Haemolytic Anaemia

Question 1

Name the protein that is a specific indicator of haemolysis and outline how it is affected by haemolysis (2½) (CPATH)

Answer 1

Haptoglobin [½]. Haptoglobin binds free haemoglobin [1] and the haptoglobin-haemoglobin complex is cleared rapidly by the reticulo endothelial system, therefore lowering haptoglobin level in haemolysis [1]

Question 2

List two (2) biochemical markers that would be useful to exclude in vitro (in tube) haemolysis (e.g. haemolysis occurring due to poor blood taking technique) (2x½ = 1) (CPATH)

Answer 2

Unconjugated bilirubin, haptoglobin, urine urobilinogen

Question 3

List three (3) laboratory features (biochemical and peripheral blood smear) that may be found in active intravascular haemolysis/ confirm diagnosis of haemolytic anaemia. Explain why these finding occur in each example (3) (Super NB)

Answer 3

Coombs test: Confirm immune basis of haemolysis

Features of increased red cell breakdown
Decreased Haemoglobin due to red cell destruction
Increased Serum unconjugated bilirubin: Red cell destruction with Hb breakdown into heme & globin. Heme broken down to bilirubin.
Increased Serum Lactate dehydrogenase (LDH): Release of red cell isoenzyme by destroyed red cells
Decreased Serum haptoglobin: Binding of released Hb
Increased Urine urobilinogen
Increased Aspartate aminotransferase

Features of increased red cell production

Reticulocytosis (Reticulocyte count)/ polychromasia: Bone marrow/erythroid response to anaemia
Peripheral blood smear for Red Cell Morphology may show polychromasia or specific red cell abnormalities e.g.
Spherocytes (extravascular haemolysis due to IgG coating or hereditary membrane defect with splenic trapping)
o Fragments (microangiopathic or macrovascular red cell destruction)
o Bite cells (oxidative stress)
o Nucleated red blood cells (bone marrow stress)
o Sickle cell etc.
Red cell survival is shortened (measured by 51Cr labelling of the red cells)
Haemosiderinuria, Haemopexin, Methaemoglobin

Question 4

State whether the following will be increased, decreased or unchanged in urine in haemolysis. Explain answer (3) (CPATH)

Urobilinogen

Unconjugated bilirubin

Answer 4

a. Urobilinogen. Increased [½] due to increased flux through the bilirubin pathway due to haemolysis [1]
b. Unconjugated bilirubin. Unchanged [½] because not water soluble [1]

Question 5

Classify the haemolytic anaemias and state one (1) example for each type (½x16 = 8) [Ess haem p60] (Super NB)

Answer 5

Hereditary or Intracorpuscular defects
Membrane disorders - Hereditary spherocytosis (HS)
Haemoglobinopathies – Sickle cell (SCA)
Enzymopathies – G6PD

Acquired or Extracorpuscular defects
Immune – Allo, auto, drugs
Mechanical – Fragmentation syndromes
Infections – Malaria
Drugs – Dapsone

Question 6

Classify the type of haemolytic anaemia caused by HS [1]

Answer 6

Intracorpuscular defect, a membrane disorder

Question 7

Describe the underlying defect causing his disorder [2]

Answer 7

S is caused by defects in the vertical interactions between the red cell membrane skeleton and the lipid bilayer of the membrane. As a result, parts of lipid bilayer are lost and as a result the RBCs become spherical (loss of surface area relative to volume)

Question 8

List two (2) causes of spherocytes other than HS [1]

Answer 8

Autoimmune haemolytic anaemia, Clostridium perfringens infection, Burns, Incompatible blood transfusion, Haemolytic disease of newborn

Question 9

Explain the use of the Coombs test in spherocytic haemolysis [1] & Describe the principle of this test [2]

Answer 9

To distinguish AIHA from other causes of spherocytes. The Coombs reagent is an Ab or mixture of Abs which will react with Ig or complement which is adherent to the red cell. It will cause agglutination of red cells which have the relevant substance on their surface.

Question 10

Name two (2) red cell enzyme deficiencies that may lead to chronic haemolytic anaemia (1)

Answer 10

G6PD & Pyruvate kinase

Question 11

Explain the pathogenesis and pathophysiology of Glucose-6-phosphate dehydrogenase (G6PD) deficiency and name two (2) precipitating factors for acute haemolysis in this disease (7)

Answer 11

X-linked genetic disease. Enzyme malfunction/deficiency involved in pentose phosphate pathway causing decreased NADPH production. Unable to maintain reduced form of glutathione stores - inability to protect the cell against oxidative stress/ free radicals. Infection, Oxidant drugs, Fava beans

Question 12

This child is unlikely to have G6PD deficiency. Give the most important reason why this is unlikely (2)

Answer 12

It is inherited as an X-linked recessive condition, and therefore would be unusual in a female child.

Question 13

Name the metabolic pathway affected in pyruvate kinase deficiency, and explain why the oxygen dissociation curve is shifted (3)

Answer 13

Anaerobic glycolysis; there is an accumulation of 2,3-DPG and right shift of O2 dissociation curve, decreasing O2 affinity of Hb and anaemia better tolerated.

