W3L5 - Haemolytic Anaemia: Introduction & Intrinsic Defects Flashcards
Haemolytic Anaemia
Physiological haemolysis removes senescent RBC as part of the normal turnover of cells
Haemolytic anaemia refers to the anaemia caused by the increased & premature destruction of RBCs
Typically, initially normocytic normochromic
Complicated by
- increased erythropoiesis
- specific pathophysiology, e.g. spherocytes formation may ↓ MCV
Anaemia may not be evident if the rate of haemolysis is compensated (balanced) by increased erythropoiesis
Senescent Cell-Specific Antigen
Part of physiological haemolysis
Changes in the conformation of the membrane domain of band 3 lead to the appearance of a senescent cell-specific antigen
Recognised by specific, autologous immunoglobulin G
This marks senescent erythrocytes for removal by macrophages
Sites of Haemolysis
Intravascular
- occurs within the vasculature; i.e. while cells are circulating in the blood
Extravascular haemolysis
- occurs outside the vasculature; i.e. after cells have been removed from the circulating blood
Intravascular Haemolysis
Occurs within the circulation
Minority of cases (~10%)
‘Free’ Hb binds to haptoglobin or haemopexin
Transport proteins take Hb to liver for degradation
Excess Hb oxidises to methaemoglobin (MetHb, Fe3+ ) as no longer ‘protected’ by cell
MetHb dissociates to haemin and globin
Globin chains are broken down & AAs recycled
Haemin binds to haemopexin or albumin
Degraded in liver
Intravascular Haemolysis - What happens if Mechanisms are Overwhelmed
Free haemoglobin in the blood (plasma)
Haemoglobin filtered through the kidney => hb in the urine (haemoglobinuria)
Also methaemoglobinuria (Fe3+ in Hb in urine)
Haemosiderin may form from the haemoglobin present and be detected in the urine (haemosiderinuria)
- as excess haemoglobin is filtered by the kidney some is reabsorbed in the proximal convoluted tubule
- the iron is removed and converted to hemosiderin
- the tubule cells of the proximal tubule with the hemosiderin are lost into the urine
- can be demonstrated with Prussian blue stain
Mechanisms of Intravascular Haemolysis
Activation of complement on RBC membrane
Physical or mechanical trauma to RBC
‘Toxic’ plasma factors which act to disrupt the integrity of the cell
Extravascular Haemolysis
Occurs outside of circulation
Occurs within macrophages (spleen, liver, BM)
Majority of cases (~90%)
Hb degraded within phagocyte to haem & globin
Haeme degraded to iron, biliverdin and CO
Biliverdin converted to bilirubin, binds to albumin, & degraded by liver
Eventually degraded to urobilinogen & excreted in faeces
Causes of Extravascular Haemolysis
Inherited RBC Defects - e.g. Thalassaemias Acquired RBC Defects - e.g. Spur cell anaemia Autoimmune haemolytic anaemia (AIHA)
Difference between Extravascular and Intravascular Haemoglobin Degradation
Extravascular haeme degeneration differs from intravascular as it has:
- no release of Hb into plasma
- no haemoglobinaemia
- no haemoglobinuria
- no haemosiderinuria
Clinical Features of Haemolytic Anaemia
Primary clinical signs due to anaemia
- e.g. fatigue, pallor
Other clinical signs associated with increased RBC destruction
- e.g. jaundice, darkened urine (IV haemolysis)
- pigment gallstones may occur
Other changes:
- extra-medullary haematopoietic masses
- splenomegaly
- increased active (‘red’) marrow (may result in bone pain)
- tissue necrosis due to hypoxia
Laboratory Investigation of Haemolytic Anaemia
Haematological findings typically reflect:
- increased destruction of RBCs
- increased activity of bone marrow (erythropoiesis)
Steps:
1. Full blood count
2. Peripheral blood film
3. Bone marrow
4. Other laboratory test
Initial aim to distinguish intravascular from extravascular haemolysis
Laboratory Investigation of Haemolytic Anaemia - FBC and Peripheral Blood Film
FBC: - decreased Hb (anaemia) - variable MCV (macrocytic if reticulocytosis) - altered sub-populations of RBC Peripheral blood film: - poikilocytosis e.g. spherocytes, schistocytes - polychromasia (+++) - nRBCs - increased reticulocytes
Laboratory Investigation of Haemolytic Anaemia - Reticulocytosis and Bone Marrow
Reticulocytosis - takes ~48h - reflects increased erythropoiesis - peripheral blood Bone Marrow: - erythroid hyperplasia - orderly RBC maturation - normal RBC morphology - decreased M:E ratio
