1.1 Red Blood Cells

Question 1

Red blood cells

• Red blood cells are ______________ 7.5 x 2 μm. This shape maximizes the surface area / volume ratio, allowing more effective gas exchange. The discs are also ____________, allowing passage through small blood vessels such as capillaries.
• RBC (erythrocytes) are the most common type of blood cell and the principal means of delivering O2 to tissues via the blood, transporting carbon dioxide from the tissues to the lung, and help regulate extracellular fluid pH.
• RBCs contain hemoglobin, a molecule that binds to oxygen. The red color of blood is due to the color of oxygen-rich hemoglobin.
• RBCs develop in the bone marrow and live for about ________________
• They are flexible biconcave discs and lack a _______________________ and they cannot synthesize proteins. RBCs use anerobic metabolism as an energy source.
• There are a total of ______________________ in the body containing 1 kg of hemoglobin.
• The precursors of mature RBCs are reticulocytes. They are formed in the bone marrow, enter the blood circulation, remain there for a day before differentiating into mature RBCs. These cells are _____________. Reticulocytes possess ribosomes for _____________, and mitochondria to allow ___________________________.

Answer 1

non-nucleated biconcave discs;

deformable;

120 days;

nucleus, mitochondria, ribosomes,

3 x 1013 RBCs (30 trillion);

non-nucleated;

globin synthesis;

oxidative metabolism and heme synthesis

Question 2

What is hematocrit?

Answer 2

Volume occupied by RBCs in 100ml of blood

Question 3

What is mean corpuscular volume (MCV)?

Answer 3

average volume of single RBCs

Question 4

What is mean corpuscular hemoglobin (MCH)?

Answer 4

average mass of hemoglobin in each RBC

Question 5

What is mean corpuscular hemoglobin concentration (MCHC)?

Answer 5

Average concentration of hemoglobin in a single RBC

Question 6

What is normal mcv?

Answer 6

83-99fL

Question 7

what is normal MCH?

Answer 7

28-32 pg

Question 8

what is normal MCHC?

Answer 8

32-36 g/dL

Question 9

Hematopoiesis before and after birth

• 1-3 months fetal life (first trimester): Blood cells arise in the _______________________.
• 3-7 month fetal life (second trimester): Blood cells arise primarily in the _______, with the ____________ contributing.
• 7-9 month fetal life (third trimester): As bone cavities develop, stem cells migrate into the bone marrow, and hematopoiesis begins in the bone marrow.
• After birth: Hematopoiesis occurs primarily in the bone marrow, mainly in the _______________________. Hematopoiesis occurs initially in long bones (e.g. _______________), but as walking begins, these bones become weight bearing and solid, so hematopoiesis in these bones decreases.

Answer 9

mesoderm of the yolk sac;

foetal liver; foetal spleen

pelvis, vertebrae, sternum and ribs;

tibia and femur

Question 10

Intramedullary versus Extramedullary hematopoiesis: In adults, hematopoiesis occurs primarily in the bone marrow (intramedullary hematopoiesis). In severe and chronic anemia, the body compensates by initiating additional hematopoiesis in the ___________________ (extramedullary hematopoiesis).

Bone marrow transplant: Bone marrow is removed from the _____________ of the donor under general anesthesia and then transfused into the blood circulation of the recipient. The cells home to the bone marrow of the recipient, colonize the bone marrow and begin hematopoiesis. The donor and recipient must be _________________ to ensure that graft-versus-host disease does not occur.

Stem cell transplant: Pluripotent stem cells are isolated from the bone marrow by first depleting mature and immature blood cells using _______________, and then enriching for CD34+ stem cells. CD34 is a glycoprotein found on the cell surface, which may contribute to the attachment of stem cells to the __________________. The stem cells are transfused into the blood circulation of the recipient. They colonize the bone marrow and initiate hematopoiesis.

Answer 10

liver and spleen ;

pelvis (Iliac crest);

HLA-matched;

lineage-specific monoclonal antibodies;

stroma/matrix in the bone marrow;

Question 11

Pancytopenia

• Decreased RBCs (Erythropenia): ________________
• Decreased WBCs (Leukopenia): frequent infections, __________________
• Decreased Platelets (Thrombocytopenia): mucosal and gingival bleeding, purpura, ___________, __________

Answer 11

anemia;

mouth & pharyngeal ulceration;

petechia, ecchymoses

Question 12

What is the pathway to be a RBC?

Answer 12

CD34 –> CFU-GEMM –> BFU-E –> CFU-E –> Proerythoblast –> Basophil erythoblast –> Polychromatophil erythoblast –> Orthochromatic erythoblast –> reticulocyte –> Erythrocyte (RBC)

Question 13

Pathway to be a neutrophil?

Answer 13

CD34 –> CFU - GEMM –> CFU- GM –> CFU- G –> neutrophil

Question 14

Pathway to be a monocyte?

Answer 14

CD34 –> CFU-GEMM –> CFU- GM –> CFU-G –> Neutrophil

Question 15

Pathway to be to be a basophil?

Answer 15

CD34 –> CFU-GEMM –> CFU- Bas –> Basophil

Question 16

Pathway to be an eosinophil?

Answer 16

CD34 –> CFU- GEMM –> CFU- Eos –> Eosinophil

Question 17

Three types of pluripotent stem cells that both self-renew (replenish themselves) and ___________________.
Pluripotent stem cell
- _____________ (Colony forming unit – Granulocyte, Erythroid, Monocyte, Megakaryocyte) – give rise to RBCs, neutrophils, monocytes, platelets, basophils and neutrophils.
-____________(Colony forming unit - Lymphoid) – gives rise to lymphocytes.

Answer 17

differentiate into multiple blood lineages;

CFU-GEMM;

CFU-L

Question 18

Committed stem cells: These cells do not self-renew and are committed to a particular blood lineage, examples are ??

Answer 18

BFU-E (Burst-forming Unit-Erythroid), CFU-E (Colony Forming Unit-Erythroid) CFU-GM (Colony Forming Unit – Granulocyte-Monocyte), CFU-Meg (Colony Forming Unit – Megakaryocyte), CFU-Bas (Colony forming unit – Basophil), CFU-Eos (Colony Forming Unit – Eosinophil), CFU-G (Colony Forming Unit – granulocyte), CFU-M (Colony Forming Unit – Monocyte) .

