Under graduate Lecture for Megaloblastic anaemia Definition, Pathophisiology ,clinical feature and lab Diagnosis

1. Megaloblastic Anemias

2. Concept
 An anemia in which there is a predominant number of
megaloblasstic erthyroblasts, and relatively few
normoblasts, among the hyperplastic erythroid cells in
bone marrow

3. Megaloblastic Anemia
 Megaloblastic anemia is an anemia (of macrocytic classification) that results
from inhibition of DNA synthesis during red blood cell production.
 When DNA synthesis is impaired, the cell cycle cannot progress from the G2
growth stage to the mitosis (M) stage. This leads to continuing cell growth
without division, which presents as macrocytosis.
 Megaloblastic anemia has a rather slow onset, especially when compared
to that of other anemias
 anemia or pancytopenia caused by impaired DNA synthesis commonly due to
deficiency of vitamin B12 or folic acid

6. Etiology
 95 % of cases due to:
 VIT B12 deficiency
 Folic Acid Deficiency
 5% of cases due to:
 Ant metabolic Drugs
 Inborn error of metabolism

7. Cobalamin (vitamin B12)
 It is solely synthesized by microorganisms and the only
source for humans is food of animal origin like meat, fish
and dairy products.
 Adult daily losses (mainly in urine and feces) are 1-3 ug
and so daily requirements are also 1-3 ug.
 Body stores are of the order of 2-3 mg, sufficient for 3-4
years if supplies are completely cut off.

9. Transportation path of Vit B12Diet Stomach
B12 IF
B12+
IF
Ileum
B12+IF
TCII
B12 +TCII
Circulation
Bone marrow
)DNA synthesis(
Liver )storage(
Other tissues
)DNA synthesis(

10. Important for DNA synthesis, nervous tissue
and fat metabolism in the liver
an intermediate of the citric acid
cycle, porphyrin synthesis
(Heme synthesis)

11. Vit B12 plays important role in two reactions:
• Necessary in the synthesis of methionine from homocysteine.
1. In this reaction both vit B12 and folic acid are involved.
2. B12 acts as a co-enzyme (methyl cobalamin) for methyltransferase.
Homocysteine methionineMethyl B 12
Methyl THF
THF
• Vit B12 is important in conversion of methylmalonyl CoA to succinyl CoA in Krebs
cycle.
1. In this reaction B12 acts as co-enzyme for methylmalonyl Co A mutase.
Methylmalonyl Co A Succinyl Co A
Adenosyl Cb
Ado B12

14. Pernicious Anemia
 Most common cause of cobalamin deficiency
 caused by failure of the gastric mucosa to
secrete intrinsic factor
 Immune Abnormalities
1. Anti–parietal cell antibodies
2. Anti–intrinsic factor antibodies -Two types
 Blocking” antibodies, which block the binding
of cobalamin to IF
 Binding” antibodies, which bind to the
cobalamin–IF complex and prevent the
complex from binding to receptors in the
ileum

15. The Schilling test
 Radioactive cobalamin (Cbl*) is taken orally; followed by injection of a saturating
dose of non-radioactive cobalamin.
 The level of Cbl* is measured in the urine. In pernicious anemia the excreted levels of
Cbl* are low.
 If intrinsic factor is given with the Cbl* the Cbl* levels will correct in PA, but not in
ileal malabsorption.

16. SCHILLING TEST

17. Folate
 Folic acid is parent compound of large family of natural folate
compounds.
 Its highest concentration is found in liver, yeast, spinach, other
greens and nuts.
 Folate easily destroyed by heating, particularly in large volumes of
water.
 Total folate in adult is ~10mg with liver containing the largest store.
 Daily adult requirements are ~100 μg and so stores are sufficient for
3-4 months in normal adult and severe folate deficiency may
develop rapidly.

