Week 9

• Anaemia

  Reduction in haemoglobin concentration with a concomitant decrease in oxygen carrying capacity of the blood
• Anaemia is not a diagnosis, it is an abnormal clinical finding
• The cause of anaemia must be found and treated appropriately
• Red cell

  • 8 μm in diameter
  • Oval, flexible, enucleate biconcave disc
  • Made in the bone marrow
  • 10^12 per day (erythropoiesis)
  • 120 day lifespan
  • Destroyed by macrophages in the spleen
  • Function: Gas exchange - To carry O2 from the lungs and return CO2 from the tissues to the lungs
• Haemoglobin (Hb)

  • Tetrameric structure: 2 different pairs of polypeptide chains - globin. Each attached to a haem molecule
  • Haem = Protoporphyrin + Iron
  • Globin permits variation in oxygen affinity, protects haem from oxidation, renders the molecule soluble
  • Haem Fe containing, transports oxygen
• Symptoms of anaemia

  • Tiredness & lethargy
  • Shortness of breath
  • Palpitations
  • Headache
  • In older patients: Cardiac failure, Angina, Intermittent claudication, Confusion
• Signs of anaemia

  • General: Pallor of mucous membranes (if Hb < 9-10 g/dl), Hyperdynamic circulation, Tachycardia, Bounding pulse, Cardiomegaly, Systolic flow murmur, Signs of heart failure
  • Specific: Koilonychia (spoon nails) - iron deficiency, Jaundice - haemolytic/megaloblastic anaemia, Leg ulcers - sickle cell disease, Bone deformities - thalassaemia major, Associated infections/bruising - bone marrow failure
• Investigations

  • Essential in all cases: Full blood count, Red cell indices, White cell and platelet counts, Blood film
  • Depending on FBC/Film: Reticulocyte count, Haematinics, Ferritin, Iron studies, Vitamin B12, Folate, Hb Electrophoresis, Bilirubin/LDH/haptoglobin, Bone marrow
• Classification of anaemia

  • Microcytic: MCV<80fl (Iron deficiency, Thalassaemia)
  • Normocytic: MCV 80-95fl (After acute blood loss, Anaemia of chronic disease, Bone marrow failure)
  • Macrocytic: MCV>95fl (Megaloblastic: vitamin B12, folate deficiency, Non-megaloblastic: Alcohol, Liver disease, Myelodysplasia)
• Hb Electrophoresis

  1. Electrophoresis on cellulose acetate at alkaline pH

  2. Proteins applied to a membrane are exposed to a charge gradient and separate from each other, then visualised by either a protein or haem stain
• High Performance Liquid Chromatography (HPLC)

  1. Adsorption: A mixture of molecules (such as normal and variant Hbs) with a net positive charge separated into its components by adsorption on to a negatively charged stationary phase in chromatography column

  2. Elution: Mobile phase (liquid) with an increasing concentration of cations compete with the adsorbed proteins for their anionic binding sites. Therefore the adsorbed proteins are eluted at a rate related to their affinity for the stationary phase.

  3. Detection: Molecules detected optically in the eluate, provisionally identified by elution time and quantified by computing the area under the corresponding peak in the elution profile
• Microcytic Anaemias

  • Iron deficiency
  • Anaemia of Chronic Disease
  • Thalassaemia (later!)
• Microcytic Anaemia - Iron deficiency

  • Most common cause of anaemia affecting 500 million people worldwide
  • 5-10% pre-menopausal women
  • 2-5% adult men / post-menopausal women
• Microcytic Anaemia - Iron deficiency

  • Limited ability to absorb iron
  • Excess loss due to bleeding is common
• Distribution of Body Iron

  • Haemoglobin (65%)
  • Ferritin and haemosiderin (30%)
  • Myoglobin (3.5%)
  • Haem enzymes (eg cytochromes etc) (0.4%)
  • Transferrin - bound iron (0.1%)
• Daily iron requirement goes up in pregnancy, adolescence and in menstruating females
• Microcytic Anaemia - Iron deficiency

  • Reticuloendothelial stores (haemosiderin and ferritin) completely depleted first before anaemia develops
  • General signs and symptoms of anaemia
  • Painless glossitis
  • Angular stomatitis
  • Koilonychia
  • Brittle, ridged or spoon-shaped nails
  • In children: Irritability, Poor cognitive function, Decline in psychomotor development
• Microcytic Anaemia - Iron deficiency: causes

  • Chronic blood loss (Uterine, Gastrointestinal, Peptic ulcer disease, Aspirin/NSAIDs, Malignancy, Inflammatory bowel disease, Hookworm, Haematuria)
  • Increased demands (Prematurity, Growth, Pregnancy)
  • Malabsorption (Coeliac disease, Gastrectomy)
  • Poor diet
• Rarely the sole cause in the developed world except in infants and children
• Microcytic Anaemia - Iron deficiency: investigations