Question 19

Growth factors: A complex hierarchy of growth factors drives hematopoiesis.

• Recombinant GM-CSF (Granulocyte-Monocyte Colony Stimulating Factor, Leukine) is approved for clinical use to accelerate hematopoiesis following autologous BM transplant. GM-CSF acts on many cells in the lineage pathway and induces the differentiation of ___________________________
• ______________ is clinically approved to drive RBC development.
• ___________________ is used to drive granulocyte differentiation. It acts on CFU-G and to some extent on BFU-E

Answer 19

CFU-GEMM, BFU-E, CFU-GM, CFU-Meg, CFU-Bas and CFU-Eos;

Recombinant EPO (erythropoietin;

Recombinant G-CSF (Granulocyte Colony Stimulating Factor, Filgrastim)

Question 20

ome definitions

• _______________: decreased numbers of white blood cells, red blood cells and platelets.
• ________________: decreased white blood cells.
• __________________: decreased neutrophils.
• Monocytopenia: decreased monocytes.
• Eosinopenia: decreased eosinophils.
• _________________: decreased basophils.
• ________________: decreased lymphocytes.
• ________________: decreased platelets

Answer 20

Pancytopenia;

Leukopenia;

Neutropenia or granulocytopenia;

Basocytopenia (basopenia);

Lymphopenia;

Thrombocytopenia

Question 21

Some causes of Pancytopenia

• Classical aplastic anemia: Failure of _________________ causes pancytopenia.
• Autoimmune: autoantibodies destroy ____________.
• Microbes (_________________) damage stem cells.
• Radiation & Chemotherapy destroy stem cells.
• Fanconi’s Anemia is a rare genetic disease due to mutations of at least 15 different genes. Patients have pancytopenia as well many other abnormalities.

Answer 21

pluripotent stem cells;

stem cells;

Hepatitis C, HIV, Epstein Barr virus, Parvovirus, Mycobacteria

Question 22

Erythropoiesis occurs in the bone marrow

• Burst forming Unit-Erythroid (BFU-E) are erythroid stem cells that exhibit low sensitivity to the growth factor ______________ and differentiate into ______________________. CFU-Es respond to erythropoietin and give rise to erythroblasts.
• Hemoglobin is synthesized during the _____________________.
• _____________________ takes place in the bone marrow at the last erythroblast stage. The cells reseal themselves after expulsion of nuclei. Losing the nucleus unloads dead weight and transforms the RBC from a rigid spheroidal cell to a supple biconcave cell that can navigate the blood stream.
• Reticulocytes are immature RBCs that leave the BM and enters the blood stream. They stay in the blood for 1 day before maturing into RBC. Reticulocytes contain _______________ which stain with supravital dyes giving a reticular pattern, hence the name reticulocyte. The rate of RBC production can be determined by assessing reticulocyte levels in the blood. Decreased reticulocytes suggests decreased hematopoiesis in the bone marrow, while increased reticulocytes suggests that the bone marrow has increased hematopoiesis, often as a compensatory response to ___________.

Answer 22

erythropoietin; Colony forming unit-erythroid (CFU-E) cells

erythroblast stages;

Denucleation;

cytoplasmic RNA;

anemia

Question 23

EPO or Erythropoietin (glycoprotein) is a hormone made primarily by the kidney in response to cellular hypoxia. Decreased oxygenation causes EPO synthesis and release within minutes, peaking in 24 hours. EPO-producing and oxygen-sensing cells are located in the ________________. EPO stimulates erythropoiesis by binding to EPO receptors on CFU-E and erythroblasts. This triggers a _________________, which leads to differentiation, survival and proliferation of the erythroid cell. Low levels of EPO are constantly secreted that are sufficient to compensate for normal RBC turnover.

Stimuli that trigger the oxygen sensor and induce EPO release

• Decreased cardiopulmonary function due to heart failure or pulmonary fibrosis or emphysema results in reduced oxygen uptake from the lung.
• Decreased hemoglobin concentration due to anemia results in less oxygen carrying capacity.
• Abnormal hemoglobins with ______________ hold on to the oxygen more tightly and less is released in tissues.
• Decreased blood volume e.g. after a bleed.
• Decreased oxygen tension e.g. at high altitude.
• Alkalosis

In chronic renal failure, less EPO is made by the kidney. The EPO made by the liver is not adequate to sustain erythropoiesis. The result is a____________________. These RBCs have a slightly shorter survival time. The anemia of chronic renal failure is treated with recombinant EPO.

Some carcinomas of the kidney or liver synthesize and release excessive EPO, even when there are no adequate environmental stimuli. These patients have _____________ (increased RBC counts, increased hematocrit).

EPO Doping: Some athletes (e.g. Floyd Landis, the cyclist) inject themselves with recombinant EPO to increase the RBC mass and thereby enhance their oxygen-carrying capacity.

Answer 23

juxtamedullary cortex;

JAK2 signaling cascade;

increased oxygen affinity;

normocytic and normochromic anemia;

polycythemia

Question 24

[iron]
- Hemoglobin is made of _______________________. Iron is required at the final stage of heme synthesis. An iron atom is part of each heme molecule.

• Iron is absorbed in the duodenum. It is reduced from ____________ to ___________ and transported across the apical surface of the duodenal enterocyte. It is then transported across the basolateral surface of the enterocyte, oxidized to ferric, bound to ______________, and transported via the circulation to tissues where it is utilized.
• _______________ is a membrane protein that transports ferrous from the inside of duodenal enterocytes across the basolateral surface of these cells to the outside. Ferrous is oxidized to ferric by ferro-oxidase, and ferric is then bound to plasma transferrin and transported in the blood to tissues for usage. Ferroportin is also present on the surface of other cells which use or store iron including __________________________.

_______________ (protein) produced by the liver inhibits iron uptake by blocking ferroportin. Hepcidin’s inhibitory effect on ferroportin ensures that excessive iron is not absorbed or released into the bloodstream.

Erythroferrone: Erythroferrone is a hormone produced by erythroblasts in the bone marrow in response to a trigger by ______________. Its main function is to suppress _____________-. By doing this, it removes hepcidin’s inhibitory effect on ferroportin and allows more iron to be absorbed and/or enter the bloodstream (e.g. from the liver).

