Hemolytic Anemia Notes

Hemolytic Anemia

Objectives

• Define hemolysis and explain the term 'compensated hemolysis.'

• List hematological and biochemical indicators of red cell hemolysis.

• Classify the principal causes of red blood cell hemolysis (e.g., extracorpuscular and intracorpuscular) and cite examples from each category.

• List and briefly discuss laboratory tests used to differentiate between intravascular and extravascular hemolysis.

• Explain the principles of the osmotic fragility test.

• Compare the inheritance pattern of hereditary spherocytosis with that of glucose 6 phosphate dehydrogenase deficiency.

• Explain the difference between alloimmune and autoimmune hemolysis.

The Normal Removal Of The RBCs

• RBC life span is approximately 120 days.

• RBC destruction occurs after this mean life span.

• Removal happens in the extravascular space by the reticuloendothelial system (phagocytic system) in the bone marrow, spleen, and liver.

Hemolysis

• Hemolysis is the accelerated destruction of red blood cells (RBCs), leading to decreased RBC survival.

• The bone marrow's response to hemolysis is increased erythropoiesis, reflected by reticulocytosis.

• Compensated Hemolysis: If the rate of hemolysis is modest and the bone marrow can completely compensate for the decreased RBC life span, the hemoglobin concentration may be normal.

• Incompletely Compensated Hemolysis: If the bone marrow cannot completely compensate for hemolysis, anemia occurs.

• A normal hemoglobin value does not necessarily denote the absence of hemolysis.

Hemolysis Classification

Multiple ways to classify hemolysis:

• Inheritance: Inherited vs. Acquired

• Site of RBC destruction:

  • Intravascular: inside blood vessels (e.g., Paroxysmal nocturnal hemoglobinuria (PNH))

  • Extravascular: macrophages in the reticuloendothelial system (liver and spleen); more common (e.g., hereditary spherocytosis)

• Origin of RBC damage:

  • Intrinsic: to RBC, usually inherited (but not always, e.g., PNH)

  • Extrinsic: to RBC, usually acquired (but not always, e.g., congenital Thrombotic Thrombocytopenic Purpura (TTP))

Clinical Manifestations

• History/Physical:

  • Fatigue

  • Pallor

  • Jaundice

  • Splenomegaly (in some cases)

• Free hemoglobin scavenges nitric oxide, so chronically this can result in:

  • Esophageal spasm

  • Non-healing skin ulcers

  • Pulmonary hypertension

• Chronic intravascular hemolysis:

  • Hemosiderinuria

• Chronic extravascular hemolysis:

  • Pigmented gallstones

Haptoglobin

• Haptoglobins are proteins in normal plasma which bind hemoglobin if either intravascular or significant extravascular hemolysis is present.

• The hemoglobin–haptoglobin complex is removed by the RE system, leading to low haptoglobin levels in hemolysis.

Hemolysis Biomarkers

• Haptoglobin binds with Hb

• (Haptoglobin + Hb) Complex is removed by the mononuclear phagocytic system

• This leads to low haptoglobin levels

Indicators of Red Cell Hemolysis

1. Features of increased red cell breakdown:

   • (a) Serum bilirubin raised, unconjugated and bound to albumin

   • (b) Urine urobilinogen increased

   • (c) Serum haptoglobins absent because the haptoglobins become saturated with hemoglobin, and the complex is removed by RE cells.

2. Features of increased red cell production:

   • (a) Reticulocytosis

   • (b) Bone marrow erythroid hyperplasia – the normal marrow myeloid : erythroid ratio of 2:1 to 12:1 is reduced to 1:1 or reversed.

3. Damaged red cells, visualized by:

   • (a) Routine blood film morphology (e.g., microspherocytes, elliptocytes, fragments)

   • (b) Flow cytometry after eosin-maleimide (EMA) staining

   • (c) Specific enzyme, protein, or DNA tests.

Intravascular vs. Extravascular Hemolysis

• Intravascular hemolysis: erythrocytes are destroyed in the blood vessel itself.

• Extravascular hemolysis: occurs in the hepatic and splenic macrophages within the reticuloendothelial system.

Intravascular Hemolysis Details

• Red cell destruction occurs primarily in the vascular space.

• Damage to the RBC membrane is significant.

• Can be detected by blood test → Hemoglobinemia.

• Free hemoglobin in the urine. Hemoglobinuria → Acute renal failure.

• Hemosiderin → Iron deficiency anemia. The byproduct of hemolysis → free hemoglobin.

Causes of Intravascular Hemolysis

• Micro-angiopathic hemolytic anemia (MAHA): hemolytic anemia, thrombocytopenia, and microvascular thrombosis.

• Thrombotic thrombocytopenic purpura (TTP).

• Shear stress: aortic stenosis, defective prosthetic valve.

• Acute hemolytic transfusion reactions (e.g., ABO incompatibility).

• Infections: clostridia sepsis, severe malaria.

• Paroxysmal cold hemoglobinuria & cold agglutinin disease (cold AIHA).

• Paroxysmal nocturnal hemoglobinuria (PNH).

Extravascular Hemolysis Details

• Red cell destruction primarily in the Reticuloendothelial system.

• Examples:

  • Warm Autoimmune hemolysis

  • Delayed hemolytic transfusion reactions

  • Hemoglobinopathies (sickle cell and thalassemia)

  • Hereditary spherocytosis

AIHA (Autoimmune Hemolytic Anemia)

• Shortened survival of RBCs due to auto-antibodies.

