MEGALOBLASTIC ANAEMIA Muhammad Asif Zeb Lecturer Hematology Khaybar Medical University Pershawar
MEGALOBLASTIC
ANAEMIA
Muhammad Asif Zeb
Lecturer Hematology
Khaybar Medical University
Pershawar
DEFINITION
The megaloblastic anaemias are a group of disorders characterized
by the presence of distinctive morphological appearances
of the developing red cells in the bone marrow and Pancytopenia in the
peripheral film.
The cause is usually deficiency of either cobalamin (vitamin B 12 ) or folate
but megaloblastic anaemia may arise because of inherited or
acquired abnormalities affecting the metabolism of these vitamins
or because of defects in DNA synthesis not related to
cobalamin or folate1
Megaloblastic anaemia is characterized by the appearance
in the bone marrow of morphologically abnormal
nucleated red cell precursors called megaloblasts and
also by hypersegmented or multi-segmented
neutrophils.
Megaloblasts are abnormal in functional as well as
appearance with the result that the mature red cells
formed from them are abnormal in size and shape; the
most prominent abnormality being macrocytosis.
Megaloblast:
Megaloblast describes a bone marrow cell that is part of the factory making red cells that has become abnormally large and has an abnormal appearance. It is a counterpart of erythroblast. These are usually large oval shaped cells and are fully haemoglobinized due to neuclear cytoplasmic asynchronisim .
CAUSES
Cobalamin deficiency or abnormalities of cobalamin metabolism.
Folate deficiency or abnormalities of folate metabolism.
Therapy with antifolate drugs. (e.g. methotrexate)
Independent of either cobalamin or folate deficiency and refractory to cobalamin and folate therapy.
Therapy with drugs interfering with synthesis of DNA. (e.g. cytosine arabinoside)
TRANSPORT: There are two major vitamin B12 binding proteins
in plasma.
TC 1: it is an α1 globulin which carries from 70-90% of circulating vitamin B12.
It is primarily a storage protein and its absence doesn’t lead to clinical signs of B12 deficiency.
Functionally dead
TC 2: It is a β-globulin and it is essential for transport of vitamin B12 from one organ to the other and in and out of cells. Congenital deficiency of TC2 leads to severe megaloblastic anaemia.
TC 3: similar to TC1, binds only a small quantity of circulating B12.
FUNCTIONS
Vitamin B12 acting as co-enzyme for two important
biochemical reactions in humans:
The conversion of methylmalonyl-CoA to succinyl-CoA.
Synthesis of methionine from homocystine.
Vitamin B12 is required for maintenance of the
integrity of nervous system.
MEGALOBLASTIC ANAEMIA DUE TO
VITAMIN B12 DEFICIENCY MECHANISM DISORDER
Decreased intake Nutritional deficiency
Impaired absorption
i). Gastric causes Pernicious anaemia
Gastrectomy (total or partial)
ii). Intestinal causes Lesions of small intestine
Coeliac disease
Tropical Sprue
Tapeworm Infestation
Zollinger -Ellison Syndrome
Abnormalities of cobalamin metabolism
i). Transport protein defects Inherited TC 2 deficiency
ii). Congenital intrinsic factor Present before the age of two
deficiency years
iii). Congenital methylmalonic acidaemia Infants are ill from birth
and aciduria
METABOLISM OF FOLATE
Folic acid was synthesized as a yellow crystalline powder. Folic acid doesn’t exist as such in nature, but is a parent compound of a large number of derivatives referred to as folates which plays an important role as co-enzymes in cellular metabolism. Reduction to dihydro - and tetrahydrofolate derivatives is necessary to participate in metabolic reactions.
Contents in food green Vegetables: rich
Meat: moderate
Effect of cooking 60-90% loss
Adult daily requirements 200 micro gram
Adult daily intake 100-500 micro gram
Site of absorption Duodenum & jejunum
Body stores 5-20 mg
SOURCES:
Widely distributed in plants and animals tissues. Richest sources are liver, kidney, yeast and fresh green vegetables such as spinach and cabbage.
