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PA16.1-3 | Haemolytic Anaemia — Definition, Classification & Lab Markers — Part 2

Classification Axis 3 — Defect Location: Intracorpuscular vs Extracorpuscular

The third axis localises the defect to inside or outside the red cell.

Intracorpuscular defects (the abnormality is intrinsic to the red cell)
• Membrane defects: Abnormal spectrin, ankyrin, or band 3 proteins → loss of membrane deformability → spherocytes or elliptocytes → trapped and destroyed in splenic sinusoids.
• Enzyme defects: Deficient G6PD → failure of glutathione reduction → oxidant damage to Hb → Heinz body formation → 'bite cells' on smear; deficient pyruvate kinase → failure of ATP generation → rigid cells → extravascular destruction.
• Haemoglobin defects: Abnormal β-globin (sickle mutation) → sickling under deoxygenation → vaso-occlusion + haemolysis; quantitative globin defects (thalassaemia) → chain imbalance → precipitation, inclusion bodies, ineffective erythropoiesis.

Extracorpuscular defects (the red cell itself is normal at the start; the environment destroys it)
• Immune: Antibodies (IgG in warm AIHA, IgM in cold AIHA) coat the RBC surface → opsonisation → extravascular destruction in spleen.
• Mechanical: Fibrin strands in microvasculature (DIC, TTP, HUS) or mechanical heart valves shear passing red cells → fragmentation → schistocytes on smear.
• Toxic/infectious: Malaria parasite directly lyses the erythrocyte from within; Clostridium toxins degrade membrane phospholipids.
• Hypersplenism: Excessively active spleen (portal hypertension, storage diseases) → non-selective destruction of RBCs, WBCs and platelets.

The diagnostic shortcut: The Ham test (acidified serum lysis test) — now largely replaced by flow cytometry for CD55/CD59 — was classically used to identify PNH, the one condition that straddles both axes: it is acquired (somatic GPI-anchor mutation) but intracorpuscular (the defect is within the cell's membrane proteins).

General Clinical Features of Haemolytic Anaemia

The clinical presentation of haemolytic anaemia reflects three converging processes: reduced oxygen-carrying capacity (anaemia), excess haem catabolism (hyperbilirubinaemia), and compensatory erythroid hyperplasia (marrow expansion and splenomegaly).

1. Anaemia syndrome (when decompensated)
• Pallor, fatigue, exertional dyspnoea, tachycardia, haemic murmur — shared with all anaemias.
• In children with chronic haemolysis: growth retardation, delayed puberty.

2. Jaundice
• Predominantly pre-hepatic (unconjugated bilirubin from haem catabolism). Mild-to-moderate, often lemon-yellow ('acholuric jaundice' — urine not dark unless there is also haemoglobinuria).
• Urine: normal colour (extravascular) or red/brown (intravascular haemoglobinuria).
• Stool: normal or dark (urobilinogen overflow).
• NOT a sign of liver disease — LFTs (ALT, AST) are typically normal unless there is concomitant hepatic pathology.

3. Splenomegaly
• Results from: (a) macrophage hyperplasia in the red pulp (phagocytic work), (b) extramedullary erythropoiesis in severe chronic haemolysis.
• Splenomegaly, in turn, can worsen haemolysis (hypersplenism) — a vicious cycle.

4. Pigment (bilirubin) gallstones
• Chronic bilirubin overload → supersaturation of bile with bilirubin → precipitation as calcium bilirubinate stones.
• Found in 30–60% of patients with chronic hereditary haemolytic anaemias (hereditary spherocytosis, sickle-cell, thalassaemia intermedia).
• May present as cholecystitis or biliary obstruction — sometimes the first clinical event that leads to a haemolysis diagnosis.

5. Leg ulcers
• Seen in sickle-cell disease and, less commonly, other haemolytic anaemias.
• Mechanism: local sickling + capillary occlusion + tissue hypoxia (sickle); also chronic ↓haptoglobin → free Hb scavenges NO → vasoconstriction → ischaemia.

6. Acute haemolytic crisis
• Sudden acceleration of haemolysis — fever, rigors, severe back/abdominal pain, haemoglobinuria, acute kidney injury (from Hb cast nephropathy).
• Triggers: infection (G6PD crisis, sickle acute chest), drugs (dapsone in G6PD), transfusion mismatch.

7. Aplastic crisis (parvovirus B19)
• B19 infects and kills erythroid progenitors → sudden halt of red cell production → haemoglobin crashes over 7–10 days.
• A patient in compensated haemolysis depends on high reticulocyte output; removing that output is catastrophic.
• Resolves spontaneously in 1–2 weeks as immune clearance of B19 occurs.