• Warm AIHA:

  • Antibodies bind to RBCs optimally at body temperature (37^{\circ}C).

  • Almost always due to IgG.

  • 80-90% of AIHA.

• Cold AIHA:

  • Antibodies bind to RBCs optimally at temperature < 37^{\circ}C (optimum temperature of 3 to 4^{\circ}C).

  • Almost always due IgM.

  • Cold reacting IgG

  • 10-20% of AIHA.

Alloimmune Hemolytic Anemia

• Antibodies are directed against transfused RBCs/foreign RBC antigen:

  • Incompatible blood transfusion:

    • Acute hemolytic transfusion reaction.

    • Delayed hemolytic transfusion reaction.

  • Pregnancy.

Laboratory Testing for Hemolytic Anemia

Labs to Order - Intravascular vs Extravascular Hemolysis

Intracorpuscular vs. Extracorpuscular Hemolysis

• Intracorpuscular Causes:

  • Hereditary:

    • Enzyme defects:

      • G6PD deficiency

      • Pyruvate kinase deficiency

    • Defects in the RBC membrane:

      • Hereditary spherocytosis

      • Hereditary elliptocytosis

    • Hemoglobinopathies:

      • Thalassemia syndrome

      • Hb S, Hb C

  • Acquired:

    • Paroxysmal nocturnal hemoglobinuria

    • Infections from Clostridia and P.falciparum

• Extracorpuscular Causes:

  • Immune hemolytic anemia

  • Chemicals and toxins

  • Physical agents

  • Infections

  • Hypersplenism

  • Microangiopathic hemolytic anemia

  • Macroangiopathic hemolytic anemia

Intracorpuscular - Intrinsic Defect

• Membrane defect: Hereditary spherocytosis.

• Enzyme defect: G6PD deficiency.

• HB defect: Hemoglobinopathies.

Intracorpuscular vs. Extracorpuscular - Key Points

• Intrinsic hemolysis most of the time is due to congenital / inherited causes.

• Inherited TTP→ extrinsic hemolysis.

• Extrinsic hemolysis most of time is due to acquired causes.

• PNH (acquired)→ intrinsic hemolysis

RBC Membrane Disorders

• Include:

  • Hereditary spherocytosis

  • Hereditary elliptocytosis

  • Hereditary pyropoikilocytosis

• Characterized by:

  • Deficiency or dysfunction of one or more membrane proteins

  • Shortened RBC survival by hemolysis

Hereditary Spherocytosis

• Epidemiology:

  • Common in Northern Europeans

• Inheritance pattern:

  • 75% autosomal dominant

  • 25% sporadic

• Pathophysiology:

  • Aberrant interaction between the skeleton and overlying lipid bilayer

  • “Vertical interactions”

• Spherocytes

  • Don’t deform easily making it difficult to travel through the splenic vascular walls à further membrane loss

  • sphErocytes = Extravascular hemolysis

  • Causes: defect in RBC membrane:

    • Ankyrin

    • spectrin

    • Band 3 mutation

    • Protein 4.2

Osmotic Fragility Test

• Osmotic fragility testing: Increasing hypotonic saline solutions

• Increased RBC lysis compared with normal RBCs

• Sensitivity enhanced by 24 h incubation at 37^{\circ}C

• Not specific for HS (spherocytes of any cause will be positive)

Osmotic Fragility Test for HS

• Osmotic fragility is a test to measure red cell hemolysis

• Cell resistance to hemolysis when exposed to a series of increasingly dilute saline solution

• The sooner hemolysis occurs, the greater the osmotic fragility of the cells.

G6PD Deficiency

• Most frequently encountered abnormality of RBC metabolism

• Affects >200 million people worldwide

• Survival advantage with P. Falciparum

• Pathophysiology:

  • Hemolysis = due to failure to generate adequate NADPH

  • Leads to decreased levels of reduced glutathione

  • RBCs susceptible to oxidation of Hb by oxidant radicals

  • Denatured Hb aggregates form intraerythrocytic Heinz bodies

  • Heinz bodies bind to RBC cytoskeleton leads to reduced cellular deformity

  • Cells with Heinz bodies are trapped in the spleen, cell membrane becomes pitted and is lost, hemolysis

• Inheritance:

  • Gene for G6PD is on X-chromosome

Hemolysis Triggers in G6PD Deficiency

• Infections and other acute illnesses (e.g., diabetic ketoacidosis)

• Drugs:

  • Antimalarials (e.g., primaquine, pamaquine, chloroquine, Fansidar, Maloprim, quinine)

  • Sulphonamides and sulphones (e.g., co-trimoxazole, sulfanilamide, dapsone, sulfasalazine)

  • Other antibacterial agents (e.g., quinolones, nitrofurans, nalidixic acid, chloramphenicol)

  • Analgesics (e.g., aspirin); moderate doses are safe

  • Antihelminths (e.g., β-naphthol, stibophen)

  • Miscellaneous (e.g., vitamin K analogues, rasburicase, glibenclamide, naphthalene (mothballs), probenecid)

• Fava beans

• Chemical oxidants

N.B. Many common drugs have been reported to precipitate hemolysis in G6PD deficiency in some patients (e.g., aspirin, quinine, and penicillin), but not at conventional dosage.