Megaloblastic anemia is caused by a deficiency of vitamin B12 or folate, which inhibits DNA synthesis during red blood cell formation. This leads to large, abnormally-developed red blood cells. Symptoms include anemia, neurological issues, and gastrointestinal problems. Diagnosis involves examination of blood smears to find megaloblasts, along with low vitamin B12 or folate levels. Treatment consists of vitamin B12 injections and oral folic acid supplementation. Pernicious anemia, a specific type, results from gastric atrophy and lack of intrinsic factor, requiring lifelong B12 injections.

1. MEGALOBLASTIC ANEMIA
BY SANJAY ARALIKATTI

2. MEGALOBLASTIC ANEMIA
Definition
Megaloblastic anaemia is a red blood cell disorder due to the inhibition
of DNA synthesis during erythropioesis.
Mitotically, the inhibition of the DNA synthesis impaires the
progression of the cell cycle development from G2 to (M)
stage.

3. MEGALOBLASTIC ANEMIA
Macrocytic blood picture caused by
1. Nutritional deficiency of the vitamin B12 or folate or both
2. Deficiency of intrinsic factor of castle causing a condition called
pernicious anemia.

5. Vitamin b12 is available from foods of animal
origin

13. • VITAMIN B12 DEFICIENCY
• Inadequate intake: This is common among pure vegetarians,
old and bed ridden patients
• Inability to absorb vitamin B12: after gastric surgery, lack of
hydrochloric acid in gastric juice, lack of intrinsic factor due to
auto antibodies to parietal cells.
• Competition for intestinal vit.B12 : Competitive absorption
of the vitamin by fish tape worm (Diphyllobothrium latum)
and bacteria overgrowth in blind-loop syndrome, intestinal
stasis.
• Drugs Inhibition: Metformin, Proton pump inhibitors

14. FOLATE METABOLISM
Vitamin b9 water soluble. Daily requirement 100-200 mcg
Sources
Folate exists in different plants, bacteria and
animal tissues. Its main dietary sources are fresh green leafy
vegetables, fruits, liver, kidney, and to a lesser extent, muscle
meats, cereals and milk. Folate is labile and is largely destroyed
by cooking and canning. Some amount of folate synthesised by
bacteria in the human large bowel is not available to the body
because its absorption takes place in the small intestine. Thus,
humans are mainly dependent upon diet for its supply.

15. FOLATE METABOLISM
• Folate is normally absorbed
• from the duodenum and upper jejunum and to a lesser
• extent, from the lower jejunum and ileum

16. FOLATE STORAGE
The liver and red cells are the main storage sites of folate, largely as
methyl THF polyglutamate form. The total body stores of folate are about
10-12 mg enough for about 4 months. Normally, folate is lost from the
sweat, saliva, urine and faeces.

17. Pathophysiology
When vitamin B12 or folate is deficient, thymidine synthase function is
impaired and DNA synthesis is interrupted but RNA synthesis remains
unimpaired. The inability to synthesize DNA leads to ineffectual
erythropoiesis resulting in excess hemoglobin and enlarged erythroid
precursors being produced. The developing red cell has difficulty in
undergoing cell division but RNA continues to be translated and
transcribed into protein leading to growth of the cytoplasm while the
nucleus lags behind. Often one or more cell division are skipped
leading to a larger than normal cell.
There is often erythroid hyperplasia in the marrow but most of these
immature cells die before reaching maturity leading to - elevated
Lactate Dehydrogenase (LDH) and hyperbilirunemia.

18. CLINICAL PRESENTATION
Anaemia symptoms
Neurological symptoms
Gastro- intestinal complain
Symptoms of Anemia
weakness, palpitation, fatigue, lightheadedness,,
shortness of breath, premature graying
of hair, jaundice and pallor.
Severe pallor and slight jaundice combine to produce
a telltale lemon-yellow skin in patient with
megaloblastic anemia.

19. Neurological symptoms
The syndrome usually begins with paraesthesia
(numbness and tingling) in the feet and fingers,
difficulties in balance and walking.
Vitamin B12 deficiency causes a demyelinization
of the peripheral nerves, the spinal cord, and the
brain, resulting in more severe neurological
symptoms.
When it affects the spinal cord it causes spastic
ataxia( stiffness of the muscles with uncoordinated
movement). At the brain it results in dementia,
psychotic depression and paranoid schizophrenia.
This has been termed “megaloblastic madness.”

20. • Gastro- intestinal complains : symptom include loss of
ppetite, glossitis (red, sore, smooth tongue) and diarrhoea

21. General Laboratory Findings
• Haemoglobin- decreased
• RBC MORPHOLOGY- megaloblasts and marked anisocytosis,
poikilocytosis and presence of macroovalocytes. Basophilic stippling
and occasional normoblast may also be seen.

22. Bone marrow smear: Bone marrow examination
reveals myeloid cell changes (giant bands,
metamyelocytes and hypertsegmentation) and
megakariocytes are decreased and show abnormal
morphology.

