Page 13 of 20

PA16.1-3 | Acquired Haemolytic Anaemias & Smear Morphology — Part 1

CLINICAL SCENARIO

A 28-year-old woman presents with sudden onset pallor, jaundice, and dark urine after a viral upper respiratory illness. Her Hb is 6.2 g/dL, reticulocyte count 12%, bilirubin elevated. The peripheral smear shows numerous small, densely stained red cells with no central pallor.

Two questions cut through the differential immediately:
1. What does the smear tell you about the mechanism?
2. Is the process immune-mediated?

The answer to both sits in a single tube of blood. This module is about reading that evidence.

WHY THIS MATTERS

Acquired haemolytic anaemias test pattern-recognition skills that will serve you in:
• Emergency medicine — TTP (schistocytes + thrombocytopenia + fever) is a haematological emergency requiring urgent plasmapheresis.
• Internal medicine — Autoimmune haemolysis complicates SLE, lymphoma, and many drugs.
• Tropical medicine — Malaria remains the most common infectious cause of haemolytic anaemia worldwide.
• Surgery — Mechanical valve haemolysis and post-cardiac-surgery DIC both produce schistocytes.

PA16.3 tests your ability to describe aetiology, pathogenesis, haematologic indices, and peripheral blood picture — the smear is not optional knowledge.

RECALL

Before proceeding, recall from the previous SDL (Hereditary Haemolytic Anaemias):
• Extravascular haemolysis — destruction in splenic/hepatic macrophages; produces unconjugated hyperbilirubinaemia + splenomegaly; haptoglobin mildly reduced.
• Intravascular haemolysis — direct lysis in vessels; produces haemoglobinaemia, haemoglobinuria, haemosiderinuria; haptoglobin markedly reduced or absent.
• General lab signature of any haemolytic anaemia: ↓Hb + ↑reticulocyte count + ↑LDH + ↑indirect bilirubin + ↓haptoglobin.

Acquired causes simply add a new layer — the trigger is extrinsic, not a genetic membrane or enzyme defect.

Classification Framework: Acquired Haemolytic Anaemias

A three-panel classification diagram organizes acquired haemolytic anaemias by mechanism, highlights immune versus non-immune clinical decision-making, and illustrates Warm AIHA pathogenesis. — **Panel A:** Main classification framework showing Acquired Haemolytic Anaemias divided into immune-mediated, MAHA, direct membrane injury, hypersplenism, and physical/mechanical mechanisms with key examples.. **Panel B:** Clinical decision split comparing immune haemolysis, where immunosuppression may help, with non-immune haemolysis, where treatment targets the underlying cause.. **Panel C:** Warm AIHA mechanism showing IgG-coated RBCs, Rh system surface antigen, 37°C antibody binding, macrophage Fc receptor interaction, and extravascular haemolysis..

Acquired causes are best organised by mechanism:

Mechanism	Key examples
Immune-mediated	Warm AIHA, Cold AIHA, Drug-induced, Alloimmune (transfusion reaction, HDN)
Microangiopathic (MAHA)	TTP, HUS, DIC, mechanical prosthetic valves, malignant hypertension
Direct membrane injury	PNH (complement), malaria, Clostridium toxin, snake venom
Hypersplenism	Portal hypertension, infiltrative splenomegaly
Physical/mechanical	March haemoglobinuria, thermal injury

The immune vs. non-immune split is immediately clinically actionable because immunosuppression helps one group and harms the other.

Warm Autoimmune Haemolytic Anaemia (Warm AIHA)

Diagram of Warm AIHA showing smear spherocytes and reticulocytes, comparison of normal RBCs with spherocytes, and IgG-mediated splenic extravascular haemolysis. — **Panel A:** Peripheral smear showing normal RBCs with central pallor, spherocytes, polychromatic macrocytes / reticulocytes, and optional nucleated RBC in severe haemolysis.. **Panel B:** Morphological comparison between normal biconcave erythrocyte with central pallor and dense rigid spherocyte with loss of central pallor and reduced surface-to-volume ratio.. **Panel C:** Warm AIHA pathogenesis showing IgG coating at 37 degrees Celsius, Fc gamma receptor-mediated partial splenic macrophage phagocytosis, spherocyte formation, splenic trapping, extravascular haemolysis, and DAT-positive result..

Warm AIHA is the most common form (~80% of AIHA). Autoantibodies (predominantly IgG, subclasses 1 and 3) bind red cell surface antigens (typically Rh system) optimally at 37°C.

Pathogenesis:
1. IgG coats the red cell (opsonisation).
2. Splenic macrophages bear Fcγ receptors — they partially ingest the IgG-coated membrane.
3. Partial phagocytosis reduces surface area without removing haemoglobin → the cell reseals as a spherocyte (reduced surface-to-volume ratio, no central pallor).
4. Spherocytes are rigid and are trapped + destroyed in the splenic sinusoids → extravascular haemolysis.

