are organized into an operon, they can be coordinately expressed.
• Each operon has a single promoter, controlled by one or more control elements.
• Binding of a specific regulatory protein to the control element can either enhance or inhibit
initiation of transcription at the promoter by RNA polymerase.
• Some operons are controlled by induction, i.e. transcription is turned on when it is needed. (default=off)
• Other operons are controlled by repression, i.e. transcription is turned off when not needed. (default=on)
• Each structural gene in the operon and the poly-cistronic mRNA transcribed from the operon
has its own translation start signal to attract ribosomes.
Induction/Repression
Lac Operon: Induction by lactose. When present, lactose binds to repressor and causes a change in the
repressor shape which prevents the repressor from binding to the operator. This allows the operon to be
transcribed. When the lactose is exhausted, the repressor binds to the operator and blocks transcription.
Lac Operon: Repression by glucose:
· binding site for a protein called CAP (catabolite activator protein) in the promoter.
· When CAP binds to the promoter, it helps to recruit RNA polymerase.
· cAMP must bind to CAP and change its shape so that it can bind to the lac promoter.
· When glucose is present, cAMP level is low. When glucose is exhausted, cAMP level increases.
Page 12 of 29
CS&F B6 B7 Transcription & Control of Transcription Page 2
3. Discuss the roles of transcriptional activator proteins, enhancer elements, coactivators, and
chromatin in regulation of eukaryotic transcription.
Transcriptional activator proteins recruit proteins called “coactivators” which help to:
· Alter chromatin structure to make the promoter more accessible (unfold)
· Recruit RNA polymerase II and its basal transcription factors
· Many transcriptional activator proteins can also bind directly to basal transcription factors associated with
RNA polymerase II to help recruit it to the promoter.
Function of enhancer/silencer elements
•
binding site for a specific transcription factor.
•
Binding of a transcription factor to its enhancer/silencer element causes activation/repression of
transcription.
•
may be controlled by environmental conditions
Remodeling of chromatin structure by coactivators recruited to a promoter by transcriptional activator factors
(TF); open chromatin is more accessible to RNA polymerase.
4. Describe the cellular response (or signal transduction) pathway used by steroid hormones and
list the major hormones which interact with members of the nuclear receptor family.
• Some hormones (e.g. insulin, catecholamines) bind to receptors on the cell surface
• Activate protein kinases and/or phosphatases in the cell
• Release 2nd messengers (cAMP, inositol triphosphate, calcium, diacylglycerol)
• Other hormones (steroid, thyroid, retinoid, vitamin D) bind to intracellular receptors
• These “nuclear receptors” are hormone-regulated transcriptional activator proteins
which bind DNA and activate transcription of target genes
Response pathway for steroid hormones
• Steroid enters cell and binds receptor in cytoplasm or nucleus, changing shape of receptor
• Hormone-activated receptor forms dimers and binds one specific type of enhancer element
(hormone response element, or HRE) found in promoters of target genes
• DNA-bound receptor recruits coactivators which remodel chromatin and recruit RNA
polymerase II and its basal transcription factors, to activate initiation of transcription.
Steroids
Glucocorticoids
cortisol
mineralocorticoids
aldosterone
estrogens
estradiol
androgens
testosterone
progestins
progesterone
Steroid-related
vitamin D
retinoic acid (vitamin A)
Amino acid derivative
thyroid hormones
thyroxine, triiodothyronine
Others???
5. Explain why agonists promote gene activation by steroid receptors, but antagonists inhibit steroid receptor fx.
Function lies in the conformational change in the receptor when the (ant)agonist binds to the receptor.
• Receptor conformation in presence of agonist can interact productively with coactivators
and/or other components of the transcription initiation complex
• Receptor conformation in the presence of antagonist cannot interact with coactivators or
other components of the transcription initiation complex e.g. Tamoxifen (breast cancer, estrogen antagonist),
e.g. prostate cancer, tx with androgen antagonists.
Page 13 of 29
CS&F B6 B7 Transcription & Control of Transcription Page 2
6. Discuss the roles of steroid receptors and their agonists/antagonists in the etiology and/or
treatment of a few diseases. CLINICAL
Cancer
· Breast
tumor growth depends on estrogen; estrogen antagonists used in therapy
coactivator gene AIB1 is amplified and/or over-expressed in many tumors
· Prostate
tumor growth depends on androgen; androgen antagonists used in therapy
Genetic diseases
· Hormone resistance due to deletion or mutation of nuclear receptor gene (thyroid,
androgen, vitamin D, glucocorticoid)
· Spinal and bulbar muscular atrophy (mutation in androgen receptor)
7. Explain how the cAMP signaling pathway can regulate transcription of specific genes.
Regulation of transcription by cAMP via CREB
· Many hormones (not those in the steroid and nuclear hormone group) have receptors on the cell surface.
· Binding of hormone to cell surface receptor causes release of second messengers, like cAMP, into the
cytoplasm (through activation of adenylate cyclase, catalyst for rxn ATPàcAMP)
· cAMP binds to and activates the cAMP-dependent protein kinase (protein kinase A or PKA).
· PKA phosphorylates many proteins to change their activities.
· One protein which is phosphorylated by PKA is the cAMP Response Element Binding protein (CREB).
· CREB binds to a specific enhancer element (cAMP-responsive element or CRE) which is associated with
genes that are regulated by cAMP.
· Phosphorylation of CREB allows it to bind and recruit coactivators, which help to activate transcription.
Page 14 of 29
8.26.03
RNA Processing
CS&F B8
Dr. Laird-Offringa
1. Describe the different kinds of RNA in the mammalian cell and their functions.
2. State the steps in the processing of mRNA.
Transcription initiation
Transcription termination and polyadenylation
Splicing. GU…AG flanking nucleotides. GU splice site. Attach to branchpoint A to form lariat. Ligation. C/U
rich region. AG end.
3. Describe how RNA processing could help regulate gene expression and stimulate biological diversity. Explain
how mistakes in RNA processing could lead to human disease.
Export
Localization
Translation
Destruction
Page 15 of 29
08-26-03
Translation
CS&F B9
Dr. Laird-Offringa
Lecture Outline:
A. Translation: the decoding of mRNA
B. The Degeneracy of the Genetic Code
C. The Mechanism of Translation
D. Splicing and Translation: the Need to Maintain the Correct Reading Frame
E. Nonsense-Mediated mRNA Decay
F. Diseases related to mRNA Translation
Learning Objectives:
1. Principle of mRNA translation & degeneracy of genetic code
Translation: decoding of mRNA
RNA à protein
DNA/RNA = nucleotides, Protein = amino acids, tRNAs translate codons into specific AAs
Degeneracy of the Genetic Code
64 codons / 20 AAs
2. Summarize general steps of translation
A: Scanning & Initiation
Initiation: mRNA w/cap (AUG), 40S ribosomal subunit, met-tRNA, initiation factors, GTP, Mg2+, ATP
40S-cap, accAUGg = Kozak sequence = start, tRNA, 60S subunit joins for protein synthesis
B: Elongation
read in triplets
C: Peptide Bond Formation
D: Translocation
shift peptidyl-tRNA to P site, new aminoacyl-tRNA to A site
2 GTP / AA addition
E: Termination
stop codon = UAA, UAG, UGA
3. Aberrant translation
splicing mutation:
splice at wrong site, could mean cut out 1 or 2 nucleotides instead of a multiple of 3
this would result in a frameshift
nonsense-mediated mRNA decay
nonsense mutation = stop codon too soon. cell detects downstream exon/exon junctions. degrade
mRNA cuz most introns not in this region
aminoglycoside antibiotics
otoxic antibiotics. interfere with mitochondrial translation cuz mito mutated and more sensitive
Page 16 of 29
08-26-03
CS&F B10
DNA Mutation & Repair
Dr. Laird
1. Describe the relationship between DNA damage, DNA repair, DNA replication and mutagenesis.
DNA Mutation = alteration in DNA sequence
Can arise from
Error in replication process
DNA Damage by chemicals or irradiation
Reversible by DNA repair unless DNA replication has occurred
Fate of a pre-mutagenic lesion depends on the race between DNA replication and DNA repair.
2. State the major sources of DNA damage and the major types of DNA repair.
B. Sources of DNA Damage
1. Endogenous Sources
• DNA Replication Errors (misincorporation, slippage)
• Deamination (cytosine to uracil, 5-methylcytosine to thymine)
• Depurination (creates abasic site)
• Reactive Oxygen Species (strand breaks, base damage)
• DNA Recombination Errors
2. Environmental Sources
• Ionizing Radiation (IR) increases reactive species (Indirect Mechanism)
double strand breaks in DNA very dangerous
• Ultraviolet Radiation (UV) generates pyrimidine (T/C) dimers (Direct Mechanism)
• Chemical Mutagens (eg alkylation by MNNG, MNU–›O6-methylguanine (Pairs like A)
C. Types of DNA Repair (mild --> more severe)
1. Lesion is a normal base, but mispaired
• Proofreading exonuclease activity of DNA polymerases can backtrack and correct
misincorporated nucleotides. structural abnormality e.g. distort double helix trigger proofreading
• Postreplication mismatch excision repair.
Known steps in the process are: Mismatch Recognition –› Strand Discrimination –›
Excision –› Resynthesis –› Ligation.
which strand correct? new strands have nicks (euk), parent strand methylated (e.coli)
e.g. HNPCC, Sporadic MSI positive colorectal cancer
2. Lesion is recognizable as a base, but abnormal
• Base excision repair. DNA glycosylase (base flipping and excision) –›
AP endonuclease –› DNA polymerase –› Ligase. abnormal bases (Examples: uracil, hypoxanthine,
3- methyladenine).
Creates an abasic site during the repair process. potentially premutagenic lesion
• Direct reversal of damage: O6-Guanine Methyltransferase (MGMT) (protein commits suicide)
3. Lesion is a damaged base or bulky adduct
• Nucleotide excision repair. Most important including bulky adducts and UV damage that
destroy the recognizability of the base. repair more active on transcribed strand in genes. >30 gene
products attributed fx in nucleotide excision repair. The
mechanism proceeds in the following steps:
Damage recognition –› Nuclease cleavage –› Removal (Helicase involvement) –›
DNA polymerase delta/epsilon –› DNA ligase.
e.g. Xeroderma Pigmentosum
4. DNA Strand Breaks
• Single -strand nicks: DNA ligase.
• Double-strand breaks: Double strand break repair.
Page 17 of 29
CS&F B10 DNA Mutation & Repair Page 2
3. Describe the clinical consequences of mutagenesis and of defects in DNA repair.
Clinical significance of mutagenesis:
Germline mutations: Hereditary disease
Somatic mutations: Cancer
Clinical significance of DNA repair defects:
Defective Repair
Human Disease
Clinical Features
Molecular Defect
Process
Mismatch Excision
HNPCC (Hereditary
Hereditary Colorectal
MLH1, MSH2 Germline
Repair
Non-Polyposis
Cancers, Other Cancers
Mutations (mismatch
Colorectal Cancer) (1repair proteins)
2% of CC)
heterozygous, else too
severe
Mismatch Excision
Sporadic MSI positive
Sporadic Colorectal
Inactivation of MLH1
Repair
colorectal cancer (15%
Cancer
by DNA methylation
of CC)
(why colorectal? Active
cell turnover)
Nucleotide Excision
Xeroderma
Sun Sensitivity, Cancer
7 genes (A-G), many
Repair
Pigmentosum
steps
Page 18 of 29
09/10/03 + 09/11/03
Carbohydrate Metabolism
CS&F B11 & B12
Dr. Kalra
1. Explain how glucose is metabolized and stored by various tissues in the body.
Starch __________ à disaccharides __________ à glucose à [epithelial cell] à [capillary]
Amylase
disaccharidase
What happens to glucose
Tissues using this method
Glycolysis
à pyruvate
brain
adipose
àLactic acid
RBC
àpyruvate & lactate
liver
muscle/heart
R5P
RBC
brain
liver
adipose muscle/heart
Oxidative phosphorylation of pyruvate
brain
Glycogenesis
liver
adipose muscle/heart
Gluconeogenesis
liver
Drug Detox
liver
RBC:
Glu -> lactic acid (glycolysis)
Glucose -> G6P -> Pyruvate -> Lactate
Glu -> Pentose phosphate (yield NADPH, keep glutathione (GSH) in reduced state (not GSSH))
Glucose -> G6P -> R5P
Brain:
Glu -> G6P -> pyruvate
Glu -> G6P -> pentose phosphate pathway R5P (NADPH for lipid biosynthesis)
Glu -> pyruvate -> oxidative phosphorylation -> ATP
Muscle & Heart:
Glu -> G6P -> glycolysis -> pyruvate and lactate
Glu -> G6P -> R5P, NADPH for lipid biosynthesis
Glu -> G6P -> pyruvate -> oxidative phosphorylation -> ATP
Glu -> G6P -> [glycogenesis] -> glycogen
Fat/Adpiose:
Glu -> G6P -> [glycolysis] -> pyruvate -> acetyl CoA -> fatty acids (fat storage)
Glu -> G6P -> R5P, NADPH for lipid biosynthesis
Glu -> G6P -> [glycogenesis] -> glycogen
Liver:
Glu à G6P à glycolysis -> pyruvate and lactate
Glu à G6P à [glycolysis] -> pyruvate -> acetyl CoA -> fatty acids
Glu à G6P à R5P, NADPH for lipid biosynthesis
Glu à G6P à [glycogenesis] -> glycogen
[gluconeogenesis] – synthesize glucose from long CHO chain
Drug detoxification
Glycogenolysis
Glycogen phosphorylase breaks down a[1-4] link of glycogen à G1P
Glucose --- (Hexokinase) ---> G6P
F6P ---- (PFK1) -----> F1-6BP
PhoPyr ----- (Pyruvate Kinase) ----> Pyruvate
2. Describe how high glucose (hyperglycemia) and low glucose (hypoglycemia) in the circulating blood cause
release of hormones from pancreas, which affect key enzymes involved in glycolysis, gluconeogenesis and
glycogen synthesis and its breakdown.
Hypoglycemia – glucagon released by alpha cells of pancreas
Glucagon inhibits glycolysis, promotes glycogenolysis, inhibits gluconeogenesis
Page 19 of 29
CS&F B11 B12 page 2
Glucagon binds to receptor on plasma memb rane of liver cell
Activate adenylate cyclase à cyclic AMP à
a) protein kinase (R2C2) à dissociate subunits, active C2 free à
Glycolysis inhibition
à (i) PFK2 ~P à no site on PFK2~P for F-2,6BP synthesis from F6P
à (ii)activates F-2,6Bpase -> hydrolyse F-2,6BP (à F6P) à PFK1 less effective à
àglycolysis inhibited
b) (high cAMP) à pyruvate kinase~P(inactive) à glycolysis inhibited
Hyperglycemia – insulin released from beta cells of pancreas
Insulin promotes glycolysis, inhibits glycogenolysis, promotes gluconeogenesis
Phosphatase activity inactivated à increase in intracellular F-2,6BP and stimulation in glycolysis
Hormone Present
Insulin
Glycogen
Phosphoprotein phosphatase
Adenylate cyclase
Protein kinase A (R2C2)
Protein Kinase A (C2)
PFK2 (phosphatase action)
PFK2 (kinase action)
F26BPase
F26Bpase
/\ glucokinase production
Glycogen phosphorylase
Glycogen phosphorylase~P
Glycogen synthase a
Glycogen synthase b~P
Phosphorylase b (inactive)
Phosphorylase a~P
PDH (E1)
PDH (E1~P)
Pyruvate kinase
Pyruvate kinase~P
cAMP phosphodiesterase
Pathway
Lots
Glycolysis regulation
Glycogenolysis
Glycogenesis
Glycogenolysis
(glycolysis) Pyr-> acetyl CoA
Glycolysis
3. Describe how glucose, needed by the brain, is synthesized during prolonged starvation.
Glycogen breakdown in liver (some kidney) ---à g6p à glucose (g6pase)
Prolonged, gluconeogenesis from non-carb sources. I.e. lactate (muscle/RBC) go thru Cori cycle, alanine (protein in
muscle) through alanine cycle, glycerol (fat)
4. describe how genetic deficiency of g6pase leads to enlarged liver or glycogen storage disease. (Von Geirke’s
disease)
G6pase converts g6p to glucose for release into blood. Not making glucose, not releasing it, so you’re stuck with a
bunch of g6p which enlarges liver.
5. explain how genetic deficiency of g6p dehydrogenase in RBCs leads to hemolytic anemia.
G6p deH2Oase converts g6p à r5p and uses NADP+ à NADPH to do so. NADPH in the RBCs is used to prevent
GSH from converting to GSSH. If GSSH is present, the RBC can’t oxidize peroxide and the cell will get rigid and
hemolyse which is anemia.
GSSG à GSH
NADPH à NADP+
6. Describe how hyperglycemic conditions generate glucose-protein adducts (AGE), which are deleterious to
cells.
Glucose will freely bind to proteins. Need no catalyst. When much glucose (in hyperclycemia) this will happen
more often. The AGE’s composition/structure changed so no longer functional. Enzymes not working is not good.
7. explain how AGE molecules (HbAlc ) are used as metabolic index of diabetes control.
High glucose à more glucose bind to hemoglobin = HbAlc. Measure this level. Normal = 4-6%, diabetes = 7-16%
Page 20 of 29
CS&F B11 B12 page 3
REVIEW:
Glycolytic steps at which ATP NADH form (in order in glycolysis)
ATP Consumption:
Glucose à g6p , f6p à f1,6bp
NADH production:
G3P à 1,3,bis -phosphoglycerate
ATP Production:
1,3,bis -phosphoglycerate à 3-phosphoglycerate
PEP à pyruvate
NADH Consumption:
Pyruvate à Lactate
Page 21 of 29
09/11/03
Bioenergetics & Oxidative Metabolism I
CS&F B13
Dr. Mosteller
1. Describe the role of the ATP cycle in anabolic and catabolic pathways.
Catabolic: energy production.i.e. ADP + Pi à ATP carb/lipid/protein breakdown
Anabolic: energy utilization i.e. ATP à ADP + Pi biosynthesis of macromo lecules, muscle
Contraction, active ion transport, thermogenesis
2. Name the general classes of substances that are oxidized in order to generate ATP.
All biological energy (of mammals) is derived from oxidation of foodstuffs (carb,prot,fat)
3. write an equation relating Gibbs free energy (G) to enthalpy (H) and entropy (S) and describe how changes in
Gibbs free energy change (∆G) are related to exergonic and endergonic chemical reactions and to equilibrium.
∆G = ∆H - T∆S
∆G < 0 = exergonic = spontaneous .: favored by \/ H and /\ S
∆G > 0 = endergonic = nonspontaneous, do not occur in nature
∆G = 0 = at equilibrium
4. Explain the importance of pyruvate and acetyl CoA in oxidative metabolism.
Key intermediates.
Pyruvate intermediate in glycogenolysis
Acetyl CoA derived from pyruvate, directly from some amino acids and fatty acids
Substrate for oxidation in Krebs cycle
5. Discuss the role of pyruvate dehydrogenase (PDH) in oxidative metabolism and describe its regulation. Name
five cofactors utilized by this enzyme.
PDH catalyzes the oxidative decarboxylation of pyruvate forming acetylCoA
Pyruvate + CoA + NAD+ à Acetyl CoA + NADH + H+ + CO2
3 catalytic subunits (E1,E2,E3), 2 regulatory subunits (PDH Kinase, phosphoprotein phosphatase)
regulation:
1) acetyl CoA and NADH inhibit E1 and stimulate PDH Kinase
2) pyruvate, CoA, NAD and ADP inhibit PDH kinase
3) dephosphorylated PDH (E1) is the active form of the enzyme
5 cofactors
2 soluble (CoA, NAD) and 3 enzyme-bound (thiamine, lipoic acid, FAD)
6. Name the most important function of the Krebs cycle and list three other functions.
** oxidize Acetyl CoA to CO2 **
synthesis of some amino acids
oxidation of some amino acids
oxidation of odd-chain fatty acids
heme (porphyrin) synthesis
ketone body oxidation
7. Identify three energy-rich products produced by the Krebs cycle and discuss their role in bioenergetics
3 NADH, 1 FADH2, 1 GTP
NADH à through complexes in mitochondrial matrix create proton gradient to generate ~7.5 ATP in
F0 F1 ATPase
FADH2 à through complexes… ~1.5 ATP
GTP ~ ATP
Page 22 of 29
CS&F B13 page 2
8. Recognize the names of the enzymes and intermediates in the Kreb’s cycle
acetyl coa (citrate synthase)>
citrate (aconitase)>
cis -aconitase (aconitase)>
isocitrate (isocitrate dehydrogenase)>
oxalosuccinate (isocitrate dehydrogenase)>
a-ketoglutarate (a -ketoglutarate dehydrogenase)>
succinyl coa (succinyl coa synthetase)>
succinate (succinate dehydrogenase)>
fumarate (fumarase)>
l-malate (malate dehydrogenase)>
oxaloacetate
9. Discuss how Krebs cycle intermediates are generated.
Regenerated in cycle.
Anaplerotic = new intermediates
Pyruvate + co2 + atp à oxaloacetate + adp + pi (pyruvate carboxylase, biotin)
Glutamate + NAD à a-ketoglutarate + NADH + H+ + NH4+ (glutamate dehydrogenase)
10. name the intracellular location of the krebs cycle and oxidative metabolism
krebs occurs totally in the mitochondrial matrix (enveloped by inner membrane)
electron transport chain complexes within inner membrane of mitochondria
Page 23 of 29
9.16.03
Bioenergetics and Oxidative Metabolism II
Mitochondrial Electron Transport System (ETS) and ATP Synthesis
CS&F B14
Dr. Mosteller
1. Name the intracellular location of the electron transport system
(ETS) and identify the tissues where it is found.
ETS is in the inner mitochondrial matrix. It is found wherever there
is mitochondria, i.e. everywhere. Esp liver, heart, …
2. Describe the roles of the cytochromes, non-heme iron, CoQ (ubiquinone), flavin-linked
dehydrogenases and molecular oxygen (O2 ) in the electron transport system (ETS).
Shuttles:
overall seq of e- carriers:
(Transport reducing
equivalents (2H+,2e -)
flavin -linked complexes à CoQ à cytochromes à O2
from cytoplasmic
cytochromes
NADH,FADH2)
Any of a class of iron-containing proteins important in cell respiration as catalysts of
Into mito out o’ mito
oxidation-reduction reactions.
Malate-Aspartate
contain heme (protoporphyrin ring + iron). Each iron can carry one electron
Cytochrome c inner surface of inner mito membrane. Transports e-s between cytochrome
α-Glycerol
bc1 (Complex III, cytochrome reductase) and cytochrome aa3 (Complex IV, cytochrome
Phosphate
oxidase)
reduces FAD in
dehydrogenase, which’ll
This is most conserved in nature to preserve function
reduce CoQ in mito
non-heme iron
non-heme iron proteins (containing iron-sulfur centers) found in the flavin-linked
complexes (Complex I,II,III) and cytochrome bc1. each iron can carry 1 electron (Fe3+ + 1e - ßà Fe2+)
CoQ
Lipid-soluble cofactor transports electrons from flavin-linked complexes to cytochrome bc1. CoQ is mobile
in the lipid bilayer or inner membrane and carries 2 H+s and 2 e-s (reduced form)
Hydrophobic polyisoprenoid chain (50 carbons)
Flavin -linked dehydrogenases
Flavin -linked complexes (containing FMN or FAD) accept e-s from NADH, succinate, glycerol-P or fatty
acids. Complexes include:
FMN-linked:
NADH dehydrogenase (Complex I)
FAD-linked:
Succinate dehydrogenase (Complex II, Krebs cycle)
Glycerol-P dehydrogenase (shuttle system)
Fatty acyl CoA dehydrogenase (β-oxidation of fatty acids)
Molecular oxygen
Final acceptor of e-s. 4 e-s and 4 H+s combine with O2 to form 2 molecules of water
4e- + 4H+ + O2 à 2 H2 O
3. Describe the relationship of the proton-electrochemical gradient to electron transport and ATP synthesis.
Protons pumped out of mito matrix (into inner-membrane space) during e- transport at Complex I, Complex III,
and Complex IV. Proton gradient generates an electrochemical gradient that stores the energy derived from
electron transport, which is used to drive ATP synthesis.
4. Discuss how ATP is generated and released into the cytoplasm.
ATP synthase (F0 F1 ATPase) in inner mitochondrial membrane.
