Anemia – Part 7 – Hereditary Spherocytosis, Glucose-6-phosphate Dehydrogenase Deficiency (G-6-PD)

December 19, 2020HematologyLab Tests

Hereditary Spherocytosis:

1. Definition:
   1. Hereditary spherocytosis anemia is quite common and transmitted as an autosomal dominant trait in the caucasian population.
   2. Rarely it may be autosomal recessive.
   3. This is the most common hereditary hemolytic anemia in northern Europe, 1 in 5000.
   4. Inheritance is dominant in 75% of the cases.
2. Pathogenesis:
   1. Molecular abnormality of the skeletal proteins has been identified in some cases.
   2. This is usually caused by the defect in the proteins involved in the vertical interaction between the membrane skeleton and the RBCs’ lipid bilayers.
   3. The basic defect is the loss of RBCs’ membrane, caused by the release of parts of the lipid bilayer that is not supported by the skeleton, resulting in decreased surface area.
   4. This will produce RBCs with the lowest surface area- to – volume ratio called spherocytes.
   5. The marrow produces normal biconcave RBCs, but these lose membrane and become more spherical where there is the loss of surface area compared to volume.
      

      Hereditary spherocytosis RBC mechanism

   6. These RBCs can not change the shape easily, so they can not move through the microcirculation, particularly through the spleen reticuloendothelial system, where they will die permanently.
   7. In one of the articles, hereditary spherocytosis is classified into four subtypes.
      1. Minor HS.
      2. Moderate HS.
      3. Moderate to severe HS.
      4. Severe HS.
         

         Clinical parameters	Minor HS	Moderate HS	Moderate to severe	Severe HS
         Hemoglobin	Normal	>80%	60% to 80%	<60%
         Reticulocytes	<6%	6% to 10%	>10%	>10%
         Peripheral blood smear	Few spherocytes	Spherocytes	Spherocytes	Microspherocytes and poikilocytosis
         Osmotic fragility at 37 °C	Increased	Increased	Increased	Increased
         Splenectomy	Rarely needed	Depending upon certain cases	Necessary >5 years old	necessary > 2 to 3 years old
         Inheritance	Autosomal dominant	Autosomal dominant, de novo	Autosomal dominant, de novo	Autosomal recessive

3. Signs and symptoms:
   1. The prominent features of hereditary spherocytosis:
      1. Chronic hemolysis.
      2. Jaundice.
      3. Splenomegaly.
   2. Anemia is mild and usually is un-noticed and diagnosed at an adult age.
   3. Anemia may be present at any age, from infancy to old age.
   4. There is an increased tendency for the increase in bilirubin level.
      1. The jaundice is fluctuating. It is marked if it is associated with Gilbert’s syndrome.
   5. Chronic hemolysis leads to pigment gall stones are quite common in these patients.
   6. In the spleen, these cells, by removing the membrane, changed into microspherocytes, and ultimately sequestered and will lead to splenomegaly.
      1. Spenemegally is quite common in these patients.
   7. These patients ultimately develop anemia, splenomegaly, and ulcer legs.
   8. Aplastic crises, usually seen in patients with parvovirus infection. This will leads to the severity of anemia.
   9. Megaloblastic anemia is due to folate depletion owing to the overactivity of the bone marrow.
   10. Autohemolysis is increased, and this can be corrected by glucose.
4. Lab. findings:
   1. There is mild to moderate anemia (8 to 12 G/dL).
   2. MCV normal or slightly low.
   3. MCHC is high.
   4. Increased reticulocytes, 5 to 7% (another reference 5% to 20%).
   5. Osmotic fragility is very high, and this is a confirmatory test.
      1. This test needs 24 hours of incubation at 37 °C to become prominent.
   6. Bilirubin slightly increased.
   7. Raised LDH level.
   8. Urine urobilinogen is increased.
   9. Haptoglobin low.
   10. Peripheral blood shows prominent spherocytes ( mircospherocytes).
       1. Spherocytes are densely staining with a smaller diameter than the normal RBCs.
       2.  Spherocytes are uniform round RBCs with more intensely staining hemoglobin, and there is no central pallor.
          

          Peripheral blood showing spherocytosis.

       3. There is increased polychromatophilia.
       4. Reticulocytes are seen.
   11. Direct coomb’s test is negative.
   12. Differential diagnosis of anemia: This hereditary spherocytic anemia needs to differentiate from:
       1. Immune hemolytic anemia.
       2. G6PD deficiency.
       3. Thermal injury.
       4. Toxins due to infection of clostridium.
       5. Snake venom.
       6. Bee and spider venom.
5. Treatment :
   1. For the treatment of symptomatic patients, splenectomy is the choice.
      1. The patient should be kept on folic acid prophylactically to prevent aplastic crises.

