Anemia:- Part 5 A – Sickle Cell Anemia, and Sickle Cell Trait, Discussion and Workup

June 30, 2021HematologyLab Tests

Sickle Cell Anemia

Background of the disease:

1. This condition was described in 1910 in Chicag0 in black patients.
   1. The carrier state in the USA is around 8%, while it is very high in blacks.
   2. This is also found in Mediterranean countries, like Greece, Italy, Israel, Saudi Arabia, India, and Pakistan.
   3. The sickle gene has some advantages, and it protects from malarial infection (Plasmodium falciparum).

To understand Sickle anemia, we need to discuss and understand the structure of the Hemoglobin:

1. 1. The normal globin, which is part of the hemoglobin, consists of 2 alpha chains and 2 beta chains.
      

      Hemoglobin normal structure

   2. Genetic code is located on chromosomes 11, are γ, δ, ε, and β-chains.
   3. While on chromosome 16, there are α and ζ loci.
      

      Globin genes location for hemoglobin formation

      Various types of hemoglobin:

      Type of hemoglobin	Genotype of hemoglobin	Functions of the hemoglobin
      Hb A	α2β2	This is the main adult Hb
      Hb A2	α2/δ2	This is present in a small amount
      Hb F	α2/γ2	Main fetal Hb in late stages
      Hb gower1	ζ2/ε2	This Hb is present in the early life of the fetus
      Hb gower2	α2/ε2	This Hb is present in a small amount in the early fetal life
      Hb portland	ζ2γ2	This is found in the embryos
      Hb H	β4	It is seen in the α-thalassemia
      Hb Bart’s	γ4	It is seen in the α-thalassemia

Sickle cell syndrome includes all the hemoglobinopathies where HbS is present, e.g:

1. 1. Sickle cell trait: AS heterozygous (a combination of HbA+HbS = α2 β^Aβ^S).
      1. Hb A = ≅ 60%    Hb S = 40%
   2. Sickle cell disease: SS homozygous (a combination of HbS+HbS = α2 β^S β^S).
      1. No Hb A  Hb S = >80  Hb F = <1 to 20%  Hb A2  = 1% to 4%.
   3. HbSC disease = α2 β^S β^C.
   4. HbS-beta thalassemia disease = α2 β^S β⁰.
   5. HbS- beta thalassemia = α2 β^S β⁺.
      

      Sickle cell disease and its combinations

Definition of Sickle cell disease:

1. 1. Sickle cell disease occurs with the autosomal recessive inheritance of abnormal β-chain of the hemoglobin, where glutamic acid is replaced by valine. The end result is precipitation and polymerization of the Hb-S under the conditions of deoxygenation and hypertonicity.
      1. There is rigid crystal formation that will deform the RBCs called sickling, and this will leads to occlusion of the microvasculature and hemolysis.
   2. Sickle cell disease is defined where HbA is absent, and only HbS is found. There is a homozygous expression of the S hemoglobin.
   3. The basic defect: The amino acid defect is in the β-chain at the 6th position, where the glutamic acid is replaced by the valine, giving the Hb less negative charge than HbA.
   4. This is the genetic mutation of Glutamic acid to valine.
      

      Sickle cell Hb structure

   5. Sickle cell RBCs have been transformed into rigid, inflexible cells with at least one prominent projection.
      

      Hb S (Homozygous) structure

Sickle cell trait (A/S):

1. This is a benign condition with no anemia and a normal appearance of the RBCs on the blood smear.
2. Hematuria is a common symptom and is thought to be due to small interacts in the kidneys.
3. Sickle cell trait usually asymptomatic.
4. This is a heterozygous condition Hb S (α2/β⁺β^S).
5. The peripheral blood smear shows normal CBC or maybe mild normocytic or normochromic anemia.
6. These patients are at risk of splenic infarcts at high altitudes.
7. These patients also have the risk of renal medullary carcinoma.
8. Peripheral blood smears also do not show sickle cells.
9. Avoid or take care of anesthesia: pregnancy and high altitude.
10. Electrophoresis shows:
    1. Hb S = 35% to 45%,
    2. Hb F = <1%
    3. Hb A2 = 1% to 3%
    4. Hb A = 50% to 55%.
       