Protons go through pores in F0 F1 ATPase (oligomycin inhibits), which catalyses ATP synthesis: ADP + Pi à
ATP + H
Enzyme brings substrates (ADP,Pi) close to each other, effectively ?[ADP]?[Pi]
ATP released by ATP synthase into mito matrix, then exchanged for cytoplasmic ADP by ATP-ADP
translocator. (inhibited by atractyloside)
Page 24 of 29
9.23.03
Lipid Metabolism I: Synthesis of Fatty Acids and Triacylglycerols
CS&F B15
Dr. Mosteller
1. Describe the general structure of fatty acids and discuss where and when they are made.
Fatty acids are straight-chain aliphatic carboxylic acids with(out) double bonds. Common FAs have an even
number of Carbon atoms and most double bonds are in the cis configuration (vs trans).
Long, non-branching.
FAs synthesized in well-fed state from excess carbohydrate and excess protein (amino acids)
FA synthesis under dietary and hormonal control (insulin and glucagon)
FA synthesis occurs in cytoplasm of cells. Liver major site. All tissues (but RBCs) can synthesize
FAs stored in adipose tissue as triacylglycerols (triglycerides)
2. Recognize the names of common fatty acids and the two essential fatty acids.
• Palmitic acid C16:0
CH3-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-COOH saturated
• PalmitoleicAcid C16:1(9) CH3-CH2-CH2-CH2-CH2-CH2-CH2=CH2-CH2-CH2-CH2-CH2-CH2-CH2-CH2-COOH monounsaturated
• Stearic acid C18:0
• Oleic acid C18:1
(note, from stearic, to oleic, there’s a double bond. Oleic. From palmitic to palmitoleic, there’s a double bond. Oleic.)
Essential (linoleic, linolenic)
• Linoleic acid C18:2 (9,12)
CH3-CH2- CH2-CH2-CH2-CH = CH -CH2-CH = CH - CH2-CH2-CH2-CH2-CH2-CH2-CH2-COOH
CH 3(CH 2)4C=CCH 2C=C(CH 2)7COOH
• Linolenic acid C18:3 (9,12,15) CH CH C=CCH C=CCH C=C(CH ) COOH
• Arachidonic acid C20:4 (eicosa-tetraenoic acid, ETE) (made from linoleic acid)
3
2
2
2
2 7
Precursor of the eicosanoids (prostaglandins, thromboxanes, etc)
3. Name the precursors of the fatty acid synthesis.
Precursors: Acetyl CoA + Oxaloacetate + HCO3- + ATP
Acetyl CoA made in mito matrix by PDH, exported to cytoplasm via citrate/malate transporter
First step: synthesis of Cytoplasmic Malonyl CoA
• [Acetyl CoA + Oxaloacetate]mitoch àCitrateà [Acetyl CoA + Oxaloacetate]cyto (malate/citrate shuttle)
• Acetyl CoA + HCO 3- + ATP à [Acetyl CoA Carboxylase]enzyme à Malonyl CoA + H 2O + ADP + Pi
Rate limiting step
Enzyme regulated by diet and hormones, citrate, (de)phosphorylation
MalonylCoA also inhibits FA oxidation
Second step: Synthesis of Palmitic Acid
• Acetyl CoA + Malonyl CoA à[FA Synthestase (FAS)]enzyme à Butyryl ACP (C4) (acyl carrier protein) + CO2 + 2 CoA
• Butyryl ACP + Malonyl CoA à Hexanoyl (C6) ACP + CO 2 + CoA (MalonylCoA sub to +2Cs /time)
Third step: Elongation and Unsaturation
4. Name the two enzyme complexes responsible for fatty acid synthesis and identify their intracellular location.
Acetyl CoA Carboxylase, Fatty Acid Synthetase in cytoplasm
5. Discuss the regulation of fatty acid synthesis.
Acetyl CoA Carboxylase regulated by diet and hormones, citrate, (de)phosphorylation
MalonylCoA also inhibits FA oxidation
6. Describe how fatty acids are made longer and how double bonds are generated.
Elongate: add to palmitic acid: + 2C’s from MalonylCoA (in cytoplasm) or Acetyl CoA
(mitochondria)
Double bonds: desaturase enzyme complexes. cis Double bonds are created by the ?4, ?5, ?6
or ? 9 (stearoyl CoA) desaturase (SCD) systems located in the endoplasmic reticulum (ER)
7. Describe the general structure of triacylglycerols (triglycerides) & discuss where & when they are
made.
3-carbon backbone (from glycerol, G3P or DHAP) see fig.
O
||
H2 C-O-C-R
|
|
O
|
||
HC-O-C-R’
|
|
O
|
||
HC-O-C-R”
triacylglycerol
Page 25 of 29
Synthesized in intestine (from dietary fat) and liver (excess carbs,AAs). Cytoplasmic pathway. All tissues (but
RBCs) can. TAG synthesis active in well-fed state.
Page 26 of 29
9.24.03
Lipid Metabolism II: Fatty Acid Oxidation and Ketone Body Synthesis
CS&F B16
Dr. Mosteller
1. Discuss when and how fats are mobilized from adipose tissue.
In fasting state, low blood glucose à low insulin, high glucagon, stimulates mobiliz’n of TAGs from adipose
TAGs broken down stepwise. Form diacylglycerol, monoacylglycerol, glycerol and free fatty acids
First rate-limiting step catalyzed by hormone-sensitive lipase. Regulated hormonally (insulin, glucagon, adrenaline)
Glycerol released. Transported to liver for gluconeogenesis
2. Describe how free (unesterified) fatty acids are transported in the blood.
FFAs transported in the blood bound to albumin.
3. Identify tissues where fatty acid oxidation occurs.
FFAs taken up by liver, muscle and other tissues for oxidation.
4. Explain how fatty acids are transported into mitochondria(l matrix).
Carnitine-palmitoyl acyltransferase (CPT) I & II and translocase system.
Inhibited by Malonyl CoA.
See figure à
5. Describe β-oxidation and discuss how energy is generated from this
pathway.
Fatty acyl CoA oxidized in mitochondrial matrix
Palmitoyl CoA + 7 CoA + 7 FAD à 8 Acetyl CoA +
+ 7 NAD + 7 H2O
7 FADH2 + 7 NADH
Electron transport system Krebs cycle
ATP synthesis from NADH/FADH2 thrown into Electron Transport System, creating H+ gradient,
stimulating ATP synthetase. ~108 gross ATP equivalents from C16 Palmitic Acid
6. Name the three ketone bodies and discuss when are where they are made and utilized.
Three:Acetoacetate, β-hydroxybutyrate, acetone
Where made: In fasting state, ketone bodies synthesized in liver mito from Acetyl CoA derived from βoxidation of FAs
Acetoacetate from AcetylCoA: AcetylCoA à AcetoacetylCoA à HMG-CoA à Acetoacetate
Other 2 from acetoacetate:
Acetoacetate + NADH + H+ à β-hydroxybutyrate + NAD+
Acetoacetate à Acetone + CO2 (non-enzymatic rxn)
Where used: Ketone bodies (acetotate, β-hydroxybutyrate) taken up by peripheral tissues (mainly muscle)
converted to AcetylCoA which is oxidized in Kreb’s cycle (try to get some energy)
Acetone lost in urine, breath, or sweat.
Prolonged fasting, brain takes up ketone bodies.
Use of ketone bodies inhibits use of other materials, such as muscle, during fasting.
7. Describe events that occur during starvation or in untreated diabetes when excess ketone bodies are formed.
During starvation and untreated diabetes, excess fat mobilization can lead to excess ketone body formation
(more than can be oxidized by the body). The liver will continue to make ketone bodies from fatty acids even
though the body is unable to oxidize them at the rate at which they are produced. High levels of ketone bodies in the
blood (ketosis) can result in excess excretion in the kidneys leading to an imbalance (acidosis) Left untreated the
combined affect (ketoacidosis) can have serious deleterious effects on the patient.
Page 27 of 29
Things Mosteller said
to *star* in lecture:
HMG-CoA reductase,
serum lipoproteins/
transport cholesterol
9.25.03
Lipid Metabolism III: Cholesterol Metabolism and Lipoproteins
CS&F B17
Dr. Mosteller
1. Describe the properties of cholesterol and discuss why cholesterol is important.
• 27 carbon atoms
• planar, hydrophobic ring structure. (packs tightly)
• hydrophilic (polar) hydroxyl group (C3)
• amphipathic (polar & nonpolar)
• cholesterol esters (storage of cholesterol w/in cell when not in use)
completely hydrophobic. Find in ER and lipoproteins
• “cholesterol” = cholesterol + cholesterol esters
why impt
cholesterol is an essential component of membranes and lipoproteins and is the precursor of bile acids and
steroid hormones. Precursor for Vitamin D3 , impt in calcium and phosphorous metabolism.
too little cholesterol à myopathy
2. Describe the sources of cholesterol (dietary and de novo synthesis) and discuss the role of bile acids in
cholesterol metabolism.
sources
Cholesterol comes from food, absorbed in intestine with other lipids. Doesn’t absorb at 100% efficiency
De novo synthesis from acetyl CoA
bile acid
• stored in gall bladder. Released w/meals.
• Conjugated contain glycine/taurine = bile acid + AA
• Emulsify dietary lipids (strong amphipathic detergents)
• Enterohepatic cirulation: 98% recycled from intestine to liver in a day
• Fecal sterols generated by intestinal bacteria from cholesterol & bile acids (only sig way lose cholesterol)
3. Discuss when and where de novo cholesterol synthesis occurs in the body.
Cholesterol is made in the ER in the liver, intestine (both busiest) and other tissues in well, fed state. (28 steps!)
Non-hepatic tissues make much of their own cholesterol, can’t use the liver’s
4. Identify the precursors of cholesterol synthesis.
Acetyl CoA, HMG-CoA reductase. Some NADPH + H+’s
5. Name the enzyme that catalyzes the rate-limiting step in cholesterol synthesis.
HMG-CoA reductase.
3-Hydroxy 3-Methyl Glutaryl CoA
Rate-limiting step: HMG-CoA + 2NADPH + 2H+ à Mevalonic Acid + CoA + 2NADP+
Target of statins (inhibit cholesterol synthesis)
6. Discuss the regulation of cholesterol synthesis.
1. Diet and hormones (insulin, glucagon)
2. Phosphorylation/dephosphorylation of HMG-CoA reductase
3. Synthesis and degradation of HMG-CoA reductase (in liver)
4. Uptake of HDL and IDL and cholesterol-rich chylomicron remnants (in liver)
5. Uptake of LDL by LDL receptors located in clathrin-coated pits on cell surface (non-hepatic cells)
6. Transcriptional repression of LDL receptor and HMG-CoA reductase by SREBP-1 (Sterol Regulatory
Element Binding Protein).
After translation, SREBP-1 is cleaved by SCAP producing a 68kDa transcription factor. Goes to
nucleus, activates transcription of genes with SRE (sterol regulatory element) DNA sequence
Page 28 of 29
CS&F B17 page2
7. Describe how cholesterol is transported between tissues.
cholesterol transported via serum lipoproteins (large particules in blood made of many molecules, lipids +
proteins)
transportation mostly from liver/intestines to other tissues.
E.g.s of lipoproteins and what they tranport
Chylomicrons – dietary lipids from intestine
VLDL – lipids made in liver
LDL – cholesterol made in liver
HDL – return cholesterol to liver
8. Recognize the names of intermediates in cholesterol synthesis: mevalonic acid, isopentenyl pyrophosphate
(PP), geranyl PP, farnesyl PP, squalene, lanosterol. (è rate limiting step)
Acetyl CoA à à HMG-CoA è Mevalonic Acid à Isopentenyl PP (5) à Geranyl PP à Farnesyl PP à
Squalene à Lanosterol à Cholesterol
Page 29 of 29
Embryology Objectives
I
Describe how fertilized egg develops from fertilization to just prior to gastrula stage embryo
Repeated cell division à 32 cell = blastocyst
Inner cell mass + trophectoderm
Implantation
Bilaminar Embryonic Disk (epiblast + hypoblast) + Amniotic cavity
Chorionic Cavity
Embryonic disc + amnion + yolk sac ---connecting stalk/chorionic cavity---trophectoderm
Describe from where the amnion, amniotic cavity, chorion, and chorionic cavity arise
Amniotic Cavity - from fluid accumulating between cells of inner cell mass
Amnion - roof of amniotic cavity from inner cell mass
Chorionic cavity - forms between layers of trophectoderm
Chorion - from the trophectoderm
Understand spatial organization of embryonic disc with respect to the amniontic cavity, yolk sac, connecting stalk
(umbilical cord), chorionic cavity, and placenta
(see lecture I figures D/E on page 4)
Developmental terms
Zygote: sperm + egg à single nucleus è zygote
Blastocyst: hollow sphere of zygote at 32 cell stage, fluid begin to accumulate
Inner Cell Mass: 2-3 cell thick region of blast cyst (I.e. all except 1-cell thick wall)
Yolk sac: blastocyst cavity = primary yolk sac. Sac pinched off by chorionic cavity = yolk sac
Implantation: penetration of uterine endometrial epithelium and invasion of endometrial stroma by
growth of the trophectoderm
Amnion: roof of amniotic cavity (derived from inner cell mass)
Chorion: membrane surrounding amnion
II
Understand the significance of the process of gastrulation.
Gastrulation: process of formation of the three definitive germ layers.
Understand that neural tissue is induced to form from the ectoderm under the influence of the notochord, (OK) and how
the neural tube forms.
Condensation of mesoderm along longitudinal midline of embryo forms notochord
Notochord direct events cause ectoderm become neural plate
(18th day) groove forms midline neural plate to delimit neural folds
Neural folds bend and fuse à neural tube
Starts in middle, progresses caudally and cranially
Know which portions of the nervous system are derived from the neural tube, and which are derived from neural crest;
understand the diverse range of neural and nonneural tissues to which neural crest cells contribute.
Neural tube: à spinal cord: dorsal and ventral regions à sensory and motor neurons
Neural crest: entire PNS (cell bodies located outside neural tube) -- spinal ganglia (drg), portions
of cranial nerves, sensory neurons, Schwann cells, portions of meninges
Craniofacial derivatives (nonneural, nonmuscle elts of head)
Pigment cells
Chromaffin cells in medullary layer of adrenal gland
Smooth muscle in cardiovascular outflow region
Know the organization of the embryo after inoculation, in particular the spatial relationship between the epidermis and
neural tube (ectoderm), the three types of mesoderm (paraxial, intermediate, and lateral), and the endoderm.
(see lecture II figures A-D page 3)
Developmental Terms from this lecture which are of particular importance:
Gastrulation: 2 layer (epiblast/1ary ectoderm + hypoblast/1ary endoderm) à 3 layer
(endo/meso/ecto-derm)
Ectoderm: remainder of epiblast after migration. Outer layer of three. Gives rise to epidermis &
neural tissue
Mesoderm - migrates in 2nd wave. Middle layer.
Notochord separates mesoderm. Paraxial closest to neural tube. Lateral most distal.
Paraxial: segmentally cubical into somites = mesenchyme (sclerotome) à bone &
cartilage of ribs and vertebral column + dermomyotome (dermotome à dermis + myotome à muscle)
HO: Embryology Page 1 of 4
Intermediate: urogenital system (kidney, gonads, ducts, etc.)
Lateral: somatic (dorsal) à body wall + splanchnic (ventral) à visceral organs
Endoderm - migrates in 1st wave. Innermost layer. Gives rise to gut, liver, pancreas
Oropharyngeal membrane - from the prochordal plate. pos’n of future mouth
Cloacal membrane - pos’n of future anus.
OM + CM = direct transition ectoderm à endoderm & corresponding transition vascular/nerve
supply. (à gag reflex at OM)
Notochord: induce events of neural plate formation. Later surrounded by paraxial mesoderm to
form nucleus pulposus of vertebral discs
Neural tube: folding of neural folds induced by neural plate induced by notochord. à spinal cord
Neural crest: transient population of cells originate from dorsal-most portion of neural folds at all
axial levels. Contribute to variety of structures (see list above) à PNS
Paraxial Mesoderm: (See Mesoderm above)
Intermediate Mesoderm: (See Mesoderm above)
Lateral Mesoderm: (See Mesoderm above)
Somites: cuboids of paraxial mesoderm. è mesenchyme/sclerotome à spinal cord,
dermotome à dermis, myotome à muscle
III
Understand the reorganization achieved by ventral folding, including the partitioning of the embryonic coelom into the
pericardial and peritoneal cavities by the septum transversum (ST).
Ventral folding - dorsal neural tube grows faster than notochord.
Tissue of future heart & diaphragm (septum transversum) brought into thoracic region,
ST separates pericardial/peritoneal cavities.
Caudal end: connecting stalk (à umbilical cord) brought into abdominal region
Partition embryonic endoderm into 3 domains: foregut (OM to just below ST)
Midgut (endoderm open to yolk sac cavity)
Hindgut (connecting stalk back to cloacal membrane (anus))
Intraembryonic coelom (I.e. coelom) à body cavities (pleural, pericardial, peritoneal)
(See Lecture III page 2 figure A2-D2)
Understand how lateral folding forms the lateral and ventral body wall, and creates the coelomic cavity.
Lateral folding - dorsal ectoderm and amnion overgrow slowly dividing mesoderm & endoderm.
Ventral & lateral portions of body wall form when opposing sides of embryo come closer
and fuse. Somatic lateral mesoderm à ventral/lateral body wall + epidermis ; paraxial
mesoderm + overlying epidermis à dorsal body wall, endoderm brought together in a
circle à
epithelial lining of gut. Coelom brought together à body cavities.
(See Lecture III page 2 figure A3-D3)
Understand how growth of the lungs creates the pleural cavity from the pericardial cavity, and how the location of the
phrenic nerve is changed as a consequence.
Lung buds from ventral side of foregut. Surrounded by splanchnic mesoderm. Pleural cavity created from pericardial
cavity by growth of lungs & tissue from lateral body wall (pleuropericardial folds). Lungs enlarge, folds extend
inwards à separation of pleural & pericardial cavities. PF tissue à fibrous pericardium
Phrenic nerve - somatic nerve, initially runs along lateral embryonic body wall towards ST. pos’n = loc‘n of
pleuropericardial folds. As PF come in to separate cavities, phrenic nerve carried along. .: phrenic nerve lies in
visceral pos’n on fibrous pericardium instead of within body wall like other somatic nerves . Motor innervation of the
diaphragm via phrenic nerves.
Developmental terms from this lecture which are of particular importance:
Septum transversum: à portion of future diaphragm, called ST at stage of ventral folding
Coelom: (aka intraembryonic coelom) - fluid filled space which separates lateral mesoderm into somatic (dorsal)
mesoderm and splanchnic (ventral) mesoderm. à body cavities.
Somatic mesoderm (dorsal): mesoderm associated with body wall
Splanchnic mesoderm (ventral): layer of lateral plate mesoderm adjacent to endoderm
08.19.03
Chromosomes & Molecular Diagnostics
HO G1
Dr. Reichardt
1. Describe the molecular components of human chromosomes and the arrangement of these
components into nucleosomes and higher order structures.
The self-replicating genetic structures of cells containing the cellular DNA that bears in its proteins.
Chromosomes are made up of genes and are supercoiled around nucleosomes
Nucleosome:
Repeating units of organisation of chromatin fibres in chromosomes, consisting of around 200
base pairs and two molecules each of the histones H2A, H2B, H3 and H4. most of the DNA
(around 140 base pairs) is believed to be wound around a core formed by the histones, the
remainder joins adjacent nucleosomes, thus forming a structure reminiscent of a string of beads.
2. Define the normal human karyotypes for males and females.
The complete set of chromosomes of a cell or organism. Used especially for the display prepared
from photographs of mitotic chromosomes arranged in homologous pairs.
Females: 22 pairs(1-22), XX
Males: 22 pairs (1-22), XY
3. Define the following terms:
· Diploid:
A cell with a full set of genetic material, consisting of chromosomes in homologous pairs and
thus having two copies of each autosomal genetic locus. A diploid cell has one chromosome from
each parental set.
· Haploid
Describes a nucleus, cell or organism possessing a single set of unpaired chromosomes. Gametes
are haploid.
· Autosome
A chromosome not involved in sex determination
· Centromere
The region in eukaryote chromosomes where daughter chromatids are joined together.
· Telomere
The end of a chromosome
· p and q arms of chromosomes
p (petite): short arm
q (next after p): long arm
· Metacentric, submetacentric, and acrocentric chromosomes
metacentric
Descriptive of a chromosome that has its centromere (kinetochore) at or near the middle of the
chromosome
submetacentric chromosome
A chromosome with the centromere so placed that it divides the chromosome into two arms of
strikingly unequal length.
acrocentric
A chromosome or chromatids with a non-centrally-located centromere, producing chromosome
arms with unequal lengths.
4. Differentiate the following human cytogenetic abnormalities and list at least one example of
a human disease for each:
· Numerical abnormalities
abnormality where their exists a different number of chromosomes than normal
e.g. trisomy 21 = 3 of chromosome 21
· Structural abnormalities
abnormality within the structure of the chromosome. e.g. Tay-Sachs, deletion of 1 of 300 AAs
HO Genetics: Page 1 of 8
HO G1 page 2
5. Describe situations in which genetic testing is medically useful and explain some of the
limitations and ethical questions associated with testing.
Medically useful when results can be used to prevent severe damage/consequences
i.e. amniocentisis to exclude PKU, galactosemia, or hypothyroidism. First two can be controlled
by diet and the last by hormones to prevent damage to the fetus
limitations
can not detect everything
ethical questions
what should we do with the genetic information. Will employers/banks/health insurance
agencies/etc. find out this information and discriminate against the one with the genetic
predisposition?
08.27.03
Genomic Medicine
HO G2
Dr. Hacia
1. Define the following terms
• Deleterious mutation
o Change in normal base pair sequence
o Usually alters protein function
• Neutral allele
o DNA sequence changed but doesn’t affect fitness of organism
• Silent mutation
o A change in nucleotide sequence that doesn’t change amino acid sequence (type of
polymorphism)
• Missense, nonsense, frameshift, splice-site mutation
o Missense: change to codon of another amino acid
§ Can be harmful mutation or neutral polymorphism
o Nonsense: change from AA codon to stop codon à shortened protein
§ UAG = stop codon Ä
o Frameshift: insert or delete basepairs.
§ Produce stop codon downstream and usually shortens protein
o Splice-site mutation
§ Change alters RNA sequence.
§ Intron not properly identified. Splice around. Can cause frameshift or lose key
components for protein
• Dominant negative mutation
o Nonfunctional allele interferes with the function of the wild type alele
o E.g. CFTR mutation produces nonfunctional CFTR protein. Also inhibits activity of wildtype CFTR protein
• Other types
o Mutation in regulatory region. Promoters, enhancers
o Large deletion or insertion (10s-100s kilobases) anywhere in gene or regulatory region
o Chromosomal translocation or inversion. Break and fuse with different chromosome
• DNA methylation
o Conversion of cytosine to 5-methylcytosine
o DNA methylation of CG-rich sites in promotors can silence gene expression
• Single nucleotide polymorphism (SNP)
o Differences between individuals. Occur ~1 in 1000 sites.
• DNA microarray technology
o Gene expression analysis.
o Simultaneously measure mRNA levels for over 12000 genes.
2. Explain the genetic basis for cystic fibrosis as well as inherited predispositions to breast and ovarian
cancers.
CF
∆F508: deletion of 3 nucleotides CTT and loss of phenylalanine causes mutation in nucleotidebinding domain 1 of protein. Autosomal recessive
2 severe alleles can cause pancreatic insufficiency
•The CFM1 gene affects complications with meconium
ileus
Breast Cancer
BRCA1 (20-45% inherited brease ca. 5-10% new cases
are inherited mutations. .: 90-95% of new cases of breast
cancer are due to a mutation acquired in life) and BRCA2 (10-35% inherited breast ca)
HO G2 page 2
Female carriers have a 50-90% lifetime risk of breast
cancer (depending on family history) and an increased risk of
ovarian cancer
Even without gene, 1/10 chance in U.S.A.
Lots of sequence mutations
3. Identify situations where it is best to
(1) screen for specific mutations
a. mutations already identified in family history.
Checking for presence
(2) screen for all possible mutations in a disease-associated gene.
a. Pt presents with s/s of genetically linked disease. No
family history. Unknown mutation.