IMPORTANT: The clinical triad of anaemia + jaundice + splenomegaly in the absence of hepatocellular disease should always raise the suspicion of haemolytic anaemia.

Haematologic Indices and Laboratory Markers of Haemolysis

The laboratory diagnosis of haemolysis rests on demonstrating increased RBC destruction and compensatory erythropoiesis simultaneously.

Markers of increased red cell destruction:

1. Serum unconjugated (indirect) bilirubin ↑
• Haem → biliverdin → unconjugated bilirubin (in macrophages). Carried on albumin to liver. ↑ in both intra- and extravascular haemolysis, but more markedly elevated in extravascular.
• Typically 2–4 mg/dL in mild/compensated haemolysis; may exceed 5–6 mg/dL in acute haemolytic crises.
• Conjugated bilirubin is normal (or only mildly ↑) unless secondary hepatic dysfunction exists.

2. Serum lactate dehydrogenase (LDH) ↑
• Released from lysed red cells (predominantly LDH-1 and LDH-2 isoforms).
• A sensitive but non-specific marker — ↑ in any cause of cell lysis (MI, liver disease, haemolysis).
• Very high LDH (>10× normal) is a hallmark of intravascular haemolysis, especially PNH and MAHA.

3. Serum haptoglobin ↓
• Haptoglobin is a plasma protein that binds free haemoglobin 1:1 (alpha–beta subunit binding). The Hb-haptoglobin complex is rapidly cleared by hepatocytes (T½ ~minutes).
• Even small amounts of intravascular haemolysis deplete haptoglobin rapidly → ↓serum haptoglobin.
• Extravascular haemolysis also causes mild haptoglobin depletion (trace free Hb released during splenic phagocytosis).
• Sensitivity ~83%, specificity ~96% for haemolysis when <25 mg/dL. Note: haptoglobin is an acute-phase reactant — may be spuriously normal or elevated in active infection/inflammation despite concurrent haemolysis.

4. Haemoglobinuria (intravascular haemolysis)
• Free plasma Hb exceeds renal threshold (~130–140 mg/dL) → passes glomerular filtration → haemoglobinuria (red/brown urine, dipstick positive for 'blood' with no RBCs on microscopy).

5. Urinary haemosiderin (chronic intravascular haemolysis)
• Renal tubular cells absorb filtered Hb, catabolise it, and accumulate ferritin/haemosiderin granules → exfoliate into urine → Prussian blue-positive cells in urine sediment.
• Appears 2–3 days after onset of intravascular haemolysis; persists for weeks after the acute episode. Highly specific for chronic intravascular haemolysis.

6. Urine urobilinogen ↑
• Conjugated bilirubin → gut bacteria → urobilinogen → partly reabsorbed → excreted in urine. ↑ in haemolysis because of bilirubin overflow into gut.

Markers of compensatory erythropoiesis:

7. Reticulocyte count ↑ — the hallmark marker of marrow response
• Reticulocytes are immature red cells released from marrow 1–2 days after erythropoietic stimulus. Normal: 0.5–1.5% (absolute count ~50,000–100,000/µL).
• In haemolysis: reticulocyte % typically 5–20% (absolute count 200,000–500,000/µL).
• The corrected reticulocyte count (or reticulocyte production index, RPI) accounts for the degree of anaemia: RPI = (Reticulocyte % × patient Hct / 45) ÷ maturation factor. RPI >3 = hyperproliferative (haemolysis or blood loss); RPI <2 = hypoproliferative.
• Polychromasia (bluish-staining large cells on Romanowsky stain) is the smear correlate of reticulocytosis.

8. Nucleated red blood cells (nRBCs) on peripheral smear
• Released from marrow in response to severe haemolytic stress (marrow 'push').
• Normally not seen in peripheral blood after the neonatal period.

Peripheral smear morphology clues:

Morphology	Associated disorder
Spherocytes	Hereditary spherocytosis, warm AIHA
Elliptocytes/ovalocytes	Hereditary elliptocytosis
Sickle cells (drepanocytes)	Sickle-cell disease
Target cells	HbC disease, thalassaemia, liver disease
Schistocytes/helmet cells	MAHA (DIC, TTP, HUS) — intravascular
Bite cells/blister cells	G6PD deficiency (oxidant haemolysis)
Polychromasia	Any haemolysis (reticulocytes)
Nucleated RBCs	Severe haemolysis, extramedullary erythropoiesis

Morphological Clues to Hemolysis: Peripheral Blood Smear Features