1. Dr.Bharathi Sengodan.M.D,
Hemolytic anemia
All are characterized by
1) An increased rate of RBC destruction
2) A compensatory increase in erythropoeisis that results in reticulocytosis
3) The retention of products of RBC destruction (iron, Hb, indirect bilirubin). Heamoglobin breaks into Heme and globin. Heme cleaved by heme oxygenase into iron and biliverdin which is further catabolized to bilirubin.Bilirubin is excreted through the bile into GIT, converted to urobilinogen by bacterial reduction and a small fraction excreted in urine and stool which are indicators of hemolysis
4) Features of damaged red cells :Shortened RBC survival, morphological changes in RBC’s, osmotic fragility ect
5) Decrease in serum haptoglobin -protein that binds to free haemoglobin (Hb escapes into the plasma ,bound to haptoglobin. Hb + Haptoglobin complex is cleared and taken to the liver (if large amounts are formed haptoglobin in plasma will reduce)
Dr.Bharathi Sengodan.M.D,

2. Intrinsic or extrinsic hemolytic anemias
Intravascular hemolysis Intrinsic
(1) Hemolysis occurs within blood vessels.
Causes of hemolysis
Enzyme deficiency (e.g., deficiency of glucose-6-phosphate dehydrogenase)
(b) Complement destruction (e.g., IgM-mediated hemolysis)
(c) Mechanical damage (e.g., calcific aortic valve stenosis)
Extravascular hemolysis or Extrinsic RBC phagocytosis by macrophages in the spleen (most common site) and liver
Reasons for phagocytosis
RBCs coated by IgG with or without C3b (immune destruction)
(b) Abnormally shaped RBCs (e.g., spherocytes, sickle cells)

3. Dr.Bharathi Sengodan.M.D,
Intravascular Hemolysis Destruction in the vascular compartment
Increased plasma and urine Hb (Hemoglobinuria)
Hemosidenuria(Renal tubules convert iron in Hb into hemosiderin.)
Methaemoglobinaemia
Levels of serum haptoglobin,a protein that binds to free Hb is characteristically LOW
Increased serum LDH from hemolyzed RBCs
Extravascular Hemolysis
Destruction by RES namely spleen and liver e.g. damage to cell membrane/altered surface protein causing hemolysis.
Increase in serum unconjugated bilirubin (End-product of macrophage degradation of Hb)
Increased serum lactate dehydrogenase (LDH) from hemolyzed RBCs
Jaundice and gall stones seen in long standing cases
Serum haptoglobin reduced
Not associated with hemoglobinemia or hemoglobinuria
Dr.Bharathi Sengodan.M.D,

4. RBC Haem (iron+protoporphrin) + Globin
Extravascular hemolysis
Intravascular hemolysis
RBC Haem (iron+protoporphrin) + Globin
RBC
Globin Iron Protoporphyrin
LYSIS AA
Bilirubin
Unconjugated bilirubin
Binds to transferrin Hb
MethHb
Bilirubin glucornidase
reabsorbed
Hemoglobinuria
Heamosiderinuria
Urobilinogen
Stercobilin

5. Dr.Bharathi Sengodan.M.D,
Lab findings
Features of increased red cell break down:
Serum bilirubin raised, unconjugated and bound to albumin
Urine urobilinogen increased
Fecal stercobilin increased
Serum haptoglobins absent (bcoz they are saturated with Hb and the complex is removed in the RE cells)
Features of increased red cell production
Reticulocytosis
Bone marrow Erythroid hyperplasia
M:E ratio is normally
2-12:1 but reversed to 1:1
Damaged red cells
a) Microspherocytes,
elliptocytes, fragments ect
b) Osmotic fragility, Auto hemolysis test ect
C) Red cell survival shortened - tested by Cr labeling
Dr.Bharathi Sengodan.M.D,

6. Important morphological changes occurring in hemolytic anemia
Tissue hypoxia due to anemia causes increased erythropoeisis
Extramedullary heamopoeisis in the spleen and liver leading to enlargement of organs
Expansion of bone marrow causing pressure atrophy of the inner aspect and neo-osteogenesis of the outer aspect of bones mainly of ribs,facial ect
Elevated levels of bilirubin leading to pigment stones in the gall bladder
With chronicity,the phagocytosis red cells are broken down to contribute to accumulation of iron in the phagocytic cells of the RE system thereby leading to hemosiderosis