4. Recognize the challenges of interpreting genetic test results and their medical relevance.
Just cuz know there’s a mutation, doesn’t mean we’ll know what the effect’ll be. Some more
mild/severe than others. Genetic results doesn’t tell us phenotypic
5. Summarize basic applications for DNA mic roarray-based gene expression analysis.
Testing a genome of carcinogenic tissue to see what mRNAs are being produced at a higher level
might indicate the components of a tumor
Discriminate between genes that are
85% identical on the nucleotide level
Determine up to 1,000-fold differences
in transcript levels
Identify transcripts whose levels
correlate with a given phenotype
9.2.03
Human Inheritance I
HO G3
Dr. Maxson
1. Define the following genetic terms and concepts
• Gene
o Unit factor on chromosome
• Locus
o Genetic location on a chromosome. Loci = 2 different genes/locations on chromosome
• Allele
o Different versions of a gene (same locus)
• Genotype
o The genetic constitution of an organism; factors (alleles) responsible for a trait(PP,Pp,PP)
• Phenotype
o The physical appearance of a trait
• Homozygous (homozygote)
o PP (dominant alleles) or pp (recessive alleles)
• Heterozygous (heterozygote)
o Pp (two different alleles that can be distinguished from one another
• Proband
o Family member first brought to attention of clinician
• Penetrance
o % of individuals with appropriate genotype express traits
• Expressivity
o Variable expressivity = variable phenotypic effects; variable severity
• Consanguineous
o Carrier Parents. At increased risk for having children with rare autosomal recessive
diseases
• Random (disassortative) mating
o Mating random
• Founder effect
• Polygenic inheritance
o Disease involves mutiple genes
• Genetic anticipation
o Severity of the disease increases with successive generations
o E.g. simple sequence repeats (e.g. huntington, myotonic dystrophy)
• Genomic imprinting
• Mosaicism
o a condition in which patches of tissues of unlinked genetic constitution are mingled in an
organism
• Chimerism
o Organism contains cells derived from two different strains
• Mitochondrial inheritance
o Only from mom through mito dna
o • Mitochondria are all maternally inherited. All offspring of affected females will be
affected.
o • Males do not transmit (unlike situation with X-linked dominant--all daughters of
affected male are affected).
o • Phenotype may depend on proportion of abnormal mitochondria (heteroplasmy versus
homoplasmy) in a particular tissue.
o E.g. Leber’s hereditary optic neuropathy
o • Aminoglycoside induced deafness
HO G3 page 2
2. Use a punnett square to solve genetics problems.
P
P
P
P
P
PP
PP
p
Pp
Pp
P
Pp
Pp
p
Pp
Pp
P
P
P
p
p
Pp
Pp
P PP
Pp
p
Pp
Pp
p
Pp
pp
3. Recognize pedigree symbols and be able to draw a pedigree.
4. State and define the four modes of Mendelian inheritance
autosomal dominant
disease shows if PP or Pp
autosomal recessive
disease shows if pp
X-linked dominant
Locus in ? on x-chromosome.
• Rare.
• Trait can be transmitted by both males and females, but still no male to male
transmission.
• All sons of affected males are unaffected, all daughters of affected males are affected.
Example X-linked hypophosphatemic rickets. (Ability of kidney tubule to
reabsorb filtered phosphate is impaired).
X-linked recessive
Disease fully expressed in males; may be expressed to some extent in females due to
X-inactivation.
Hallmark of X-linked disease is no male to male transmission.
Hemophilia
e.g. Cuchenne muscular dystrophy
9.3.03
Human Inheritance II
HO G4
Dr. Maxson
1. Define the following genetic terms
• polymorphism
o the occurrence of two or more genetically determined alternative genotypes (or
phenotypes) polymorphisms may affect gene activity, or may be ‘silent’.
• Mutation
o A change in DNA nucleotides
• centiMorgan (cM)
o genetic distance between 2 markers, based on recombination rate btn the markers
o 1 cM = 1% chance of recombination
• Lod score
o Statistical measure of evidence for linkage; performed as a computer analysis
o (logarithm of the odds) = log (likelihood ratio)
o if lod = 3, there is a 1/1000 chance that genes are not linked
2. Write and be able to apply the Hardy-Weinberg equation
p 2 + 2pq + q 2 = 1
2pq = proportion of heterozygotes
p 2 = proportion of homozygotes for the common allele, usu A
q2 = “
“
“
less common allele, usu a
p+q=1
3. Describe in general terms how genetic linkage analysis is done.
To identify disease gene, study segregation of disease with polymorphic markers on each
chromosome.
Ideally start with large familiy
Id marker that consegregates with disease
Use statistical approaches to evaluate the quality of linkage data
Genome – informed candidate gene approach
Msx2 Boston craniosynostosis maps to 5q terminus
9.9.03
Models of Human Disease
HO G5
Dr. Maxson
1. Explain the importance of animal models in medical research.
Types of models
A. Models of genetic regulatory systems.
• Genetically tractable animals such as Drosophila or zebrafish.
• Set stage for work in mammalian experimental models —notably mouse—and ultimately in
humans.
B. Models of disease processes.
• A human genetic defect is recreated in experimental animal, usually a mouse, resulting in
phenotype that resembles a disease.
1. Naturally occurring. (mutant)
• One example is the “Splotch” mutant mouse:
Discovered over 50 years ago,
Splotch is caused by mutation in a gene encoding a transcription factor, Pax3.
Splotch is a model for Waardenburg syndrome, (ex of heterogeneity)
Characterized by sensorineural deafness, craniofacial and pigmentary defects.
2. Directed.
• Investigators can recreate human genetic defects in experimental animals —usually mice.
• There are now literally hundreds of examples of mice that have been engineered so that they have
a genetic defect.
2. Describe in general terms the approaches used to create genetically altered mice to model human disease.
1. alter copies of a gene. produce targeting vector
markers inserted
2. vector introduced into embryonic stem cells (pleuripotent & indifferentiated) isolated from
mouse embryo
3. ES cells inserted into young embryos (typically embryos that would acquire a totally black coat
from lack of agouti gene
4. embryos grow to term in surrogate mothers
chimera - brown shading intermixed with black indicates ES cells survived and proliferated in
an animal. chimera because contain cells derived from two different strains of mice.
5. chimeric males are mated to black (non-agouti) females
6. examine genes of brown mice reveal which inherited target mutation
7. males/females carrying mutation mated to each other to produce mice whose cells carry the
chosen mutation in both copies of the target gene and .: lack a functional gene. directly analyze
DNA to confirm.
8. examined for physical/behavioral abnormalities
Human Organism Objectives: Gross Anatomy
GA1
08/21/03
Intro to Gross Anatomy and Dissection
Exam material from:
•
Lab guide
•
Lecture
•
Lachman’s
•
Study guide
Describe the structure and innervation of the thoracic body wall.
Thoracic body wall = bony framework (thoracic cavity) + muscular layers + lining by endothoracic fascia
and pleura
Thoracic cavity = thoracic vertebrae + sternum + ribs + diaphragm
Cells, Tissues & Organs Objectives: Microanatomy
08-18-03
Cells, Tissues, and Organs
CS&F MA1
Dr. Schechter
1. Cells and their products form the fundamental units of organization of tissues & organs.
A. Be able to define the terms cell, tissue and organ.
cell
small membrane-bounded compartment filled with a concentrated aqueous solutions of chemicals.
tissue
cells that are organized in distinctive patterns to perform specific functions
organs
tissues organized in associations with other tissues to perform certain functions
B. Be able to list and characterize the 4 basic types of tissues.
epithelial
tightly packed cells. cover free surfaces of body or line spaces and fx as selective barriers
connective
not close together. relatively abundant amts extracellular materials btn cells. fx in support (physical, metabolic, vascular,
immunological)
contractile/muscle
cells w/abundant amts actin and myosin filaments. fx in force generation and movement
neural
tightly packed. specz'd to conduct electrical responses and integrate info for coordination and control of organ fxs.
2. Be able to define what is meant by a cellular domain, and what is meant by polarity.
domain
Used to describe a part of a molecule or structure that shares common physico chemical features, for
example hydrophobic, polar, globular, helical domains or properties for example DNA binding domain, ATP binding
domain.
polarity
In epithelial cells, the polarity meant is between apical and baso lateral regions, in moving cells, having a
distinct front and rear. Some cells seem to show multiple axes of polarity (which will hinder forward movement).
3. Be able to define organelle and distinguish organelles from inclusions.
organelle
components of cells that are essential for survival and function
e.g. mitochondria, ER, nucleus
inclusion
substances within a cell which are not necessarily present at all times
e.g. glycogen, lipid droplets
C. Explain the importance of membranes in the evolution of cells.
Membranes allow for the isolation of functions and allow a cell to specialize and regulate cellular fxs
4. Complete self-study lesson and membrane handout prior to session on 8/20/03
CS&F: MicroanatomyPage 1 of 36
08-18-03
Introduction to Cells
Microanatomy Lab
CS&F MA1
1. Cellular dimensions:
In today's lesson state the criteria you will use to assess the sizes of cells in a given field and the
component parts of cells.
Red blood cells are ~7-8µs in diameter and .: are frequently a useful guide to help determine size of cells
in a specific field.
2. Fluid-mosaic membranes as the basis of cellular compartmentation:
Be able to explain the fluid mosaic model of cytomembranes, and specifically what is meant by fluidity and what is meant
by mosaic.
Fluid mosaic model
A model used to conceptualise cell membranes, in it, the membranes are described as a structually
and functionally asymmetric lipid bilayer studded with embedded proteins that aid in crossmembrane transport.
Fluidity
lipids, most phospholipids “sea of lipids”
A term to characterize the motility of components of the cell membrane, specifically nonrigid and
mobile, often moving laterally within the plane of the membrane and occasionally flipping to the
opposite side.
mosaic
proteins
A composite of the components of a membrane
3. Intracisternal space within cells:
Make a simple diagram of a cell and its organelles, and indicate which sites are cytosolic and which
are within an intracisternal space.
The cell gains biological advantages by creating distinct compartments to perform unique functions. For example, it uses one
compartment, the intracisternal compartment, to isolate and stabilize hydrolytic
enzymes that would otherwise digest the cell itself.
CS&F MA1 Introduction to Cells
4. Nuclei are heterogeneous organelles:
For each slide in today's lesson demonstrate to your lab partner examples of variations in nuclei
5. Cellular domains and polarity:
Using cells from today's lesson be able to explain what is meant by polarity and domains within individual cells.
domain
<molecular biology> Used to describe a part of a molecule or structure that shares common physico
chemical features, for example hydrophobic, polar, globular, helical domains or properties for
example DNA binding domain, ATP binding domain.
polarity
<cell biology> In epithelial cells, the polarity meant is between apical and baso lateral regions, in moving
cells, having a distinct front and rear. Some cells seem to show multiple axes of polarity (which will
hinder forward movement).
08-20-03
Membranes
CS&F MA2
MDLs
Self Study Objectives:
1. Be able to describe the common general structure of cellular membranes, a bilayer of lipids associated with various
integral and peripheral proteins.
Cellular membrane
is a lipid and protein structure which functions to segregate and localize cellular activities. Planar stx pacts as interface btn
compartments and provides a surface or matrix upon which proteins or other molecules (such as hydrophobic molecules) are positioned for
proper function in activities such as cell-cell interactions, cell-extracellular matrix interactions, enzymatic activities, signaling (sensing, signal
transduction, etc.), transport, stabilization of cytoskeleton, etc.
The structure enveloping a cell, enclosing the cytoplasm and forming a selective permeability barrier.
It consists of lipids, proteins and some carbohydrates, the lipids thought to form a bilayer in which integral proteins are embedded to
varying degrees.
Integral Protein
Require detergent solubilization techniques to separate the proteins from the membrane. Many integral proteins contain one or
more sequences of hydrophobic amino acids that intercalate into
the lipid bilayer
Peripheral Protein
More loosely associated with the bilayer and can be isolated from the membrane by relatively mild biochemical treatment.
A water-soluble protein that is loosely bound (by hydrogen bonds orelectrostatic forces) to a membrane.
2. Explain how membrane lipids, proteins, and carbohydrates are asymmetrically distributed. This heterogeneity subserves
different functions, such as: recognition; adhesion; communication; and directed transport.
The lipids are not covalently linked to one another in a rigid matrix but form a
fluid bilayer.
Membrane proteins are often anchored to the cytoskeleton and/or extracellular
matrix (ECM) in a manner that establishes functional domains in the membrane.
Membrane carbohydrates net negative charge on the cell surface serves to maintain water
at the cell's surface and facilitates fluid uptake.
3. Define Fluidity as it relates to membranes and explain how the degree of fluidity can be highly variable.
Generally accepted model for membranes in cells. In its original form, the model held that proteins floated in a sea of
phospholipids arranged as a bilayer with a central hydrophobic domain. Although it is now recognised that some proteins are
restrained by interactions with cytoskeletal elements and that the phospholipid annulus around a protein may contain only
specific types of lipid, the model is still considered broadly correct.
1. One measure of fluidity is related to the membrane lipids. However, many factors govern the interactions of membrane
lipids, e.g., extent of double bonds (unsaturated fatty acids) and length of fatty acid tails.
2. The ultimate fluidity of biological membranes will also be affected by protein
content and surface properties, such as charge and surface hydration
properties.
CS&F MA2 Membranes
4. Explain how the Fluid Mosaic model stresses both the heterogeneous distribution of components and the fluid nature of the
bilayer.
A model used to conceptualise cell membranes, in it, the membranes are described as a structually and functionally asymmetric
lipid bilayer studded with embedded proteins that aid in cross-membrane transport.
5. Describe how membranes form selective barriers that segregate functional
compartments of the cell. Also explain how membranes form a three-dimensional
matrix upon which cellular activity may occur.
Membrane trafficking (solid arrows) distributes new membrane from ER to other
membranes. Membrane organelles not in "continuity" with ER, such as mitochondria and peroxisomes, receive membrane
lipids via cytoplasmic carriers called phospholipid
exchange proteins or chaperone proteins
Membranes help establish compartments and domains. The function of
compartments is to optimize cellular activities and to separate incompatible activities.
Also, these compartments serve to direct in a vectorial manner the synthesis and
processing of molecules. In many cases there are specific molecular signals to target
materials to specific cellular compartments.
The cell plasma membrane and the closed membrane-bounded organelles
separate the cytosol from the extracellular space or "intracisternal space." The cytosol is
a major cellular compartment organized by cytoskeletal elements and contains both
"soluble" proteins and proteins bound to the cytoplasmic surfaces of membranes.
Lab
1. Fluid-mosaic membranes as the basis of cellular compartmentation:
Be able to explain the fluid mosaic model of cytomembranes, and specifically what is meant by fluidity and what is meant by
mosaic.
Fluid mosaic model
A model used to conceptualise cell membranes, in it, the membranesare described as a structually
and functionally asymmetric lipidbilayer studded with embedded proteins that aid in crossmembrane transport.
Fluidity
lipids, most phospholipids “sea of lipids”
A term to characterize the motility of components of the cell membrane, specifically nonrigid and
mobile, often moving laterally within the plane of the membrane and occasionally flipping to the
opposite side.
mosaic
proteins
A composite of the components of a membrane
2. Membrane domains:
Be able to explain what is meant by a membrane domain and the mechanisms cells may use to maintain or stabilize these
domains.
domain
<molecular biology> Used to describe a part of a molecule or structure that shares common physicochemical
features, for example hydrophobic, polar, globular, helical domains or properties for example DNA binding domain, ATP
binding domain.
CS&F MA2 Membranes
3. Glycoproteins, glycolipids and integral membrane proteins
Be able to summarize the various functions served by these components of membranes.
Glycoproteins, glycolipids
Sugars are present on proteins, lipids, and proteoglycans. Collectively, these
molecules are called glycoconjugates. The expression of carbohydrate is almost
exclusively on the non-cytosolic side of membranes. Due to the sialic acid-rich
oligosaccharides of the surface membrane glycoconjugates, the cell surface
membrane maintains a net negative charge. Since water is a polar molecule with a
positive charge, the net negative charge on the cell surface serves to maintain water
at the cell's surface and facilitates fluid uptake. Glycoconjugates are involved in cellcell interactions, including cell-cell recognition, and cell-matrix interactions.
Integral Membrane Protein
A protein that is firmly anchored in a membrane (unlike a peripheral membrane protein). most is known about the integral
proteins of the plasma membrane, where important examp les include hormone receptors, ion channels and transport proteins.
An integral protein need not cross the entire membrane, those that do are referred to as transmembrane proteins.
08-21-03
Nucleus: Structure & Function
CS&F MA3
Dr. Ying
1. Make a simplified schema of the basic structural components of the nucleus (nuclear
envelope, pores, nucleolus, chromatin, nuclear matrix).
2. Explain the structure and function of the nuclear envelope and nuclear matrix.
Nuclear envelope
A 2 membrane system that encloses a perinuclear cisterna (space) and consists of inner nuclear membrane and outer nuclear
membranes that are continuous at the NPCs (nuclear pore complexes)
Nuclear matrix
Act as as a scaffold that aids in organizing the nucleoplasm. Stxl’ly the nuclear matrix includes nuclear lamina and a residual
ribonucleoprotein network, and fibrillar elements. Fxlly, the nuclear matrix is associated with dna replication sites, rRNA and
mRNA transcription and processing, steroid receptor-binding sties, carcinogen-binding sites, heat shock proteins, DNA virus
and viral proteins.
3. Explain the importance of the structure of nuclear pore complexes and how this plays a
role in nucleocytoplasmic exchange.
Transport of macromolecules between the nucleus and cytoplasm takes
place via large proteinaceous channels through the nuclear envelope, the nuclear pore complexes. The nuclear pore complexes
act as selective barriers that allow some molecules (e.g. messenger RNA's and ribosomal subunits) to move from nucleoplasm
to cytoplasm and other molecules (e.g. enzymes, DNA-associated proteins, ribosomal proteins) to move from the cytoplasm
into the nucleus.
(1) Structure (Figure 3-6)- The NPCs, formed by fusion of the inner and outer nuclear membranes,
average 80-100 nm in diameter and number from dozens to thousands per nucleus. It is a 3 ring-like proteinaceous structure.
First, the cytoplasmic ring consists of 8 subunits with fibrils protruding from the cytoplasmic side of the NPC, which are RNAbinding proteins and may mediate import of materials into the nucleus. Second, the middle ring is a set of 8 transmembrane
proteins projecting into the lumen and perinuclear space of the nuclear envelope. These octagonal-shaped glycoproteins,
nucleoporins, are responsible for transporting proteins into and out of the nucleus via receptor-mediated transport. In the
middle ring, there is a luminal subunit, glycoprotein gp210, that anchors the NPC in the nuclear envelope. In addition, there are
gated channels for passive diffusion. Third, the nucleoplasmic ring consists of thick fibrils to form a nuclear basket protruding
into the nucleoplasm.
CS&F MA3 Nucleus Structure & Function
(2) Function - Numerous molecules are transported in and out of the nucleus (bidirectional) by
passive diffusion (small molecules) and active transport (energy-required, temperature-dependent, receptor-mediated &
selective barrier). For the nuclear import, nuclear localization segments (NLS) of proteins serve as signals for transport which
bind to importin a in the cytoplasm. The NLS-protein-importin a finds importin b which is docked with the NPCs, then, nuclear
translocation takes place and importin a is recycled. For the nuclear export, there is the nuclear export signal (NES) coupled to
exportins. In addition, transporin also facilitates the transport mechanism in the cytoplasm. M9 is a specific peptide sequence
that acts as both NES and NLS signals for proteins, i.e., ribosomal proteins.
4. Describe the organization of chromatin and its role in synthesis, processing and storage of DNA and RNA.
DNA and associated proteins in the nucleus are present as heterochromatin
and euchromatin. The degree of packing or condensation of DNA is governed by the DNA-associated proteins
(histones and non-histone proteins).
5. Have a general concept of the cell cycle and its control as well as apoptosis.
M-> G1->S -> G2
q The cell cycle is influenced by a variety of factors including hormones and growth factors and is regulated at the
molecular level by proteins called cyclins and cyclin-activated protein kinases.
q The cell cycle is regulated by (a) cyclin, (b) cdc2 which is a type of Cdk (cyclin-dependent protein kinase), (c) M-phase
promoting factor, (d) Rb gene regulatory proteins, (e) p53, and (g) growth factors.
q Apoptosis, occurs in fetal and adult cells such as mature blood cells and cells attacked by pathogens, is regulated by
numerous regulatory genes which lead to activation of a family of enzymes, caspases.
8-21-03
Nucleus and Nuclear Compartment
CS&F MA3
Lab
1. For each slide in today's lab. demonstrate examples of variations in nuclei
2. Describe the mechanisms used by nuclear pores to regulate the exchange of materials between the cytoplasm and the
nucleus.
(1) Structure (Figure 3-6)- The NPCs, formed by fusion of the inner and outer nuclear membranes,
average 80-100 nm in diameter and number from dozens to thousands per nucleus. It is a 3 ring-like proteinaceous structure.
First, the cytoplasmic ring consists of 8 subunits with fibrils protruding from the cytoplasmic side of the NPC, which are Ranbinding proteins and may mediate import of materials into the nucleus. Second, the middle ring is a set of 8 transmembrane
proteins projecting into the lumen and perinuclear space of the nuclear envelope. These octagonal-shaped glycoproteins,
nucleoporins, are responsible for transporting proteins into and out of the nucleus via receptor-mediated transport. In the
middle ring, there is a luminal subunit, glycoprotein gp210, that anchors the NPC in the nuclear envelope. In addition, there are
gated channels for passive diffusion. Third, the nucleoplasm ic ring consists of thick fibrils to form a nuclear basket
protruding into the nucleoplasm.
(2) Function - Numerous molecules are transported in and out of the nucleus (bidirectional) by
passive diffusion (small molecules) and active transport (energy-required, temperature-dependent, receptor-mediated &
selective barrier). For the nuclear import, nuclear localization segments (NLS) of proteins serve as signals for transport
which bind to importin a in the cytoplasm. The NLS-protein-importin a finds importin b which is docked with the NPCs, then,
nuclear translocation takes place and importin a is recycled. For the nuclear export, there is the nuclear export signal (NES)
coupled to
CS&F MA3 Lab: Nucleus Structure & Function
exportins. In addition, transporin also facilitates the transport mechanism in the cytoplasm. M9 is a specific peptide sequence
that acts as both NES and NLS signals for proteins, i.e., ribosomal proteins.
3. Normal cells in the blood demonstrate considerable variations in nuclei. Therefore this lab session is a good opportunity to
study nuclear diversity and to learn to identify the cells in normal blood and their functions.
Leukocytes = WBCs
Granulocytes
Display cytoplasmic granules.
Neutrophils
Eosinophils
Basophils
AKA polymorphonuclear leukocytes. From irregular multilobulated nuclei.
Fx: phagocytosis, inflammation
Agranular
Lack prominent visible granules
Lymphocytes
Monocytes
Never Let Monkeys Eat Bananas
Neutro Lympho Mono Eosino Baso
60-70 20 4-10 1-3 0-1 % of total WBC
Blood = Plasma + RBC + WBC = 55% + 45% + 1%
Granulocytes
Neutrophil
Numerous small bluish purple granules
Phagocyte, engulf & kill bacteria
Forms of lysosymes
Eosinophil (Acidophil)
Kill parasites. Phagocytose bacteria. Allergic response phagocytose
Cytoplasmic granules. Dark red -> crimson
Basophil
Scarce
Deep violet cytoplasmic granules often cover nucleus
Cytoplasmic granules -> histamine.
allergic rxn
Agranular
Monocyte
Central, ovoid, u-shaped, or indented nucleus
Cytoplasm: small vacuoles & fine granules
Store lysosymes
Source of phagocytic cells
Lymphocytes
Round nucleus. Thin-to-moderate cytoplasmic rim, no svisible granules so don’t know if it’s B or T yet. Humoral & all
mediated immunity
Leukocytes:
Eosinophils > (slightly) neutrophils > basophils (smallest)
Monocytes (largest of all cells seen in normal blood films) (usually) > lymphocytes
neutrophils: phagocytes
eosinophils: kill parasites, modulate allergic, inflammatory responses by phagocytosis of antigen-antibody complexes
basophil: granules contain histamine, vasodilator, and heparin (anticoagulant)
monocytes: cytoplasm contains fine particulate granules, which store a substantial supply of lysosomes in degradation of
engulfed cells, etc. mature into macrophages
lymphocytes: B cells & T cells (from bone marrow)
B cells -> antibody-secreting plasma cells
T cells -> subtypes help other cells in immune rxns or cytotoxic lymphocytes kill
targeted cells
8-25-03
Protein Synthesis & Secretion
CS&F MA4
Lab
Be able to evaluate the cytological features of a cell in order to diagnose whether or not the cell is likely to be releasing large
quantities of a protein secretory product.
Lots of RER, golgi, secretory granules (likely), much euchromatin (maybe)
Be able to trace the pathways of protein synthesis
Dna à RNA à mRNA à small ribosomal subunit à large ribosomal subunit à (if not cytosolic) SRP à bind to porphorin
à enter RER à continue polypeptide addition à vesicle à cis -Golgi à trans-Golgi
-- à condensing vacuole à secretory granule
-- à clathrin coated vesicle (lysosome) (M6P receptor)
Lecture
1. Describe the general mechanism of protein synthesis, e.g., lamins vs. CFTR, and where it takes place in
epithelial cells.