       Hemoglobin electrophoresis of Sickle cell

Sickle cells disease (S/S):

1. This is a homozygous condition for Hb S (α2/2β^S) disease.
2.  These patients have moderate to severe normochromic and normocytic anemia, and hemoglobin is 5 to 11 g/dL.
3. Circulating nucleated RBCs are seen.
4. Due to splenectomy, peripheral blood smear may show Howell-Jolly bodies, anisopoikilocytosis are present because of repeated spleen infarction (autosplenectomy).
5. Indirect bilirubin is raised.
6. Sickle cell preparation is positive.
7. Electrophoresis shows:
   1. Hb S = >80%
   2. Hb F = <20%
   3. Hb A2 = 3% to 8%
   4. Hb A = absent

Pathophysiology of Sickling:

1. 1. Hb S is soluble and usually causes no problem when properly oxygenated.
   2. So sickling depends upon the degree of oxygenation.
   3. When the HbS is deoxygenated in vitro under near physiologic conditions, it becomes relatively insoluble compared to HbA and aggregates into long rigid polymers called tactoids.
      

      The sickle cell formation mechanism

   4. Hb S, called sickle hemoglobin, reacts to deoxygenation and dehydration by solidifying and stretching the RBCs into an elongated sickle -shape, producing hemolytic anemia.
      1. These sickled RBCs settle in the small blood vessels, giving vaso-occlusive crises, as abdominal crises, painful crises, and stroke.
      2. These cells have shortened life in RBCs’ blood with an average of 17 days, where the normal half-life of RBCs is 120 days.
   5. These cells appear crescent in shape on stained blood film.
      

      sickle cell phenomenon

1. 6. These cells return to normal shape when they are oxygenated.
   7. After several sickling episodes, irreversible damage occurs to these RBCs; now, these RBCs will be phagocytosed by the macrophagic cells in the liver, spleen, or bone marrow, which will lead to anemia.
   8. This process of phagocytosis and hemolytic crises will depend upon the number of damaged RBCs in the circulation.
   9. The rigid sickle RBCs cannot deform and circulate through the small capillaries, resulting in these capillaries’ blockage.
   10. This will lead to hypoxia and extreme pain and ultimately cause the death of the affected tissue. Infarction in the spleen is common.
   11. In this case, repeating this process in the spleen will lead to necrosis, scarring, and the result is a nonfunctional spleen called autosplenectomy.

Clinical findings:

1. Clinical manifestation of sickle diseases does not occur until the infant is 6 months of age. Now decrease in Hb F occurs, and Hb S rises.
2. When sickling takes place, which leads to hemolytic anemia, and the patient will have:
   1. Pallor and fatigue.
   2. Jaundice.
   3. Irritability.
   4. In the case of acute crises, the patient will have the manifestation of  sickle crises, which includes:
      

      Organs involved	Clinical manifestations
      Bone	Bone pains are the most common.
      Lungs	There is pleuritic pain Infarction of the lung
      Brain	There are fits  Hemiparesis
      Kidneys	Papillary necrosis Hematuria Lake of urine concentration
      Spleen	There are painful infarcts.
      Liver	Pain Abnormal liver function test
      Penis	There is priapism

   5. Thrombotic crises (vasoactive changes). This will takes place due to obstruction of the microcirculation by sickled cells. There are thrombosis and infarction. Vasoactive crises are very painful and may last for days or weeks, with an average duration of 4 to 6 days.
      

      Sickle cells leading to ischemia

   6. Aplastic crises are due to decreased erythropoiesis instead of increased need because of hemolytic anemia. This is due to the short life of RBCs, which is just 10 to 20 days. Normally there is compensatory erythropoiesis, 5 to 8 times normal. When this compensatory mechanism is compromised, aplastic crises develop.
   7. Pooling crises are due to a large pool of blood in the liver and spleen.  This type of crisis is seen only in early life. The spleen can hold 1/5th of the body’s blood. There may cause death due to cardiovascular collapse.
   8. Hemolytic crises are unusual but may occur with some drugs or infections.
   9. Infection is the most common cause of death. Sepsis and meningitis develop in as many as 10% of the children during the first 5 years of life and a mortality rate of 25%.
      1. Survival is unpredictable, but many patients die in their twenties.
         