Answer 24

heme and globin chains;

ferric, ferrous; serum transferrin

Ferroportin;

hepatocytes, macrophages and adipocytes;

Hepcidin;

erythropoietin; hepcidin

Question 25

Following a bleed (hemorrhage) a number of changes occur in the body to ensure that more red blood cells are produced to replace the cells that are lost.

• Increase __________________.
• EPO increases ___________________.
• Erythroferrone suppresses __________________.
• Reduced hepcidin results in reduced inhibition of iron absorption from the _________ via ferroportin.
• More iron absorption results in increased iron in the blood.
• More Iron is incorporated into heme.
• More RBCs are produced to replace the ones lost by bleeding.

Answer 25

erythropoietin (EPO) release;

erythroferrone release;

hepcidin production;

duodenum

Question 26

Causes of iron deficiency anemia

• Diet: Inadequate consumption of iron in the diet.
• Decreased absorption: Gastric acid is required to release iron from food and helps to keep iron in the soluble ferrous state. ______________ (reduced gastric acid) in the elderly or ____________ (chronic inflammation of the gastric mucosa leads to atrophy and less acid production) may decrease iron absorption, leading to iron deficiency anemia. Iron is mainly absorbed in the duodenum and upper small intestine. ___________, which affects this region of the GI tract may lead to less iron absorption and iron deficiency anemia.
• Increased utilization: During ___________________, and during growth spurts.
• Increased loss: Chronic blood loss in individuals with bleeding peptic ulcers, or women with excessive bleeding during menstruation, or individuals with hookworm infestation. Patients with bleeding peptic ulcers, alcoholic gastritis will often complain of passing __________________, which is caused by oxidation of iron in hemoglobin during its passage through the ileum and colon.

Answer 26

Hypochlorhydria;

atrophic gastritis;

Celiac disease;

pregnancy and lactation;

black stool (melena)

Question 27

Iron deficiency anemia

• Deficiency of iron results in less heme synthesis which in turns reduces hemoglobin and ______________ RBCs, a hallmark of iron deficiency anemia.
• Iron is required by other tissues, particularly in tissues with _________________ e.g. tongue, stomach, nails.
• __________ (reddened, swollen, smooth, shiny, tender tongue),
• _____________ (erosions at the corners of the mouth)
• Gastric atrophy
• ____________ (spoon-shaped nails)

Answer 27

Microcytic (small) hypochromic (pale);

rapid cellular turnover;

Glossitis;

Angular stomatitis;

Koilonychia

Question 28

Microcytic and hypochromic RBCs are characteristic of iron deficiency anemia and develop because of the following:

• Decreased iron leads to _____________, which in turn results in decreased hemoglobin.
• Hemoglobin is the main oxygen carrying molecule in blood. A decrease in hemoglobin results in a decrease in total arterial oxygen concentration.
• EPO levels rise to drive the BM to produce more erythroid progeny, but with each division, the failure of hemoglobin synthesis creates an increasingly altered _______________________.
• The end result is a non-viable or a barely viable RBC that is both pale (hypochromic) because of reduced hemoglobin, and small (microcytic). ____________________ are decreased in iron deficiency anemia.

Treatment is iron replacement. Since ferrous is absorbed better than ferric, iron tablets are ferrous. Iron injections are given when iron deficiency is severe.

Answer 28

reduced heme synthesis;

nuclear : cytoplasmic balance;

MCV and MCHC

Question 29

Vitamin B12 (Cobalamin) and folate

• Important for final maturation of RBCs.
• Essential for the synthesis of DNA, each in a different way being required for the synthesis of ______________, one of the building blocks of DNA.
• Lack of either vitamin causes diminished DNA synthesis and consequently leads to alterations in tissues with the greatest rate of cell turnover (e.g. hematopoietic and gastrointestinal systems, skin).
• Vitamin B12 stores in the body are large and daily utilization low, so it takes at least ________________ to deplete stores and cause clinically evident vitamin B12 deficiency.

Causes of Vitamin B12 deficiency

1. Dietary deficiency especially in pure vegans
2. Decreased absorption – stomach defect (Pernicious anemia, gastrectomy)
3. Decreased absorption - upper small intestine defect (pancreatic insufficiency, usurpation of vitamin B12)
4. Decreased absorption - terminal ileum defect (Crohn’s disease, ileal resection)

Causes of folate deficiency

1. Dietary deficiency
2. Over-utilization: pregnancy
3. Drugs: _____________________
4. Malabsorption: Celiac disease

Answer 29

thymidine triphosphate;

three - five years;

methotrexate, contraceptives, phenytoin

Question 30

Pernicious anemia

• Most common cause of vitamin B12 deficiency in developed countries.
• Mechanism – Autoimmune disease:
• Anti-parietal cell autoantibody → gastric atrophy → decreased _____ → Decrease Vitamin B12 absorption
• Anti-IF → autoantibodies → decreased absorption of ____________ in the terminal ileum.
• Associated with other autoimmune diseases: _________________), vitiligo, hypoparathyroidism, Addison’s disease

Answer 30

IF;

IF– Vitamin B12 complex;

Hashimoto’s thyroiditis, Grave’s disease (thyroid

Question 31

Cobalamin deficiency leads to folate deficiency: In cobalamin deficiency, _______________ is not converted adequately to _________________, the form of folate that gets conjugated to __________ and is retained in tissues. Consequently, there is leakage of folate out of the cell, leading to deficiency of tissue folate.

Megaloblastic anemia: The reduction in tissue folate leads to reduction in DNA synthesis (thymidine synthesis) relative to RNA synthesis (_______________ normal) resulting in unbalanced cell growth. In RBCs, the deficiency of tissue folate results in the formation of _____________, which are fully hemoglobinized RBCs that result from omitted cell divisions during erythropoeisis. The result is megaloblastic anemia.

Neurological findings: Seen in cobalamin deficiency, but NOT folate deficiency. Features include demyelination of the ___________________; glove and stocking paresthesia in extremities; reduced vibration sense in the extremities; dementia; optic atrophy; autonomic neuropathy causing sexual dysfunction, and bowel and bladder incontinence.

