Haemolytic Anaemia

• Hemolytic anemias (HA) cause an increase in the rate of red cell destruction.
• In hemolytic disorders, red cells are destroyed prematurely.
• Red cell destruction occurs usually after 120 days.
• Red cells are removed extravascularly by macrophages in the reticuloendothelial (RE) system, especially in the marrow, liver, and spleen.
• The breakdown of red cells liberates iron for recirculation via plasma transferrin to marrow erythroblasts.
• Protoporphyrin is broken down to bilirubin, circulates to the liver, is conjugated to glucuronides, and excreted into the gut via bile, then converted to stercobilinogen and stercobilin, which is excreted in feces.
• Stercobilinogen and stercobilin are partly reabsorbed and excreted in urine as urobilinogen and urobilin.
• A small fraction of protoporphyrin is converted to carbon monoxide (CO) and excreted via the lung
• Globin chains are broken down to amino acids which are reutilized for general protein synthesis. -Haptoglobins are proteins present in normal plasma can bind hemoglobin, which is then removed from plasma by the RE system.

Laboratory Diagnosis Criteria

• Reticulocytosis.
• Serum level of unconjugated/conjugated bilirubin.
• Serum level of lactic dehydrogenase (LDH) is elevated.
• Serum haptoglobin level is decreased.
• Peripheral blood smear shows morphologic changes in the red blood cells consistent with hemolysis.
• Spherocytes suggest hereditary spherocytosis or immunohemolytic anemia.
• Sickle cells suggest sickle cell syndromes.

Hemolysis Sites

• Intravascular hemolysis: Red blood cells are destroyed directly within the circulatory system.
• Extravascular hemolysis: Destruction of red blood cells occurs within mononuclear phagocytic cells, often in the spleen and is more common.

Classification of HA

• Extrinsic HA: Originates from a source outside the red cell and disorders extrinsic to the RBC are usually acquired; the RBC may get damaged by chemicals, mechanical or physical agents or immune destruction, and Hemolysis occurs either intravascularly or extravascularly.
• Intrinsic HA: Due to defects of the RBC itself and are usually inherited; the abnormality can be in the membrane, cell enzymes, or the hemoglobin molecule, and the cells that show intrinsic defects are hemolyzed extravascularly because the RBCs are not severely damaged.

Extrinsic Hemolytic Anemias

• Immune Hemolytic Anemia (IHA) is mediated by the immune response, especially humoral antibodies, resulting in a shortened RBC lifespan; IHA has 3 main types.

Autoimmune HA (AIHA)

• Autoimmune HA (AIHA): Patient makes an autoantibody against their own red cells and includes Warm AIHA and Cold AIHA.
• Alloimmune: The patient's antibody is directed against foreign red cells; includes Hemolytic transfusion reactions and Hemolytic disease of the fetus and newborn.
• Drug-induced: A drug-dependent or related antibody is responsible for hemolysis.

Autoimmune Hemolytic Anemia (AIHA)

• The immune system loses its ability for self-recognition and antibodies are made to own RBCs, which initiates hemolysis.
• AIHA is characterized by premature RBC destruction caused by autoantibodies that bind to the RBC surface.
• In AIHA, the loss of T suppressor cell control results in the production of autoantibodies against RBC self‐antigens.
• Autoimmune disease occurs because of genetic predisposition, exposure to infectious agents that induce antibody production, and defects in the mechanism regulating immune tolerance are all contributing factors.

Warm AIHA

• The majority of warm autoantibodies are of the IgG class and are optimal at 37°C.
• Extravascular hemolysis of the RBC.
• Red cells are coated with IgG alone, IgG and complement, or complement alone.
• The complement component detectable is C3d, the degraded fragment of C3.
• The disease may occur at any age or sex and presents as a hemolytic anemia of varying severity.
• Approximately 60% of cases are idiopathic, with the remaining percentage due to another underlying disease, such as lymphoma, chronic lymphocytic leukemia, or viral infections and immunodeficiency syndromes; AIHA in SLE is typically of the IgG + complement type.
• Anemia is moderate to severe.
• Normochromic, normocytic with Polychromasia.
• Spherocytes and schistocytes may be seen.
• Reticulocytosis increases.
• Autoantibody presence
• Direct Coombs test (DAT) is positive, testing for RBCs sensitized with IgG antibody or complement.

Cold AIHA

• The majority of cold autoantibodies are of the IgM class and optimal below 37°C.
• Cold AIHA has 2 main types: Cold Agglutinin Disease (CAD) and Paroxysmal Cold Haemoglobinuria.

Cold Agglutinin Disease (CAD)

• Occurs predominantly in older individuals with peak incidence after 50 years of age
• Affects men and women equally
• The antibodies in this disorder show the greatest reactivity with antigens at temperatures below 30° C; antibodies are usually of the IgM class and they bind complement
• Cases of cold agglutinin disease can be idiopathic or secondary to another underlying condition, such as infections with Mycoplasma pneumoniae or Infectious mononucleosis.
• Hemolytic syndromes of varying severity may occur depending on the titer of the antibody in the serum, its affinity for red cells, its ability to bind complement, and its thermal amplitude.
• Cold IgM antibody dissociates from red cells when they pass to the warmer central circulation, however complement remains bound leading to the destruction of red cells in the RE system, particularly the liver, giving rise to a chronic haemolytic anemia
• DAT remains positive for complement.

Paroxysmal Cold Haemoglobinuria (PCH)

• PCH is found associated with viral infection or syphilis, both chronic conditions.
• Autoantibodies to red blood cells bind to the cells in cold temperatures and fix complement, which can cause intravascular hemolysis upon warming.
• PCH is caused by biphasic IgG antibody that can bind at low temperature.