7. Hereditary spherocytosis
Membrane cytoskeleton disorder
Autosomal dominant inheritance, Rarely autosomal recessive, M=F
Intrinsic defect, extravascular hemolysis
Membrane protein defect results in the loss of RBC membrane and spherocyte formation.
(1) Mutation in ankyrin is the most common defect. (2) Mutation in band 2, spectrin (α and β), or band 3 account for other defects.
Dr.Bharathi Sengodan.M.D,

9. Dr.Bharathi Sengodan.M.D,
Pathogenesis
Cytoskeletal abnormality leads to reduced membrane stability due to loss of membrane surface area relative to intracellular volume
Increased permeability of spherocytes to sodium due to membrane defect and dysfunctional Na+/K+-ATPase pump
This leads to the spheroidal shape and decreased deformability of the cell
The loss surface area results from increased membrane fragility
This spherical RBC’S get trapped in splenic cords, get destroyed
Shortened life span
Dr.Bharathi Sengodan.M.D,

10. Dr.Bharathi Sengodan.M.D,
Diagnosis
(1)Family history (2)Clinical features (3)Lab
Clinical features
Anemia
Splenomegaly
Jaundice due to increased unconjugated bilirubin
Increased incidence of black calcium bilirubinate gallstones (Due to increased concentration of conjugated bilirubin in bile)
Aplastic crisis: parvovirus induced [In children after a viral infection (e.g., parvovirus)]
Dr.Bharathi Sengodan.M.D,

11. Laboratory Diagnosis
Decreased Hb, Increased MCHC , Reticulocytosis- 5-20%
PBS : Normocytic anemia with Microspherocytes which are densely staining with smaller diameters than normal red cells, Lack the area of central pallor
Osmotic fragility test (measures the sensitivity of the red cells to lysis in vitro to swelling, caused by incubation in increasingly hypotonic saline solutions)
Increased RBC osmotic fragility (1) Increased permeability of spherocytes to sodium and water (2) Spherocytes rupture in mildly hypotonic salt solutions.
Bilirubin raised

12. Dr.Bharathi Sengodan.M.D,
SICKLE CELL ANEMIA
Geographic distribution: Africa, Mediterranean regions
Americas and Asia (SA, India)
Basic defect is production of an abnormal globin chain
Hereditary disorder : Both Autosomal recessive and dominant inheritance
Homozygous: Inheritance of two genes from each parent coding for the same type of abnormal hemoglobin, e.g., Hb SS
Heterozygous: Inheritance of genes from each parent which code for a different type of abnormal hemoglobin each, e.g., Hb SC
Protective against Plasmodium falciparum malaria
Dr.Bharathi Sengodan.M.D,

13. Pathophysiology of sickle cell anemia.
Missense point mutation
HbS results from a single amino acid substitution
Substitution of valine for glutamic acid in the sixth position of the beta chain

14. Causes of sickling
(1) HbS molecules aggregate and polymerize into long needle-like fibers. RBCs assume a sickle shape
(2) Sickle Hb (HbS) concentration greater than 60% is the most important factor for sickling.
(3) Increase in deoxyhemoglobin (correlates with decreased O2 saturation) increases the risk for sickling due to
(a) Acidosis : Right-shifts the OBC causing O2 release from RBCs
(b) Volume depletion : Intracellular dehydration
(c) Hypoxemia Sickling: Decrease in arterial Po2 decreases O2 saturation of Hb.

15. Clinical features :Key pathologic processes in HbSS
Reversible and irreversible sickling
(1) Initial sickling is reversible with administration of O2.
(2) Recurrent sickling causes irreversible sickling due to membrane damage.
(3) Irreversibly sickled cells have increased adherence to endothelial cells in the microcirculation causing Microvascular occlusions (vaso-occlusive crises) produce ischemic damage.
HbF prevents sickling. (1) Increased HbF at birth prevents sickling in HbSS for 5 to 6 months.
Clinical features :Key pathologic processes in HbSS
(1) Severe hemolytic anemia
(2) Painful vaso-occlusive crises