Effects on tissues that rapidly turn over: The inadequate DNA synthesis in rapidly growing tissues (e.g. tongue) leads to glossitis, which is characterized by a ___________________

Answer 31

N5-methyl-tetrahydrofolate;

tetrahydrofolate;

polyglutamates;

uracil levels;

macrocytes;
posterior columns and corticospinal tracts of the spinal cord;

blackened smooth sore tongue with atrophy of papillae

Question 32

Features of Vitamin B12 and Folate deficiency

• General: Weakness, shortness of breath, pallor, tachycardia.
• Blood: Megaloblastic anemia (macrocytic anemia), ______________ neutrophils (5 or more nuclear lobes; normal 3-4 lobes), Hb ____________, MCV ____________.

GI tract:

• Glossitis (Smooth sore tongue with atrophy of papillae, hyperpigmented – blackened tongue)
• nausea, epigastric pain, loss of appetite
• Skin: ______________
• Neurological: subacute combined neurodegeneration in vitamin B12 deficiency

Treatment

• Vitamin B12:
• Parenteral: Intensive therapy
• Maintenance: __________. per 3 months. Oral Vitamin B12.

Folate

• Folate supplements will rapidly normalize patient with only folate deficiency.
• Folate administration will correct hematological abnormalities of Vitamin B12 deficiency because decreased tissue folate contributes to these changes. Folate will NOT treat or stop the development of neurological complications
• If vitamin B12 + folate deficiency occurs simultaneously, replace _________ first to avoid neurological damage.

Answer 32

hypersegmented;

decreased;

increased;

hyperpigmented macular lesions.

1000 µg i.m;

vitamin B12

Question 33

Polycythemia: Increased hematocrit (>55%) due to increased RBCs or decreased plasma volume; increased ___________; clogging of ________, thrombotic complications, strokes, blindness.

• Physiological Polycythemia: High altitude living increases EPO and causes physiological polycythemia
• Secondary Polycythemia occurs when EPO rises in _________________ , EPO doping, or due to chronic hypoxia (e.g. lung or heart disease).
• Polycythemia Vera is due to over-production of RBCs because of clonal expansion of _________________. The abnormal erythroblasts may be hypersensitive to EPO.
• Hematocrit is the volume occupied by RBCs in ________________

Answer 33

blood viscosity;

capillaries;

renal or hepatocellular carcinoma

“abnormal” pluripotent stem cells;

100 ml of blood ~45%.

Question 34

1. Hemoglobin genes

Hemoglobin

• Hb, Hgb is an iron-containing oxygen-transport ___________ in RBCs.
• Normal levels are: Men: _____________; Women: ____________
• Hb is made up of four globin proteins and four heme molecules (each with a single iron). Each globin chain harbors one heme moiety. A single globin chain combined with a single heme is called a monomer of hemoglobin.
• The proteins, which are held together by polar bonds, are able to move with respect to each other to assume a taut (__________________) or a relaxed (_______________) form of the hemoglobin tetramer.

Globins:

• Each Hb molecule typically contains two chains belonging to the alpha globin family and two belonging to the beta globin family.
• The alpha globin gene family consisting of the zeta, alpha1 & alpha2 genes reside on ______________.
• The beta globin gene family consisting of the epsilon, GammaG, GammaA, delta and beta genes reside on ________________.

Chromosomal arrangement: The alpha and beta-globin gene clusters are arrayed on their chromosomes in the same 5’ to 3’ order that they are expressed during development.

• a family: z is located at the most 5’ end, followed by ___________________.
• b family: e is located at the most 5’ end, followed by _________________are at the 3’ end.

Answer 34

metalloprotein;

14 -16 g/dL;

12 -15 g/dL;

deoxyhemoglobin;

oxyhemoglobin;

chromosome 16;

chromosome 11

a2 and a1 genes;

gG and gA, and d and b

Question 35

2. Different types of hemoglobins

Hb Gower I (z2e2) (Embryonic hemoglobin): The zeta and epsilon genes turn on during the first trimester of pregnancy. The zeta and epsilon globin proteins combine to form Hb Gower I.

Hb Portland (z2g2) (Embryonic hemoglobin): The gamma genes turn on during the first trimester, peaking at the _________________. g globin proteins associate with z globin proteins to form Hb Portland.

Fetal Hb (a2g2): The alpha genes turn on during the first trimester and reach peak levels at the _______________. a globin proteins associate with g globin proteins to form Fetal Hb. gG or gA can be used.

HbA (adult Hb) (a2b2): The b globin gene turns on during the first trimester and reaches peak levels at about ________________. b globin proteins combine with a globin proteins to form HbA, the commonest adult Hb.

HbA2 (minor type of adult Hb) (a2d2): The d gene turns on during the _________________. d globin proteins combine with a proteins to form HbA2, a minor adult form of Hb.

Answer 35

beginning of the second trimester;

beginning of the second trimester;

6 months after birth; third trimester of pregnancy

Question 36

3. Alpha Thalassemia- abnormal Hbs

First three months in utero:
Normal because _______________________ made normally

Disease begins in the ______________________

• In alpha thalassemia, one to four of the alpha globin genes may be deleted or mutated. Consequently, the switch from Embryonic Hb (Hb Gower I and Hb Portland) to fetal hemoglobin (HbF) does not occur normally.
• Hb Barts (γ4): Due to loss of alpha genes in alpha thalassemia, free gamma chains form gamma chain tetramers called Hb Barts, which ___________________ and contributes to hypoxia. ________________ is seen in alpha thalassemias and NOT in beta thalassemia.

After birth:

• Because one to four alpha genes are deleted or mutated in alpha thalassemia, the formation of _______________________ are impaired or absent.
• HbH (β4): Due to loss of alpha genes in alpha thalassemia, free beta chains form beta chain tetramers called HbH, which are susceptible to oxygen denaturation leading to HbH precipitates in RBCs called __________.

Answer 36

embryonic hemoglobin (Hb Gower-1);

Second and Third trimesters in utero;

binds oxygen tightly;

Hb Barts;

adult HbA and minor adult HbA2;

Heinz bodies

Question 37

3. Alpha Thalassemia- abnormal Hbs

Carrier state: One of four a genes deleted. Carrier state is an advantage in an _____________.

a thalassemia minor (types 1 & 2): Deletion / mutation of 2 of the 4 a genes. Because there are 50% less a chains, the formation of fetal Hb (HbF), Adult Hb (HbA) and minor Adult Hb (HbA2) is impaired. Hb Barts (g4) is seen during the ___________________. Patients develop ____________.