Glucose-6-Phosphate Dehydrogenase (G-6-PD) Deficiency anemia:

1. Definition: This is X-linked glucose-6-phosphate dehydrogenase deficiency, which is an inborn error of the RBCs.
   1. The gene for G6PD is localized to chromosome Xq28 near the factor VIII gene.
   2. The main clinical S/S are seen in the male.
   3. This will protect against the Plasmodium spp infestation.
   4. Reticulocytes have normal G6PD activity than the older RBCs.
   5. Deficient RBCs of G6PD are more fragile and short-lived.
   6. This disease is prevalent in Africa, South Europe, the Middler east (around 20%), South East Asia (around 49% in some regions), and Oceania.
2. Basic mechanism:
   1. There are two types of disease-causing mutation are:
      1. Mediterranean  = G6PD _med^. 
         1. This mutation is more severe and is due to every new RBCs are deficient in the G6PD enzyme.
         2. The G6PD enzyme deficiency is 10% of the normal.
      2. African = G6PD_A^.
         1. Young RBCs (these are reticulocytes) maintain the adequate G6PD enzyme level for a longer time.
         2.  G6PD activity is around 20% to 60%.
   2. G-6-PD is the initial enzyme involved in the pentose-phosphate pathway of RBCs metabolism.
   3. It catalyzes the removal of H⁺ ions from glucose-6-phosphate to produce 6-phosphogluconate and require cofactor NADP.
   4. NADP is reduced to NADPH, and this will prevent the effect of oxidants on RBCs.
      1. G-6-PD produces NADPH, which maintains the Glutathione and other proteins when the RBCs are exposed to oxidants.
         

         G-6-PD enzyme role for RBC membrane maintenance

   5. In G-6-PD deficiency, there is a shortage of NADPH, and RBCs can not neutralize the oxidant stress and are hemolyzed.
      

      G-6-PD deficiency mechanism for hemolysis

   6. When RBCs are exposed to oxidant stress, it damages the RBCs membrane and gives rise to Heinz bodies’ formation (this is denatured hemoglobin) formation.
      

      G-6-PD enzyme deficiency leads to Heinz bodies formation

   7. A gene on the X-chromosome determines the structure of  G-6-PD, which gives remarkable polymorphism in the human population.
3. Causes of G-6-PD deficiency:
   1. Vegetables like Fava beans.
   2. Drugs:
      1. Antimalarial drugs like:
         1. Primaquine.
         2. Pamaquine.
         3. Chloroquine.
         4. Fansidar.
         5. Maloprim.
         6. Quinine.
      2. Antibacterial drugs like:
         1. Sulphonamide.
         2. Sulphones like co-cotrimoxazole, sulfanilamide, dapsone, and salazopyrin.
         3. Nitrofurans.
         4. Chloramphenicol.
         5. Ciprofloxacin.
         6. Dapsone.
      3. Analgesics like:
         1. Asprin.
         2. Acetanilide.
         3. Phenacetin
      4. Antihelminths like stibophen and β-naphthol.
      5. Other agents like vitamin K, probenecid, naphthalene, nalidixic acid, dimercaprol, and phenylhydrazine.
4. Clinically there is variable presentation.
   1. Usually, these patients are asymptomatic and more common in the male than females.
   2. Hemolytic anemia starts:
      1. Acute hemolytic anemia in response to oxidant stress like fava beans or infections.
         1. There is intravascular hemolysis,
         2. There is hemoglobinuria.
      2. Neonatal jaundice.
      3. Rarely, there is congenital non-spherocytic hemolytic anemia. This may be because of some other enzyme deficiency.
   3. G6PD can be divided into:
      1. Type A  is present in about 20% of the black African with reduced G6PD activity.
      2. Type B is present in almost all Caucasians and 70% of the black  African with normal activity.
   4. The commonest presentation is neonatal jaundice.
   5. There is congenital hemolytic anemia.
   6. The patients may have drug-induced hemolysis.
   7. Mostly hemolysis starts after eating the fava beans called favism.
   8. Most of the patients do not have any symptoms.
   9. RBCs number is normal, functions and survival are normal unless exposed to oxidative stress.
   10. Antimalarial drugs primaquine directly lead to hemolysis.
5. Lab findings:
   1. In between the crises, the blood picture is normal.
   2. Typically following the drug exposure, there is an acute drop in the Hemoglobin level (Hb is low in the hemolytic crises).
   3. There is an increase in the reticulocyte number. These may go as high as 50%.
   4. The peripheral blood smear shows poikilocytosis. During the attack the peripheral blood smears show:
      1. RBCs with punched-out defects in their shapes are seen.
      2. There are fragmented RBCs, and more typically, there are blister cells.
      3. Spherocytes may be seen.
      4. Heinz bodies may be found in the RBCs.
         

         G-6-PD blood film in an acute attack

   5. Diagnosed by a screening test for G-6-PD deficiency.
      1. A definite diagnosis by enzyme assay.
      2. In the acute attack, the screening test is normal.
      3. But in the asymptomatic phase, the screening test will be abnormal, indicating deficiency.
6. Treatment:
   1. Stop the offending agent.
   2. Keep a high urine output.
   3. Blood transfusion if there is severe anemia.
   4. Babies having a deficiency of G-6-PD deficiency will have neonatal jaundice.
      1. These babies need phototherapy and maybe blood transfusion.