See above
2. Diagram and list the basic functions of the endoplasmic reticulum (RER and SER) and Golgi apparatus in epithelial cells.
RER
Synthesize proteins for lysosomes, cell membrane, or secretion
SER
Lipid synthesis, detox, Ca sequester, steroid hormone synthesis
Golgi
Package, transport, modification of proteins (glycosylates
3. In your diagram indicate where post-translational modifications of CFTR protein may occur in epithelial cells.
SER, RER, Golgi
4. Distinguish between regulated and constitutive secretory pathways
Regulated
“on demand” and a stimulus is required; a good example is the secretion of hormones.
Constitutive
Default pathway. Always on. Can be up/down regulated
9-02-03
CS&F MA5
Cytoskeleton
For each filament system:
Describe the organization of a filament, including the subunits and how they are combined.
Describe the location of filaments within the cell. Describe the functions in different cellular domains.
Describe how the cytoskeleton contributes to cytokinesis.
Microfilaments (3-6nm diameter)
Organization
Actin (globular protein) in twisted double strand
Polarity. Actin adds to (+) end
Location
In microvilli and around periphery of cell
Functions
Stability. Cytoplasm viscosity. Motility (actin contractile). Anchoring of cytoplasmic proteins.
Structural rigidity
Cytokinesis
Pinches/cleaves daughter cells after mitosis
Clinical Relevance
Muscular Dystrophy
Intermediate Filaments (10nm diameter)
Organization
Intermediate filament proteins arrange in coiled coil homodimer. Dimerism, then tetramize à protofilaments. Octets of the
tetramers (I.e. 8 protofilaments = intermediate filament)
Location
Basket around nucleus. Desmosomes. Cytoplasm.
Functions
Space filler. Structural strength
Cytokinesis
No fx
Microtubules (20-25 nm diameter)
Organization
Alpha/beta tubule subunits combine in circular fashion.
13 dimers in a circle. Longitudinal rows of dimers = protofilaments.
Cilia/Flagella: dimers of protofilaments, arranged 9 dimers in a circle + a dimer in middle
Centriole: triplets of protofilaments, arranged 9 in a circle, zero in middle
(-) / (+) ends
(-) disassembly. .: dyneid microtubule based motor protein moves from + to - end
(+) extend to periphery. Kinesis mbmp moves from - to + end. Subunits added to + end.
Location
Microtubule Organizing Center near nucleus, radiating outwards. Extend to plasma membrane
Cilia/Flagella, Centrioles
Functions
Structural rigidity. "Road map" for intracellular movement. Cytokinesis. Motility of cilia and flagella
Cytokinesis
Formation of the mitotic spindle, and separation of sister chromatids.
9-02-03
CS&F MA6
Endocytosis, Lysosomes, and Trafficking
1. Distinguish between trafficking of cellular materials by vesicular or membranous components and
trafficking that does not involve vesicles.
Trafficking that does not involve vesicles: e.g. active transport of a material across a concentration gradient using active
transport molecules within the membrane. Some passively diffuse down [] gradient, others by facilitated transport or
molecular pores in membrane.
Pinocytosis, vesicles rich in caveolin
Receptor mediated endocytosis: receptors bind ligand à clathrin coated pits
Any movement across cellular membranes can be classified as trafficking. Therefore active transport as well as facilitated
diffusion can be thought of as non-vesicular modes of transport. Any type of movement involving the use of membranebound vesicles however falls under the category of vesicular transport.
2. Define endocytosis and distinguish (structurally and functionally) between the various types of
endocytosis.
Endocytosis: the process of uptake and recovery from the environment. Involves a vesicular component.
Pinocytosis: “cell drinking”.
Fluid-phase pinocytosis. Plasma membrane invaginates. Caveolae project into cell. Non-selective. But more favor basic
molecules.
Receptor-mediated endocytosis. Specific ligands bind and initiate uptake.
Phagocytosis = “cell eating”
Large insoluble particles ingested via phagosomes. Often specific, require particle bind to cell surface. Extend
pseudopods. E.g. Antibodies. Bind to infectious organism, tail interacts with receptors trigger phagocytosis.
Endocytosis is a category of vesicular transport that uses vesicles to import material into the cell. Endocytosis can further be
broken down into 1) pinocytosis and 2) phagocytosis.
There are two types of pinocytosis. The first, fluid-phase pinocytosis, uses small invaginations of the cell membrane called
caveolae, to non-specifically “drink” parts of the surrounding extra-cellular fluid. The protein caveolin has been linked to this
process and although little is known about the mechanism, it is hypothesized that more basic molecules will be taken up more
readily. Fluid-phase pinocytosis is found in most tissue types in the body, but can be found frequently in smooth muscle
cells and the endothelial cells of the blood vessels.
The second type of pinocytosis is more specific. Receptor mediated endocytosis uses receptors on the surface of cell
membranes to capture ligands to be brought into the cell.
Phagocytosis is a process whereby larger insoluble molecules, microorganisms or cellular debris are brought into the cell.
This process can be mediated by cell surface receptors, such as Abs, but doesn’t have to be.
3. Be able to trace the path taken by LDL and its component parts during receptor-mediated endocytosis.
LDL binds to LDLRs à clathrin coated pits/coated vesicles à early endosome à interior acidified, ligand released from
receptor, sorted à fuse with early lysosome à lysosomal enzymes hydrolyze cholesteryl esters in LDL particles, freeing
cholesterol to move into cytoplasm to incorporate into new membranes.. LDLR recycled back to plasma membrane.
LDL is a lipoprotein that is brought into the cell via receptor mediated endocytosis.
1. LDL binds to the LDLR
2. Clathrin mediates the pinching of plasma-membrane high in [LDL-LDLR]
3. H+ -pumps decrease intravesicular pH causing both the dissociation of clathrin and the dissociation of LDL from its
receptor.
CS&F MA6 page 2
4. Be able to explain the differences between heterophagy, autophagy and residual bodies.
Heterophagy: digestion of substances imported into a cell from external environment
Autophagy: degradation of organelles in an otherwise healthy cell. Membranes of ER envelop organelle in
autophagosome, then lysosomal enzymes fuse autophagolysosome
Residual bodies: indigestible residues of lysosomal activity associated with normal wear and tear. Accumulate with advancing
age in the form of lipofuscin pigment.
These terms refer to the different lysosomal modes of digestion. Heterophagy is the lysosomal degradation of foreign
particles. Autophagy is the digestion of the cells own organelles that may have become compromised or exhausted. Finally,
residual bodies are those remnants of secondary lysosomes that never fully digest.
5. Be able to explain the basic sorting mechanisms that are used to target lysosomal enzymes properly to
lysosomes.
Mannose-6-PO4 groups added to N-linked oligosaccharides of lysosomal enzymes (prolly in cis -Golgi) Mannose-6-PO4
groups bound to Mannose-6-PO4R, transmembrane proteins in trans-Golgi. Mannose-6-PR complex packages lysosomal
enzymes into clathrin-coated transport vesicles. Later fuse with late endosome to become primary lysosome.
Lysosomal enzymes are given a distinct marker of Mannose-6-phosphate to their N-linked oligosaccharides in the cis -golgi.
These mannose groups bind to the Mannose-6-phosphate receptor in the membrane of the trans-golgi. This Mannosereceptor complex packages these lysosomal enzymes in clathrin coated vesicles. Later acidification of the intralysomal
compartment causes the separation of clathrin. These vesicles subsequently fuse with late endosomes and become primary
lysosomes.
Lysosomal storage diseases: Pompe’s disease, Gaucher’s disease, Tay-Sachs disease, Fabry’s disease
6. Be able to discuss the complexities of vesicle budding and targeting of vesicles.
Coatomer-coated proteins mediate non-selective trafficking btn golgi cisternae and golgi to plasma mem. COPII protein coat
on vesicles from ER to golgi, COPII facilitates pinching off of vesicles. Also + SNARE proteins, direct to golgi.
Cells have an enormous array of management systems for vesicle budding and targeting. Vesicles for non-specific, intra-golgi
transport in addition to transport from the golgi to plasma membrane involves a protein called coatomer. This method is
fundamentally different from clathrin-mediated transport. In addition to these two targeting proteins, COPII is responsible for
movement from ER to cis -golgi and SNARE proteins project into the cytomatrix and complex with Rab and tethering proteins.
9.4.03
Peroxisomes and Mitochondria
CS&F MA7 Lecture
Dr. Da-Yu Wu
1. Be able to discuss the structure and function of peroxisomes.
Structure:
single membrane organelle, spherical. Diameter 0.5µm.
Homogeneous fine granular matrix. Various oxidases and catalase.
Crystalline structure in non-human cells.
Located in all cell types, abundant in liver and kidney
Functions: metabolize long chain fatty acids to generate acetyl CoA and hydrogen peroxide
Regulate level of H2O2, destroy excess H2O2 by catalase. (detox)
Detox alcohol, formaldehyde, and purines. Kill some microorganisms
Other info: maternally inherited. Replicate by growth and fission. Proteins synthesized in cytosol. Defects lead to brain,
liver, kidney damage abnormal accumulation of fatty acids
40 types of enzymes in peroxisome.
2. Be able to identify and describe the structural features of mitochondria as seen by LM and EM.
LM: see mito double membrane and possibly distinguish cristae inside.
EM: see mito double membrane and differentiate cristae inside. Ribosomes. Granules in matrix.
3. Describe genetic inheritance and replication of mitochondria, and the biosynthetic pathway, transport and assembly of
mitochondrial proteins.
Maternal mito in oocyte sole source of mito in fertilized egg .: maternal inheritance. Each mito circular DNA encodes 13 mito
proteins, 2 rRNAs, 22 tRNAs. Other mito proteins encoded in nucleus.
Self-replication: mito replicate circular DNA and grow in size, then fission into daughter cells.
Mito proteins: encoded by nuclear genome. Synthesized in cytosol by free ribosomes. Transported by specific signaling
mechanisms: 1) mito signal peptide added in protein tln, 2) chaperone keeps prot unfolded, guides to mito mem receptor, 3)
prot complex binds to mem receptor/protein transporter, moves laterally contact side, 4) prot across membranes, signal peptide
cleaved off.
Goes into matrix first. If more signals, may go back to inner mem, intermem space, or outer mem. Fold.
4. Be able to define the function of mitochondria and their roles in disease and cell death.
Fxs:
• ATP production through chains of e- carriers/H+ pumps and ATP synthase complex
• Metabolism of carbs, fatty acids and amino acids
• Ca2+ storage
• Apoptosis. Trigger programmed cell death due to genetic defect or receptor/translocases medicated
signalling event.
Disease/cell death:
• Apoptosis in cancer cells
stimulated transport of TR3 and p53 into mito triggers cancer cell death. Clinical trial of gene therapy in lung cancer
• Leber’s hereditary optic neuropathy: three mitochondrial gene mutations on genes control cytochrome oxidases in
mito. Optic nerve. Hit puberty, blind.
9.4.03
Peroxisomes and Mitochondria
CS&F MA7 Lab
Dr. Da-Yu Wu
1. Be able to identify mitochondria in TEMs and variations in mitochondrial morphology and function.
ok
2. After being told that you are viewing an EM of a peroxisome, be able to discuss its function.
Functions: metabolize long chain fatty acids to generate acetyl CoA and hydrogen peroxide
Regulate level of H2O2, destroy excess H2O2 by catalase. (detox). oxidase
Detox alcohol, formaldehyde, and purines. Kill some microorganisms
Other info: maternally inherited. Replicate by growth and fission. Proteins synthesized in cytosol. Defects lead to brain,
liver, kidney damage abnormal accumulation of fatty acids
40 types of enzymes in peroxisome.
3. Be able to interpret images of cells in a variety of LMs and EMs so as to discuss all topics presented in the course to
date.
cells, tissues and organs
intro to cells
membranes
nucleus: stx & fx
nucleus and nuclear compartment
protein synthesis and secretion
cytoskeleton
endocytosis, lysosomes, and trafficking
peroxisomes and mitochondria
9.15.03
Epithelium
CS&F MA8
Dr. Wood
1. Draw and label a schematic to demonstrate cell polarity and different types of cell-cell adhesion.
2. Classify epithelium based on cell layers and cell morphology.
Simple: one layer. Stratified: multiple layers
Squamous: squashed, flat. Nucleus bulgy. Cuboidal: looks like cube.
Columnar: looks like column.
Simple squamous
thin! layer of squamous. Endothelial cells, line blood vessels, air sacs.
Simple cuboidal
one layer of cuboidal. Kidney tubules and glands.
Simple columnar
one layer of columnar. Lines digestive organs. (goblet cells, too)
Stratified squamous
top layer squamous. Skin (w/keratin), mouth, vagina, esophagus.
Stratified cuboidal
top layer cuboidal. Lines ducts of sweat glands.
Stratified columnar
top layer columnar. Lines epididymus/penile urethra, mammary
glands, larynx.
Pseudostratified columnar
appears like >1 layer. All cells contact basement mem, not all
reach surface. Aka respiratory epithelium. Cilia, goblet cells
Transitional epithelium stratified, rounded at surface. Cells stretch as bladder fills. Line
urinary system.
3. Explain replacement of epithelium.
Epithelium needs constant replacement, under constant stress. Replaced by continued mitotic division (liver cells, endothelial
cells) or mitosis of stem cells (à one differentiated cell, one stem cell so no stem cell loss) (intestines, epithelium, blood)
4. Explain different types of glandular secretion.
(M)Merocrine: vesicle fuses to (M)embrane. Contents secreted.
(A)Apocrine: vesicle surrounded by membrane and secreted. (lose Apical membrane)
(Holo)Holocrine: entire cell contents dumped out. w(Hole) cell lost.
Active transport: simple channels or transporters transport molecules in or out
9.15.03
Epithelium Lab
CS&F MA8 MA9 Lab
MDLs
1. List the general features that identify any epithelium.
• Closely apposed cells adhere by means of junctions
• Plasma membrane has domains
• Attached to a basement membrane
• avascular
•
•
•
Supported by and attached to connective tissue via specialized layer extracellular matrix material i.e. the basement membrane
Polarized
o
Basal surface abuts basement membrane
o
Apical surface acts as interface with luminal or surface environment
Arrangements of contiguous cells cover surfaces, line cavities or spaces, form ducts, form secretory portions of glands
2. List the special features that identify specific epithelia.
Number of cell layers between apical surface and basement membrane
Simple = only one cell thick
Stratified = two or more cells thick. Only basal cells contact basement membrane
Pseudostratified = appears stratified actually all cells contact basement membrane
Transitional = special category of stratified epithelium characteristic of urinary system
shape
squamo us = thin, flattened cells (look like fried egg)
cuboidal = cells have roughly same dimensions along all sides
columnar = tall, thin cells
3. Identify the basic types of intercellular junctions in epithelia. (see pictures on prev page)
Tight junction
• Aka zona occludens
• Form continuous belt surrounding cell
• Impermeable (relatively) barrier restricts mobility of transmembrane proteins and small molecules between apical
and basolateral domains
• Closest to lumen/apical surface
• Transmembrane proteins (claudins and occludins) bridge the two plasma membranes
• Other proteins, e.g. cadherins, facilitate tight junctions
• Cytoplasm clearer
Intermediate junction
• Aka adhering junctions
• Lie below tight junction
• Form continuous belt
• Membranes not in close apposition
• Cadherins (transmembrane linking proteins) span space between
• In cytoplasm, intermediate jxn anchored by proteins (a-actinin, vinculin, etc) attached to inner surface of plasma
membrane
• reinforced by actin microfilaments.
Desmosomes
• spot welds, reinforce, esp at sites of stress
• plamsa membranes do not come in contact in a desmosome
• space between occupied by transmembrane linking proteins
• reinforced within cell by desmoplakin attached to intermediate filaments
Gap Junctions
• communication connections (vs. support). Facilitate transfer of ions/small molecules. Connexons. Electrical
coupling
CS&F MA8 lab page 2
4. Identify the interface between epithelia and connective tissue in LMs and EMs.
• Connections with basement membrane = hemidesmosomes
• Inside cell, anchored by desmoplakins attached to intermediate filaments
• Within plasma membrane, hemidesmosome has integrins attach to laminin and collagen in basement membrane
• Basement membrane
Secreted by epithelium
o Laminin
o Integrins
o Type 4 collagen
o Fibronectin heparin sulfate proteoglycan
Fibroblasts contribute
o Type 3 collagen
o Fibronectin
9.15.03
Cell-Cell and Cell-Matrix Interactions
CS&F MA9
Dr. Schechter
1. Be able to give specific examples of how molecules in the cell membrane mediate cell-cell and cell-matrix interactions.
• Recognition
o Membrane glycolipids,glycoproteins
o Sialyl Lewis X (sialic acid-rich carbohydrate) binds to selectins. Cancer. Block off blood supply
o Major histocompatability Complex, I & II. Immune responses. I: all nucleated body cells. II. Specific cells
(b-cells, activated T-cells, APCs)
• Cell Adhesion
o For maintenance of body form and structure
o Homophilic interaction
§ Cell adhesion molecules (CAMs), ca2+ independent
§ Cadherins, Ca2+ dependent
o Heterophilic interactions
o Adhering-type intercellular junctions (desmosomes and intermediates)
• Indirect cell-cell interactions
o Hormones, cytokines, growth factors
o Endocrine (secrete to blood stream), paracrine (short distance, diffuse cells), autocrine(on self)
• Cell-Matrix interactions
o EC Matrix
§ Fibrous proteins in hydrated gel rich in glycosaminoglycans (GAGs)
§ Collagen (I-IV and elastin)
§ Proteoglycans/proteoglycan aggregates. Lots (-) charge, lots water. Rigid backbone (HA)
§ Adhesive glycoproteins
• Attach cells to fibrous and other (fibronectin and laminin) components
• Junctional Epidermolysis Bullosa (JEB) mutation in laminin gene, blisters
o Transmembrane receptors
§ E.g. integrins link proteins of ECM with cytoplasmic cytoskeletal elements
§ CLINICAL: Integrin αvβ3 expressed in angiogenesis. Raf-1 inhibits expression of prot. Halts
development, block tumors require angiogenesis (grow blood vessels)
2. Be able to give examples (and explain) how interactions between cells may be through direct cell-cell contact or
indirectly between cells separated by variable distances.
o Endocrine (secrete hormones to blood stream) kidney
o paracrine (short distance, diffuse cells)
o autocrine(on self)
3. Be able to explain the importance of interactions between cells, and between cells and extracellular matrix, that involve
specialized matrix molecules, membrane receptor molecules and intracellular cytoskeletal elements.
Homeostasis. Generate and maintain the orientation of cells in tissues and organs and influence cellular physiology and
gene expression.
9.17.03
Cellular Differentiation
CS&F MA10
Aaron Logan
1. Be able to explain that differentiation is a result of differential gene expression, not change in the genomic content of
the cell.
All cells have same genomic content. Difference between cells is what part of genome gets expressed. Stimulus to
differentiate signals from soluble proteins, cell-cell, cell-matrix interactions
2. Give examples to demonstrate that differentiation may or may not be associated with distinguishing morphological
characteristics.
Expression of a specific mRNA may precede the appearance of specific proteins and indicates the cells are determined, i.e.
confirmed to follow a certain path of differentiation. .: cells can be determined to follow a specific differentiation pathway
before they demonstrate overt structural characteristics of differentiation.
3. Be able to explain that differentiation may or may not be terminal.
Some differentiated cell types maintain ability to divide by mitosis (fibroblasts)
Some incapable of dividing further (plasma cells)
4. Give examples that demonstrate that differentiation involves inductive interactions mediated by cell-cell, cell-matrix
and cell-factor interactions.
Induction: stem cells poised to differentiate and cells in process of differentiating, induced down specific pathways of
gene expression by specific mediators:
a. cell-cell interaction
direct interaction btn prots in mems influence differentiation of one or both cells. E.g. during embryogenesis,
formation of the optical lens from surface ectoderm is induced by contact with underlying cells in the optic
vesicle.
b. Cell-matrix interactions
interaction mem receptors and ECM components. Transmit signals direct cell migration and/or differentiation.
E.g. fibronectin guides migration of neural crest cells to different part of embryo in embryogenesis
c. Soluble factors (i.e. hormones or growth factors)
likely majority of inductive signals in embryogenesis and tissue maintenance are soluble factors
5. Explain how normal cellular turnover and tissue regeneration often demonstrate characteristics resembling those seen
during embryonic differentiation.
Temporal specificity
Different effect depending on what point in time presented to cell
e.g. thalidomide & limb development in fetuses
Instructive Induction
Signal causes a cell to proceed down a particular path or differentiation
Permissive Induction
Cell already committed, requires additional signals to begin the process
Control of cellular differentiation critical to normal development and cell turnover. Derangements can lead to metaplasia
(reversible), dysplasia (partially reversible), neoplasia (irreversible)
Apoptosis, cells served their purpose. Die. Fetal development, web tissue between digits undergoes apoptosis
9.18.03
Connective Tissue (CTI): Loose Connective Tissue (LCT), Dense Connective Tissue (DCT) & Adipose Tissue
CS&F MA11
Dr. Ying
1. Define connective tissue and describe its origin, heterogeneity and spectral nature as well as other general features.
connective tissue:
compartments and components that provide the structural support of the body and bind
together its cells, organs, and tissues
origin: mesenchymal cells (middle layer of embryo, mesoderm)
heterogeneity: CT ranges from watery consistency to bony rigidity.
Include:
1. blood
4. cartilage and bone (skeletal tissue)
2. loose connective tissue
5. adipose tissue
(LCT, CT proper, areolar
6. hemopoietic
tissue)
7. lymphatic
3. dense connective tissue
(DCT)
2.
comprised of ECM materials (collagen,elastic,reticular-fibers) + cells (fibroblasts – secrete matrix, macrophages –
perform phagocytosis, plasma cells – secrete Abs, mast cells – produce histamine, adipocytes – store fat,
WBCs – migrate from blood in response to infections or other stimuli)
nerve supply, highly vascular (except cartilage, tendons, ligaments)
functions: support, defense, repair, storage, transport
Classify CT based on types, relative amounts, and arrangement of cells, fibers, and ground substance.
Loose connective tissue (LCT)
• One of most widely distributed CT
• Fewer fibers, more cells and non-fibrous E CM than DCT
• Viscous ground substance (GAGs, proteoglycans, collagen, elastin, glycoproteins chondronectin, fibronectin, laminin), type
IV, fibroblasts, macrophages, plasma cells, mast cells, adipocytes, wbcs
• No obvious organization
• Support, protection, transport, repairs
• Found around vessels, nerves, under epithelia, in spaces btn other tissues, other subQ layers of skin
• Repairs itself well (vascularity)
Dense regular connective tissue (rDCT)
• Regularly arranged type I, sparse ground substance, little space btn fibers
• Parallel patterns as rope-like, fiber bundles
• Fx: transmission of mechanical force
• E.g. tendons, most ligaments
• Repairs itself more slowly (less vascular)
• Cells not as numerous as LCT
Dense irregular connective tissue (irDCT)
• Extensive collagenous fibers, irregularly arranged, type I
• Sparse ground substance, few cells
• Fibroblasts predominant, others present
• Fx: protect fragile organs, where pulling forces exert various directions
• E.g. dermis, organ capsules, heart valves, perichondrium, periosteum
• Repair crappy (less vascular than lct)
Adipose tissue
• Small amt ground substance + reticular fibers
• Lobules and lobes with rich capillary networks
• Fx: reduce heat loss thru skin, energy reserve, support, protect
• White (unilocular) vs brown (multilocular)
• Found: subQ deep to skin, bone marrow, around heart & kidneys, yellow bone marrow
CS&F MA11 page 2
3. Explain the biochemical composition and the sites of synthesis of the extracellular matrix components.
comprised of ECM materials (collagen,elastic,reticular-fibers) + cells (fibroblasts – secrete matrix, macrophages – perform
phagocytosis, plasma cells – secrete Abs, mast cells – produce histamine, adipocytes – store fat, WBCs – migrate from blood in
response to infections or other stimuli) nerve supply, highly vascular (except cartilage, tendons, ligaments)
fibroblasts secrete fibril procollagen
Collagen type I,II,III,IV
Collagen and reticular secreted by fibroblasts = collagen + glycine (30%) + proline + hydroxyproline, lysine, hydroxylysine
elastic fibers = elastin surrounded by fibrillin
Marfan’s syndrome = mutation in fibrillin gene. Weak elastic fibers. Tall, long extremities & digits. Weak aorta.
