Megaloblastic anaemia
Pathology § Defect in DNA synthesis affecting rapidly dividing cells in bone marrow and other tissues § Disparity in the availability of the precursors of DNA which are required for DNA synthesis during S phase of cell cycle § Adenine, guanine, cytosine and thymidine § In B12 and folate deficiency this is due to an inability to methylate deoxyuridine monophosphate to deoxythymidine monophosphate § The methyl group is supplied by the folate co-enzyme, methylene tetrahydrofolate § Folate deficiency reduces the supply of this co-enzyme § B12 deficiency reduces its supply by slowing the demethylation of methyltetrahydrofolate and preventing cells receiving tetrahydrofolate for synthesis of methylene tetrahydrofolate § Slowed elongation of newly originated replicating segments § Small fragments accumulate and single stranded areas become points of weakness, where breakage may occur § Some cells apoptose at this stage
§ B12- folate relationship
§ In the cytoplasm, methylcobalamin serves as cofactor for methionine synthesis by allowing transfer of a methyl group from 5-methyl-tetrahydrofolate (5-methyl-THF) to homocysteine (HC), forming methionine and demethylated tetrahydrofolate (THF). § This results in reduction in serum homocysteine, which appears to be toxic to endothelial cells. Methionine is further metabolized to S-adenosylmethionine (SAM). (Methionine levels increase in B12 and folate deficiency) § THF participates in purine synthesis and the conversion of deoxyuridylate (dUTP) to deoxythymidine monophosphate (dTMP), which is then phosphorylated to deoxythymidine triphosphate (dTTP) , which is required for DNA synthesis § Therefore, in vitamin B-12 deficiency, formation of dTTP is inadequate and accumulation of 5-methyl-THF occurs, trapping folate in its unusable form and leading to retarded DNA synthesis. § RNA contains dUTP (deoxyuracil triphosphate) instead of dTTP, allowing for protein synthesis to proceed uninterrupted and resulting in macrocytosis and cytonuclear dissociation. § Cobalamin as well as a co-enzyme to methionin synthase is also a co-enzyme in the isomerization of methylmalonyl CoA to succinyl CoA
Causes of Megaloblastic anaemia § B12 and folate deficiency § Anti-folate drugs § Eg methotrexates § Some cases of AML and MDS § Drugs interfering with DNA synthesis § Cytarabine, HU, 6-MP, AZT § Orotic aciduria (AR condition UMPS deficiency – produces megaloblastic anaemia resistent to B12 and folate) § Lesch-Nyhan syndrome (XLR deficiency of HGPRT – hypoxanthine-guanidine phosphoribosyl transferase – results in build up of uric acid, severe gout, mental retardation and megaloblastic anaemia in some cases)
Clinical Features § General § Anorexia, weight loss, diarrhoea, constipation § Glossitis, angular cheilosis § Mild fever § Jaundice (unconjugated – due to death of red cells in marrow) § Reversible melanin skin hyperpigmentation § Epithelial surfaces § Affect all rapidly growing tissues § Glossitis, villous flattening with GI upset § Cervical smear abnormalities § Pregnancy § Infertility in men and women § Neural tube defects § Folic acid from conception to 12 weeks reduces incidence of neural tube defects by 70% § Cardiovascular defects § Homocysteinuria with high levels of homocysteine suffer with cardio and cerebrovascular disease (homocysteine toxic to endothelial cells) § Lesser degrees of high homocysteine have also been associated with vascular disease § Malignancy § Prophylactic folic acid during pregnancy has been found to reduce subsequent ALL in childhood § Neurology § B12 deficiency causes peripheral neuropathy with demyelination of the posterior and pyramidal tracts § Optic atrophy or cerebral symptoms
§ Peripheral blood § Oval macrocytes with anisocytosis and poikilocytosis and fragmentation § Hypersegmented neutrophils (>5) § Leucopenia and thrombocytopenia § Bone marrow § Hypercellular with accumulation of primitive cells § Nuclear; cytoplasmic asynchrony in erythroblasts § Nucleus has an open, fine lacy appearance § Nuclear fragments § Giant metamyelocytes and hyperpolypoid megakaryocytes § Can be confused with erythroleukaemia