___________________: 3/4 alpha genes are deleted / mutated. Because there are 75% less alpha chains available, the formation of fetal Hb (HbF), Adult Hb (HbA) and minor Adult Hb (HbA2) is impaired. Hb Barts forms in utero, HbH forms after birth and accounts for 5-30% of total Hb in these patients. Patients develop moderate to severe anemia.

________________ : All four a genes are deleted or mutated. Because there are NO alpha chains available, fetal Hb (HbF), Adult Hb (HbA) and minor Adult Hb (HbA2) do NOT form. Hb Barts accounts for >80% of Hb in these fetuses from the second trimester onwards.

A molecular epidemiologic study of thalassemia using newborns’ cord blood in a multiracial Asian population in Singapore (J Pediatr Hematol Oncol. 2004;26:817-9)
Chinese - 6.9% of newborns had a thalassemia mutations and deletions
Malays – 4.8% of newborns had a thalassemia mutations (none had deletions)
Indians – 5.2% of newborns had a thalassemia mutations (none had deletions).

Answer 37

endemic malaria area;

second and third trimesters in utero; mild anemia;

HbH disease (a thalassemia intermedia);

Hydrops Fetalis (Hb Bart’s disease)

Question 38

4. Beta Thalassemia

One beta globin gene per chromosome and two beta globin genes in the genome.

Beta-thalassemias are due to _______________ within beta-globin genes that result in errors of transcription, RNA processing or translation. Less frequently beta-thalassemia is caused by massive deletions. Since there are only two beta globin genes, patients can be heterozygous (beta thal minor) or homozygous (beta thal major) for the mutation.

Anemia develops after birth (unlike with alpha thalassemias when the disease begins after 1st trimester).

• 1st trimester in utero: The fetus develops normally because zeta and epsilon genes are not affected and hence embryonic hemoglobins (______________________) form normally
• 2nd and 3rd trimesters in utero: The fetus develops normally because the alpha and gamma genes are not affected and hence fetal hemoglobin forms normally.
• After birth: For the first few months after birth, there is sufficient fetal hemoglobin (HbF) to prevent anemia. Because the beta globin genes are defective in beta thalassemia, the switch from HbF to adult Hb (HbA) is impaired. In beta thal minor (one beta gene defective) less HbA is made, and in beta thal major (both beta genes defective) NO HbA is made. The patients develop severe anemia at this time. In compensation, the production of _____________________ increases to minimize the anemia.

Answer 38

point mutations, small insertions or small deletions;

Hb Gower I and Hb Portland

minor adult HbA2 and HbF;

Question 39

Beta thal major: These patients will have _________________; they can develop hemolytic anemia leading to jaundice due to increased RBC breakdown; they may have a ___________________ due to increased red blood cell production in the skull, maxilla, _____________________ because of increased red blood production in the tibia and femur, which become soft and bow-legged; Liver and spleen enlargement due to _____________________ (bone marrow is produced in the liver and spleen to compensate for inadequate production in the BM. They may develop ___________________ because of increased excretion of bilirubin through the bile ducts leads to formation of small bilirubin-containing pigment stones in the bile duct and gall bladder. They may develop cardiac failure because heart pumps faster until it fails.

Thalassemia intermedia refers to patients with beta thalassemia whom the clinical severity of the disease is somewhere between the mild symptoms of the β thalassemia trait and the severe manifestations of β thalassemia major. The diagnosis is a clinical one that is based on the patient maintaining a satisfactory hemoglobin (Hb) level of at least _____________ at the time of diagnosis without the need for regular blood transfusions. This form is not common in Singapore.

Hb Lepore: The Lepore globin gene results from a “crossover” between the d and b globin gene loci. This results in a hybrid protein in which the ___________ end of the d chain is fused to the _______________ end of the b chain.

_______________: one chromosome has normal d and b globin genes and the other chromosome has the d-b fusion. Individuals are asymptomatic.

_________________: both chromosomes have the d-b fusion. Patients develop severe anemia.

A molecular epidemiologic study of thalassemia using newborns’ cord blood in a multiracial Asian population in Singapore (J Pediatr Hematol Oncol. 2004;26:817-9)
Chinese - 2.7 % of newborns had b thalassemia mutations
Malays – 6.3% of newborns had b thalassemia mutations
Indians – 0.7% of newborns had b thalassemia mutations

Answer 39

microcytic hypochromic anemia;

Chipmunk facies;

Bow-legged;

extra-medullary hematopoiesis

gall stones;

6-7 g/dL;

amino;

carboxyl;

Hb Lepore trait;

Hb Lepore disease

Question 40

[Clinical features of thalassemia]

• _________________ due to decreased Hb production. A similar pattern is seen in iron deficiency because hemoglobin (heme) synthesis is deficient.
• Hemolytic anemia because some RBCs in thalassemias vary in shape and size and so the spleen destroys these abnormally shaped RBCs. Target cells are RBCs with ________________ .
• Jaundice due to increased RBC breakdown .
• Chipmunk facies due to increased red blood cell production in the _______________.
• Bow-legged because of increased red blood production in the _______________, which become soft and bow-legged.
• ____________________ due to extra-medullary hematopoiesis (bone marrow is produced in the liver and spleen to compensate for inadequate production in the BM).
• Gallstones due to increased excretion of bilirubin through the bile ducts leads to formation of small bilirubin-containing pigment stones in the bile duct and gall bladder.
• Cardiac failure because ________________________, until the heart fails.
• __________________ due to repeated blood transfusions leading to iron overload; brownish skin (iron deposition in the skin), elevated blood glucose (due to iron deposition in the pancreas), arthralgias (iron deposits in joints) and cardiomyopathy (iron deposits in the heart)

Answer 40

Microcytic hypochromic;

peripheral rim and central core of Hb;

skull and maxilla;

long bones;

Liver and spleen enlargement;

heart pumps faster to compensate for anemia by increasing oxygen delivery to tissues;

Secondary hemochromatosis

Question 41

6. Sickle Cell Anemia

A point mutation in β globin (bs) resulting in _______ (negatively charged) at position 6 being replaced by _______ (hydrophobic).