Reticular fibers = type III collagen coated with glycoproteins
4. Label fibers and cells in a diagram of LCT and describe their functions.
1. collagen fiber
8
5
rope-like, tensile strength, stx
2. elastic fiber
form network. Strong & stretch
3. lymphocyte
immune response
4. monocyte
immune response, à macrophage
5. macrophage
engulf bacteria/debris
6. fibroblast
secrete ECM components
7. mast cell
histamine and heparin
8. undifferentiated mesenchymal cell
can differentiate into lots o’ stuff
9. plasma cell
antibody-producing, extensive RER
10. capillary
blood vessel. Supply stuff to ECM
11. adipose cell
insulate, heat, energy reserve
5. Compare the structure and function of LCT, DCT, and
adipose tissue by preparing schematics of each. (see adipose above)
tendons, ligaments. Transmit mech forces | protect fragile organs, forces various directions, support, protection, transport, repairs
dermis, organ capsules, heart valves
around vessels,nerves,under
epith,subQ
9.18.03
Connective Tissues I: Cartilage
CS&F MA11
Dr. Ying
1. List the characteristics (structurally and functionally) of the cells and extracellular matrices in cartilage.
chondrogenic cell
mesenchymal cells --> ch-enic --> chondroblasts
Chondroblast
young cartilage-forming cell (from mesenchymal). Synthesize constituents of cartilage ECM
Chondrocytes
mature cartilage cell important in maintenance of ECM. Surrounded completely by cartilage matrix
Lacuna
space/cavity in which chondrocytes reside
Isogenous groups
groups of 2-8 chondrocytes in one lacunae
Perichondrium fibrous layer w/dense collagen fibers, fibroblasts. Surround hyaline and elastic cartilage. Innermost layer
chondrogenic capability
Matrix
mostly Type II collagen fibrils, proteoglycan aggregates (GAGs), hyaluronic acid
Some keratan sulfate, heparan sulfate, glycoproteins
(elastic) + elastin
(fibrocartilage) mostly type I, equal chondroitin sulfate and dermatan sulfate
Hyaline cartilage most abundant
Elastic cartilage give flexibility
Fibrocartilage
fibroblasts & collagen fibers (DCT) present w/matrix and chondrocytes in rows
2. Explain the statement that cartilage is avascular.
There are no vessels that run through cartilage, hence it is avascular. There are, however, blood vessels that run alongside
the cartilage (in hyaline and elastic) that provide the nutrients from the blood through diffusion. In fibrocartilage, there are a
few blood vessels in DCT bundles. The lack of great vasculature results in an inability to repair well.
3. Explain the difference between interstitial and appositional growth.
Appositional growth
growth from the surface; an inner layer of perichondrium may contain immature chondrogenic cells
w/the potential to become chondroblasts and secrete new cartilage matrix on the SURFACE
Interstitial growth
growth from within; secretion of new cartilage matrix by the division of chondrocytes
4. Be able to discuss the limitations of cartilage repair.
See 2.
9.22.03
Connective Tissues II – Bone
CS&F MA12
Dr. Ying
1. Be able to describe the characteristic structures and functions of the cells and extracellular matrix in bone.
Osteoprogenitor cells aka osteogenic, spindle shaped, from mesenchymal to osteoblasts, in periosteum and endosteum
Cover all internal surfaces of bone. I.e. spongy, H.canal, marrow cavities.
Osteoblasts
synthesize and secrete osteoid, à osteocytes
Osteoid
uncalcified bone matrix
Osteocyte
mature bone cell in own lacunae
Osteoclasts
large, multinucleated cells from monocytes, degrade bone matrix
Canaliculi
space between osteocytes for processes. Pathway for chemical, electrical, stress-generated fluid
communication. Contain gap junctions. periosteocytic space and calcium reservoir.
Volkmann’s canals
channels transverse to bone axis. Communication btn H.systems, periosteum & marrow cavity
Osteon/Haversian system long cylinder composed of several lamellae surrounding a central canal.
Central/Haversian canal central canal of H.system. Neurovascular channels in specialized LCT
Cementing line
deposit of amorphous material(mineralized matrix + collagen fibers) around ea. H.system. sharp line
Volkman’s canal
neurovascular channels connect H.canals, periosteum, endosteum
Periosteum
thick layer specialized CT. cover external surface bone, except articular cartilage and sites lig/tendon
attachment. Inner layer = osteogenic, outer layer = irDCT. Provides blood and nerve supplies to bone,
anchorage to other CT, source osteogenic cells
Sharpey’s fibers
bundles of periosteal collagen fibers penetrate bone matrix to connect periosteum to bone
Endosteum
inner surface of long bone. Thin CT layer w/osteogenic cells. Fx: bone growth and repair.
Outer circumferential lamellae several lamellae extending around periphery of shaft immediately beneath periosteum. Form
outermost region of bone surfaces
Inner circumferential lamella
several inconstant lamellae extend around marrow cavity immediately beneath endosteum.
Inner/Outer CL
separate the collection of H systems from the endosteum (inner) and periosteum (outer)
Interstitial lamellae
residual lamellae btn H systems. Old osteons built over.
Extracellular Matrix
a calcified matrix
Consists of osteogenic cells, osteoblasts, osteocytes, osteoclasts.
Organic: Type I collagen (80-90% bone composition), GAGs, keratan sulfate, glycoproteins,
phosphoproteins, enzymes, hormones, growth factors, and tissue fluid
Glycoproteins
• Osteonectin
o High affinity for Ca and hydroxyapatite. Binds ECM to cell processes
• Osteocalcin
o Synthesized by osteoblasts indicate bone formation. Role in osteoclast recruitment and
bone formation
• Osteopontin
o Binds to matrix. Interacts with integrins on osteoclasts to encase or seal the zone of bone
destruction resorbed by osteoclasts
• Sialoprotein
o Binding sites for matrix and integrins of osteoblasts and osteoclasts
Inorganic: (65% dry weight of ECM)
Ca and Phosphorus form hydroxyapatite crystals.
Mineralization on freshly depostied osteoids in matrix vesicles. Early crystals follow type I fiber, into
gap regions of the collagen. Mineral salts added.
Clinical:
Osteomalacia
failure to mineralize matrix
Rickets (children)/Osteomalacia (adults) bone poorly calcified. Vitamin D deficiency.
Paget’s disease of bone more rapid osteoid production than mineralization
Osteoporosis
CS&F MA12 page 2
Osteogenesis imperfecta
Rheumatoid arthritis
Osteoarthritis
more bone resorption, less bone formation à low bone mineral density, bone fragility, net bone loss
abnormal maturation of collagen in bone
deterioration of joint stx and fx. Due to autoimmune condition
trauma-induced deterioration of joint stx and fx. Not autoimmune.
2. Be able to explain the statement that bone is a highly metabolically active tissue despite its structural rigidity.
Bone constantly breaking down and rebuilding. Osteoclasts, osteoblasts.
Newly synthesized bone is not-yet-calcified and .: not as strong. But it quickly gets mineral salts deposited to give bone its
characteristic rigidity and mechanical strength.
3.
Be able to explain what occurs during bone repair.
At endosteum, e.g. H canal in osteon and at surface in spongy bone, osteogenic cells becoming osteoblasts which secrete
their matrix and enclose themselves in lacunae, becoming osteocytes and building upon the lamellae of the osteon. During
bone repair this is how the bone grows. To break down the bone, the osteoblast secretes a signal to the osteoclast precursor cell (monocyte-like cells) to divide and move to bone surface. In howship’s lacunae, osteoclasts bind and begin to
convert H2O and CO2 into H2CO3 and H+. H2CO3 gets dumped into blood (cuz vasculature here) and the H+ gets pumped
out onto bone to be broken down by acidification and dissolving the mineral. Components of bone matrix enzymes digest
organic matrix.
Bone repairs faster than cartilage.
9.25.03
Contractile Tissue
CS&F MA13
Dr. Da-Yu Wu
Bolded Terms
I. Skeletal Muscle
1. Basic structure
myofibrils, muscle fasciles, muscle fibers
epimysium, perimysium, endomysium (encloses individual muscle fibers)
syncytium, myoblasts, myotubes, sarcolemma, myofibrils, sarcoplasmic reticulum, sarcoplasm, sarcomeres, myofilaments,
actin filaments, z line/z disk. Myosin filaments, M line.
2. Sarcomere
alpha-actinin, myomesin, I band, A band, H zone, titin, nebulin
3. Triad system
Transverse tubules (T tubules), terminal cisternae, “triad” or T systems.
4. skeletal muscle innervation and excitation-contraction soupling
motor neurons, bouton, neuromuscular junction, action potential, acetylcholine, troponin, tropomyosin, TnI, TnC, TnTa,
ATPase, ATP, ADP + Pi
5. skeletal muscle fiber types, connective tissue investiments and myotendinous junctions
myoglobin, red, white, and intermediate muscle fibers. Red = smaller, have rich vascular supply, high amount of myoglobin
for aerobic metabolism
myotendinous junction, tendon
II. Cardiac Muscle
1. Structure
intercalated disks, fascia adherens, desmosomes, gap junctions, diads,
2. Innervation and excitation-contraction
Purkinje fibers, SA node (pacemaker), AV node, interventricular bundle of His
III. Smooth Muscle
1. Structure
caveolae (sarcolemma indentations), desmin (intermediate filaments), electron-dense bodies (dense plaques) vimentin
2. Innervation and excitation-contraction:
gap junctions
calmodulin, myosin light chain kinase, myosin light chains, caldesmon
IV. Regeneration of Muscles
1. Skeletal Muscle
satellite cells
2. Cardiac Muscle
no
3. Smooth Muscle
retain mitotic ability
V. Muscle Spindle and Golgi Tendon Organ
1. Be able to explain the organization and cellular structure and function of skeletal muscle.
Muscle ß fascicle ß fiber ß myofibril ß sarcomere
Striated, banding.
[[Muscle]]epimysium ß[[ fascicle ]] perimysiumß fiber [[endomysium]]
multinuclear
sarcolemma = cell membrane
function: quick reaction. Voluntary movement.
CS&F MA13 page 2
2. Describe the structure of myofibril and roles of anchoring proteins in skeletal muscle.
Myofibril = up to 100 segments of sarcomeres
I line = actin, Z line ? A band = Hzone + M line + area both actin and myosin
Sarcomere = contractile myofilaments
Z-line anchors actin through binding of alpha-actinin
M-line harnesses myofilaments through myomesin
Titin elastic protein connects myosin to Z line
Nebulin courses along actin filaments
Intermediate filaments, i.e. desmin and vimentin, connect Z-lines of adjacent sarcomeres and btn sarcomere/sarcolemma
3. Be able to point out different structural features of skeletal, cardiac, and smooth muscles.
Skeletal:
Striated
Multinucleated, peripheral
Parallel fibers
1-40mm length
epimysium, perimysium, endomysium
sarcomere
triad
(4)motor neurons in spinal cord, Somatic NS
one neuron à multiple fibers
each fiber, one axon
acetylcholine
troponin, tropomyosin
“power stroke” contraction
red, white, intermediate fibers
red: small, rich vascular, long contractions, lots
myoglobin
white: large, less vascular, fatigue quickly, less
myoglobin
force at myotendinous junction: sarcomlemma
à basal lamina à reticular fiber & collagen
fiber à tendon à bone
muscle fibers don’t divide or regenerate
Cardiac:
Striated
Uni- or bi-nucleated, central boxcar,
nuclear halo
Cells branch and anastomose
Intercalated disks
Fascia adherens, desmosomes, gap
junctions
Diads
Endomysium
(4)ANS, Perkinje fibers
Force waves: SA node à AV node à
interventricular bundle of His à Perkinje
fibers
No regeneration post injury
Smooth:
Bundles/sheets spindle-shaped
contractile cells
Non-striated
Single, centrally located, elongated
nucleus
Caveolae, membrane indentations
::T-tubules
Higher actin filament ratio (vs
skeletal muscle)
Desmin intermediate filaments
Dense bodies / plaques :: Z-lines
(4)ANS. Norepinephrine,
angiotensin, vasopressin, oxytocin
Gap junctions
Calmodulin, caldesmon
Slower strat/finish reaction
Cells have mitotic ability
4. Describe the innervation and basic biochemical aspects of excitation-contraction coupling of the three types of muscles.
see references to (4) above
5. Describe the structural and functional features of connective tissue investments of muscles.
epimysium – irregular DCT surround entire muscle
perimysium – derived from epimysium, surround muscle fascicles (bundles muscle fibers)
endomysium – reticular (type III) fiber LCT, surround each individual muscle fiber
muscle transmits force through DCT
9.29.03
Neural Tissue I: Neurons and Their Myelins
CS&F MA14
Dr. Da-Yu Wu
I. Introduction: The major components and developmental origin of Nervous System
Central nervous system (CNS), peripheral nervous system (PNS), neuronal cells, glial cells, afferent nerves, efferent
1. Central nervous system
gray matter, perikarya/somata, white matter, axons, myelin sheath
2. Peripheral nervous system
somatic nervous system, autonomic nervous system, sympathetic, parasympathetic
3. Early development
neuroectoderm, neural plate, neural tube, neural crest cells
4. Neurogenesis and Neural tissue characteristics
ventricular zone, synapses
II. Neurons
dendritic arbor
1. Neuronal cell body (perikaryon, soma)
neurofilaments, neurotubules, Nissl
substance/body, lipofuscin, melanin
2. Dendrites and axons
Dendrites: microtubule associated protein (MAP2)
Axon: axon hillock, tau protein
3. Neuronal synapse and neuromuscular junctions
end bulbs/boutons, synapses
Synapse: neurotransmitters,active zone,synaptic
cleft
Signal Transduction: impulse/action potential,
axodendritic, axosomatic, axoaxonal synapses,
excitatory synapses and inhibitory synapses
Neuromuscular Junction: neuromuscular junction, junctional/subneural folds, acetylcholine receptors (AChR)
4. Axonal transport
anterograde transport via kinesin, retrograde transport via dynein, trophic factors
5. Neuronal types by morphology and function
morphology: pseudounipolar, bipolar, multipolar
function: motor neurons, sensory neurons, interneurons. MESA: motor efferent, sensory afferent
content of NT and effect: cholinergic (ACh, muscular), glutamatergic (glutamate, brain/eye), catecholaminergic (norepi,
epi, visceral organs), monoaminergic (dopamine and serotonin, movement), peptidergic (morphine, endorphins,
substance P, pain conduction)
III. Myelin
Myelinated, unmyelinated
1. Schwann cells
2. Node of Ranvier
Saltatory conduction
3. Oligodendroglia
from lab:
neural plate, neural tube, neural crest, ventricular layer, ependymal cells, ventricles, central canal, cerebrospinal fluid (CSF)
dendrites, cell body, Nissl substance, axon
1. Be able to explain how the ectodermal origin or neurons during development is reflected in characteristics of mature
neuronal cells.
Epithelia arise from same ectoderm. Neuronal characteristics similar to epithelial: tightly packed, continuous basal lamina, free
apical surface (face ventricles and central canal), joined together by specialized junctions, synapses, regulated secretion
CS&F MA14 page 2
2. Describe the basic cellular specializations of a neuronal cell, and correlate the structural specializations with the functions.
Dendrite, to receive input. Axon, to conduct information. Central, large, pale-staining nucleus cuz lots euchromatin, single
prominent nucleolus. Extensive network cytoskeleton, transport synaptic vesicles and organelles through axons and
dendrites. Lots RER (Nissl body), mito, free polyRibs cuz making lots of stuff. Boutons, at synapse to transmit signals
3. Be able to diagram the structural elements of neuronal synapses and explain the
functions they serve.
membranes of both presynaptic neuron (bouton) and post synaptic neuron
specialized, contain diffuse, electron-dense materials on either side
numerous mitochondria and aggregates of secretory vesicles at presynaptic area.
Vesicles contain neurotransmitters
active zone exists at which vesicles closely aggregate next to presynaptic membrane
between presynaptic membrane and post synaptic membrane is a 20-30nm wide
synaptic cleft. NTs diffuse through here to bind to the ligand-gated ion channels on
the post synaptic membrane to cause the channels to open and cause an influx
of Na+ and Ca2+ and outflux of K+. causes postsynaptic potential, trigger
depolarization
neuromuscular junction
synaptic cleft
neuron releases NT
NT binds to post -synaptic membrane
opens Na+ channels
depolarization
conducted through triads/T -tubules
induce sarcoplasmic reticula to release Ca2+
Ca2+ binds to troponin
tropomyosin shift
myosin head bind to actin
sarcomere contraction
4. Describe the cellular structures relevant to neuronal metabolism and axonal transport.
anteograde/retrograde transport require lots of stuff. I.e. neurofilaments, neurotubules, microfilaments to give transported
items a pathway, Nissl substance/body i.e. RER create the proteins to make the stuff to transport, i.e. NT. anterograde (from body
to axon) transport via kinesin, retrograde transport (from axon to body) via dynein, trophic factors, signals from whatever the
neuron innervates to keep the neuron alive.
5. Be able to identify and diagram the basic structures of Schwann
cells, and describe
their functions in the
PNS and CNS.
Schwann cells
myelinate nerves in
PNS. Unmyelinated
axons are still
enclosed within
invaginations of
Schwann cells, just
not to the
layered extent of the myelinated. Myelinated: wrap own plasma
membrane layer upon layer (up to 300
concentric layers) around axons in a tight spiral fashion.
Each Schwann cell myelinates one axon for about 1mm .: many
Schwann cells to one neuron. Nodes of Ranvier between Schwann cell layerings on neuron lead to saltatory conduction.
Outermost layer basal lamina separate neighboring Schwann cells.
CS&F MA14 page 3
Oligodendroglia: highly branched, small glial cells form myelin in CNS. Extend plasma membrane as short branches in
several directions. Wrap several axons at same time. Only myelinated axons in white matter, no neuronal cell bodies.
Yes cell bodies of oligodendroglial cells, epithelial line vasculature.
CLINICAL: multiple sclerosis. Demyelination in nervous system. Triggers problems/damage in nervous system. Origination,
possibly inflammation, .: autoimmune response. Other poss cause: radiation, chemical therapy. Immune system comes to clean
up. Nervous system remyelinates every few weeks, so may have periods where pt is better than at other times.
From lab: Be able to define CNS and PNS and list the major derivatives of neural crest.
CNS – nervous system consisting of the brain and spinal cord, derived from neural plate/neural tube
PNS – nervous system consisting of the various ganglia and peripheral nerves, derived from neural crest. Other things that arise
from neural crest: entire PNS (cell bodies located outside neural tube) -- spinal ganglia (drg), portions of cranial nerves, sensory
neurons, Schwann cells, portions of meninges
Craniofacial derivatives (nonneural, nonmuscle elts of head)
Pigment cells
Chromaffin cells in medullary layer of adrenal gland
Smooth muscle in cardiovascular outflow region
10.02.03
Neural Tissue II: Glial Cells, Meninges, Injury and Regeneration
CS&F MA15
Dr. Da-Yu Wu
1. Be able to identify and diagram the basic features of various glial cells, & explain their major functions in the PNS & CNS.
CNS: fibrous astrocytes, protoplasmic astrocytes, ependymal cells, microglia
Astrocytes
Many processes off cell body, highly attenuated branches. Endfeet. Regulate and maintain unique microenvironment of
nervous system. Endfeet on basal lamina of capillary form glia limitans. Assist with blood-brain-barrier by transporting
and exchanging metabolites and ions between capillaries and extraneuronal environment in CNS
Fibrous – white matter, Protoplasmic – gray matter.
Ependymal cells
Line ventricular surface of brain and central canal of spinal cord. Cuboidal, microvilli and cilia. No basal lamina.
Epitheloid. Cilia propels CSF so it flows
During development, arise from ventricular layer. Some differentiate into choroid plexus epithelial cells, covers choroid
plexus. CP filters rbcs,wbcs,proteins,and other stuff to make CSF. Choroid plexus epithelial cells form extensive tight
junctions between each other and make up a blood-CSF barrier.
Microglia
Small, ovoid cells fx like macrophages. Origin monocytes. Wont be asked to id. Injury, retract processes and proliferate.
Express MHC II antigens and interleukin-1, involved in immune response of body.
PNS: satellite cells
Satellite cells
Surround neuronal cell bodies in a ganglia. Provide a stable and specialized extraneuronal microenvironment. Smaller
than neurons. Cytoplasm (TEM) appears more electron-dense that neurons. Don’t usually see nucleolus.
2. Describe the structure and functional significance of the blood-brain-barrier.
neurons bathed in special microenviron so fxs not influenced by changes of blood content. BBB formed by tight
junctions of endothelial cells line capillaries in brain. Dynamic active and passive transport of nutrients, ions and water
through endothelial cells. Astrocytes terminate their endfeet on basal lamina of endothelial cells. Endfeet cover entire
capillary surface, glia limitans. Assist and influences BBB in transport/exchange metabolites and ions btn capillaries and
extraneuronal environ in CNS
3. Be able to explain the specialization of ependymal cells lining the inner surface of the CNS, the cellular features of choroid
plexus, and the origin of cerebrospinal fluid (CSF).
Ependymal cells arise from ventricular layer in embryo. Tight junctions create blood-brain-barrier.
Choroid plexus: protrudes out of the ventricular wall into the ventricles of the brain. Highly folded thin sheet-like connective
tissue richly vascularized, filtering blood to create CSF. Epithelial tight junctions create blood-CSF barrier.
Origin of CSF: Cerebrospinal fluid (CSF) is made from the blood. Special cells that make up the walls of some collections of arteries in the
brain called the choroid plexus filter the blood. Red, white, and platelet cells are too big to pass through the filter. So are most of the proteins
(immunoglobulins, albumin) and most drugs, that circulate through the blood. The filtered CSF has no cells to give it color or to make it opaque
so it is colorless and transparent. Ions and glucose are small enough to pass through the filter that makes CSF but their concentrations in blood
and CSF are not equal because of special regulatory channels and transport mechanisms.
4. Be able to point out and diagram the basic cellular features of meninges lining and protecting the outer surface of the CNS
and PNS.
CNS
Dura: irDCT, beneath bone, continuous with periosteum of skull,
venous sinuses (vascular spaces, collect blood exiting brain and
back to veins) epidural space with adipose and venous plexuses
Arachnoid: middle layer. Tenuously attached to dura one side, other
somewhat fused with pia. Avascular. Blood vessels go across it.
Dural side layer of epitheloid cells tightly bound by desmosomes
And tight junctions, creates subarachnoid space with CSF. CSF
Collected into arachnoid villi at superior sagittal sinus.
Arachnoid trabeculae, collagen/elastic fibers
Pia: innermost layer, LCT, lines outer surface brain and spinal cord.
Layer of simple squamous epitheloid cells. Highly vascularized
Perivascular space continuous with subarachnoid, filled with CSF
Origin mesenchymal, no tight junctions. Supports free exchange
of materials between CSF/CNS.
PNS
Endoneurium: LCT outside basal lamina of Schwann cells
Rich with collagen fibers. Contains CSF, maintain environ.
Vascularized.
Perineurium: surrounds many endoneuria form nerve fascicle.
Fibroblasts covered by basal lamina w/tight jxs & desmosomes
Fibroblasts layers alternate collagen bundles
Capillaries continuous with those of endoneurium à blood-neural
Barrier. Perineurium :: arachnoid
Epineurium: outermost sheath of irDCT covers PNS nerves.
5. Be able to explain chromatolysis, Wallerian degeneration, and axon
regeneration in PNS.
chromatolysis: The disintegration of the granules of chromophil substance (Nissl bodies)
in a nerve cell body which may occur after exhaustion of the cell or damage to its peripheral
process; other changes considered part of chromatolysis include swelling of the perikaryon and
shifting of the nucleus from its central position to the periphery.
Wallerian degeneration: A form of anterograde (from injury to axon terminal)
degeneration occurring in nerve fibres as a result of their division. Microglia and macrophages move in and engulf the debris.
Axon regeneration: Schwann cells proliferate and form tube -like cords in endoneurium. Sprouting axons guided to enter the Schwann cell
tubes and eventually extend all the way to the original target. Schwann cells remyelinate. Slow axonal transport limiting factor. Retrograde transport
of trophic factors from target organs and muscles needed to induce regrowth and help axon find right target.
Duchenne muscular distrophy dystrophin is an ABP
CF
actin needed for Cl- channel function
Lysosomal storage diseases
Tay-Sach’s
Gauche’s
Fabrys
Pompe’s
Lack of gene coding for phosphotransferase for mannose. Lysosomal enzymes secreted
I cell
Zellwegger
Cancer/Leber’s
JEB. Mutation in laminin gene
Raf-1/avb3 avb3 expressed by newly growing blood vesels
syndactyly
Human Organism - Nutrition Objectives
Online Modules
Proteins
Describe the differences between essential, conditionally essential, and nonessential amino acids.