Vitamin B12 § Only source is food of animal origin § Meat, fish, dairy § Body stores are sufficient for 3-4 years § Absorption § Passive § Duodenum and ileum § Inefficient; <1% of an oral dose is absorbed § Active § Ileum, mediated by intrinsic factor § Dietary B12 binds to R-protein (salivary glycoprotein) § B12 – R complex is digested by pancreatic trypsin § B12 transferred to intrinsic factor – resistant to enzyme digestion § Intrinsic factor is produced by gastric parietal cells in fundus and body of stomach § Intrinsic factor attaches to a receptor on brush border of ileal cells and B12 enters the cells, whilst intrinsic factor remains in the lumen § Ileum has restricted capacity, because of limited receptor sites § B12 then transported by transcobalamin II to all cells of the body § In plasma, B12 is mainly bound to transcobalamin I § Some B12 enters bile, binds to intrinsic factor and is reabsorbed § Transport § R-protein § Transcobalamin · Synthesised by liver, also macrophages, ileum and endothelium · Readily gives up B12 to marrow, placenta and others § Deficiency § Dietary deficiency · Vegan § Excess degradation · Nitric oxide § Gastric causes · Pernicious anaemia § Lack of IF due to gastric atrophy § Peak age of onset 60 years (younger in black, Latin Americans) § Association with AI disease, premature greying, blue eyes and vitiligo, Blood group A and hypogammaglobulinaemia § IF antibodies · Type 1 – prevents binding of IF to B12 · Type 2 – prevents IF attaching to ileal mucosa § GPC Antibodies · Sensitive but not specific · Congenital IF deficiency § Present at toddler age, when stores accumulated in utero have been used up · Gastrectomy § Inevitable after total § 10-15% after partial gastrectomy § Intestinal · Stagnant loop syndrome § Bacterial overgrowth of upper small intestine by faecal organisms § Consume cobalamin · Ileal resection · Selective malabsorption of cobalamin with proteinuria § AR § Most common cause of megaloblastic anaemia due to caolabmin deficiency in children in Western countries § Congenital defect of the ileum · Tropical sprue § Usually improves after antibiotic therapy · Fish tapeworm (Diphyllobothrium latum) · Gluten induced enteropathy § 30% of untreated · Severe chronic pancreatitis § Lack of trypsin results in cobalamin –protein R complex unavailable for absorption · HIV · Zollinger-Ellison syndrome § High acidity inactivates trypsin · Radiotherapy and GVHD · Drugs § Esp long term use of H2-antagonists § Abnormalities of cobalamin metabolism · Congenital transcobalamin deficiency § Present within a few weeks of birth § Responds to massive injections of B12 which allows passive transfer of B12 into marrow cells · Congenital methylmalonic acidaemia and aciduria § Ill from birth § Diagnosis § Serum B12 · Radioisotope dilution or ELISA · High levels usually due to elevated haptocorrin (cobalamin-binding protein) § MPD, hepatoma, increased granulocytes (IBD, liver abscess), eosinophilia § Methylmalonic acid level · Increased due to impairment of methylmalonyl CoA to succinyl CoA · Levels fluctuate in renal failure § Homocysteine levels · Increased, but poor specificity, therefore not used § Schilling test · Not done currently · Radiolabelled oral B12 and measure urinary excretion. If malabsorption, give the same but with IF. If abnormal = intestinal rather than gastric
Folate § Found in most foods, but easily destroyed by cooking (unlike B12) § Stores are only sufficient for about 4 months § Absorption § Upper small intestine – about 90% of an oral dose is absorbed § Enterohepatic circulation § Transport § 1/3 bound to albumin, 2/3 unbound § Rate of uptake in cell is dependant on rate of DNA synthesis § Deficiency § Dietary § Tropical sprue § Gluten-induced enteropathy § Specific malabsorption of folate § Excess utilisation or loss § Pregnancy § Prematurity § Haemolysis § Inflammatory disease/ chronic infections § Homocysteinuria § Haemo and peritoneal dialysis § CCF/ liver disease § Release of folate from damaged liver cells § Anti-folate drugs § Diagnosis § Serum folate § Radioassay or ELISA § Very sensitive, can be affected by weekly dietary changes § Red cell folate § Useful test of folate stores § Less affected by recent diet § False normals if recent transfusion or raised reticulocytes
Management of megaloblastic anaemia § B12 § Should treat if haematological abnormalities even in the absence of anaemia § Routinely to all who have undergone total gastrectomy or ileal resection § Uusually given IM § Because a small amount can be absorbed passively, large daily oral doses (1-2mg) of cyanocobalamin can be given § Most need to continue for life, unless treatable cause such as tropical sprue § Folate § Exclude B12 deficiency § Can treat for 4 months or long-term if likely to recur § Food is often fortified § Pregnency – 400mcg is an adequate dose § If previous NTD, then 5mg § Folinic acid – stable form of reduced folate § To overcome toxic effects of methotrexate |
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