16. (1) Hemolytic anaemia
(2).Painful vascular-occlusive crises
Precipitated by infection, acidosis, dehydration or deoxygenation (altitude,obstetric delivery, stasis of circulation, exposure to cold, violent exercise)
Infarcts may occur in the bones, lungs and spleen and the most serious one is of the brain or spinal cord
Hand foot syndrome ( painful dactylitis due to infarcts of the small bones) is common in children 6-9 months old
Increased susceptibility to infections :Due to dysfunctional spleen (Impaired opsonization of encapsulated bacteria) Streptococcus pneumoniae sepsis & osteomyelitis due to Salmonella paratyphi;

17. 2.Visceral sequestration crises :Sickling within organs and pooling of blood
3.Aplastic crises: Due to infection with parvovirus and/ folate deficiency & Reticulocytopenia present
4.Hemolytic Crises: Increased rate of hemolysis
5. Autosplenectomy
(1) Spleen is enlarged but dysfunctional by 2 years of age. Nuclear remnants (Howell-Jolly bodies) appear in RBCs indicating loss of macrophage function
(2) Spleen is fibrosed and diminished in size in young adults. ( autosplenectomy)
5.Others :
Lower leg ulcers due to vascular stasis and ischemia
A chronic damage to liver and pigment gall stones
Renal findings : microhematuria due to infarctions & Renal papillary necrosis may occur.

19. LABORATORY FINDINGS LOW Hemoglobin
PBS: Red cells are normochromic .Sickle cells (in sickle cell disease), target cells, polychromasia indicative of reticulocytosis, & Howell-Jolly bodies (splenic atrophy)
WBC and platelet count is elevated
Sickling test are positive
Raised Serum bilirubin (unconjugated) and LDH
Haptoglobin levels are reduced
Hemoglobin electrophoresis : HbSS profile-HbS 90% to 95%, HbF 5% to 10%, no HbA
Prenatal screening Target cells: excess RBC membrane; Analysis of fetal DNA to detect the point mutation

20. Malaria and sickle cell anemia
Malaria is a mosquito borne parasite (Plasmodium falciparum) that invades RBC's.
Persons who have the sickle-cell trait have an unusually high resistance to infection by the parasites
the HbS in their RBC's interferes with the ability of the parasite to reproduce
Dr.Bharathi Sengodan.M.D,

21. Dr.Bharathi Sengodan.M.D,

22. Dr.Bharathi Sengodan.M.D,
G6PD deficiency Intrinsic defect with predominantly intravascular hemolysis
Most experience little or no anemia unless exposed to precipitating event or drug
precipitants:
infections
sulfa, primaquine, dapsone
fava beans
Laboratory diagnosis
Bite cells
Heinz bodies
measure G6PD level
Most common inherited red cell enzymopathy
X-linked
Hemolysis is due to increased oxidative damage to red cells
Dr.Bharathi Sengodan.M.D,

23. Decreased synthesis of reduced form of nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione (GSH) in the pentose phosphate pathway
(1) GSH normally neutralizes hydrogen peroxide, an oxidant product in RBC metabolism .
(2) In G6PD deficiency, in the absence of GSH peroxide oxidizes Hb, which precipitates in the form of Heinz bodies.
(a) Heinz bodies damage the RBC membranes causing intravascular hemolysis.
(b) Heinz bodies removed from RBC membranes by splenic macrophages produce bite cells.

24. Paroxysmal nocturnal hemoglobinuria (PNH)
Pathogenesis : loss of anchor for DAF due to mutation [Normally DAF destabilizes C3 and C5 convertase adhering to RBCs, platelets, and neutrophils (Prevents activation of the membrane attack complex and subsequent lysis of RBCs, neutrophils, and platelets)]
In PNH:Intravascular complement-mediated lysis of RBCs, neutrophils, and platelets

25. Peripheral blood findings :Normocytic anemia with pancytopenia
Clinical findings :
Episodic hemoglobinuria ,Increased incidence of vessel thrombosis (e.g., hepatic vein) Due to the release of aggregating agents from destroyed platelets
Peripheral blood findings :Normocytic anemia with pancytopenia
Decreased leukocyte alkaline phosphatase & serum haptoglobin
Increased serum/urine Hb
Identify the defect on hematopoietic cells -sensitive test
Older tests PNH:
Screening test is the sucrose hemolysis test (sugar water test).-Sucrose enhances complement destruction of RBCs.
Confirmatory test is the acidified serum test (Ham test). -Acidified serum activates the alternative pathway causing hemolysis.