Essential - AAs that cannot be synthesized by the body and must come from the diet
Nonessential - AAs that can be synthesized by the human body
Conditionally essential - AAs are essential under certain conditions (e.g. cys essential if low met,
tyr essential if phe low)
List essential, conditionally essential*, and nonessential amino acids
Essential: thr
Ile
Nonessential
ala
gln
Trp
Met
arg
gly
Val
Phe
asp
pro
Leu
His
asn
ser
Lys
cys*
Tyr*
glu
Calculate the recommended protein intake for a healthy adult
0.8g protein/kg healthy body weight
= 56g protein/day for 70 kg male
= 44g protein/day for 55 kg female
Explain the difference between animal and plant proteins and the concept of a limiting amino acid
Animal: tend to be complete proteins (except gelatin)
Includes meat, poultry, fish, eggs, dairy products
Plant: tend to be incomplete proteins (except soy)
Includes grain, nuts, seeds, legumes, vegetables
Discuss the adequacy of vegetarian diets and the concept of complimentary proteins
Limiting AA in plants
Food
Limiting AA
Complementary plant source
Beans (legumes) met, ile
grains, nuts, seeds
Grains
lys, thr
legumes
Nuts & seeds
lys
legumes
Vegetables
met
grains, nuts, seeds
Corn
trp, lys
legumes
Can combine plants to get all essential AAs
State the primary function of protein
Tissue maintenance and growth
Describe the purpose of a nitrogen balance study
Measurement of nitrogen provides a measure of protein
Describe the possible consequences of high protein intake
Source of extra fat
Low-protein diets slow decline in kidney function in people with developing kidney disease
Osteoporosis, increased urinary Ca loss
Define PEM, Kwashiokor, and Marasmus
PEM: Protein-energy malnutrition = protein deficiency usually accompanied by a deficiency of
energy intake and other nutrients.
Kwashiokor: “the disease that the first child gets when the new child comes” diet characterized
by low-protein density foods. Moderate kcal deficit, severe protein deficit. Symptoms: apathy,
listlessness, withdrawal, growth failure, edema, fatty infiltration of the liver, death. Tx infection,
diet w/protein
Marasmus: “to waste away” condition where the individual (esp. infants) starves to death.
Characteristics of diet: several kcal deficit, severe protein deficit. Symptoms: starvation, retarded
brain growth, “skin and bones” appearance (wasting), opportunistic infections, death. Tx: large
amounts of energy and protein, tx infections
HO Nutrition: Page 1 of 7
Online Modules
Lipids
Provide an operational definition for dietary lipids
- water-insoluble (hydrophobic); some exceptions
- soluble in fat solvents (benzene, chloroform, ether)
- utilizable in metabolic reactions by humans
Describe how lipids are classified
Triglycerides (TG)
- Glycerol
- Fatty acids
- saturated
- monounsaturated
- polyunsaturated
- trans
- storage form of fat in the body
Phospholipids
Sterols
- cholesterol
- vitamin D
- sex hormones
Essential Fatty Acids
- linoleic acid
C18:2, ω6
- linolenic
C18:3, ω3
Describe the association between LDL, HDL, and heart disease
Cardiovascular disease: elevate LDL, decrease HDL
List the essential fatty acids and identify dietary sources of essential fatty acids
Essential Fatty Acids
Linoleic acid
Linolenic acid
Dietary sources
Peanut butter
corn oil
walnuts/pumpkin seed oil
canola oil
olive oil
soybean oil
lard
vegetable shortening
fish oil
Margarine
polyunsaturated vegetables
butter fat
flaxseed oil
Describe recommendations for dietary fat intake
See Arnold’s book
Discuss the role of lipids in health
Lipoproteins: proteins + phospholipids. Act as emulsifiers so fat and fat-soluble substances can be
transported in the blood
Discuss the nutritional significance of lipids (I.e. what are the functions of lipids)
Source of energy: 9kcal/g
Insulate and protect the body
Biologically important molecules
Transporting fat-soluble vitamins
Provide flavor and satiety
9.8.03
Nutrition and Body Weight
HO N1
Dr. Koprowski
1. Define overweight and obese.
v Overweight – BMI 25.0 – 29.9
v Obese – BMI >=30.0
2. Describe three methods of assessing body fat.
v Body Mass Index (BMI)
§ Kg/m2
v Waist-to-hip circumference (WHR)
§ Body fat storage
§ WHR>1.0 = “risky”
v Waist circumference
§ Body fat storage
§ Gender specific guidelines
• Men >40 inches at risk
• Women >35 inches at risk
3. Calculate and interpret body mass index (BMI).
v 2.2 kg = lb
v 1 in = 2.54 cm
v kg/m2 = (2.2/(2.54*100)2 )lb/in2 = 3.41e-5
v see (1) for assessment of this measurement
4. Determine energy intake requirements.
v 35kcal/kg (16kcal/lb)
5. Describe the risk factors that increase the need for weight reduction.
v Smoking
v Hypertension
v Elevated LDL
v Presence of other medical conditions (comorbidities)
§ Coronary heart disease
§ Type 2 diabetes
6. Discuss lifestyle modifications for treating overweight and obesity.
v Diet
§ Slow rate of weight loss. 1-2lb/week. 500-1000Cal/day reduction
§ Very low calorie diets (VLCD). E deficit >1000Cal/day
§ Meal replacement. Usu. Liquid formula. Replace 1-2meals/day
§ Macronutrient composition
§ “popular diets” atkins, sugar busters, the zone
v Physical activity
§ Activity+diet modif’n = slightly better results than diet modif’n alone
§ Impt for long-term maintenance of weight loss
v Behavioral therapy
§ Group v. individual
§ Self-monitor
§ Stimulus control
§ Relapse prevention strategies
§ Assess, evaluate, define, develop
7. Describe how diets can be modified to promote weight loss and evaluate the effectiveness of these
modifications.
v Need to create an energy deficit.
v 500-1000 Cal/day reduction for 1-2lb/week weight loss
v see (6) – Diet above
9.10.03
Medical Nutrition Therapy, part 1
HO N2
Dr? Koprowski
1. Define medical nutrition therapy.
Guidelines that provide step-by-step instructions regarding nutritional care
Diet prescription (order)
2. List the five steps of the nutrition care process and recognize what happens at each step.
AIDiE assess, identify, develop, implement, evaluate
Assessment of nutritional status
Identify Nutritional needs
Develop nutrition care plan with specific objectives
Implement nutrition care plan
Evaluate the effectiveness of the nutrition care plan and make changes prn.
3. State the goal of medical nutrition therapy.
To supply needed nutrients to the body in a form it can handle.
Meeting the nutritional needs of a patient who require some type of diet modification as a result of
illness or disease.
4. Describe seven methods for modifying a normal diet.
Consistency, energy content, type of foods, omission of foods, energy distribution, number
and frequency of meals, route of delivery of nutrients
1. Change in consistency of foods.
i. Liquid diet, soft diet, low-fiber diet
2. Change in energy content.
i. High-calorie diet, weight reduction diet
3. Change in type of foods consumed
i. Na-restricted diet, lactose-restricted diet, high-fiber diet
4. Omission of specific foods
i. Gluten-free diet, allergy diet
5. Change in energy distribution (%kcal from CHO, fat, and protein)
i. Diabetic diet, renal diet, low-fat diet
6. Change in number and frequency of meals
i. Diabetic diet, postgastrectomy diet
7. Change in route of delivery of nutrients
i. Enteral and parenteral nutrition
ii. Enteral: provide nutrient solutions into GI through tube
iii. Parenteral: direct entry of nutrients into systemic circulation
9.22.03
Medical Nutrition Therapy, part 2
HO N3
Dr. Koprowski
1. Describe how changes over the lifespan affect nutrition therapy.
Ø Pregnancy and lactation
o Pregnancy Requires + 300kcal/day during 2 nd-3 rd trimesters
o Lactation requires + 800kcal/day
Ø Infancy
o Increasing energy requirements until the age of ~14
Ø Childhood
o Initially sharp decrease in protein req’s until age 2, then level off to 0.8g/kg/day
Ø Adolescence
o High energy req
Ø Adult
Ø Aging (older adults)
o Progressive loss of lean body mass, decrease caloric needs
2. Explain how nutrient needs can change based on how disease affects the digestion, absorption,
utilization, and excretion of nutrients.
Modification of diet to treat disease
Anything that impairs how a nutrient functions within the body will impair nutritional status.
Includes factors associated with food choices.
? Food choices
? Lack of intrinsic factor (req’d for
? Oral health
vitamin B12 absorption)
? Food preferences
? Utilization
? Culture, religious
? Cachexia
? Availability
? Muscle wasting, increase
? resources
BMR, e.g. in some cancers
? Digestion
? Diabetes mellitus
? Lactose intolerance
? Excretion
? Pancreatic insufficiency (e.g. in
? End-stage renal disease
CF)
? Liver disease and Zn
? Absorption
? Vitamin/mineral metabolism
? Inflammatory bowel disease
easily viewed in this sit’n
3. Describe how changes in clinical status may require changes in nutrition therapy.
Modification of diet to treat disease based on how disease affects nutrient use.
e.g. tumor affects nutritional status.
e.g. diagnose diabetic, must change eating habits.
4. Assess how nutrient-drug interactions can affect nutrient needs and/or disease treatment.
Interaction between a drug and a nutrient(food) that would not occur with the nutrient or drug
alone
? Food can affect drug
? Drug can affect nutrient
? Drug absorption
? Nutrient absorption
? Drug distribution
? Nutrient distribution
? Drug metabolism
? Nutrient metabolism
? Drug excretion
? Nutrient excretion
? Modification of drug action
? Food intake
? Enhancement
? Oral and taste/smell effects
? Gastrointestinal
♦ Grapefruit/cholesterol
? Appetite changes
? Antagonism
♦ Cipro/calcium
9.23.03
Enteral and Parenteral Nutrition
HO N4
Dr. Koprowski
1. Distinguish between enteral and parenteral nutrition support.
Enteral (EN)
Method of providing nutrient solutions into a GI tube
Use when patients cannot ingest or digest enough amounts of food
Use when patients have adequate absorptive capactiy (100cm small bowel)
Parenteral (PN)
Direct entry of nutrients into systemic circulation
Peripheral venous access
Central venous access
Bypass GI tract and first circulatory pass through liver
Use when patient cannot be adequately nourished by oral or enteral feeding methods
2. Describe the rationale and criteria for nutrition support
Indications for EN
Ø At risk for, or existing, malnutrition
Ø Maldigestion or absorption
Ø Poor appetite or anorexia
o Pancreatic insufficiency
o Side effects of chemotherapy
o Chron’s disease
o Mental illness
Contraindications for EN
Ø Inability to ingest food
Ø Obstruction in GI tract that cannot be
o Ventilator dependent
bypassed with a feeding tube
o Dysphagia (difficulty
Ø Protracted diarrhea or vomiting
swallowing)
Ø Acute bowel ischemia
Ø Gastroparesis (GI dysmotility)
Ø GI inflammation
Ø Acute severe pancreatitis
3. List the complications associated with enteral and parenteral nutrition.
Complications of Enteral Nutrition (EN)
Mechanical
GI
Metabolic
Sore throat
Nausea
Hypoglycemia
Esophagitis
Vomiting
Glycosuria
Mucosal damage
Diarrhea
Edema
Aspiration
Distention
Electrolyte imbalances
Nasal tube removed by pt
Bloating
Dehydration
Clogged feeding tube
EFA (Essential fatty acids)
deficiency
Hepatic and renal
respiratory
Ø
Ø
Ø
Complications of Parenteral Nutrition (PN)
Technical
problems develop e.g. ulcers
o Placement of catheter
Ø Metabolic
Septic
o Hepatobiliary
o Infections can be life
* Overfeeding and underfeeding
threatening
(à weigh patients)
Gastrointestinal
o Hyperglycemia
o Don’t use GI tract à more
o Refeeding syndrome (feed
o
o
o
after chronically starved)
Hyperlipidemia
Electrolyte and mineral
imbalances
Dehydration or fluid overload
4. Define Transitional feeding and explain what happens during this process
transitional feeding is the period after being taken off of total parenteral nutrition and before going
back to entirely oral feeding. Body readjusting to the entire process, so want to wean pt off nutritional
support. I.e. TPN à enteral + parenteral à total enteral nutrition à enteral + oral à oral
1. Distinguish between food-based and nutrient-based guidelines.
2. Using the U.S. Dietary Guidelines and the Food Guide Pyramid, describe
the general characteristics of a healthy diet—
a. Contributions of various food groups
b. Good common sources of individual nutrients
c. Foods to be consumed in limited amounts
d. Distribution of calories (carbohydrate:protein:fat ratio)
3. Define the following terms:
a. RDA (Recommended Dietary Allowance)
b. DRI (Dietary Reference Intake)
c. AI (Adequate Intake)
d. EAR (Estimated Average Requirement)
e. UL (Upper Limit)
4. Define HEI (Healthy Eating Index) and discuss its use in evaluating
dietary intake.
food guide pyramid, total fat, saturated fat, cholesterol, sodium, variety
5. Describe the limitations associated with using the Food Guide Pyramid as
a method of planning and evaluating dietary intake.
Prevention and Treatment of Disease – Pharmacology
9.10.03
Drug Development, Evaluation & Control
P&T Pharm1
Dr. Miller
1. Identify and describe the major stages of development
(preclinical, 4 stages of clinical).
• Drug Discovery
o Chemical modification of known molecules
o Recombinant technologies
o Random screening of natural or synthetic
products
o Targeted discovery: Rational drug design
o Therapeutic monoclonal and ScFvantibodies
o Genome -based approaches: Genomics,
proteomics
o Serendipity
• Drug Screening and Non-Clinical Research Phase
o Investigative New Drug (IND) investigational
exemption submitted
o Non-clinical screening & development, test
pharmacologic and toxic drug effects through
in vitro and in vivo short-term lab animal
testing (if evidence compound producing
desired biological effect)
o FDA asks for:
§ Pharmacological profile to determine
probable therapeutic action
§ Acute (single dose) toxicity
§ Subacute (short-term) toxicity
o How much drug absorbed in blood. How
chemically broken down. Acute/short-term
toxicity. How quickly drug and metabolites excreted
o Application for clinical trials
• Phase I Clinical Studies
o Is the drug safe in humans?
o Initial introduction to determine its Safety and a Minimum Effective Dose
o Open (“non-blind”) studies. 6-12 months
o Healthy volunteer subjects
o 20-80 subjects
o determine metabolic and pharmacologic drug actions, adverse effects,(some early evidence of)effectiveness
• Phase II Clinical Studies
o Preliminary data on effectiveness for a particular indication in patients with the target disease. I.e. to
determine efficacy in relation to safety. Develop therapeutic index (ratio therapeutic dose:lethal)
o Determine common short-term side effects and risks, including fertility and teratogenesis
o Several months to years
o Several hundred subjects
o Study designs
§ Cross over and randomized control studies
control group matched by age, disease state, sex and other factors
switch which pt receives drug and placebo
§ Single and double blind studies
single blind: subjects either get placebo or the drug
double blind: subjects don’t know, nor the researcher
§ Placebo response (20-40%)
• Phase III Clinical Studies
o After Phase II suggesting drug effectiveness of drug
P&T Pharmacology: Page 1 of 12
P&T Pharm1 page 2
o Gather additional information about effectiveness and safety to evaluate overall benefit-risk relationship,
including less common or rare adverse reactions
o Develop dosage regimens to extrapolate results to general population
o One to four years
o Multi-center testing
o Double-blind and cross-over studies
o Several hundred to several thousand subjects
• NDA Filing and Approval
o New Drug Application.
o Average two years
• Phase IV Postmarketing Surveillance
o After DNA approval
o Long-term safety
o Fine-tune aspects of delivery and dosage
o Active clinical studies or postmarketing surveillance
o Crucial to detect unexpected and rare drug safety issues (1/10,000 or less)
•
•
•
Accelerated Development/Review
o Speed up drug development of drugs significant benefits for serious or life-threatening illnesses
Treatment IND
o Process make promising new drugs available to desparately ill pts asap. Typically during phase III
Orphan Drugs
o Drugs treat rare diseases, i.e. affecting < 200,000 Americans
Number of patients
Length
Purpose
I
II
20-100
Up to several hundred
Several months
Several months to 2 years
III
Several hundred to several thousand
1-4 years
Mainly safety
Some short-term
safety, but mainly
effectiveness
Safety, dosage,
effectiveness
% of drugs
successfully
tested
70%
33%
25-30%
2. List essential information to be developed in Phase I,II,III, and IV clinical studies.
I
determine metabolic and pharmacologic drug actions, adverse effects,(some early evidence of)effectiveness
II
Preliminary data on effectiveness for a particular indication in patients with the target disease. I.e. to determine efficacy in
relation to safety. Develop therapeutic index (ration therapeutic dose: lethal. = lethal dose / effective dose)
Determine common short-term side effects and risks, including fertility and teratogenesis
III
Gather additional information about effectiveness and safety to evaluate overall benefit-risk relationship, including less
common or rare adverse reactions
Develop dosage regimens to extrapolate results to general population
IV
Long-term safety
Fine tune aspects of deliveray and dosage
Detect unexpected and rare drug safety issues
9.10.03
Pharmacodynamics: Drug -Receptor Interactions
P&T Pharm2
Dr. Miller
1. Explain the difference between pharmacodynamics and pharmacokinetics.
Pharmacodynamics – effect of a drug on the body, or, the relationship of drug concentration and biologic
(physiological or biochemical) effect
Pharmacokinetics – effect of the body on a drug, or, the way the body handles a drug (absorption, distribution,
metabolism, elimination, etc.)
2. Be familiar with pharmacodynamic terms including drug, receptor, ligand and effector
drug
chemical substance that acts on living systems at the molecular level
drug receptors the specific macromolecular components of the organism, usually complex proteins, that interact
with the drugs to create the ultimate effects, whether therapeutic or toxic
ligand
any molecule (hormone, NT, drug, messenger molecule) that binds to a receptor
effector
molecules that translate drug-receptor interactions into a change in cellular activity
3. List five transmembrane signaling mechanisms with examples of endogenous liquids or drugs and receptors by which
drug-receptor interactions exert their effects.
1
2
3
4
5
Binding
binding of lipidbinding of ligands binding of ligands binding of
binding of
soluble agents to
to extracellular
to extracellular
ligands directly
ligands to sevenintracellular receptors
domains of
domains of
to ion channels,
transmembranetransmembrane
transmembrane
regulating their
domain receptors
proteins:activating proteins:activating open state
that are linked by
intrinsic enzyme
separate,
G proteins to
activity of a
associated
various effector
cytoplasmatic
cytoplasmatic
systems
receptor domain
proteins
Ligands
Guanylyl cyclases
Insulin, EGF,
Cytokines, growth Acetylcholine
Adrenergic
(nitric oxide/NO)
PDGF,
hormone,
(n), glutamate
amines,
Corticosteroids, sex
TGFb, ANF
erythropoetin,
and
serotonin,
steroids,
interferons
other excitatory
acetylcholine
mineralocorticoids ,
amino acids,
(m),
vitamin D,
glycine,
odorants
thyroid hormone,
GABA-A
many P450
inducers
Receptors
Cytoplasmic/nuclear
Cell surface,
Cell surface,
Cell surface,
Cell surface,
transmembrane
transmembrane
transmembrane
transmembrane
(one-pass, often
(one-pass)
Multi-subunit
(seven-pass)
dimeric)
ion channels
(ligand-gated)
Mechanisms
Binding to DNA
Ligand binding to Ligand binding to Ligand binding
G proteinresponse elements,
extracellular
extracellular
to channel
mediated
regulation of gene
domain activates
domain activates
directly
interaction with
expression (onset lags
intracellular
separate,
regulates open
enzymes
30 min – many hrs,
serine/threonine,
loosely associated probability and
(adenylyl
long duration of
tyrosine
proteins
ion flux
cyclase,
action)
protein kinase or
(mostly adaptor
E.g., influx of
phospholipases)
phosphatase
proteins and
Na+ ions,
or ion
or guanylyl
protein tyrosine
depolarization
channels (K+,
cyclase domain
kinases)
in millisecond
Ca2+)
Protein
Protein
time scale;
Protein
phosphorylation
phosphorylation
regulation of
phosphorylation
cascade or
cascade or
Ca2+
cascade or
formation of
formation of
concentration
formation of
cGMP
second messenger
second (cAMP)
as second
molecules
or third (Ca2+)
messenger
messenger
molecules
Targets
exploited
Cytosolic and nuclear
receptors of
endogenous regulatory
ligands
-guanylyl cylclases
(NitricOxide NO)
-DNA binding
proteins/transcriptional
regulators
(corticosteroids,
mineralocorticoids,
sex steroids, vitamin
D, thyroid hormone)
Other Examples
1:Activation of soluble
(cytoplasmic) guanylyl
cyclase by nitric oxide
(NO) and
nitrovasodilator drugs.
--G protein-linked
receptors (ACh,
norepinephrine,
serotonin)
--ligand-regulated
transmembrane
enzymes
including tyrosine
kinases and
phosphatases
(insulin, EGF,
PDGF, serine
kinases (TGF β),
and guanylyl
cyclases (ANP))
--cytokine
receptors
(interferons,
growth hormone,
erythropoietin)
--ligand-gated
ion channels
(ACh, GABA,
glycine)
P&T Pharm2 page 2
1)
binding of lipid-soluble agents to intracellular receptors
binding of ligands to extracellular domains of transmembrane proteins: (2,3)
2)
activating intrinsic enzyme activity of a cytoplasmatic receptor domain
3)
activating separate, associated cytoplasmatic proteins
4)
binding of ligands directly to ion channels, regulating their open state
5)
binding of ligands to seven-transmembrane-domain receptors that are linked by G proteins to various
effector systems .: generating various chemical intracellular 2nd and 3rd messengers (cAMP, cGMP, Ca 2+,
many lipids, others)
9.17.03
Pharmacodynamics: Quantification of Drug Action
P&T Pharm3
Dr. Miller
1.
Define a graded dose response in both linear and logarithmic terms.
Response to a low dose of a drug usually increase in direct proportion to the dose (guarded response), but as doses
increase, the response increment diminishes.
Drugs chosen according to relative pharmacological potency and maximal efficacy in relation to the desired therapeutic
effect (response).
Potency:
concentration (EC50) or dose of a drug req’d to produce 50% of that drug’s maximal effect. One drug
more potent than another when produces an effect at lower concentration that other drug. (Graded dose-response
curves illustrate the concepts of potency and efficacy. Quantal dose-response curves show only the frequency of occurrence of a specified response in a
population at a given dose. )
Maximal efficacy: limit of the dose-response relation
Linearly, see a curve. Logarithmically, the curve’s steepness will give an indication as to the potential to cause an OD
by a slight increase in dosage.
2.
Define the terms affinity, efficacy, full and partial agonist, full and partial inverse agonist, competitive and noncompetitive antagonist.
Affinity
the probability of the drug occupying a receptor at a given time
Efficacy
the effect a drug can produce
Agonist
drugs that interact with receptors and cause effects as a result of alterations of the
functional properties of these receptors
Full agonist
a drug capable of fully activating the effector system when it binds to the receptor.
Efficacy = 1.0
Partial agonist
a drug that produces less than the full effect, even when it has saturated the receptors.
Efficacy <1.0.Acts as competitive antagonist in the presence of a full agonist!
Full inverse agonist
Produces an effect opposite to the agonist. Efficacy = -1.0
Partial inverse agonist
Produces an effect opposite to the agonist. 0 > Efficacy > -1.0
Antagonist
interact selectively with receptors. Inhibit the action of agonists while initiating no effect
Themselves. Possess affinity, lack intrinsic efficacy.
Competitive antagonist
Drug that blocks the action of a drug at its receptors by occupying those receptors
without activating them. Bind reversibly. Effect can be overcome by increasing the
concentration of an agonist to achieve maximum efficacy of agonist.
Non-competitive antagonist cause inhibitory effect resulting from irreversible interaction of the antagonist with the
receptor to prevent the binding of agonist or result from interactions prevent the initiation
of effects following agonist binding. Agonist/antagonist can be bound at same time, but
antagonist binding reduces or prevents the effect of the agonist. Max efficacy of agonist
cannot be reached.
3. Explain the concept of spare receptors and its biological consequences.
Don’t have to have enough drug to fill up all receptors to get the desired effect.
Due to the normally high efficiency of receptor-effector coupling, a maximal pharmacological response can often be
elicited by an agonist at a (lower) concentration that does not result in complete occupancy of all available receptors (i.e.
there are said to be spare receptors)
How? Receptor activation may trigger downstream responses that last longer than initial drug-receptor interaction.
Coupling receptor occupancy to response limited by [] of other signals
downstream, so max response w/o all receptors occupied
Biological consequences: agonists with low affinity for receptors (high KD ) can
produce full pharmacologic responses at low concentrations. Low affinity drugs
dissociate more rapidly, .: allow rapid reversal of a biologic response.