The mutant beta globin gene is called βs. Mutant βs protein is detected clinically by hemoglobin electrophoresis. There is survival value in carrying the sickle cell mutation in endemic malaria areas.

Sickle cell trait (HbAS)

• These individuals have _________________________.
• RBCs contain a mixture of adult HbA (α2β2) and abnormal HbS (α2βs2). Since α chains bind more effectively to β chains than βS, each RBC contains a _______ ratio of HbA and HbS.
• Cells sickle only when O2 saturation falls below 40% (e.g. hiking at high altitudes or in low pressurized planes.

Sickle cell disease (HbSS): These individuals have two βs globin genes. RBCs only contain HbS (α2βs2).

Polymerization and sickling: In the ______ state, the hydrophobic valine at position 6 in mutant bs permits intermolecular bonding between adjacent HbS molecules. Intermolecular bonding leads to the build-up of _____________________, which deforms the RBC causing sickling.

Disease starts a few months after birth: For the first few months after birth, there is sufficient fetal hemoglobin (HbF) to prevent anemia. Once HbS starts getting made, the patient develops sickle cell anemia.

Hemolytic anemia because sickled RBCs are removed by the spleen. This leads to jaundice and gallstones.

Vaso-occlusive crises: Sickled RBCs obstruct capillaries and restrict blood flow to different organs, resulting in ischemia, pain, stroke, necrosis of bone, skin ulcers, kidney failure. In ____________________, sickled RBCs obstruct capillaries and restrict blood flow to the spleen causing it to die

Answer 41

glutamate, valine;

one normal beta globin gene and one mutant beta globin gene (βs) ;

60:40;

deoxy;

long polymerized chains of Hb;

autosplenectomy

Question 42

Oxygen-hemoglobin Dissociation Curve

• SaO2 is oxygen saturation of ___________
• PaO2 is the partial pressure of oxygen in arterial blood.
• Oxygen-hemoglobin dissociation curve: Progressive increase in the percentage of hemoglobin bound with oxygen as the PaO2 increases.

Physiological advantages of sigmoid shape of curve

• The plateau allows oxygen saturation of arterial blood to remain >90% even if PaO2 drops from 100 mmHg to 60 mmHg.
• The steep section allows a large amount of oxygen to be delivered with only a small drop in PaO2

P50 value is the PaO2 at which ____________________. For adult hemoglobin (HbA), P50 = 27 mmHg. Fetal Hb (HbF) and Hb Barts with ______ affinity for oxygen, have _____ P50 values (i.e. 50% saturation occurs at lower PaO2).

• ______ mL oxygen is bound to 1g Hb. Normal Hb concentration in human blood is 15 g/dL. This means (15 x 1.34) 20 mL of oxygen are carried by Hb in 100 mL of blood.
• Hb tense conformation is when no oxygen is bound. Hb relaxed conformation is when the four oxygens are bound to Hb.

The curve can be shifted to the right or left.

• Shift to the right: Hb has lower affinity for oxygen and therefore Hb releases oxygen (as happens in tissues); P50 _________.
• Shift to the left: Hb has higher affinity for oxygen and therefore Hb binds more oxygen; P50 ___________

Answer 42

arterial blood;

hemoglobin is 50% saturated with oxygen;

higher-than-normal;

lower;

1.34;

increases;

decreases

Question 43

Effect of pH

• Acidic pH: When the pH decreases (H+ increases), the oxygen-hemoglobin dissociation curve shifts to the right and more O2 is released from Hb.
• Alkaline pH: When the pH increases (H+ decreases), the oxygen-hemoglobin dissociation curve shifts to the left and less O2 is released from Hb.
• _____________: CO2 released from tissues into the blood decreases pH and thereby causes a shift to the right promoting O2 release into tissues.

Effect of 2,3-diphosphoglycerate (DPG)

• DPG binds with greater affinity to ___________________.
• During ________________, DPG in blood rises. The oxygen-hemoglobin dissociation curve shifts to the right so more O2 is released from Hb.
• Banked blood contains lower levels of - DPG. The oxygen-hemoglobin dissociation curve shifts to the left so less O2 is released from Hb. Be careful when you transfuse old banked blood because the patient could become hypoxic.

Effect of temperature

• ____________ shifts the oxygen-hemoglobin dissociation curve to the right, so more O2 is released to tissues
• Decrease in blood temperature shifts the oxygen-hemoglobin dissociation curve to the left, so Hb binds more O2.

Effect of exercise

• In exercising muscle more ____________ are generated, making the blood acidic, which shifts the oxygen-hemoglobin dissociation curve to the right, and so more O2 is released to tissues.
• During exercise, the ____________ of muscle increases, which shifts the oxygen-hemoglobin dissociation curve to the right, and so more O2 is released to tissues.

An adaptation to living at high altitude is a right shift in the oxygen-hemoglobin dissociation curve, and so more O2 is released to tissues.

Answer 43

Bohr effect;

deoxygenated Hb than to oxygenated Hb;

prolonged hypoxia (oxygen deprivation);

Increase in blood temperature;

CO2 and lactic acid;

temperature

Question 44

Oxygen release in tissues

In tissues

• When blood reaches tissue capillaries, CO2 enters the blood from tissues, thereby increasing pCO2.
• CO2 combines with H2O to form carbonic acid (H2CO3). This reaction is catalyzed by the enzyme _____________
• Carbonic acid dissociates into ____________
• H+ increases. H+ combines with Hb to form ____. The O2 displaced from Hb diffuses from blood to tissues.
• The HCO3- produced is exchanged for ________ by the anion exchanger.

In the lungs (opposite of above)

• CO2 diffuses from the blood into the lung alveoli leaving less CO2 and H+ in RBCs (pH increases).
• _______________shifts to the left.
• Hb binds more O2

Answer 44

carbonic anhydrase;

H+ and HCO3-;

HbH+;

Cl-;

Oxygen-hemoglobin dissociation curve

Question 45

Maternal-fetal oxygen-hemoglobin dissociation curve

The fetal curve is shifted to the left because fetal hemoglobin (α2γ2) has a higher affinity for oxygen than HbA (adult Hb), and also because of poor binding of ________ to gamma chains of fetal Hb.

