HAEM: Haemolytic anaemias Flashcards by Kishan P

What is the normal RBC lifespan?

120 days

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Define haemolysis. How is it categorised?

Shortened RBC survival, may be predominantly:

Intravascular - within circulation
Extravascular - removal/destruction by reticuloendothelial (RE) system
Inherited or acquired

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What are the causes of extravascular haemolysis?

Autoimmune
Alloimmune
Hereditary spherocytosis

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What are 4 intravascular causes of haemolysis?

Malaria (most common worldwide)
G6PD deficiency
Mismatched blood transfusion (ABO)
Cold antibody haemolytic syndromes
Drugs
Microangiopathic haemolytic anaemia e.g. haemolytic uraemic syndrome, thrombotic thrombocytopenic purpura
Paroxysmal nocturnal haemoglobinuria

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What are the main causes of herediatry haemolytic anaemias? Divide these into categories.

Membrane

Cytoskeletal proteins
Cation permeability

Red cell metabolism

Haemoglobin

Thalassaemia
Sickle cell syndromes
Unstable Hb variants

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What are the consequences of haemolysis?

Anaemia(+/-)
Erythroid hyperplasia with increased rate of RBC production and circulating reticulocytes
Folate demand increased
Parvovirus B19 susceptibility
Gallstones(cholelithiasis)

Increased risk of:

Iron overload - due to increased intestinal iron absorption
Osteoporosis

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What is shown in this BM slide?

Parvovirus B19 infection - presents as an aplastic crisis.

Erythroid progenitors in the bone marrow shown which are early forms with no differentiation.

Immunity is lifelong after infection .

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This shows a liver slide of pyruvate kinase deficiency with Pearls stain. There is an increased amount of iron in the parenchyma of the liver and Kupffer cells.

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What is the genotype of Gilbert’s?

UGT 1A1 TA7/TA7 genotype

The UGT1A1 gene belongs to a family of genes that provide instructions for making enzymes called UDP-glucuronosyltransferases

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In haemolytic anaemia, what is the patient more at risk of if they also have Gilbert’s?

Coinheritance of Gilbert syndrome further increases risk of cholelithiasis in chronic haemolytic anaemia

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What are the clinical features of haemolytic anaemia?

Pallor
Jaundice
Splenomegaly
Pigmenturia
Family history

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What are the laboratory findings in a haemolytic anaemia?

Anaemia
Increased reticulocytes - esp in PB19 infection
Polychromasia
Hyperbilirubinaemia
Increased LDH
Reduced/absent haptoglobins
Haemoglobinuria
Haemosiderinuria

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RBC membrane - what are labels A, B, C and D?

A (band 3) and B (rhesus) = transmembrane proteins

C (junctional) and D (spectrin) = skeletal proteins

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What are defects in RBC membrane categorised into?

Hereditary spherocytosis
Hereditary elliptocytosis

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What is the pathophsyiology of most herditary spherocytosis vs elliptocytosis? Name 3 locations of defects that may cause each.

Spherocytosis = Vertical interaction

Band 3
Protein 4.2
Ankyrin
b Spectrin

Elliptocytosis = Horizontal interaction

b Spectrin
Protein 4.1
a Spectrin

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What is the pattern of inheritance in hereditary spherocytosis?

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FH in 75% - typically autosomal dominant

25% recessive or de novo mutation

How do RBCs react to hypotonic saline in vitro in hereditary spherocytosis?

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Osmotic fragilty test - In vitro red cells show increased sensitivity to lysis in hypotonic saline

Reduced binding of dye eosin-5-maleimide

What is shown in this slide of hereditary spherocytosis?

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RBCs ack central pallor which usually arises from the biconcave shape of RBCs
Smaller than normal
MCHC is increased (seen on blood count) so you see hyperchromic cells with higher Hb concentration
Polychromatic cells (indicative of young RBCs)

What does this dye binding test in hereditary spherocytosis show?

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Dye binding test uses flow cytometry to look at the mean cell fluorescence after incubation with the dye

You can see that the second plot shows that of a patient with herediatry spherocytosis (cut off used is 0.8, 97% sensitive test)

What condition is shown?

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Hereditary elliptocytosis

What condition is shown? What is seen? What is the inheritance of this condition?

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Hereditary pyropoikilocytosis - fragmentation of RBCs, budding or vesciculation of the membrane, large variation in shape of the RBC (poikilocytosis)

NB: this is the homozygous state for hereditary elliptocytosis i.e. more severe which may sometimes require transfusion.

What is the most common haemolytic anaemia caused by problems affecting RBC metabolism?

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Glucose-6-phosphate dehydrogenase deficiency

Where is G6PD most common? What is the inheritance pattern?

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Prevalent in areas of malarial endemicity - selection as there is some protection against plasmodium falciparum malaria

X-linked - clinical effects seen predominantly in hemizygous males and homozygous females

What is the role of G6PD?

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Enzyme catalyses first step in pentose phosphate(hexose monophosphate) pathway - generates NADPH required to maintain intracellular glutathione(GSH) which is needed to protect the cell against oxidative stress

So G6PD deficiency causes haemolytic anaemia in response to oxidative stress.

What are the clinical manifestations of G6PD deficiency?