Alloimmune Hemolytic Anemia (AHA)

• AHA happens because of Alloantibodies development to an erythrocyte antigen that the individual lacks through transfusion or pregnancy.
• Recipient lymphocytes recognizes these RBCs are foreign which stimulates antibody production.
• Antibodies coat the foreign RBCs and shortens RBC survival with a positive DAT result.
• The two main types are Haemolytic transfusion reactions and Haemolytic disease of the fetus and newborn.

Haemolytic transfusion reactions

• Acute transfusion reaction causing Intravascular hemolysis occurs due to the complete activation of complement mediated by IgM antibodies.
• Delayed transfusion reaction: Occurs days to weeks after the transfusion with RBCs experience the process of extravascular hemolysis as they are coated with IgG antibodies and get remove by the macrophages, without complement being involved.
• Alloimmune Hemolytic Anemia (AHA)

Haemolytic disease of the fetus and newborn (HDN)

• Fetus RBCs are destroyed by maternal IgG antibodies crossing over the placenta.
• The newborn often presents with jaundice and anemia.

Autoimmune Hemolytic Anemia Drug-Induced AIHA

• Immune complex mechanism, the drugs bind to plasma proteins and Abs are made against the drugs and Abs made bind to the drug to form an immune complex which adsorbs non-specifically to patients' RBCs, activating complement causing intravascular hemolysis.
• Drug adsorption mechanism- Drugs bind unspeciifically to proteins on RBC triggering IgG Abs bind to the drug and cause extravascular haemolysis.
• Methyldopa induced mechanism- Drug induces autoantibodies causing extravascular haemolysis.
• Membrane Modification Mechanism

Non-immune Hemolytic Anemia

• Chemicals or antagonists in blood cause abnormalities in plasma lipids
• Animal venoms , such as bees and spiders
• Infectious agents , Malarial parasites and babeiosis

Intrinsic Haemolytic Anaemias

• Intrinsic HA has 2 main types, there are Membrane Defects and Enzyme Deficiency

Membrane Defects

• Defects are due to abnormalities in membrane proteins or lipids
• Defects alter the membrane's stability, shape, deformability and permeability
• Hemolysis occurs extra-vascularly

Skeletal protein abnormalities

• Separates the lipid bilayer from the skeletal lattice
• Results in the decrease in surface area-to-volume ratio ..spherocyte
• Disrupts the skeletal Lattice
• Destabilizes Membrane
• Cell fragmentation

Lipid composition abnormalities

• This presents as an excess of cholesterol accumulation in the outer bilayer of the RBC, which indicates Acanthocyte.

Hereditary spherocytosis (HS)

• A defect is in the ankyrin & spectrin
• Results in the formation of fragile spherocytic red cells
• Spherocyte becomes less flexible and more permeable to Na+
• Tends to affect Northern Europeans and is Inherited.

CBC for Hereditary spherocytosis (HS)

• Mild anemia
• MCV is usually normal (77-87f L)
• MCH normal
• MCHC is >36% which is the only condition in which an MCHC can be truly increased.
• RDW Increased and has a Spherocyte morphology.
• It presents with Varying degrees of polychromasia, anisocytosis and poikilocytosis.

Hereditary spherocytosis (HS) Bone Marrow results

• Presents as Normoblastic erythroid hyperplasia and Increased iron storage

Hereditary spherocytosis (HS) Chemistry Results

• Presents as Increased Bilirubin and Fecal urobilinogen and LD/LDH
• Shows a Decreased Haptoglobin

Immunohematology for Hereditary spherocytosis (HS)

• Is seen indicated by Negative DAT results

DIAGNOSTIC TESTS FOR HS

• Patients express Osmotic fragility that shows Cells incubated in decreasing concentrations of NaCl, since Spherocytes lyse sooner than normal red cells which indicates an increased value.
• Autohemolysis test involves Red cells being incubated at 37 C for 48 hours, increased hemolysis is visible due to the pretense of spherocytes.
• Red cell membrane studies presents the Membrane proteins being analyzed using gel electrophoresis.

Hereditary Elliptocytosis (HE)

• Occurs due to a skeletal proteins defect.
• Results in the formation of the fragile, elliptocytic red cells are sensitive to mechanical stress.
• More permeable to Na+ and has Increased sensitivity to heat.
• More commonly found in Africa and the Mediterranean
• Not evident Hemolysis
• Mild Anemia
• There is a Presence of Elliptocytes or ovalocytes.

Glucose-6-Phosphate Dehydrogenase Deficiency (G6PD)

• Disease is Sex-linked(X) disorder, Patients are Africans, Middle East, some Americas, and Mediterraneans
• Is the Most common enzyme disorder.
• Causes Denatured hemoglobin precipitates in the RBC when exposed to oxidative stress with hemolysis.
• Causes Oxidative stress (reduced GSH)triggers response and unable to generate NADPH to reduce GSH causing the System to overwhelm.
• Oxidized Hgb accumulates as Heinz bodies.

Causes of Oxidative Stress leading to G6PD

• Infections from hydrogen Peroxide release, which is sometimes not clear.
• Ingestion of oxidative drugs such Aantimalaria drugs (i.e Primaquine) or Quinine ,quinidine or even Analgesics.
• Ingestion of fava beans that is found in the Mediterranean area: Person eats or inhales fava bean or its pollen and is Hereditary There is a Laboratory Findings of Laboratory Findings:
• Decreased H&H (hemoglobin and hematocrit) and Hemoglobinuria.
• Presents as increased bilirubin and LD with Reticulocytosis.