24. TESTS FOR VIT B12 DEFICIENCY-
SERUM VITAMIN B12 ASSAY Assay of vitamin B12 in blood can be done by 2
methods—microbiological assay and radioassay.
Microbiological assay In this test, the serum sample to be assayed is added
to a medium containing all other essential growth factors required for a
vitamin B12-dependent microorganism. The medium along with
microorganism is incubated and the amount of vitamin B12 is determined
turbimetrically which is then compared with the growth produced by a
known amount of vitamin B12. Several organisms have been used for this
test such as Euglena gracilis, Lactobacillus leichmannii, Escherichia coli and
Ochromonas malhamensis. E. gracilis is, however, considered more sensitive
and accurate. The addition of antibiotics to the test interferes with the
growth and yields false low result.

25. Radioassay Assays of serum B12 by radioisotope
dilution (RID) and radioimmunoassay (RIA) have been
developed.
These tests are more sensitive and have the
advantage over microbiologic assays in that they are simpler
and more rapid, and the results are unaffected by antibiotics
and other drugs which may affect the living organisms

26. SCHILLING TEST (24-HOUR URINARY EXCRETION TEST) Schilling test is done to detect
vitamin B12 deficiency as well as to distinguish and detect lack of IF and malabsorption
syndrome. The results of test also depend upon good renal function and proper
urinary collection. Radioisotope used for labeling B12 is either 58Co or 57Co. The test
is performed in 3 stages as under:
Stage I: Without IF The patient after an overnight fasting is administered oral dose of 1
mg of radioactively labelled vitamin B12 (‘hot’ B12) in 200 μl of water. At the same
time, 1 mg of unlabelled vitamin B12 (‘cold’ B12) is given by intramuscular route; this
‘cold’ B12 will saturate the serum as well as the tissue binding sites. The patient is kept
fasting for a further period of 2 hours, following which urinary excretion of B12 is
estimated: ”
. In normal individuals, 24-hour urinary excretion is >10% of the oral dose
of ‘hot’ B12. ”
. Patients with IF deficiency excrete lower quantity of ‘hot’ B12 which is
further confirmed by repeating the test as in stage II
Stage II: With IF If the 24-hour urinary excretion of ‘hot’
B12 is low, the test is repeated using the same procedure as
in stage I but in addition high oral dose of IF is administered
along with ‘hot’ B12.

27. If the 24-hour urinary excretion of ‘hot’ B12 is now normal, the low value
in first stage of the test was due to IF deficiency (i.e. pernicious
anaemia). Patients with pernicious anaemia have abnormal test even
after treatment with vitamin B12 due to IF deficiency. However,
abnormal 24-hour urinary excretion of ‘hot’ B12 is further investigated in
stage III for a cause in intestinal malabsorption of ‘hot’ B’12.
Stage III: Test for malabsorption of vitamin B12 Some
patients absorb vitamin B12 in water as was stipulated in
the original Schilling test. Modified Schilling test employs
the use of protein-bound vitamin B12. In conditions
causing malabsorption,
the test is repeated after a course of
treatment with antibiotics or anti-inflammatory drugs.

28. TESTS FOR FOLATE DEFICIENCY
The normal range of serum folate is 6-18 ng/ml. Values of 4 ng/ml or
less are generally considered to be diagnostic of folate deficiency
URINARY EXCRETION OF FIGLU Folic acid is
required for conversion of formiminoglutamic acid (FIGLU)
to glutamic acid in the catabolism of histidine. Thus, on oral
administration of histidine, urinary excretion of FIGLU is
increased if folate deficiency is present.

29. SERUM FOLATE ASSAY
Microbiological assay This test is based on the principle
that the serum folate acid activity is mainly due to the
presence of a folic acid co-enzyme, 5-methyl THF, and that
this compound is required for growth of the microorganism,
Lactobacillus casei. The growth of L. casei is inhibited by
addition of antibiotics.

30. Radioassay The principle and method of radioassay
byradioisotope dilution (RID) test are similar to that for serum
B12 assay. The test employs labelled pteroylglutamic
acid or methyl-THF. Commercial kits are available which permit
simultaneous assay of both vitamin B12 and folate.

31. PRINCIPLES OF TREATMENT
This includes: hydroxycobalamin as
intramuscular injection 1000 μg for 3 weeks and oral folic acid
5 mg tablets daily for 4 months

34. 1.The most characteristic
pathologic finding in PA is gastric atrophy affecting the acid and
pepsin-secreting portion of the stomach and sparing
the antrum.
CLINICAL MANIFESTATIONS ARE MAINLY DUE VITAMIN B12
DEFICIENCY

35. PRINCIPLES OF TREATMENT FOR PERNICIOUS
ANEMIA
1. Parenteral vitamin B12 replacement therapy.
2. Symptomatic and supportive therapy such as physiotherapy
for neurologic deficits and occasionally blood transfusion.
3. Follow-up for early detection of cancer of the stomach.