  1. History (Diet, Bleeding symptoms, Medication, Family history)

  2. Examination (Abdominal mass, Rectal examination)

  3. FBC and blood film

  4. Serum iron studies (Ferritin, Iron, Total iron-binding capacity (TIBC), Transferrin saturation (TS))

  5. Consider: Upper and lower GI endoscopies, Transvaginal ultrasound, Urine dipstick
• Blood film in iron deficiency

  • Pencil cells
  • Hypochromic, microcytic anaemia with aniso/poikilocytosis
• Bone marrow iron assessed by Perls' stain
• Microcytic Anaemia - Differential diagnosis

  • Iron deficiency
  • Anaemia of Chronic Disease
  • Thalassaemia Trait (a or b)
• Differences in investigations between the differential diagnoses

  MCV, Ferritin, Serum iron, TIBC, TS, Bone marrow iron stores, Hb electrophoresis
• Microcytic Anaemia - Iron deficiency: treatment

  1. Treat underlying cause

  2. Oral iron (Ferrous sulphate 200mg TDS on empty stomach, If side-effects then give with food or use lower iron content preparation)

  3. Hb should rise by 2g/dl every 3 weeks, Continue for 3 months once FBC normal to replenish iron stores

  4. Parenteral iron (IV venofer/monofer), Side-effect: hypersensitivity/anaphylaxis, Haematological response no faster than with oral iron but stores replenished faster
• Microcytic anaemia - Anaemia of Chronic Disease (ACD)

  • Due to chronic inflammatory and malignant diseases
  • Decreased release of iron from macrophages to plasma
  • Reduced red cell lifespan
  • Inadequate erythropoietin response to anaemia
• Microcytic anaemia - Anaemia of Chronic Disease (ACD)

  • Normocytic or mildly microcytic anaemia (MCV rarely < 75fl)
  • Mild, non-progressive anaemia
  • Serum ferritin normal or raised
  • Serum iron, TIBC and TS all low
  • Bone marrow storage iron normal but erythroblast iron reduced
• Diagnosis of ACD: Careful history and examination, Elevated inflammatory markers (ESR/CRP), Treat the underlying cause
• Megaloblastic anaemia

  Erythroblasts in the bone marrow show a characteristic abnormality: Maturation of the nucleus is delayed relative to the cytoplasm
• Megaloblastic anaemia is associated with delayed maturation of the nucleus relative to the cytoplasm in erythroblasts in the bone marrow
• Haemolytic anaemias

  Anaemia resulting from an increase in the rate of red cell destruction
• Haemolytic anaemias

  • Bone marrow compensates by increasing production of red cells to 6-8 x normal rate
  • Marked reticulocytosis
  • Anaemia may not become apparent until red cell lifespan < 30 days
• Classification of haemolytic anaemias

  • Hereditary
  • Acquired
  • Immune
  • Non-immune
  • Intravascular
  • Extravascular
• Hereditary haemolytic anaemias

  • Red cell membrane defects (Hereditary spherocytosis, Hereditary elliptocytosis)
  • Red cell enzyme deficiencies (G6PD deficiency, Pyruvate kinase deficiency)
  • Abnormal haemoglobins (HbS, HbC, unstable Hb)
• Acquired non-immune haemolytic anaemias

  • Red cell fragmentation syndromes (Heart valves, Microangiopathic - TTP, HUS, DIC, Pre-eclampsia, meningococcal sepsis)
  • March haemoglobinuria
  • Infections (Malaria, clostridia)
  • Chemical and physical agents
  • Secondary (Liver and renal disease)
  • Paroxysmal nocturnal haemoglobinuria (PNH)
• Acquired immune haemolytic anaemias

  • Alloimmune (Haemolytic transfusion reactions, ABO incompatible blood, Haemolytic disease of the newborn)
  • Autoimmune (Warm antibody type, Cold antibody type)
• Causes of intravascular haemolysis

  • Mismatched blood transfusion
  • G6PD deficiency with oxidant stress
  • Red cell fragmentation syndromes
  • Some autoimmune, drug-induced, infection-induced haemolytic anaemias
  • PNH
  • March haemoglobinuria
  • Unstable haemoglobin
• Investigations for haemolytic anaemias

  • Proof of haemolysis (Decreased Hb, Increased unconjugated bilirubin and LDH, Decreased haptoglobins, Urinary haemoglobin and haemosiderin)
  • Features of increased red cell production (Increased reticulocytes)
  • Blood film
  • Specific tests (Direct antiglobulin test, HPLC and Hb electrophoresis, Red cell enzyme assays, Flow cytometry for CD55 and CD59)
• Mutations in the globin genes are the most prevalent monogenic disorders worldwide
• Hb disorders represent a significant health problem in 71% of 229 countries