ABSORPTION:
Normally absorbed from duodenum and upper jejunum and to a lesser extent from lower jejunum and ileum. Absorption of folate is a rapid active process 80% is absorbed unchanged. Synthetic polyglutamates are absorbed as well as monoglutamates. Polyglutamates are cleaved to monoglutamates by the enzyme pteroylpolyglutamate conjugase. Monoglutamates than undergo reduction and enters in circulation as methyltetrahydrofolate.
TRANSPORT:
Folate circulates in plasma as methyltetrahydrofolate monoglutamate, either in a free form or weekly bound to a variety of proteins.
FUNCTIONS
Folate co-enzymes are required for several bio-
chemical reactions in the body involving transfer of
one-carbon units from one compound to another.
Two reactions that are important in the context of
clinical folate deficiency are:
Methylation of homocysteine to methionine.
Synthesis of pyrimidine nucleotide, thymidylate
monophosphate from deoxyuridylate
monophosphate in the DNA synthesis pathway.
MEGALOBLASTIC ANAEMIA DUE TO
FOLATE DEFICIENCY
MECHANISM DISORDER
Decreased intake Nutritional deficiency
Impaired absorption Coeliac disease
Tropical sprue
Increased demand Pregnancy, haemolytic anaemia, myeloproliferative
disorders, leukaemia & lymphoma, carcinoma, Inflammatory disorders
Dihydrofolate reductase Methotrexate
Inhibitors Trimethoprim
Pathophysiology In megaloblastic anaemia, the anaemia results from failure of
the megaloblastic bone marrow to compensate for a moderate reduction in red cell life span. Red cell survival studies have shown the presence of mild haemolysis.
Lack of vitamin B12 or folate causing slowing of DNA synthesis in developing erythroblasts with an accumulation of cells in premitotic phase of cell cycle. Some of these cells die within the marrow.
The neutropenia and thrombocytopenia also appear to result from ineffective production by abnormal precursor cells in the marrow.
PATHOGENESIS Both vitamin B12 and folic acid are required for ordered DNA
synthesis. But due to deficiency of vitmain B12 & folic acid
in megaloblastic anaemia DNA synthesis is impaired or
blocked in rapidly dividing cells.
As a result the cells proliferates abnormally and increases in
size and become megaloblasts which are fully
heamoglobinized cells and abnormal in appearance and
function.
Because of their increased size they occupy much of the space
in marrow and they disturb other cell lines too.
MASKED MEGALOBLASTIC
ANAEMIA In certain megaloblastic anaemias , iron deficiency is sometimes
present at the same time as folate or B12 deficiency.
Associated iron deficiency may partly mask the typical
haematological features of megaloblastic anaemia.
PF showes double population (dimorphic blood picture) some red
cells being oval and well haemoglobinized, and others small
and poorly haemoglobinized.
In other cases masking takes the form of a lesser degree of
macrocytosis, so that most cells are of normal size and
MCV is normal or even mildly reduced. However, careful
scrutiny of blood film shows a small no. of oval
macrocytes and hypersegmented neutrophils.
Other disorders:
Thalassaemia, Infection, chronic renal
disease, rheumatoid arthritis.
CLINICAL MANIFESTATIONS
Features of anaemia: pallor, anorexia, weight loss, diarrhoea.
i). Macrocytic megaloblastic anaemia.
ii). Glossitis.
iii). Peripheral neuropathy and subacute combined degeneration (demyelination) of spinal cord.
NOTE: Deficiency of folate doesn’t produce sub-acute combined degeneration of spinal cord, but peripheral neuropathy is occasionally seen.