• So although the fetal pO2 is lower than in the adult, the fetus uses its higher oxygen-saturation of Hb to cope.
• P50 is lower in the fetus.

Changes across the placenta:

• The maternal blood gains CO2, H+ increases / pH falls, the oxygen-hemoglobin dissociation curve shifts to the right releasing additional oxygen.
• On the fetal side, CO2 is lost, H+ decreases / pH increases, the oxygen-hemoglobin dissociation curve shifts to the left allowing additional oxygen binding to Hb

Other important factors in delivery of oxygen to the fetal tissues are:

• A high maternal intervillous blood flow (almost double the fetal placental flow)
• The high fetal hemoglobin (16 - 17 g/dl)
• The high fetal cardiac output
• Fetal metabolic acidosis, which shifts the curve to the right and thus aids delivery of oxygen to the tissues.
• High oxygen affinity of fetal blood could limit oxygen unloading to the tissues although this is minimized by the greater steepness of the curve.

Answer 45

2,3-DPG

Question 46

Carbon monoxide-hemoglobin dissociation curve

• Carbon monoxide has 250-fold greater affinity for Hb than O2.
• A pCO of ______ results in maximum Hb saturation with CO
• At a pCO of 0.4 mmHg, 50% of Hb will be bound to CO rather than O2. This causes tissue hypoxia and death

Carbon monoxide poisoning: A patient severely poisoned with CO can be treated by administering pure oxygen, because oxygen at _______________ can displace CO rapidly from hemoglobin. A patient can also benefit from ______________________, because this strongly stimulates the respiratory center, which increases alveolar ventilation and reduces alveolar carbon monoxide (CO is breathed out more rapidly).

Answer 46

0.4 mm Hg;

high alveolar pressure;

simultaneous administration of 5% carbon dioxide

Question 47

Hemolysis: premature expiry of RBCs - excessive destruction either extravascularly or intravascularly. Shortened RBC survival (normal 120 days).
- Hemolytic anemia is anemia due to hemolysis. Hemolytic anemia represents approximately 5% of all anemias.
Hemoglobin level decreases. Normal level ~15g/dl.
- Hematocrit decreases: proportion of blood volume occupied by RBCs. Normal level = 38-49%
- _____________: free hemoglobin in blood once haptoglobin is depleted.
- Hemoglobinuria: free hemoglobin in urine. The _____________ in the kidney reabsorb free hemoglobin filtered by the kidney to conserve iron. When tubular capacity is saturated, free hemoglobin appears in the urine. The color of the urine depends on the pH. In alkaline urine, Hb is oxygenated to oxyhemoglobin, giving urine a _________ In acidic urine, Hb is oxidized to ___________ (Ferrous to Ferric) giving urine a ______________.
Increased serum bilirubin, jaundice:
- Depletion of ___________________
- Raised _______ released from destroyed RBCs

Compensatory hematopoiesis

• Intramedullary hematopoiesis: In response to hemolysis, the body compensates by releasing erythropoietin from the kidney and liver. EPO acts on erythroblasts and increases erythropoiesis in the bone marrow. The bone marrow becomes ________. The majority (70%) of BM synthesis normally takes place in the _________________. In severe chronic hemolytic anemia, hematopoiesis extends into the marrow of long bones and skull. In the skull, this leads to a chipmunk-like face, frontal bossing (protruding forehead and maxilla).
• Extramedullary hematopoiesis: In severe chronic hemolytic anemia, hematopoiesis extends into spleen and liver (outside the bone marrow). These organs will enlarge as a consequence. Hepatosplenomegaly may also occur because of increased extravascular hemolysis.
• Reticulocytosis: Reticulocytes are RBC precursors that leave the BM, enter blood and stay for 1 day before maturing into RBC. Reticulocytes contain cytoplasmic RNA which stain with _______________ giving a reticular pattern, hence the name reticulocyte. Increased erythropoiesis in the bone marrow as compensation for hemolysis leads to an increase in reticulocytes in the blood.

Answer 47

Hemoglobinemia;

proximal tubular cells;

pink color;

methemoglobin;

dark brown color;

haptoglobin and hemopexin;

LDH;

hypercellular;

pelvis, vertebrae, sternum, shoulder blade and ribs;

supravital dyes

Question 48

Intravascular hemolysis
- RBCs are destroyed in the bloodstream (intravascular destruction), which destruction accounts for about 10% of destruction of aged and/or abnormal RBCs.
- Hemoglobin-haptoglobin complex: Broken RBCs release hemoglobin into the plasma where it is bound to a protein called _________ (synthesized in the liver) to form an Hb-haptoglobin complex, which is cleared by the liver.
- Hemopexin-Heme complex: Broken RBCs may also release heme into the plasma where it is bound to a protein called _________ (synthesized in the liver) to form the heme-hemopexin complex, which is cleared by the liver. Recycling by both these complexes reduces loss of iron.
- Depletion of haptoglobin and hemopexin with excessive usage: If intravascular hemolysis is severe and prolonged, haptoglobin and hemopexin levels are depleted (synthesis cannot keep up with usage).
Consequences of haptoglobin and hemopexin depletion:
- Hemoglobinemia- ____________ in the blood once haptoglobin and hemopexin are depleted.
- _______________: free Hb in urine once the capacity for tubular reabsorption is saturated.

Answer 48

haptoglobin;

hemopexin;

free Hb;

Hemoglobinuria

Question 49

Extravascular haemolysis

• More extravascular hemolysis occurs in hemolytic anemia. Aged RBCs are phagocytosed by macrophages lining _____________________.
• Total ___________ in blood increases in hemolytic anemia.
• Unconjugated bilirubin in blood increases in hemolytic anemia.
• Conjugated bilirubin does not increase because all the unconjugated bilirubin that enters hepatocytes is converted to water soluble conjugated bilirubin and excreted in bile.
• ___________ level in blood increases in hemolytic anemia because a proportion of the bilirubin excreted into the intestine is absorbed into the circulation.
• Jaundice and hyperbilirubinemia: Clinically detectable jaundice is apparent when serum bilirubin is >2 mg/dL. The excess bilirubin is unconjugated in hemolytic anemia

Answer 49

sinusoids of the spleen, liver and bone marrow;

bilirubin level;

Urobilinogen

Question 50

2. RBC membrane and hereditary spherocytosis

RBC membrane and cytoskeleton: The RBC membrane is tethered to the underlying cytoskeleton, and this interaction determines the shape of RBC and their response to and recovery from the shearing forces of the circulation. A key membrane protein is the anion exchanger that exchanges _______________. The anion exchanger is bound to a cytoskeletal protein called __________, which is bound to a noodle shaped protein called ___________ that forms a cytoskeletal network.