Peripheral blood smear showing spherocytes and polychromatic cells in warm autoimmune hemolytic anemia with comparison diagrams of normal versus abnormal red blood cells. — **Panel A:** Peripheral blood smear (100× oil immersion, Leishman stain) showing spherocytes (dense, round cells lacking central pallor) and polychromatic macrocytes (reticulocytes) with labeled arrows. **Panel B:** Morphological comparison between normal biconcave red blood cells with central pallor versus spherocytes showing loss of biconcave shape and central pallor. **Panel C:** Pathophysiology flowchart of warm AIHA showing IgG antibody coating, splenic sequestration, spherocyte formation, and positive DAT result.

Peripheral smear: Spherocytes (densely staining, small, no biconcave disc shape) + polychromasia (reticulocytes) + nucleated RBCs in severe cases.

Causes (secondary warm AIHA): Lymphoma (especially CLL, diffuse large B-cell), SLE, drugs (methyldopa, penicillin), solid tumours.

Lab: DAT positive (IgG ± C3d), indirect Coombs variably positive.

SELF-CHECK

A patient with chronic lymphocytic leukaemia (CLL) develops progressive anaemia. Peripheral smear shows spherocytes and polychromasia. Which finding on the direct antiglobulin test (DAT) best confirms immune haemolysis?

A. Positive for IgG alone

B. Positive for C3d alone

C. Positive for IgM alone

D. Negative DAT

Reveal Answer

Answer: A. Positive for IgG alone

Warm AIHA (the type associated with CLL) is mediated by IgG antibodies coating red cells. The DAT (direct Coombs) detects these antibodies bound to the patient's own red cells. IgG positivity, often with C3d, is the hallmark. Cold AIHA (IgM) activates complement, leaving C3d on the cell surface but not IgM (IgM dissociates at 37°C). A negative DAT in this scenario would argue against immune haemolysis (though rare DAT-negative AIHA exists).

Cold Autoimmune Haemolytic Anaemia (Cold AIHA)

Diagram showing cold AIHA pathogenesis with IgM binding RBCs in cool peripheral vessels, complement deposition, hepatic haemolysis, RBC agglutination, DAT C3d positivity, and common causes. — **Panel A:** Cool peripheral vessels, IgM cold agglutinins, RBC I/i antigens, classical complement pathway, C3b deposition, Kupffer cell destruction in liver, C5b-9 membrane attack complex, intravascular haemolysis, 37°C IgM dissociation, persistent C3d.. **Panel B:** Temperature comparison showing separated RBCs at 37°C and clumped RBC agglutination at 4°C or room temperature.. **Panel C:** Peripheral blood smear showing irregular grape-like RBC clumps, RBC agglutination, spuriously high MCV, and spuriously low RBC count.. **Panel D:** DAT pattern with C3d positive and IgG negative, plus causes: Mycoplasma pneumoniae anti-I, infectious mononucleosis anti-i, lymphoma, and idiopathic..

Cold AIHA is mediated by IgM antibodies (cold agglutinins) that bind red cell surface antigens (typically I/i antigens) maximally at 4°C and dissociate at 37°C.

Pathogenesis:
1. In peripheral vessels (extremities, ears, nose — cooler temperature), IgM binds to red cells and activates the classical complement pathway.
2. C3b deposition opsonises red cells for hepatic (Kupffer cell) destruction → extravascular, predominantly hepatic haemolysis.
3. If complement activation proceeds to C5b-9 (membrane attack complex), intravascular lysis also occurs.
4. At 37°C (central circulation), IgM dissociates — but C3d remains bound. Hence DAT is positive for C3d, negative for IgG.

Peripheral smear hallmark: RBC agglutination — clumps of red cells visible on smear (spuriously high MCV, spuriously low RBC count on automated analyser).

Blood smear showing red cell agglutination in cold autoimmune hemolytic anemia with grape-like clumps of RBCs and comparison of normal versus clumped cells. — **Panel A:** Peripheral blood smear (40× objective, Leishman stain) showing irregular grape-like RBC clumps with IgM cold agglutinin labeling. **Panel B:** Temperature-dependent RBC behavior comparison - normal single cells at 37°C versus clumped agglutination at room temperature.

Causes: Mycoplasma pneumoniae (anti-I), infectious mononucleosis (anti-i), lymphoma, idiopathic (elderly).

Clinical: Acrocyanosis, Raynaud's phenomenon in cold exposure; haemoglobinuria after cold exposure.

SELF-CHECK

An automated blood analyser reports MCV 130 fL and RBC count 1.8 × 10¹²/L, but on smear the cells look normocytic and the cell count appears much higher. What is the most likely explanation?

A. Megaloblastic anaemia (B12 deficiency)

B. Cold agglutinin disease causing RBC clumping

C. Hereditary spherocytosis

D. Multiple myeloma with rouleaux

Reveal Answer

Answer: B. Cold agglutinin disease causing RBC clumping

Cold agglutinins (IgM) cause red cells to clump at room temperature — the temperature at which automated analysers run. The analyser counts each clump as one large 'cell', producing a spuriously high MCV and spuriously low RBC count. The discrepancy between the machine result and smear appearance is itself a diagnostic clue. Warming the blood sample to 37°C before re-running resolves the artefact.