19. Absorption
 Absorption: during absorption in the upper small
intestine, folates are converted to MTHF. Once inside the
cells they are converted to folate polyglutamates.
 Location: duodenum and jejunum.
 Transport: folate found in circulation as mono-glutamate
MTHF, transported to the liver and B.M by bound loosely
to proteins including albumin and α2-macroglobulin.

22. Biochemical basis of megaloblastic
anemia
 The folate is needed as coenzyme 5,10-methylene THF polyglutame
for conversion of dUMP to dTMP.
 In deficiencies of folate or cobalamin, there is failure to convert of
dUMP to dTMP; precursor of dTTP.
 The availablity of 5,10-MTHF is reduced in either cobalamin or folate
deficiency.
 in cobalamin deficiency due to failure of formation of THF, the
substrate on which folate polyglutame are build, MTHF
accumulates in plasma and intracsellular concentration fall; termed
as THF starvation or methylfolate trap.

23. Pathogensis
 Megaloblastic anemia are group of disorders characterized by distinct morphological pattern in
hematpoitic stem cells.
 A common feature is a defect in DNA synthesis with lesser alterations in RNA and protien synthesis,
leading to state of unbalanced cell growth and impaired cell division.
 Most RBCs are not resting but vainly engaged in attempting to double their DNA,with frequent arrest
in S phase and lesser degree of arrest in other phases of cycle.
 An increased percentage of RBCs have increased DNA content is expressed morphologically as
larger than normal “immature nucli” with finely particulate chromatin whereas the relatively
unimpaired RNA and protein synthesis results in larger cells with greater “mature” cytoplasm and cell
volume.
 The microscopic appearance of this nuclear-cytoplasmic asynchrony is morphologically discribed as
“megaloblastic”.

24. Clinical features
 Many patients are diagnosed through finding of a raised MCV on rotuine CBC.
 The main clinical features in more severe cases are those of anemia.
 Anorexia is usually marked and there may be weight loss, diarrhea or
constipation.
 Glossitis, angular cheilosis and mild fever in more severely anemic patients,
Juandice(unconjugated) and reversible skin melanin hyperpigmentation also
may occur with deficiency of cobalamin or folates.
 Thrombocytopenia sometimes leads to bruising and this may be aggrevated by
vitamin C deficiency or alcohol in malnourished patients.
 Anemia and low leukocyte count may predispose to infections, particularly of
respiratory and urinary tract.
 Cobalamin deficiency has also been associated with impaired bactericidal
function of phagocytes.

27. Diagnosis of megaloblastic anemia
 Hematological findings
1. Peripheral blood
2. Bone marrow
3. Ineffective hematopoiesis
4. Chromosome abnomalities
 Diagnosis of cobalamin and folate deficiencies
1. Serum cobalamin
2. Serum folate
3. Red cell folate
4. Serum methylmalonate and homocysteine

28. Laboratory findings
 Hb: low
 RBCs count: low
 PCV: may not reflect the actual decrease in erythrocytes
because of the enlarged size of the RBCs.
 MCV: high
 MCH: high
 MCHC: normal
 Leucocyte count: low
 PLT count: usually low

29.  Blood film:
RBCs:
• Moderate to marked anisocytosis and poikilocytosis with
many macrocytic ovalocytes.
• Tear drop and fragmented cells may be seen.
• Megaloblasts may seen in the P.B.
• Red cell inclusions: Howell-jolly bodies, Cabot ring and
basophilic stippling may present.
WBCs:
• Hyper-segmented neutrophils is classical finding
PLT:
• Giant form is usually present.

30.  Reticulocyte count: low
 B.M:
• Hypercellular with increased M:E ratio.
• Increased ratio of megaloblasts.
• Giant metamyelocytes are characteristics.
• Megakaryocytes: reduced in number.
 Biochemical tests:
• Serum B12: low
• Serum folic acid: low
• Serum bilirubin: increased
• Serum LDH: increased.

32. PB MorphologyPB Morphology
Macro-ovalocytes
Howell-Jolly bodies
Hypersegmented
neutrophils Bi-nucleated RBCs

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