Osmotic Fragility
Gives an indication of the surface area:volume ratio of cells
Principle
- small amount of blood is mixed with large excess of buffered saline solution at various concentrations
- the fraction of RBC lysed at each saline solution is determined colorimetrically
- usually performed at room temperature
RBC that are spherocytic take up less water in hypotonic solution before rupturing
Intrinsic Haemolytic Anaemias
Reflect a deficit in membrane, enzymes, or protein characteristic of RBC that shorten the lifespan of the cell May be inherited or acquired Include: - hereditary spherocytosis - hereditary elliptocytosis - hereditary acanthocytosis - glucose-6-phosphate dehydrogenase deficiency - pyruvate kinase deficiency - paroxysmal nocturnal haemoglobinuria
Hereditary Spherocytosis
Membrane defect of main structural protein (spectrin)
Abnormal interactions between cytoskeletal proteins & membrane => microvesicles of membrane bud off
Loss of RBC shape: biconcave to spherical
Mild to moderate haemolysis
Osmotic fragility: increased
DAT: negative
Reticulocytosis
Hereditary Elliptocytosis
Defect in glycophorin C
Defective spectrin dimer dimer association
~1 in 4000 persons affected
Milder than hereditary spherocytosis
Cells acquire elliptical shape in circulation
Elliptocytes >25% & often >60%
Reticulocytes ~4%
Hereditary Stomatocytosis (Hydrocytes)
Membrane is abnormally permeable to Na+ & K+
Gain of Na+ > loss of K+ => excess H2O enters the cell
=> overly hydrated cells = ‘overhydrated hereditary stomatocytosis (OHS)’
Mild-moderate haemolytic anaemia
- 10-15% stomatocytes
- macrocytosis
- positive osmotic fragility test
- right shift of osmolarity curve
Hereditary Stomatocytosis (HSt)
HSt can be classified into:
- syndromic forms that show extra-hematologic signs
- non-syndromic forms with selective involvement of the erythroid system
Hereditary Stomatocytosis (HSt) - Syndromic Forms
Stomatin deficient cryohydrocytosis with mental retardation, seizures and hepatosplenomegaly
Phytosterolemia nonleaky stomatocytosis with macrothrombocytopenia
Dehydrated hereditary stomatocytosis with perinatal edema and/or pseudohyperkalemia
Hereditary Stomatocytosis (HSt) - Non-Syndromic Forms
Overhydrated hereditary stomatocytosis
Cryohydrocytosis
Dehydrated hereditary stomatocytosis
Familial pseudohyperkalemia
Hereditary Acanthocytosis
Inherited abnormality of lipids Absence of - serum β-lipoprotein - low serum cholesterol - low triglyceride - increased ratio of cholesterol to phospholipid Absence of anaemia Mild haemolysis Clinical features: - steatorrhea - retinitis pigmentosa - neurological abnormalities
Enzyme Deficiences
Mature RBCs cannot synthesise proteins
- limits life-span of normal RBC
- shortened life-span with enzyme deficiency
RBC depend on anaerobic glycolysis for ATP production by the Embden-Meyerhof pathway and Hexose monophosphate shunt
- maintains reduced GSH levels
- protects RBC from oxidative damage
Glucose-6-Phosphate Dehydrogenase Deficiency
Enzyme of hexose monophosphate shunt Deficiency or functional abnormality => inability to resist oxidative damage Sex linked inheritance Blood film - Heinz bodies - blister cells - bite cells - eccentrocytes/‘hemighost cells’ - fragments (crisis) Screening tests - fluorescent screening test - ↓ NADPH, ↓ fluorescence Definitive test - G6PD enzyme assay
Pyruvate Kinase Deficiency
Enzyme of glycolytic pathway Deficiency => - decreased ATP levels - Na/K ATPase pump activity - K+ loss, cell dehydration - echinocyte formation - splenic phagocytosis by macrophages Variable anaemia RBC Mx: echinocytes “prickle cells” Autosomal recessive inheritance Reticulocytosis (polychromasia) is prominent Bone marrow - erythroid hyperplasia
Paroxysmal Nocturnal Haemoglobinuria
Rare acquired stem cell disorder (~1 in 100,000 people)
Deficiency of glycosylphosphatidylinositol (GPI)
- GPI anchors surface proteins to membrane
- => absence of GPI linked proteins e.g. CD55, CD59
- CD 55: decay accelerating factor (DAF)
- CD 59: membrane inhibitor of reactive lysis (MIRL)
=> RBCs have abnormal sensitivity to complement
- inability to inhibit non specific C activation
Haemolysis occurs while sleeping (haemoglobinuria)
DAT & OF normal
Haemoglobinuria/Haemosiderinuria
Ham’s Test; RBC lysis at low pH