DIAGNOSIS
Diagnosis is usually made on:
Patient history
Clinical features
Lab diagnosis
LAB DIAGNOSIS HAEMATOLOGICAL FINDINGS
CBC:
Haemoglobin Decreased
RBC Decreased
WBC Leukopenia Neutropenia
Platelets Mild, usually symptom less thrombocytopenia
Hct Decreased
MCV Increased (125 fl is diagnostic of megaloblastic anaemia)
MCH Increased
MCHC Normal RDW Increased
Reticulocytes usually reduced
Peripheral blood picture:
Macrocytic Normochromic anemia
Anisocytosis: (Increased variation in RBC size)
Oval Macrocytes
Poikilocytosis: (Increased variation in RBC shape)
oval macrocytes, target cells, Tear drop cells
Schistocytes
Inclusions:
Howell-jolly bodies
Cabot rings
WBC: hypersegmented neutrophils
Platelets: Giant platelets
Bone Marrow Examination
Erythropoiesis
Hypercellular
increased erythroid /myeloid ratio
erythroid cell changes (megaloblasts,
RBC precursor a abnormally large with
nuclear- cytoplasmic asynchrony)
Thrombopoiesis
magakaryocytes may
be decreased, normal,
or increased.maturation,
however, is distintly
abnormal.some larger
than normal forms can
be found with separation
of nuclear lobes and
fregments
Grannulopoiesis
is abnormal large as
typical granulocytes giant
metamyelocytes(30µm)
and bands with loose,
open chromatin in the
nuclei are diagnostic.
Myelocytes show poor
granulation more mature
stages
BIOCHEMICAL FINDINGS
Serum vitamin B12 Decreased
Serum folic acid Decreased 2-15micro g/l
Red cell folate level Decreased 160-640 micro g/l
Serum bilirubin Increased 0.1-1.0 mg/dl
Homocysteine level Increased
Methylmalonic acid Increased
LDH Increased
FIGLU Increased
Serum Iron Increased
Serum ferritin Increased
Serum heptoglobin Decreased
Haemosiderinuria
Serum methylmalonic acid and homocysteine Increased
SPECIAL TESTS
Serum vitamin B12 assay:
It is performed by two ways:
Microbiological assay.
Radio-isotope assay.
Radioactive vitamin B12 absorption test:
The ability of body
to absorb vitamin B12 can be assessed by measuring the
absorption of a small dose of Co- labelled vitamin B12.
The test is called Schilling test.
SCHILLING TEST
An oral dose of 1 micro gram radioactive vitamin B12 is administered to the fasting subject followed two hours later by a large parenteral injection of unlabelled B12(1000 micro gm). The injection flushes out about one-third of absorbed radioactive B12 into the urine in the next 24 hours. Normal subject excretes 10% of 1 micro gm dose. Patients with pernicious anaemia excrete less than 5%. If the patient absorbs normal amounts of vitamin B12 no further testing is required.
UNSATURATED B12 BINDING CAPACITY
Measurement of unsaturated B12 binding capacity which in the normal subject reflects the amount of TC2 and to a lesser extent TC1 & TC3, may be diagnostically useful. Normal range is 500-1200 ng/l.
Deoxyuridine Suppression Test
In normal bone marrow, dU considerably suppresses
the uptake of radioactive thymidine into DNA.
This is thought to be due to conversion of dU to
thymidine triphosphate via dUMP, which inhibits
thymidine kinase, on which thymidine uptake
depends. Deoxyuridine suppresses radioactive
thymidine incorporation less effectively in meg.
Anaemia due to folate or cobalamin deficiency
because of the block in dUMP methylation to
dTMP.
Tests for the cause of
cobalamin deficiency
Clinical History :diet, drugs,operation etc.
Cobalamin absorption using radioactive cobalamin: Alone, with food, with intrinsic factor.
Tests for tissue specific antibodies in serum (e.g. IF, parietal cells etc)
Endoscopy with gastric biopsy.
Measurement of intrinsic factor in gastric juice after maximal stimulation. (rarely performed)
Small intestinal studies.
Stool for fish tapeworm ova.
SERUM FOLATE ASSAY
Microbiological Assay.
Radio-isotope Assay.
(Levels below 3micro gm/l suggest clinically significant
folate deficiency).
RED CELL FOLATE ASSAY
Red cells contain 20-50 times as much folate as serum. It is
usually a more reliable indicator of tissue folate stores than
the serum folate. It reflects mean folate that existed in
plasma during maturation of precursors.
Parietal cell antibodies:
Serum antibodies to surface membrane and
cytoplasmic antigens of gastric parietal cells are found in at least
85% of patients with pernicious anaemia.
Intrinsic factor antibodies:
Two types of antibodies are found:
Blocking antibodies.
Binding antibodies.
THANX!