Question 14

A number of mechanisms exist to explain how drug-induced haemolysis occurs, name two (2)

Answer 14

Drug adsorption (hapten), Immune complex formation, Autoantibody production, Red cell membrane modification

Question 15

List four (4) causes of fragmentation haemolysis (2)

Answer 15

DIC, TTP, HUS, Cardiac lesions, AV malformations, Malignant hypertension

Question 16

List two (2) examples of micro-angiopathic haemolytic anaemias (MAHA) (1)

Answer 16

TTP, HUS, DIC, vasculitis, disseminated malignancy, vasculitis, GPH

Question 17

Name two (2) causes of micro-angiopathic haemolytic anaemia (MAHA) (1)

Answer 17

DIC, Thrombotic thrombocytopenic purpura, Haemolytic uremic syndrome, Vasculitis, Malignant hypertension

Question 18

Name the test used to distinguish between immune and non-immune haemolytic anaemia and explain the test principles (3)

Answer 18

Coombs’ test: sensitised Ig/ complement coated RBCs agglutinate on addition of antihuman globulin (Coombs) reagent.

Question 19

Anaemia may be caused by haemophagocytic lymphohistiocytosis. State which cell type is hyperactivated in this condition (½) (IMMUNO)

Answer 19

Macrophages

Question 20

Outline the causes of this hyperactivation (3) (IMMUNO)

Answer 20

There is uncontrolled secretion of cytokines (e.g. IFNg, TNFa, IL-2, GM-CSF) from CD8+ T cells and natural killer cells because there is an underlying gene defect in perforin production. These cells cannot kill infected cells and there is an increase in systemic levels of pro-inflammatory cytokines and promotes excessive activation of macrophages.

Question 21

Briefly describe the sequence of events leading to Macrophage Activation Syndrome (2½) (IMMUNO)

Answer 21

When CD8+ T cells fail to clear infections, there is an increased secretion of systemic inflammatory cytokines, such as IFNg or TNFa (either), that causes activation of tissue resident macrophages.

Question 22

Name three (3) soluble factors that are involved in increased vascular permeability during secondary haemophagocytic lymphohistiocytosis (1½) (IMMUNO)

Answer 22

IL-1 (alpha or beta), TNFa and IL-6.

Question 23

Name two (2) familial conditions that leads to haemophagocytic disease (1) (IMMUNO)

Answer 23

Any of: Chediak-Higashi Syndrome, Griscelli syndrome, X-linked Lymphoproliferative syndrome or Wiskott-Aldrich syndrome

Question 24

Define AIHA and name the most important test that would confirm this diagnosis (2)

Answer 24

Autoimmune haemolytic anaemias are caused by antibody production by the body against its own red cells. Characterized by positive direct coombs test.

Question 25

Outline the mechanism whereby IgM coated red blood cells (RBCs) are destroyed in AMHA (2) (IMMUNO)

Answer 25

IgM/ antigen complexes (½) (IgM coated RBCs) are very effective at activating complement (½) and the ‘membrane attacked complex’ of complement (MAC) (½) destroys the RBCs by osmotic lysis (½)

Question 26

Outline the mechanisms whereby IgG coated RBCs are destroyed in AMHA (6x½ = 3) (IMMUNO)

Answer 26

IgG opsonization/ coating of RBC leads to phagocytosis (½) by macrophages (½) and intracellular lysis (½) of RBC. Macrophages have Fc receptors which bind to antibody (½).

Natural Killer (NK) cells (½) can also play a role in lysing antibody coated RBCs by the release of perforins (½)/ or by Antibody-dependent cell cytotoxicity.

Question 27

State which is clinically more severe, IgG or IgM mediated haemolytic anaemia, and explain why (4x½ = 2) (IMMUNO)

Answer 27

IgM is more dangerous (½). Much more rapid (½) and dangerous intravascular (½) hemolysis occurs as a result of complement activation. IgG coated RBCs are more gradually destroyed in spleen/ liver (½)

Question 28

Name a group of antigens which may be responsible for haemolytic disease of the newborn (½) (IMMUNO)

Answer 28

Rhesus group of Ag

Question 29

State where the antibodies that mediate haemolytic disease of the newborn come from (1) (IMMUNO)

Answer 29

Across the placenta (½), from the mother (½)

Question 30

Name the organ in a newborn infant where RBCs are destroyed as a result of AMHA (½) (IMMUNO)

Answer 30

Liver

Question 31

Explain how Rhesus haemolytic disease of the newborn develops (8)

Answer 31

Rh neg woman and Rh pos man conceive a child. Rh neg woman carries a Rh pos foetus. Cells from the foetus enter the woman’s blood stream. The woman becomes sensitized, antibodies form against Rh pos RBCs. With next pregnancy of a Rh pos foetus, maternal antibodies attack foetal RBCs

Question 32

Describe the pathogenesis of haemolytic disease of the newborn and list two (2) common blood group systems that evoke this response (½x9 = 4½)

Answer 32

Discrepancy in blood group between mother and baby, Feto-maternal bleed, Ab response in mother against blood group of baby, Second pregnancy (Ab response IgG), Cross placenta, Target red cells of baby, Haemolysis/ removal in spleen. ABO & Rh