Download biochem 44 [4-20

Biochemistry Chapter 44
Learning Objectives
1. Which cells are granulocytes?
Neutrophils, Eosinophils, Basophils
2. Which cells are mononuclear leukocytes?
T-Cells, B-Cells, NK-Cells
3. How does Flow Cytometry work in a CBC?
Cells flow through the machine where a laser shines light at the cell, which leads to a predictable
pattern of light scattering and absorbance depending on the cell type. The machine keeps track
of the results of each cell and records the total number of red cells, amount of hemoglobin,
hematocrit, MCV, total white blood cell count, and differential of all the white cells.
4. What is the energy production pathway in the erythrocyte? What is the role of 2,3-BPG
Glycolysis, which uses the rapaport-luebering shunt to generate 2,3-bisphosphoglycerate. 2,3BPG is a modulator of oxygen binding to hemoglobin that stabilizes the deoxy form of
hemoglobin.
5. What state must hemoglobin’s iron be in to bind oxygen? How does it get to that state?
The iron of hemoglobin must be in the ferrous state (+2). Reactive oxygen species can oxidize
the iron to the ferric state (+3) producing methemoglobin. But, it is converted back to the O2
binding state by the NADH-cytochrome b5 methemoglobin reductase system, in which
cytochrome b5 reduces the Fe3+ of methemoglobin.
6. What determines the lifetime of the RBC?
G6PD (glucose 6-phosphate dehydrogenase) activity. This enzyme catalyzes the first step in the
hexose monophosphate shunt. Because RBCs lack ribosomes, they can’t produce more of it, so
as G6PD activity decreases, oxidative damage accumulates, leading to lysis of the erythrocyte.
7. Why does an inherited deficiency in pyruvate kinase lead to hemolytic anemia?
The amount of ATP formed from glycolysis is decreased by 50% in the red blood cell with
pyruvate kinase deficiency. This means that the cell ion ransporters can’t function effectively, so
the RBC tends to gain Ca2+ and lose K+ and water. Water loss causes internal viscosity of the
cell to increase to the point that the cell becomes rigid and susceptible to damage. Once
damaged, the red blood cells are removed from circulation.
8. What is congenital methemoglobinemia?
The presence of excess methemoglobin, found in people with enzymatic deficiency in
cytochrome b5 reductase or people who have inherited hemoglobin M. In hemoglobin M, a
single amino acid substitution in the heme-binding pocket stabilizes the ferric oxygen.
Individuals with this may appear cyanotic, but few have clinical problems.
Methemoglobinemia can also be caused by ingestion of oxidants like nitrates, quinones, aniline,
and sulfonamides. It is treated by administration of reducing agents, like ascorbic acid or
methylene blue.
9. Describe G6PD Deficiency?
It is the most common enzyme deficiency in humans. Individuals with G6PD deficiency are
resistant to malaria. These red cells have a shorter life span and are more likely to lyse under
conditions of oxidative stress.
10. Pyridoxine (Vit B6) deficiency is associated with microcytic, hypochromic anemia. Why?
In Vitamin B6 deficiency, the rate of heme production is slow because the first reaction in heme
synthesis requires pyridoxal phosphate. Thus, less heme is synthesized, causing blood cells to
be small and pale.
11. How does lead poisoning affect heme synthesis?
In lead poisoning, the enzyme δ-ALA dehydratase and ferrochetalase are inactivated, so δ-ALA
and protoporphyrin IX accumulate, and the production of heme is decreased. This leads to lack
of hemoglobin, loss of energy production (because of lack of cytochromes for electron transport
chain).
“Think: Romans using lead to make their water pipes. “
12. What are porphyrias?
This is a group of rare inherited disorders resulting from deficiencies of enzymes in the pathway
for heme biosynthesis. Intermediates of the pathway accumulate and may have toxic affects on
the nervous system causing neuropsychiatric symptoms. Porphyrinogens may be converted by
light to porphyrins, which react with O2 to form oxygen radicals. This may cause severe damage
to the skin and thus individuals are photosensitive. There may also be scarring and increased
growth of facial hair in some porphyrias.
13. How and where is iron stored? How is it transported?
Stored in most cells but especially: liver, spleen, and bone marrow. It is stored using ferritin.
When excess iron is absorbed from the diet, it is stored as hemosiderin, a form of ferritin
complexed with additional iron that can’t be readily mobilized.
Transported in the blood as transferrin (+3).
14. Describe the breakdown and products of heme degradation:
Hemoglobin phogocytosed. Globin is cleaved to constituent amino acids and iron is returned to
body’s iron stores.
Heme oxidized  biliverdin (reduced to)  Bilirubin (transported to liver). Bilirubin conjugated
to glucuronate to form bilirubin monoglucuronide, which is converted to bilirubin diglucuronide.
This is excreted into bile. Bacteria deconjugate bilirubin digluronide and convert it to
urobilinogens. Some of this is excreted in urine. Other is oxidized to urobilins like stercobilin 
excreted in feces (gives it brown color)
15. What do you suspect in a man eating a Western diet who has iron-deficiency anemia?
Bleeding from GI tract as a result of ulcers or colon cancer
16. What happens when drugs like phenobarbital induce cytochrome P450?
What happens to heme specifically?