1. Neonatal jaundice - probably most common cause of kernicterus worldwide 2. Acute haemolysis(triggered by oxidants/infection) 3. Chronic haemolytic anaemia(rare)

What can trigger acute haemolysis in G6PD?

**Drugs:** * Anti-malarials - Primaquine * Antibiotics - Sulphonamides, ciprofloxacin, nitrofurantoin * Other drugs - dapsone, Vitamin k **Infections** **Fava beans** **Moth balls**

What is the condition shown? What is seen?

G6PD deficiency * Contracted RBCs * Nucleated RBCs * Bite cells - some cytoplasm removed * Hemi ghosts - haemoglobin retracted to one side of the cell

The G6PD slide has been stained with Methylviolet. What is now seen?

Peripheral inclusions within the RBC called **'Heinz bodies'** formed from deformed haemoglobin. Usually only transiently seen as the spleen will remove these cells.

Apart from G6PD, name some other metabolic pathways within the RBC that may cause haemlytic anaemia if defective.

*Defects in nucleotide and glutathione metabolism etc:* **Embden-Meyerhof** = ATP, NADH **Hexose monophosphate shunt** = NADPH (pentose phosphate) **Rapoport-Luebering shuttle** = 2,3-DPG **Nucleotide metabolism** * Adenine salvage * Adenosine phosphate equilibrium * Removal of pyrimidine nucleotides **Glutathione biosynthesis** = GSH **Cytochrome b5 reductase** = Methemoglobin reduction

What is the most common defect in tye glycolytic pathway of RBC metabolism causing haemolytic anaemia? (condition shown below - what features are seen?)

**Pyruvate kinase deficiency** (post splenectomy shown) * **Echinocytes**- RBCs have short projections. The number of these increases post-splenectomy * **Spherocytes/spheroechinocytes** - as there is shrinkage due to dehydration there *Pyruvate kinase enzyme is near the end of the pathway*

What condition is shown below?

**_Pyrimidine 5'-nucleotidase deficiency -_** defect in a nucleotide metabolism pathway. *Pyrimidine is toxic to the RBC if it accumulates. But purines need to be retained so there should be a mechanims for retaining these which invovles the above enzyme.* ## Footnote **Relatively uncommon** **Basophilic stippling** - basophilic inclusions in RBCs; also seen in lead poisoning as lead is a potent inhibitor of this enzyme.

What investigations are important in patients presenting with haemolysis?

* **Direct antiglobulin test (DAT) -** detects Ig on surface of RBC in AI disease * **Urinary haemosiderin/haemoglobin** - tells us whether intravascular haemolysis is present; haptoglobin levels low in intravascular haemolysis * **Osmotic fragility** - if considering membrane defect * **G6PD +/- PK activity**- if considering enzyme deficiency; but G6PD is acute and PK chronic usually. * **Haemoglobin separation A and F%** - electrophoresis or HPLC * **Heinz body stain** - for oxidative haemolysis * **Ham’s test/Flow cytometry of GPI-linked proteins** - for paroxysmal nocturnal haemoglobinuria * **Thick and thin blood film** - Malaria

What are the principles of management of haemolytic anaemia?

**Conservative:** * Avoidance of precipitating factors e.g. oxidants in G6PD deficiency * Immunisation against blood borne viruses e.g. hepatitis A and B * Monitor for chronic complications **Medical:** * Folic acid supplementation * Red cell transfusion/exchange **Surgical**: * Cholecystectomy for symptomatic gallstones * Splenectomy if indicated

What are the conditions in which splenectomy may be indicated?

Beneficial in: * PK deficiency and some other enzymopathies * Hereditary spherocytosis * Severe elliptocytosis/pyropoikilocytosis * Thalassaemia syndromes * Immune haemolytic anaemia

What are the complications/risks of splenectomy?

* Capsulated bacteria eg Pneumococcus * Penicillin prophylaxis and immunisation

What clinical features are indications for splenectomy in haemolytic anaemia?

Criteria * Transfusion dependence * Growth delay * Physical limitation Hb * Hypersplenism

When can splenectomy be carried out i.e. age? What age is it most beneficial?

**After 3 years only -** as spleen is important in immune function in infants and toddlers **But before 10 years** - most beneficial as helps to maximise prepubertal growth

_Do not learn_ Case: Haemoglobin Hammersmith discover 1967 * Female infant aged18 months from UAE; Presented at 6 months of age - anaemia jaundice and pigmenturia; Sudden onset after respiratory illness; Chronic haemolysis ensued; Parents (first cousins) and 2 siblings well * Examination - pale and icteric with palpable spleen tip * Hb 7.0 reticulocytes 880 (37.4%) i.e. raised; Film shown with supravital stain - methyl violet * Heat stability and isopropanol precipitation tests strongly positive - 17% Hb precipitation (less than 1% in parental and control samples) * Cellulose acetate electrophoresis testing did not show anything which is not unusual but HPLC showed other peaks so new Hb variant and this was done for the rest of the family. * Mass spectrometry was then used to separate molecules based on their mass - named Hb Hammersmith

* G6PD deficiency suspected based on blood film * Unstable Hb variant seen by heat stability testing * Haemoglobin Hammersmith - severe electrophoretically silent, Heinz bodies present, mutation disrupts haem contact, reduced oxygen affinity

HAEM: Haemolytic anaemias Flashcards

(38 cards)