4. Compare potency and maximal efficacy of drugs on the basis of their dose-response
curves.
Potency: dose of a drug to produce 50% of that drug’s maximal effect
Maximal efficacy: greatest attainable response of a drug.
Drug B >(potent)> than A,C,D. DrugA =(efficacy)=C. C>(potent)>D, but
C=(efficacy)=D.
P&T Pharm3 page 2
5. Define a frequency distribution curve and a quantal dose response
curve.
Quantal dose response curve: based on population-based
determination of drug dose response. Lognormal distribution of
doses required to produce a specified quantal effect in individuals.
Indicates variability of drug responsiveness among individuals.
Quantal events are either-or, e.g. had seizures or not. Summation in a
plot of cumulative frequency distribution of responders versus log
dose.
Frequency distribution: Provides information about the standard
deviation of sensitivity to the drug in the population studied.
ED50 =median effective dose = dose at which 50% of inds exhibit specified quantal effect.
TD50 =median toxic dose. LD50 =median lethal dose.
Therapeutic index = LD50 /ED50 . estimate safety of drug.
6. Define the terms ED 50 , TD50 , LD50 , and calculate the therapeutic index of a drug.
See above
9.17.03
Pharmacokinetic Principles
P&T Pharm4
Dr. Miller
1. Define and calculate the most important pharmacokinetic parameters, including bioavailability, half-life, volume of
distribution, and clearance.
LADME
L
liberation
A
absorption
D
distribution
M
metabolism
E
excretion
Bioavailability
(F)
the amount of a given dose of drug that gets into the systemic blood circulation
IV, bioavail = 100% (F=1.0)
PO, bioavail <100%. Depends on amt drug released/absorbed & amount metabolized by GI/liver
Salt form (S) is fraction of drug salt/ester that is parent compound.
Half-life
time req’d for drug concentration to decrease by ½ after absorption/distribution complete
Value completely depends on values of clearance(Cl) and volume of distribution (Vd )
Use to predict how long it takes for a dosing regimen to achieve steady-state concentrations in
blood and determine dosage intervals. 4-5 half-lives “ “ “ “ “
t1/2 = (0.693 x Vd ) / Cl
Volume of distribution
a proportionality constant that relates the amount of drug in the body to the serum or
plasma concentration. Describes relationship btn []s of drug results when certain dose
administered to a pt. Apparent body space into which a drug can diffuse or the size of the space
where a drug distributes
water
total body water
0.60L/kg
extracellular water
0.20L/kg
blood
0.08L/kg
plasma
0.04L/kg
fat
0.2-0.35L/kg
bone
0.07L/kg
drug concentration achieved = ( dose x (FxS) )/ V d
Clearance
volume of space from which all drug is removed per unit time. 100ml/min means that all drug can
be removed (by distribution, metabolis m, and/or excretion) from 100ml of serum in 1 min.
Clinically, clearance is impt to determine amt drug necessary to maintain blood [] at steady state
Steady-state: Rate in = rate out. Steady-state concentration = Css
Rate in = dose x (FxS) / dosing interval τ
Rate out = Css x Cl
Css = dose x (FxS)/τ x Cl
9.24.03
Drug Absorption, Distribution & Elimination
P&T Pharm5
Dr. Roffey
1. Calculate the % absorption from the GI tract, if given the pH of the environment and the pKa of an acid or base drug.
Absorption from the G.I. tract occurs mostly via passive processes: The ionized, more polar forms
do not easily transfer across plasma membranes. Absorption is favored when a drug is in its
nonionized and more lipophilic form.
For a weak acids pKa = pH – log (concentration of ionized acid (A-)/ concentration of nonionized acid (HA))
For a weak bases pKa = pH – log(concentration of nonionized base (B)/concentration of ionized base (BH+))
2. Define terms relating to bioavailability and bioequivalence studies.
Bioavailability (F) : extent (and rate) to which a drug reaches its site of action.
Depends on drug solubility in water and fluids, dosage form (tablets, soft gelatin capsule) and route of administration.
Bioequivalence: two or more different preparations of the same drug (e.g. brand-name vs. generic)
3. Describe and understand the advantages and disadvantages of various routes of drug
administration (p.o., sublingual, rectal, i.v., i.m., s.c., topical, inhalation).
ROUTE
ABSORPTION PATTERN
ENTERAL (go through GI tract)
ORAL (PO)
Variable; depends upon
many factors
Buccal/sublingual
Rectal (suppositories)
Prompt for nonionic, lipid
soluble drugs (nitrates)
Often irregular and
incomplete
PARENTERAL (not through GI tract)
Intravenous (iv)
Absorption circumvented
Potentially immediate effects
Subcutaneous (sc,subq)
Prompt (from aqueous
solution), or slow and
sustained from repository
preparations
Intramuscular (IM)
Prompt (from aqueous
solution), or slow and
sustained (from repository
preparations)
SPECIAL UTILITY
LIMITATIONS AND
PRECAUTIONS
Most commonly used; most
convenient and economical;
usually more safe
Requires patient cooperation.
Availability potentially
erratic and incomplete for
drugs that are poorly soluble,
slowly absorbed, unstable, or
extensively metabolized by
liver/gut (first-pass effect)
Protection from first-pass
metabolism by liver
Pt vomiting or unconscious;
children
Partial protection from firstpass metabolism by liver
(50%)
Valuable for emergency use
Permits titration of dosage
Usually required for high
molecular weight protein and
peptide drugs
Suitable for large volumes
and for irritating drugs
(when diluted)
Suitable for some insoluble
suspensions and for
implantation of solid pellets
Suitable for moderate
volumes, oily vehicles, and
some irritating substances
Possible irritations of
mucosa
Increased risk of adverse
effects
Must inject solutions slowly;
as a rule
Not suitable for oily
solutions or insoluble
substances
Asepsis must be maintained
Not suitable for large
volumes
Possible pain or necrosis
from irritating substances
Not suitable in shock
Precluded during
anticoagulation medication
May interfere with
interpretation of certain
diagnostic tests (e.g. CK)
Not suitable in shock
4. Describe distribution and half-life curve in a two-compartment model and explain how the
volume of distribution will change when the elimination phase is reached.
Distribution: after drugs are absorbed or injected into the blood stream, often both
redistribution into interstitial and cellular fluids and elimination occur. First compartment
(Vi) (blood, plasma or tissue with high blood flow). Then distributes more slowly into 2nd
compartment (Vt). Sum of all compartments is apparent volume of distribution (Vd)
Initial decay half life due to drug being distributed into tissue (Vt). Second half-life due to
drug being eliminated from body. Elimination curve gets less steep at point of elimination
versus excretion.
Vd =
apparent volume into which drug distributed =
amount drug in body / plasma drug concentration
5. Discuss the effect of protein binding on drug half-life.
*Drug stays longer in circulation if highly bound.*
Only free or unbound drug can diffuse/be transported. Disease states or drug interactions can displace drugs from protein
binding: increased drug effect since more free drug accessible to tissues. Drug also then available for elimination or
distribution into non-active tissues.
After drug displacement from protein binding, new steady state where free drug concentration same as before, but TOTAL
concentration less. Less bound.
6. Define the differences between zero and first order elimination kinetics.
Zero order: drug saturates routes of elimination and .: disappears from circulation in a non-concentration dependent manner.
ABSOLUTE amount of drug disposed of is constant per unit time. Half-life t 1/2 not constant. E.g. asa, ethanol.
First order elimination: rate of elimination proportional to [drug]. Most commonly seen. Drug disappears in a concentrationdependent manner. Half of drug eliminated in a constant period of time. t 1/2 is constant, not concentration-dependent. (t 1/2 =
0.693 / k)
k = elimination rate constant = fraction of volume of distribution which is cleared per unit of time. k used to predict how
drug plasma concentration varies with time, assuming no add’l drug added. Cl = clearance. E.g. 100 ml/min = in 1 minute,
drug eliminated from 100ml of serum. Assume metabolism liver, elimination kidney.
k = 0.693 / t 1/2 = Cl / Vd
have enough of a drug of first order kinetics, it’ll change to zero order cuz the proteins are full.
7. List sites of drug elimination.
Lungs (metabolism/exhalation), saliva, tears, sweat, breast milk, plasma, most impt: Liver (biotransformation, excretion into
bile and feces) and kidney (urine)
Renal elimination by filtration
Small, unbound (unless glomerular damaged), excrete >40% drug unchanged, decreased renal fx require
dosing adjustment
Renal elimination by active transport processes
Secrete and reabsorb, separate for organic acids/bases, drug transport system in distal nephron for digoxin
elimination, competed for by spironolactone, quinidine, verapamil. Drug interaction
Renal elimination by passtive transport processes
Weak acid/base passively reabsorbed in collecting duct, influenced by urinary pH, urine flow rate
8. Calculate the ratio of nonionized to ionized drug in blood and urine, and know whether to acidify
or alkalinize urine to “force” renal excretion of a drug.
Urine pH can vary pH 4.3 – 8.0. can be made acidic or alkaline
Alkaline diuresis: force elmination of weak acidic e.g. barbiturates, salicylates
Urine made alkaline by Na bicarbonate
Acidic diuresis: ascorbic acid given. Force weak base out
pKa = pH – log ([ionized acid A -]/[nonionized acid HA])
ex: weak acid drug pKa 8.4. pH blood 7.4. will drug diffuse across membrane of kidney tubule? pH lower than
pKa, so for a weak acid, the nonionized (aka diffusible) form will predominate. How much will diffuse
log ( [A-]/[HA] ) = pH – pKa = 7.4 – 8.4 = -1. 10^-1 = 0.1, so ratio of ionized to nonionized is 1:10. .: 10/11 of
drug will diffuse into the urine.
How to keep drug from diffusing back:
If urine made alkaline (with bicarbonate) to pH 9.4, diffusion of drug away from urine back into plasma pH – pKa =
9.4 – 8.4 = 1. 10^1 = 10 = [ionized]/[nonionized] i.e. more ionized so won’t diffuse. Only 1/11 of drug will diffuse
back into blood. Acidic drug trapped in alkaline urine.
9.24.03
Multiple Dosing & Drug Accumulation
P&T Pharm6
Dr. Mogos
10.01.03
Pharmacology Lab Cases
P&T Pharm7
MDLs
The Human Organism Objectives: Physiology
08-15-03
Compartmental organization of the body:
Fluid and electrolyte homeostasis
HO Phy1
Dr. McDonough
1. Describe the major subdivisions of the body fluids and the approximate
percentage of total body water, sodium and potassium in each compartment.
TBW (total body water) = 60% body weight, = 42L for 70kg person
Name
%Total Body Water
Sodium (Na+) Potassium (K+)
ICF: Intracellular fluid
0.4 x body weight
12 meq/L
150 meq/L
= 2/3 total body water
--------------------------------------------------------------------------------------------------------------Cell Membrane
--------------------------------------------------------------------------------------------------------------ECF: Extracellular fluid
0.2 x body weight
= ISF + plasma
ISF: Interstitial fluid
¾ of ECF
145 meq/L
4 meq/L
Directly bathes the cells
= ¼ total body water
--------------------------------------------------------------------------------------------------------------Vascular capillary Endothelium
--------------------------------------------------------------------------------------------------------------Plasma
¼ of ECF
145 meq/L
4 meq/L
The ECF in cardiovascular space = 1/12 total body water
--------------------------------------------------------------------------------------------------------------Epithelium
---------------------------------------------------------------------------------------------------------------
TCF: Transcellular fluid
Highly variable input, e.g. dietary, saliva, stomach lumenal fluid
2. Be able to use the normal concentrations of Na+, K+, Cl-, and HCO3- in extracellular and intracellular
fluids to interpret electrolyte disorders, e.g. hyponatraemia
Extracellular
Intracellular
Na +
145
12
K+
4
150
Cl105
5
HCO3 27
12
hyponatraemia
Abnormal decrease in blood sodium concentration.
Normal blood sodium should be 136 to 142 milliequivalents per litre. Hyponatraemia can occur secondary
to inadequate salt intake, excessive sweating, vomiting or as a drug side effect.
3. Explain why fixed negative charges on plasma proteins leads to a difference in the concentrations of
anions and cations in plasma water vs interstitial fluid.
Plasma proteins do not move between the compartments but their charges do contribute towards a chemical
gradient, so the diffusable ions move between the plasma water and interstitial fluid to compensate for this
chemical gradient.
4. Explain why a change in total amount of NaCl in the body affects prima rily the extracellular fluid
volume rather than extracellular osmolality of the body fluids when water/drinking is not limiting.
HO: Physiology Page 1 of 13
HO Phy1 Compartmental organization of the body: Fluid and electrolyte homeostasis Page 2
Sodium is the primary determinant of ECF volume. Osmotic pressure difference across membrane, not
absolute osmolality, is the driving force for water movement. Osmolality = solute particles/1kg.
The “semipermeable” membrane allows water, but not solutes to pass through
Since water/drinking is not limiting, when the addition/deletion of NaCl occurs, water will shift to
accommodate for the change in NaCl. The water will follow the flow of NaCl, so the ECF volume will
change accordingly. The osmolality won’t change because the solutions will be at the same concentration
due to the unlimitedness of water availability.
5. Demonstrate the ability to calculate how infusions of: isotonic saline, water, hypotonic and hypertonic
fluid affect ICF and ECF volume and osmolality.
Isotonic saline
Water
Hypotonic fluid
Hypertonic fluid
volume osmolality volume osmolality volume osmolality volume osmolality
ICF
=
=
↑
↓
↑
↓
↓
↑
ECF
=
↑
↑
↓
↑
↓
↑
↑
See: osmotic pressure. The pressure required to prevent osmotic flow across a semi permeable membrane
separating two solutions of different solute concentration. Equal to the pressure that can be set up by
osmotic flow in this system.
08-19-03
Ionic Equilibria and Resting Membrane Potentials
HO Phy2
Dr. Farley
1. Distinguish between net flux and unidirectional flux
Net Flux: The difference between the two unidirectional flux's.
Unidirectional Flux: The flux of a substance from one surface of a boundary layer or membrane to the
other, disregarding any counterbalancing flux in the other direction
2. Define:
electrochemical potential:
electrochemical potential difference is what represents the thermodynamic force that is moving ions
equilibrium potential difference for ion i
(Ei) is the electrical energy equivalent of the chemical energy stored in the concentration gradient
e.g. ENa = +65 mV. Potential to change [ ] gradient into electrical energy. Maintained by pump.
The voltage difference that prevents change in the [ ] difference for an ion across a membrane
electrochemical equilibrium
the point at which the potential energy contained in the concentration difference for an ion is balanced
by an oppositely directed voltage difference
membrane potential (Vm)
a weighted average of the equilibrium potentials described by Nernst for each ion, calculated by the
chord conductance equation
E = Nernst (equilibrium) potential = (RT/zF)ln(Cout/Cin) = 25/zln(Cout/Cin)=60/z*log(Cout/Cin)
g = membrane conductance for each ion. Det’d by transport proteins in cell membrane
conductance
proportional to the rate of ion movement across cell membranes, determined by the transport proteins
embedded in the cell membrane. Related to permeability.
electrochemical gradient
A measure of the tendency of an ion to move passively from one point to another, taking into
consideration the differences in its concentration and in the electrical potentials between the two points;
commonly expressed as the additional voltage needed to achieve equilibrium.
3. Describe how voltage differences across membranes are generated by ion fluxes
Voltage differences across cell membranes arise because ions move across the membrane at
different rates.
The ions diffuse at different rates. This difference in diffusion rates is due to a difference in the size of the
hydrated ions.The difference in diffusion rates between Na+ and Cl- very rapidly results in a displacement of
some of the chloride ions ahead of the sodium ions.This negative potential in compartment B slows the
diffusion rate of the remaining negatively-charged chloride ions and increases the rate of diffusion of the
positively-charged sodium ions through electrostatic interactions. As the Na+ ions begin to catch up to the
Cl- ions, the electronegativity in compartment B decreases and the Cl- ions speed up again. Eventually, when
the NaCl concentrations are equal in A and B, there will be no voltage difference across the membrane.
Note that if some mechanism existed to pump the ions back from B to A and thereby maintain the initial Na+
and Cl- concentration differences between A and B, the ions would never equilibrate and a constant voltage
difference could be maintained across the membrane.
HO Phy2 Ionic Equilibria and Resting Membrane Potentials Page 2
4. Describe the relationship between the Nernst equation and the equilibrium potential
(RT/zF) ln (Cout/Cin) = Vin - Vout = E
The Nernst equation defines values of the electrical potential difference (Vin - Vout ) that exactly
balance the driving force due to a concentration difference (Cout/Cin) for an ion.
When an ion is at equilibrium there is no net flux
5. Write the chord conductance equation
6. List the major clinical features of cystic fibrosis
q elevated Na+ and Cl- in sweat,
q a generalized abnormality of mucus secretions that leads to chronic obstructive pulmonary disease
(à respiratory acidosis) and pancreatic insufficiency
q colonization and infection of the airways and lung parenchyma by Pseudomonas aeruginosa.
7. Describe the mechanism of sweat production in
sweat glands
The primary sweat is made by acinar cells in the
secretory coil of the sweat gland and is secreted by
these cells into the lumen of the gland in response to
cholinergic stimulation. The primary sweat is similar to
plasma except for the absence of plasma proteins, and it
is the same in CF patients and in people without the
disease. After secretion into the lumen of the sweat
gland, Na+, Cl-, and water are resorbed from the sweat
by the resorptive duct as the sweat travels to the skin
surface. The abnormality characteristic of CF affects
only the resorption of the ions in the resorptive sweat
duct and does not affect the primary sweat secretion.
8. Explain the relationship between elevated sweat NaCl in cystic fibrosis and altered
chloride conductance
The movement of chloride is passive and is driven by the movement of a positive charge
carried by sodium. Sodium transport is driven by the Na,KATPase which maintains low
intracellular sodium concentrations inside the cell and makes it thermodynamically
favorable for Na+ to enter. When the chloride permeability of the cell membrane is
reduced in cystic fibrosis, the charge brought into the cell by the entry of sodium is not
neutralized, and so the further entry of Na+ is reduced by electrostatic repulsion. Thus,
higher concentrations of both Na+ and Cl- arrive at the skin surface in patients with
cystic fibrosis than in people without the disease.
08-22-03
Membrane Transport Mechanisms
HO Phy3
Dr. Farley
Topics
Structure of Cell Membranes
Active and Passive Transport
Passive Transport Mechanisms: Facilitated Diffusion Carriers
Passive Transport Mechanisms: Ion Channels
Ion Channels and Cystic Fibrosis
Active Transport: The Na,K-ATPase
Sodium-Solute Cotransport
Regulation of Biological Transport
Coupling Solute Movement and Water Movement Across Cell Membranes
Regulation of Cell Volume
Epithelial Structure and Transport
Absorptive Epithelial Transport
Secretory Epithelial
How Does This Explain the Clinical Presentation of Cystic Fibrosis?
Objectives
1. Categorize the following as either active transport or passive carrier-mediated transport:
A: ion pumps, sodium-solute cotransporters
P: ion channels, facilitated diffusion transporters,
2. Define:
CFTR
a chloride channel, homologous to a family of proteins that actively transport small solutes in an
ATP dependent manner (ABC transporters). The regulator protein is a protein which is embedded
in the cell membrane and acts as a channel for certain ions to be transported into or out of the cell.
The disease cystic fibrosis is caused by a defect in the gene for this protein.
Gating
the opening and closing of a channel, believed to be associated with changes in integral membrane
proteins and/or voltage changes in the membrane potential. Voltage-gated, ligand gated.
cardiac glycosides
a class of drugs, including digitalis and ouabain, that specifically inhibit the α subunit of Na,KATPase
symport aka cotransporter
a mechanism of transport of the sodium-solute cotransporter when the transport of the solute
occurs in the same direction as sodium. Both go the same way. E.g. Na+/glucose.
antiport
a mechanism of transport of the sodium-solute cotransporter when the transport of the solute
occurs in the opposite direction as sodium. Molecules go opposite ways. E.g. Na+/H+,Na+/Ca2+
electrogenic
a substance that contributes to an electrical potential across a membrane. E.g. Na+/K+ATPase
isotonic
a solution that does not cause a change in the volume of a cell suspended in it
HO Phy3: Membrane Transport Mechanisms Page 2
hypertonic
a solution that causes cells suspended in it to shrink
hypotonic
a solution that causes cells suspended in it to swell
transcellular pathway
a molecule passes through the apical membrane then diffuses through the cytoplasm of the cell
and passes through the basal membrane
paracellular pathway
a molecule that penetrates the tight junction and then diffuses through the lateral intercellular
space between adjacent cells. Tightness of tight junction varies with tissue type
apical or luminal cell membrane OR brush border OR mucosal surface
the membrane of the epithelial cell that faces either the external environment or lumen of organ
basolateral OR basal OR lateral cell membrane OR serosal surface
the surface membrane of the epithelial cell that is either supported by the basement membrane or
is otherwise is contact with the interstitial fluid
brush border
see apical
amiloride
an inhibitor of the epithelial Na channel in the Colon Henle’s loop
A drug that blocks sodium/proton antiport, used clinically as a potassium sparing diuretic.
Furosemide
An inhibitor of the Na/K/2Cl Symport in Henle’s loop
Potent diuretic that increases the excretion of sodium, potassium and chloride ions and inhibits
their resorption in the proximal and distal renal tubules.
DIDS
An inhibitor of the Cl/HCO3 antiport in the small/large intestine and RBC
An inhibitor of anion conductance
3. What is the most common mutation in CFTR and what are the consequences of that mutation
for CFTR function?
∆F508
deletion of this codon results in misfolding of the protein so that it is never inserted into the
plasma (apical) membrane.
In the pancreatic ducts, the ducts are unable to secrete sufficient amounts of NaHCO3 and water.
Clogged with digestive proteins normally secreted into acini, eventually destroyed. Also, food not
digested so CF patients tend to be malnourished & failure to thrive and muscle wasting.
In airways, accumulation of mucous associated with Pseudomonas infection & pulmonary
insufficiency.
4. Explain how the activity of Na,K-ATPase can generate a membrane potential.
Na,K-ATPase binds and hydrolyzes intracellular ATP and for each ATP molecule hydrolyzed, the
Na,K-ATPase pumps out 3 Na+ ions out and two K+ ions into cell. .: pump is electrogenic
5. List two mechanisms whereby transport of small molecules or ions is regulated in animal cells.
1) Hormones/Neurotransmitters/Drugs that bind to receptors. Regulate e.g. PO4- of receptor so
hormone can’t bind
e.g. cAMP
2) Regulate number of transport proteins – transport to/from membrane
e.g. insulin recruits GLUT4
6. Describe three mechanisms that are used by animal cells to maintain a constant intracellular
volume in response to changes in extracellular osmolarity.
Na,K-ATPase: pumps out more Na+ than it pumps in K+. result: decrease in intracellular volume
Regulatory volume ↓: activation of pathways that allow for the passive eflux of K+ and an anion
Regulatory volume ↑: activation of pathways for the passive entry of Na+ and an anion
7. Explain the mechanism whereby Vibrio cholerae causes symptoms of cholera and how oral
rehydration therapy for cholera works.
Vibrio cholerae invades GI tract, secretes protein toxin: cholera toxin. Pts can lose 10-20L of
fluid. The toxin
activates kinase
which inhibits
Na/H antiporter in
intestinal villus
(absorption),
activates chloride
efflux. Net
reduction NaCl
absorption and
increase in Clsecretion leads to
diarrhea and loss of
fluids and
electrolytes.
Oral rehydration
involving isotonic
fluids with glucose
stimulate
Na/glucose
cotransporter.
Transports Na+ into
cell and water
follows.
8. Describe the mechanism of electrolyte and water
secretion by secretory epithelia and explain
how mutations in CFTR affect this mechanism to cause
symptoms of cystic fibrosis.
Exocrine gland = simple tubule = Secretory endpiece
i.e. acinus + duct. Acini secrete Na+ and Cl-, which
drives water secretion osmotically.
During normal secretion, hormone (aka secretagogue)
binds, increases 2nd messenger, activates PKA,
activates CFTR channel. In CF, no CFTR channel to
activate. So no Na+/Cl- excretion à no H2O secretion
à build up of mucus or lack of secretion of digestive
enzymes.
Name
Location
Inhibitor
Na+ entry across apical membranes of absorptive
epithelia
Cl- entry across apical membranes
Epithelial
Na
Channel
Colon
Henle’s
loop
Na/H
Antiport
Na/K/2Cl
Symport
Na/Solute
Cotransport
Cl
Channel
Na/K/2Cl
symport
Cl/HCO3
Antiport
Proximal
tubule
small/large
intestine
Henle’s loop
Small
intestine
Proximal
tubule
Henle’s loop
Small/large
intestine
RBC
Amiloride
(10-7 M)
Amiloride
(10-5 M)
Furosemide
(Lasix)
bumetanide
phlorizin
Tight
junction
Sweat
gland
Tracheal
Epithelia
?