Because heme is used for synthesis of cytochrome p450, free heme levels fall and δ-ALA
synthase is induced to increase the rate of heme synthesis
17. In an iron deficiency, what characteristics will the blood exhibit?
Microcytic, hypochromic anemia
18. What is the role of spectrin, band 3, ankyrin, and band 4.2 in the membrane?
Spectrin cytoskeleton is connected to the lipid bilayer by ankyrin, which interacts with betaspectrin and integral membrane protein, band 3. Band 4.2 stabilizes the connection. #s 2-3-4
19. Why do defects in erythrocyte cytoskeletal proteins lead to hemolytic anemia?
Shear stresses in circulation result in the loss of pieces of the red cell membrane. As the
membrane is lost, the red blood cell becomes spherical and is more likely to lyse in response to
mechanical stress.
20. What is the role of 2,3-bpg?
It binds to hemoglobin in the central cavity formed by the four subunits, increasing the energy
required for conformational changes that facilitate binding of oxygen. So, it lowers the affinity
of hemoglobin for oxygen.
21. Describe the Bohr Effect of Proton Binding in the Lungs, Tissues?
Tissues: dissociation of carbonic acid (at low pH) produces protons that react with amino acid
residues in hemoglobin, causing conformational changes that promote release of oxygen. (The
binding of protons by hemoglobin lowers its affinity for oxygen).
Lungs: oxygen binds to hemoglobin, causing a release of protons, which combine with
bicarbonate to form carbonic acid.
22. How are hematopoietic cells isolated?
Using fluorescence-activated cell sorting, based on the expression of cell-surface markers.
Increasing the population of stem cells in cells used for bone marrow transplantation increases
the chances of successful transplant.
23. What is leukemia?
Malignancy of the blood, arising when a differentiating hematopoietic cell does not complete its
developmental program but remains in an immature, proliferative state. Leukemias have been
found in every hematopoietic lineage.
24. What is X-linked SCID?
Severe Combined Immunodeficiency Disease: circulating T-cells are not formed, and B-cells are
not active. The affected gene encodes the gamma-chain of the IL-2 receptor. Mutant receptors
are unable to activate JAK3 and the cells are unresponsive to the cytokines that stimulate
growth and differentiation.
25. What is SHP-1? What does a mutant epo receptor that can’t bind SHP-1 cause?
SHP-1 is a tyrosine phosphatase found in hematopoietic cells. It is necessary for proper
development of myeloid and lymphoid lineages. Its function is to dephosphorylate JAK2,
inhibiting it. Normally it is activated due to binding of growth factor-type cytokines.
Individuals with mutant epo receptor have a higher than normal percentage of RBC in circulation
because the receptor can’t be inactivated by SHP-1. EPO causes sustained activation of JAK2
and STAT5 in these cases. SOCS and PIAS have similar effects.
26. What happens with perturbed JAK/STAT signaling?
Development of lymphoid and myeloid leukemias, severe congenital neutropenia, Fanconi
anemia (bone marrow failure and increased susceptibility to malignancy)
27. Why does sickle cell disease cause gallstones?
The amount of unconjugated bilirubin that is transported to the liver increases. If the
concentration exceeds capacity of the hepatocytes to conjugate it, both the total and
unconjugated bilirubin levels in the blood rise. The increase in unconjugated bilirubin (not-very
water soluble) results in its precipitation within the gall-bladder lumen, leading to formation of
gallstones.
28. How does HbC affect heterozygotes with HbS?
These heterozygotes show significantly more hematopathology than individuals with sickle cell
trait (HbA/HbS). Polymerization of deoxygenated HbS is dependent on HbS concentration
within the cell. The presence of HbC in the heterozygote increases the HbS concentration by
stimulating K+ and water efflux from the cell, and also makes the Hb more likely to precipitate
29. Describe severity of alpha thalassemias:
Mutations resulting in one copy of the gene deleted, the size and hemoglobin concentration is
minimally reduced. Two copies deleted, red blood cells are decreased in size and reduced
hemoglobin concentration – usually not anemic. Three copies deleted: moderately severe
microcytic hypochromic anemia with splenomegaly. Four copies deleted: hydrops fetalis. HbH
30. Describe severity of beta thalassemias:
Heterozygotes for B+ (some synth.) or B0 (no synth) are generally asymptomatic, though they
have microcytic, hypochromic red blood cells and may have mild anemia. B+/B+ homozygotes
have anemia of variable severity, B+/B0 heterozygotes are more severly affected, and B0/B0
homozygotes have severe disease. Alpha chains do not form stable tetramers, and so they
precipitate at every RBC developmental stage.
31. Give two scenarios in which you might get 2 alpha-globin genes, and populations they are
common in:
One copy of chromosome 16 lost both alpha globin genes – Asians
“asia is bigger”
Each chromosome may have lost one of their two copies – Africans
32. What is the difference between HbA and HbF?
The gamma chains of HbF results in changes that cause it to have a lower affinity for 2,3-BPG,
and thus a greater affinity for oxygen.
33. What is HPFH?
Hereditary persistence of fetal hemoglobin. Nondeletion form may have an ameliorating effect
on beta-thalassemia mutations. Deletion forms result from deletion of entire delta and beta
genes.
34. Give an additional source of variation in the level of fetal hemoglobin?
Some variation is due to the FCP (F-cell producing) locus on the short arm of the X chromosome,
in a region thought not to be susceptible to X inactivation. A high level of hemoglobin F appears
to be inherited as an X-linked dominant trait.