Mutations of membrane skeleton 
- Ankyrin (50%) 
- Spectrin (25%) 
-  Band 3 / Anion Exchanger (25%).
Spherocytes are trapped in the spleen and engulfed by splenic macrophages. This results in anemia. 

Hereditary spherocytosis: This genetic disease is due to mutations of ___________________. The RBCs have a spherical shape and are called spherocytes. The RBC membrane in spherocytes is not anchored properly to the underlying cytoskeleton. The spleen removes the abnormally spherocytes (excessive extravascular hemolysis) leading to hemolytic anemia. Patients with hereditary spherocytosis develop _________________ because of increased macrophage-phagocytosis of spherocytes.

Splenectomy: Cures the anemia by reducing removal of spherocytes by the spleen

Osmotic fragility test: This is a clinically used laboratory test. Spherical RBCs from hereditary spherocytosis patients are more prone to __________ when exposed to hypotonic solutions than normal RBCs. Splenectomy cures the hemolytic anemia in these patients.

Answer 50

HCO3- and Cl-;

ankyrin;

spectrin;

ankyrin, spectrin or the anion exchanger;

splenomegaly (enlarged spleen);

lysis

Question 51

3. RBC, Glucose-6-phosphate dehydrogenase, Hemolysis, Favism

G6PD deficiency

• Glucose-6-phosphate dehydrogenase deficiency is the most common enzyme deficiency in RBCs.
• X-linked disease: G6PD is on the _______________
• Affects 10% of the world, with a geographic distribution that parallels the malaria belt. Heterozygotes are protected from malaria parasitization

Function: RBCs are usually at substantial risk of damage from ________________ due to their role as oxygen carriers. G6PD, an enzyme pivotal in the pentose pathway, is required for the production of ____________ that protects RBCs from oxidative stress. The G6PD/NADPH pathway is the ONLY source of _____________ in RBCs that is required to mop up oxygen free radicals. When this system is poorly functioning, RBCs get damaged by oxygen free radicals and are removed by macrophages in the splenic and liver sinuses, and they are also destroyed in the circulation.

Prevalence: An estimated 400 million people worldwide have glucose-6-phosphate dehydrogenase deficiency. This condition occurs most frequently in certain parts of Africa, Asia, and the Mediterranean. It affects about 1 in 10 African-American males in the United States.

Precipitating factors: Hemolysis is precipitated by oxidant stress. Infections, certain drugs (anti-malarial - _________________), or ingesting fava beans (favism) can increase the levels of reactive oxygen species, causing RBCs to be destroyed faster than the body can replace them (hemolysis).

Answer 51

X-chromosome;

oxidizing free radicals (e.g. superoxide and hydrogen peroxide);

NADPH;

reduced glutathione;

primaquine, sulfonamides, aspirin, ciproflox

Question 52

4. Autoimmune hemolytic anemia

• Hemolysis due to destruction by anti-RBC antibodies.
  Antibodies are mainly __________; IgA and IgE rarely.
• _______________ if antibodies weakly activate complement system, destruction is in the spleen.
• Intravascular hemolysis if antibodies strongly activate complement system, destruction is within the circulatory system.
  Classification
• Warm: Anti-RBC antibodies are active at ___________. Account for 80% of immune hemolytic anemias. Usually ____ that usually react against Rhesus antigens.
• Cold: Anti-RBC antibodies are active at ______. Usually _______ that cause RBCs to agglutinate. May cause intravascular hemolysis if complement is activated. May be associated with ________________.

Direct Coomb’s test is used to detect antibodies or complement proteins that are attached to the surface of the patient’s RBCs. The patient’s RBCs are washed to remove any unbound antibodies and the patient’s own plasma. The RBCs are then incubated with anti-human immunoglobulin (“Coombs reagent”). If the _____________________ the direct Coombs test is positive, a visual indication that antibodies or complement proteins are bound to the surface of RBCs and may be causing destruction of those cells.

Indirect Coomb’s test is used to detect anti-RBC antibodies in the patient’s blood. These antibodies are usually IgG or IgM, but rarely may be IgA or IgE. Patient’s serum is collected and incubated with _________________. If anti-RBC antibodies are present in the patient’s serum, these bind to the surface of the RBCs. In step 2, anti-human immunoglobulin (“Coombs reagent”) is added. If the RBCs agglutinate, the indirect Coombs test is positive, a visual indication that the patient’s serum contains anti-RBC antibodies and may be causing destruction of those cells.

Answer 52

IgG and IgM’;

Extravascular hemolysis (spleen);

body temperature; IgG

4oC; IgM; Mycoplasma infection or Infectious mononucleosis;

RBCs agglutinate (clump together);

foreign RBCs of known antigenicity

Question 53

Intramedullary versus Extramedullary hematopoiesis: In adults, hematopoiesis occurs primarily in the bone marrow (intramedullary hematopoiesis). In severe and chronic anemia, the body compensates by initiating additional hematopoiesis in the __________________ (extramedullary hematopoiesis).

Bone marrow transplant: Bone marrow is removed from the pelvis (Iliac crest) of the donor under general anesthesia and then transfused into the blood circulation of the recipient. The cells home to the bone marrow of the recipient, colonize the bone marrow and begin hematopoiesis. The donor and recipient must be HLA-matched to ensure that graft-versus-host disease does not occur.

Stem cell transplant: Pluripotent stem cells are isolated from the bone marrow by first depleting mature and immature blood cells using lineage-specific monoclonal antibodies, and then enriching for CD34+ stem cells. CD34 is a glycoprotein found on the cell surface, which may contribute to the attachment of stem cells to the stroma/matrix in the bone marrow. The stem cells are transfused into the blood circulation of the recipient. They colonize the bone marrow and initiate hematopoiesis.

Answer 53

liver and spleen