         The Sickle cell manifestation

Long term complications:

1. There is an increased susceptibility to infection.
2. The spleen may be completely infarcted.
3. There are chronic leg ulcers.
4. There are gall stones due to repeated hemolysis.
5. There may be aseptic necrosis of the bone, particularly femoral heads.
6. Chronic renal disease.
7. There may be blindness due to retinal detachment or proliferative retinopathy.
8. There may be an infarction of the lungs.
9. Bone marrow necrosis may cause the fat-emboli syndrome.

Factors influencing the severity of Sickle cell anemia:

It depends upon various factors like:

1. Amount of the Hb-S.
2. Its association with thalassemia.
3. Influence of deoxygenation.
4. Presence of amount of  Hb-F.
5. If it is associated with G6PD deficiency.
6. There is the effect of the temperature.
7. There is the effect of dehydration.
8. It depends upon MCHC.
9. Vascular stasis also influences sickle cell disease.
10. There is a role of the pH of the body.

Lab findings are:

1. Low hemoglobin (7 to 9 G/dL).
2. The peripheral blood smear shows sickled RBCs.
3. Sickle cell preparation is positive.
4. Procedure No 1:
   1. Take one drop of blood on the slide.
   2. Put the cover glass and seal the edges with paraffin.
   3. Characteristic sickling may be seen in 6 hours or earlier.
   4. This may not be seen after 24 hours.
5. Procedure No 2:
   1. Add reducing substance sodium metabisulfite 2%.
   2. Metabisulfite is mixed with the blood.
   3. After mixing with the blood, cover with cover glass.
   4. Now seal the edges with paraffin.
   5. The result may be readable in 15 to 60 minutes.
6. Procedure No 3:
   1. There are commercially available kits based on immunoassays.
   2. There are antibodies specific for the Hb-S.
   3. This method is very sensitive for newborn screening.
   4. This method has no interference by the Hb-F or Hct.
7. Peripheral blood smears:
8. There are marked poikilocytosis and inclusion bodies.
9. May see sickle cell 6 to 18% on the peripheral blood smear.
10. Platelets are increased.
11. Reticulocytes are 10 to 20%.
12. ESR is very low.
13. Bone marrow shows marked erythroid hyperplasia.
    1. Iron stain shows low deposits of iron.
14. Bilirubin is increased.
15. serum iron is increased
    1. TIBC is normal.
    2. %saturation is normal.
16. RBC survival is decreased.
17. Electrophoresis of blood shows Hb S and no normal Hb A.
    1. This is the definitive method for the diagnosis.
       

       Hemoglobin electrophoresis of Sickle cell anemia

18. The parents of sickle cell disease will show features of the sickle cell trait.
    Laboratory findings in Sickle cell anemia:

    Type of anemia	Hemoglobin	Hct	Reticulocytes	MCV	Hb electrophoresis	RBC morphology	The severity of the disease
    Sickle cell trait (α2/β+βs)	Usually normal	Usually normal	Normal	Usually normal	HbS = 38% to 45% HbF = Normal HbA2 = 1% to 3% HbA = 55% to 60%	Normal	Mostly asymptomatic
    Sickle cell disease (α2/βsβs)	6 to 9 g/dL	18 to 30 fl	4 to 30%	93	HbS = 80% to 95% HbF = 2% to 20% HbA2 = <3.6% HbA = Nil	Many sickle cells Many target cells Frequent nucleated RBCs	It is moderate to severe

Treatment:

1. 1. Prophylaxis: Avoid those factors known to precipitate crises, especially dehydration, anoxia,  infections, stasis of circulation, and cooling of the skin surface.
   2. Folic acid 5 mg once weekly should be given.
   3. Patients should have good general nutrition and hygiene.
   4. Sickle cell trait does not affect life unless there is severe hypoxia caused by shock, vigorous exercise, high altitude, and anesthesia, leading to vasooclusive episodes.
   5. Sickle cell disease needs supportive treatment for hemolytic anemia and to avoid crises.
   6. In the case of crises:
      1. Treat by rest, warmth, rehydration by oral fluids, or I/V normal saline. Can give 3 L in 24 hours.
      2. Antibiotics are given in case of infection.
      3. Analgesic is given in case of need, paracetamol, non-steroidal anti-inflammatory agents, and opiates.
      4. Blood transfusion is given in case of anemia.
   7. Stem cell transplantation can cure the disease, and nowadays, many patients are successfully treated.
      

      Various forms of red blood cells