Furosemide
bumetanide
SITS
DIDS
9.9.03
Introduction to Endocrinology and Transmembrane Signaling
HO Phys4
Dr. Kaslow
1. Be able to define hormone, target cell, receptor.
Hormone: molecules that are the signals of the endocrine system
Target cell: cell receiving hormone as a signal
Receptor: specific recognition site to which hormone binds to create the signal. Could be on suface, in
cytoplasm, or nucleus.
2. Understand the unique characteristics of nervous, endocrine, paracrine, autocrine, and adhesion
signalling.
nervous: between neurons. Very selective, only to neuron synapse with is signal transmitted. Long/short
distances
endocrine: signals through blood and lymph. Long distances. Specificity through substances, receptors,
types cellular metabolism.
Paracrine: signals through diffusion. Short distance. Specificity cuz only small volume can receive signal
Autocrine: bounce ball off own head. Signal affects cell that secretes it. Respond to change in that cell’s
environment.
Adhesion: cell-cell and cell-matrix interactions
3. Draw a generic diagram of a hypothalamus-pituitary-target organ feedback loop and be able to use
such diagrams along with an understanding of negative feedback loops to solve problems regarding
diseases and therapies.
See discussion questions #2 for an example with
human growth hormone
4. Draw a diagram that shows common steps in
the synthesis, release, transport and action of
hormones, and use it to propose dysfunctions that might lead to disease, or be potential therapeutic
targets.
5. Understand the theory underlying the design of binding assays for hormones (e.g. RIAs) and be able
to recognize when an artifact may cause an RIA to indicate presence of an active hormone, even when
not present.
radioactivally tag stuff (either hormone, antibody, receptor) to detect presence of a hormone. Tagging
antibody = RIA. apparent increase in H may be caused by: degraded H, mutated H, degradation of binding
protein, presence of substance resembling hormone. Tagged H degraded, measuring non-bound H, so what
measuring is initial minus (degraded and bound), but if think it’s only the bound stuff, will think a lot is
bound and there’s less untagged H which would be an underestimate.
6. Clearly communicate regarding data concerning hormone sensitivity (EC50 ) and responsiveness (E max)
EC50 = concentration of H that causes 50% of Emax. (effective concentration giving 50% effect)
Decreased responsiveness à decrease in Emax. Can’t ever get full effect. Receptor says screw you.
Decreased sensitivity à increase in EC50. can get full effect, it’s just going to take more. Receptor will
do it if you just nudge it, it’s just being sensitive. It’ll just take more nudging, i.e. increase EC50.
7. Understand the terms spare receptors, efficacy, agonists, competitive antagonists, non-competitive
antagonists, and partial agonists. Understand the difference between additive and synergistic hormones.
Spare receptors: if a full agonist can get to Emax without needing all receptors to be occupied, the leftover
unoccupied receptors are spare receptors
Efficacy: how much effect the H can have.
Agonists: a molecule with the desired effect
Competitive antagonist: a competitive antagonist binds to a hormone receptor but has no effect. Will
increase EC50, but not affect Emax
Non-competitive antagonist: binds to different spot on hormone receptor than H so that H can’t bind. Emax
decreases.
Partial agonist: creates same response (but not same efficacy) as full agonist, but only after it occupies all
receptors. Can’t get to Emax of full agonist.
Additive hormones: effect of A and B is A + B
Synergistic hormones: effect of A + B is greater than the effects of A and B individually
8. Be able to recognize evidence for positive and negative cooperativity in data regarding hormone
binding to receptors, and predict how cooperativity might influence dose-response curves.
positive cooperativity,
Log Dose-Response curve, HR% higher for each [H]
Lineweaver-Burke plot, 1/[HR] lower for each 1/[H]
Scatchard, HR/[H] higher for each HR.
*Hill, HR/(HRmax-HR) smaller for log[H]<1, greater for log[H]>1 *slope = quantitave measure of
cooperativity
9. Explain why it is difficult to use data regarding the binding of a hormone to its receptor to predict
efficacious dosages of the hormone.
data rests on assay system where product linearly proportional to time reaction runs. May not be the case.
May have desensitization, which may require pulses of the drug.
10. Use an understanding of the difference between homologous and heterologous desensitization to
diagnose disease and use drugs in therapies.
homologous desensitization – one hormone. Common cause, change in receptor structure or location. For
a drug, may be able to get another drug that the receptor will respond to, or use another
hormone/receptor combo that produces the same effect but isn’t desensitized
heterologous desensitization – multiple hormones which act via distinct receptors but cause common
effects. Drug at the receptor level won’t help this. Need to effect the step farther down the line.
11. Explain why the pulsatile delivery of hormones may be more effective than constant delivery, and be
able to incorporate knowledge of this phenomena into the therapeutic use of hormones and hormone
analogs.
signal information requires changes in concentrations.
12. Understand how changes in allosteric regulation of enzyme activity can be used to assess the effects
of hormones.
mechanisms of allosteric regulation: 1) allosteric regulation of existing molecules. 2) reversible or
irreversible covalent modification 3) change physical location. 4) change number of enz molecules by
changing the rate of synthesis and/or degradation
receptors on surface generally regulate production of 2nd messengers or activity of ion channels
receptors in cytoplasm or in nucleus generally regulate gene transcription
13. Understand the terms cascade, amplification, coordinate control, adenylate cyclase, G proteins, and
A proteins.
cascade: initial effect causes lots of others events
amplification: initial effect that causes a lot of the other events to occur a lot
coordinate control: lots of stuff have to come together to get the final result
adenylate cyclase: catalyse synthesis of cAMP from ATP
G protein: inhibit or stimulate adenylate cyclases catylase activity
A protein: cAMP dependent protein kinase. Phosphorylates lots of stuff. PKA
14. Diagram the generic G-protein cycle, predict the effect
of hydrolysis-resistant analogs of GTP, and the effects of
ADP-ribosylations that block hydrolysis of GTP or release of
GDP (figure 18).
Hydrolysis -resistant, stuck in alpha-GTP state, so will
continue to stimulate/inhibit adenylate cyclase
ADP-ribosylation prevents release of GDP so stuck in alphaGDP state, so can’t do its function to either stimulate/inhibit.
Stuck in off position relative to effect on adenylate cyclase.
Pertussis toxin: blocks G-protein cycle at level of (inhibit)
GDP releasing
Cholera toxin : blocks G-protein cycle (stimulatory of
adenylate cyclase) at level of GTPase, so stuck on in
stimulating adenylate cyclase to catalyze ATP à cAMP.
15. Explain why mutations that increase cyclic-AMP
promote differentiated processes, catabolic metabolism, and can cause tumors that secrete abnormally
high amounts of hormone.
persistant signal to perform the cell’s differentiated function, leads to cell division. Lots of cell division à
tumors.
16. Explain the roles of PKA and PDE in the cAMP system, and why they might become therapeutic
targets.
PKA , upon binding of cAMP, releases regulatory catalytic subunits which phosphorylate several substrates
PDE = cAMP-phosphodiesterase. Terminates reign of cAMP.
PHYS 5
1. Use the normal value range of the constituents in Table 1 in
interpretation of simple clinical scenarios
IMPORTANT CONSTITUENTS OF EXTRACELLULAR
FLUID (ECF)
Normal range Non-lethal limits
Sodium ion 138 - 146 mM 115-175 mM
Chloride ion 103-112 mM 70-130 mM
Potassium ion 3.8 - 5 mM 1.5-9.0 mM
Glucose 75 - 95 mg/dl 20-1500 mg/dl
Bicarbonate ion 24 - 32 mM 8-45 mM
Acid-base - pH 7.3 - 7.4 pH 6.9-8.0 pH
Osmolality 280 - 290 mOsm/kg
These ranges are maintained by interactions between multiple
organs and/or hormonal systems.
2. Define feedback regulation and its importance in homeostasis
Feedback regulation
Control mechanism that uses the consequences of a process to regulate the rate at which the process occurs:
Keep the body in check.
3. Define the main organ systems responsible for rapid and for long term
homeostatic regulation of each of the following : plasma
sodium/volume, plasma potassium, plasma pH, plasma osmolality.
Kidney, plasma
Na/volume: cardiovascular, kidney, brain
K+ plasma: muscle
Plasma pH: renal. Nervous system.
Plasma osmo lality. Brain, renal
Glucose: muscle/fat
1. Learn the various buffers in the body that are important in responding to acid-base changes, and
learn the relative speed of each group of buffers.
1. Extracellular buffers
· HCO3 (kidney) - - CO2 (lungs) most important because close control by lungs and kidney possible
HCl + NaHCO3 ß à NaCl + H2CO3ß à H2O + CO2
· Plasma proteins : protein -n à Hn -protein
· Inorganic phosphate, amino acids, other.
2. Intracellular buffers
· Bone (can buffer as much as 40% of an acute acid load)
· Proteins: protein –nà Hn-protein (e.g. hemoglobin in hi concentration in red blood cells)
· Organic and inorganic phosphates: phosphate-n → Hn- phosphate
2. Write the Henderson - Hasselbalch equatio n.
pH = pK + log ([A-]/[HA])
3. Explain the unique role of the CO2 - HCO3- system in regulating body fluid pH.
Acids buffered by HCO3- → H2CO3 →CO2 + H20 → blow off the excess CO2 (rapid) and lose HCO3- in
the process (lower [HCO3-] in uncompensated metabolic acidosis ).
4. Learn the changes in blood pH, [HCO3-], and PCO2 that occur in the following uncompensated
acid-base disturbances: metabolic acidosis, metabolic alkalosis, respiratory acidosis, respiratory
alkalosis.
Uncompensated
Metabolic acidosis: ?plasma [HCO3-]
?H+ ?pH drugs, diarrhea (lose base), dm, renal failure (gain acid)
Metabolic alkylosis: ?plasma HCO3?H+ ?pH. Vomit stomach contents, overingest base. Lose acid: gastric
suction tube
Respiratory acidosis: ?CO2 (hypoventilation)
?pH ?H+ hold breath, COPD, emphysema, CHF, anesthetics, drugs,
chronic lung disease
Respiratory alkylosis: ?PCO2 (hyperventilation)
?pH ?H+ anxious, hypoxia, hike high altitude, pain
5. Explain how the kidney compensates for acid-base disturbances
induced by the respiratory system, and how the respiratory system
compensates for metabolic disturbances.
Renal compensation to restore [buffer]: excrete more H+ (requires
changing renal gene expression) which allows replenishing [HCO3- ] by
slowing: H+ + HCO3- → CO2 + H2O (more in Renal section).
Metabolic acidosis: ?plasma [HCO3-]. Respiratory: hyperventilate
Metabolic alkylosis: ?plasma HCO3-. Respiratory: hypoventilate.
Respiratory acidosis: ?CO2 (hypoventilation). Renal: ?renal net acid output/reabsorption
Respiratory alkylosis: ?PCO2 (hypervent) Renal: ?renal net acid output/reabsorption
Human Organism – Psychiatry Objectives
08-20-03
Human Behavior: An Introduction to Psychiatry
HO Psych1 Psych2
Drs. Signorelli, Horton
1. Define psychiatry and describe the typical scope of practice of a psychiatrist
• ?The branch of medicine that focuses on the diagnosis and treatment of mental disorders
• ?Primary focus is assessing and addressing problems in what humans think (cognition),
feel (emotions), and do (behavior)
2. Enumerate the diagnostic methods used for psychiatric diagnosis
• Clinical Intervie w
• Mental status examination (Standardized Pts-ICM)
• History
• Physical Examination
• Laboratory Studies
• Neuroimaging studies
• Psychological assessment
•
Transference- the feelings a
patient has toward their
caregiver
•
Countertransference- the
feelings you, the healthcare
provider, have towards your
patients
SIGE CAPS M
5 symptoms within 2 weeks
Sleep
lack or too much
Interest
lack of anhedonia
Guilt
hopelessness help/worth –lessness
Energy
usually decreased, sometimes increased (nervous, fidgety)
Concentration
memory
Appetite
increased or decreased
Psychomotor Activity speech, movements, thoughts
Suicide
Mood
3. List the commonest modalities used to treat psychiatric disorders
Psychotherapy
• Short term, long term
• Analysis
• Insight oriented
• Cognitive Behavior
• Supportive
Psychopharmacology/biological therapies
• Antidepressants
• Antipsychotics
• Mood stabilizers
• Benzodiazepines
• ECT (electroconvulsive therapy)
Environmental support
4. Define the Bio-Psycho-Social Model and be able to apply it
Biopsychosocial Model of Psychiatry
• Biological Aspects
– Constitution (personality)
– Neuropsychiatric disorders
– Medical conditions
– Pharmacologic interventions
HO Psychiatry : Page 1 of 9
HO Psych1 Psych2 page 2
• Psychological aspects
– Family functioning and dynamics
– Individual capacities
– Experience
– Personality
• Social aspects
– Culture
– Role in society
– Financial, occupational, support systems, etc
– Treatment setting
5. Discuss the patient-healer relationship in the context of culture.
Our medical beliefs and practices are part of another cultural system you are about to join
a. Western Biomedical Culture
A. illness explained in impersonal terms
B. medical rituals to reduce anxiety in the face of uncertainty and to reduce errors
C. emphasis on therapeutic activism
D. professional autonomy
E. emphasis on technique, over which physician has some control, rather than results,
over which have less control
b. Non-Western concepts
A. diagnosis involves the world of ancestral spirits or current social conflict
B. physician/healer may have influence but not control
C. illness explained in personal terms
D. typically addresses symptoms instead of seeking causes
6. Delineate the cultural determinants for normality and abnormality and the ranges of normal behavior in
human societies
• When is a behavior defined locally as abnormal or pathological?
– Breaking a code of social expectation, cultural taboo or political view
• Clinically distinguishing normality from pathology
– 1) By professional definition
– 2) By deviation from the mean
– 3) By assessment of function
– 4) By social definition
– Tseng, Handbook of Cultural Psychiatry, 2001:443-444
7. Discuss examples of the role of culture in adaptation and maladaptation to the environment
• Ex. Somatization versus psychologization
• Culture-based diagnoses
• Ex. “Dhat” (physical illness defined in Indian culture as caused by loss of too much semen)
• Culture-based treatments
Primate Behavior
• Techniques for Peaceful Coexistence
o Reassuring and Calming Behaviors
• Treatment of blind and physically impaired infant monkeys
• Adoption of orphaned monkeys and apes
• The Bonobo Chimps
o Peace and Love, Not War
• Attachment and need for nurturance
o Harry Harlow’s Monkeys; attachment disorders
HO Psych1 Psych2 page 3
8. Describe the relevance of research in primatology to psychiatry in conceptualization of
attachment and aggression
Evolutionary Origins
• Psychiatry and Anthropology: The Long View
• Primates as our closest living relatives
– Similar Social and ecological challenges
• Reasoning by Analogy
– Morphological similarities
• Reasoning by Homology
• Aggression and Peacemaking Among the Apes
– Chimpanzees have capacity for cooperation
• Sociability has its costs
– Chimpanzees have capacity for aggression
• Potential for unprovoked and violent attacks; infanticide
8/27/03
Human Life Cycle 1: Child and Adolescent Development
HO Psych3
Dr. S. Turkel
1. Define and use in context the following terms:
attachment – being bound to some object by strong and lasting ties
latency - The period from about 5 to 7 years to adolescence when there is an apparent cessation of
psychosexual development
object constancy - The tendency for objects to be perceived as unchanging despite variations in the
positions in and conditions under which the objects are observed;
stranger anxiety - the fear of strangers that infants commonly display, beginning by about 8 months of age
puberty - the period of sexual maturation, during which one first becomes capable of reproducing
adolescence - the transition period from childhood to adulthood, extending from puberty to independence
antepartum – before birth, peripartum – during birth, postpartum – after birth
perinatal cognitive - Newborn responses and capacities
temperament - The manner of thinking, behaving, or reacting characteristic of a specific person
psychoanalytic model - a subcategory of the pathology model which holds (1) that the underlying
pathology is a constellation of unconscious conflicts and defenses against anxiety, usually rooted in
early childhood, and (2) that treatment should be by some form of psychotherapy based on
psychoanalytic principles.
transactional model - a model of child development that emphasises the complex dynamic interplay
between biological factors within the child and the caretaking environment. This transactional model
postulates that developmental outcomes are the end result of a complex transaction btn intrinsic or
within child factors (eg. genes, central nervous system development, temperament) & environmental
factors (eg. parenting style, amount of stimulation, socio-economic status).
2. Describe the major developmental milestones of infancy and childhood in terms of:
age at transition
cognitive and motor function
typical behavior
interpersonal relationships and skills
1)
2)
3)
0-2
a)
b)
c)
d)
2-8
•
•
•
•
months
bonding and attachment
interactions and attention
beginnings of memory
myelination within nervous system causes jump in development
months
Cognitive, social and behavioral changes which change relationships and interactions with environ
Motor development: lift head > sit > grasp > release (sensory-motor cognitive function) > transfer
Social interactions increase (with sitting up) (no sense of who these people are)
Temperament: innate differences in infant's behavior which contribute to personality development
a. Easy temperament
b. Slow to warm (anxious, shy)
c. Difficult temperament
8-12 months
• Transition at 7-9 months with myelination of limbic system
• Cognitive development
a. Object permanence (pen stay put)
b. Intentional communication: expressive language begins
c. Problem solving begins (peek-a-boo)
d. Early symbolism: receptive language begins (what understand when people talk to you.
Before expressive)
e. Improved memory and sense of future
HO Psych3 page 2
• Social and emotional development
a. Affect sharing (ability have expression. Mean a feeling. If child can see, his first words will
be nouns; people. If blind, first words are verbs)
b. Social referencing (no sense of who these people are)
c. Separation distress (know daily routine. Adapt, wait for mom to get home)
• Motor development: sit up > crawl > stand > step > cruise > walk
4) 12-18 months
• Transition at 12-13 months
a. Complex symbolic representation: word = object
b. Improved imitation
a. Special and specific attachment to different individuals: recognition of self and family
b. Complex social referencing
c. Attempts to comfort
d. Attachment theory: practicing and exuberance
• Commu nication and object relationships
• Play
5) 18-36 months
• Transition at 18-20 months
a. Improving symbolic representations
b. Ability for simple abstraction
c. 300 words at 2 years > 1000 words at 3 years
d. Early problem solving and begin to think before do
e. Begin conceptualize rules and events
f. Ordering and sequencing (know what to expect. stories)
a. Limited interaction with peers
b. Willfulness, negatism, self-assertion
c. Increasing self reliance and limited self care
d. Relationships with siblings
• Terrible Twos: very opinionated, stubborn, assert-selves.
3. Describe the stages of adolescence (preadolescence, early adolescence, middle adolescence, and late
adolescence) in terms of:
age range
sexual development and typical behavior
cognitive function
interpersonal relationships with family and friends
IV. The school age child
A. Preschool: ages 3 to 6 years (Magic years, pretend)
1. Improving motor skills, cognition and memory
2. Socialization
3. Magical thinking, guilt, rules
4. Increasing self reliance and self care
5. Experiences separate from family (nursery school. Peers attend party. Self-reliant)
B. School age: ages 6 to 11 years
1. Motor skills and coordination smooth and integrated
2. Increasing capacity for thinking, memory, speech, conceptualization, & abstract
thought
HO Psych3 page 3
more time setting up rules than playing
expectations à rewards for meeting expectations
3. More successful management of impulses into more acceptable forms of gratification
“you hurt my feelings” instead of
BAM
V. The adolescent
A. Definitions
1. Puberty: physical manifestations of sexual maturity
2. Adolescence: psychologic process of adapting to sexual maturity
physical maturity vs. societal maturity
B. Considered time of recapitulation and opportunity for resolution of earlier conflicts and
problems in psychologic developmental process
C. Stages of adolescence
1. Preadolescent: ages 9 to 11-12 years
a. Before physical changes manifested
b. Emotional changes begin first (realize won’t be child forever)
c. Time transition from childhood begins
2. Early adolescent: ages 11-12 to 14-15 years
a. Onset of physical changes
b. Thinking becomes more abstract (cognition more complex. Sensory-motor
concrete. Not yet abstraction. Begin to think about nontangibles)
c. Importance of peer group (surrogate family)
d. Emotional separation from parents begins
3. Middle adolescent: ages 15-16 to 18-19 years
a. Physical changes essentially complete
b. Increasingly comp lex relationships
c. Time of sexual experimentation
4. Late adolescent: ages 16-17 to 20+ years
a. Phase of consolidation
b. Ability for abstract, logical thought
c. Development of internalized, individual code of ethics and conduct
d. Resolution of issues of independence and individual identity
4. List the major tasks of adolescence and describe how these tasks are usually accomplished.
1. Move from dependent to independent person
2. Establish an identity
3. Learn to relate as an adult, including ability for intimacy
9/4/03
The Medical Interview Workshop
Psych4
Dr. Spring
Incorporated in ICM workshop
9/8/03
Human Life Cycle 2: Adult Development
HO Psych5
Dr. Gross
1. Define Erikson’s Epigenetic Model of Development
Erik Erikson (1902-1994) child psychoanalyst
First psychologically integrated view of how an ind develops throughout life, growing out of the
interaction of both internal (psychological) and external (social) events
Epigenetic Principle: “game plan”
Predictable, non-random, stepwise process
Developmental issues for the individual in relation to the societal world
Moments of decision between progress and regression (push/pull). There are critical
decisions at each stage, “indichotomies”
2. Define Erikson’s Eight Stages of Man and briefly describe the psychological conflict associated with
each stage.
Eight Ages of Man
(1) Basic trust vs mistrust (birth-18mo) (freud-oral)
• Caring, nurturing usually with mom. Need consistency. Mistrust if parent not there
(2) Autonomy vs shame and doubt (18-36mo) (freud-anal)
• Autonomy – Toilet train. 1st psychologic attempt to control your body
(3) Initiative vs guilt (3-5yr) (freud-phallic)
• Interpersonal world expanding. Child takes initiative to compete with dad for mom’s
attention. Increase more force in triad. Superego. Conscious start to develop. Child
overpunish self
(4) Industry vs inferiority (6-12yr) (latency)
• Elementary school kids learning lots of skills. How to support kids with difficulties
(5) Identity vs role confusion (13-20yr) (genital)
• Develop identity in many arenas. Role change. Physiological changes.
(6) Intimacy vs self-absorption/isolation (20-40yr)
• Go in knowing some sense. Interpersonal intimacy and commitment. Danger of
isolation if cannot connect. Another individual is as important as you are. Develop
ability to have truly intimate relationships. May lose yourself. Not protecting
yourself.
(7) Generativity vs stagnation (40-60yr)
• Facilitation of next generation. Contribute back to society. Not just procreation.
(8) Integrity vs despair (60+yr)
• Ability to look back on life. Know who you are, what you do and be okay. Face the
inevitable end of life.
3. Define “Mid -life Crisis” and list six areas in a person’s life which may involve this mid-life transition.
Period of reappraisal in many areas of life. (40-45 years)
1) bodily changes
decrease biological/physiologic function. climacterium
2) changes in time perception
“how long have I lived” à “how much time do I have left”
3) career changes
4) relationship changes: spouse, children, parent
“in the middle” take care of kids & parent
5) social changes
6) financial pressures
educate kids. Take care of parents
HO Psych 5 page 2
4. Discuss the relationship between age and timing regarding smooth vs. tumultuous transitions in adult
development.
Bernice Neugarten
“Time, Age, and the Life Cycle”
timing: expected vs unexpected events
self-utilization. Accomplishments appropriate. Body monitoring. “time left to live” death
personalized interiority. “life review”
biologic timing
we’re living longer now
social timing
entry delayed. Exit early.
individualized “normal, expectable life cycle:
how am I doing for my age? No set phases.
5. Discuss the major biological, psychological, and social issues characteristic of early, middle and late
adulthood.
Bio-Psycho-Social aspects of adult development
A. Early adulthood
1. Bio: peaking of biological development
2. Psycho: intimacy vs self-absorption/isolation
3. Social: assumption of major social roles
a. Education
b. Occupation
c. Marriage
d. Parenthood
B. Middle adulthood
1. bio: climacterium (gradual decline)
2. Psych: generativity vs stagnation (menopause)
3. Social: re-evaluation of roles
a. Relationships/career
b. Divorce
c. Mid-Life Crisis
C. Late adulthood
1. bio: aging (senescence)
aging of cells. Physiological aging.
2. psycho: integrity vs despair
3. social: economics, retirement, social/sexual activity
4. General themes
a. Increased vulnerability to stress
b. Dealing with loss

Core I Objectives - Three-Dimensional Orthopaedic Animations

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