Vitamin B12 and folate deficiencies can cause megaloblastic anemia due to impaired DNA synthesis. Vitamin B12 deficiency specifically can also cause neurological manifestations like peripheral neuropathy. Diagnosis involves blood tests showing macrocytic anemia and low vitamin levels. Treatment is with high dose vitamin B12 injections initially then maintenance doses to correct the deficiency. Folate deficiency has similar blood features but no neurological involvement, and is treated with oral folic acid supplementation.

1. Megaloblastic anaemia
Prof. S c kohli

2. Macrocytic anaemias
1. Megaloblastic
Vitamin B12 deficiency
Folate deficiency
2. Nonmegaloblastic
Myelodysplasia,
chemotherapy
Liver disease
Increased reticulocytosis
Myxedema

3. Megaloblastic Anemias- etiology
1. cobalamin or folic acid deficiency,
2. by drugs that interfere with the synthesis of
DNA or
with the absorption or metabolism of
cobalamin,
3.and by genetic disorders that interfere with
DNA metabolism or with the absorption or
distribution of cobalamin.

4. Role of vit B12 cobolamin and
folate
Homocysteine-- -CONVERSION to ----methionine
folate is substrate & Cobolamin is cofactor
in this reaction –>
- tetrahydrofolate is produced -
Tetrafolate --- CONVERTED INTO -
 Thymidine monophosphate
-- Thymidine is incorporated into DNA

5. causation
Deficiency of
either vitamin B12
or folate
will therefore produce high plasma levels of
homocysteine
and
impaired DNA synthesis.

6. Morphological changes
The end result is production of cells
with arrested nuclear maturation
but normal cytoplasmic development
So-called
nucleo-cytoplasmic asynchrony.
Megalocytosis results

7. nucleo-cytoplasmic asynchrony.
Megalocytosis

8. Cell morphology
All proliferating cells will exhibit
megaloblastosis;
hence changes are evident in
the buccal mucosa,
tongue,
small intestine,
cervix, vagina and uterus

9. Morphological changes in
bone marrow
Cells become arrested in
development
and die within the marrow;
intramedullary haemolysis
results in a
raised bilirubin and lactate dehydrogenase
Iron stores are usually raised.

10. Morphological changes
bone marrow
This ineffective erythropoiesis results in an
expanded hypercellular marrow.
The megaloblastic changes are most evident
in the early nucleated red cell precursors,
The mature red cells are large and oval,
and sometimes contain nuclear remnants.

11. Morphology
bone marrow
• Nuclear changes are seen in the immature
granulocyte precursors
and a characteristic appearance is that of
'giant' metamyelocytes with a large
'sausage-shaped' nucleus.

12. Cellular morphology
The mature neutrophils show
hypersegmentation of their nuclei,
with cells having six or more nuclear lobes.
If severe,
a pancytopenia may be present in the
peripheral blood.

13. Peripheral smear
• In folic acid or cobalamin deficiency,
• the smear is characterized by
• variation in erythrocyte size
• and by distinct macrocytosis.
• Occasionally, fish-tailed erythrocytes are
present, along with
• hypersegmented neutrophils

14. Peripheral smear
variation RBCsize
hypersegmented WBCs

15. Peripheral smear
nuclear hypersegmentation
polymorphonuclear neutrophil
macro-ovalocytes

16. Common Symptoms
• Malaise (90%) Breathlessness (50%)
• Paraesthesiae (80%) Sore mouth (20%)
• Weight loss Altered skin pigmentation
• Grey hair
• Impotence Poor memory
• Depression Personality change
• Hallucinations Visual disturbance

17. Common Signs
• Smooth tongue
• Angular cheilosis
• Vitiligo
• Skin pigmentation
• Heart failure
• Pyrexia

18. Special features in vit B 12 deficiency
• Vitamin B12 deficiency----but not --- folate
deficiency ----is associated with neurological
disease in up to 40% of cases.
• The main pathological finding is focal
demyelination affecting spinal cord,
peripheral nerves, optic nerves and cerebrum.
• The most common manifestations are sensory
with peripheral paraesthesia and ataxia of
gait.

19. B12 deficiency
• Neurologic Manifestations
Cobalamin deficiency may cause
a bilateral peripheral neuropathy
or
degeneration (demyelination) of the posterior
and pyramidal tracts of the spinal cord
and, less frequently, optic atrophy
or cerebral symptoms.

20. Symptoms neurological
• The patient, more frequently male, presents
with
• paresthesias,
• muscle weakness,
• or difficulty in walking
• and sometimes dementia,
• psychotic disturbances,
• or visual impairment.

21. NEUROLOGICAL FEATURES
• Patients with severe cobalamin deficiency
initially complain of paresthesia.
• sense of touch and temperature sensitivity
The may be minimally impaired
• The disease may progress, involving the dorsal
columns, causing ataxia and weakness.
• The physical examination reveals a broad-base
gait, Romberg sign, slowed reflexes, and loss
of sense of position and feeling of vibration

22. NEUROLOGICAL FEATURES
• If the disorder is not detected and treated,
• the lateral columns become involved
resulting in weakness, spasticity,
inability to walk,
sustained clonus, hyperreflexia,
and Babinski sign.

23. NEUROLOGICAL FEATURES
• Because the peripheral nerves,
• as well as the dorsal
• and lateral columns, are involved,
• these neurologic manifestations are
sometimes termed
• subacute combined degeneration

24. SUMMARY OF NEUROLOGICAL
FINDINGS IN VIT B12 DEFICIENCY
• Peripheral nerves Glove and stocking
paraesthesiae
• Spinal cord Subacute combined degeneration
• Posterior columns-diminished vibration and
proprioception
• Corticospinal tracts-upper motor neuron signs
• Cerebrum Dementia
• Optic atrophy
• Autonomic neuropathy

25. VIT B12 ABSORPTION
• The average daily diet contains 5-30 μg of
vitamin B12,
• mainly in meat, fish, eggs and milk
• - 1 μg is daily requirement
ABSORBTION OF VIT.B12
• In the stomach, gastric enzymes release
vitamin B12 from food
• At gastric pH it binds to a carrier protein
• ----termed R protein.-------

26. Vit B12 absorption (contd) step 2
• The gastric parietal cells produce
intrinsic factor, a vitamin B12-binding protein
which optimally binds vitamin B12 at pH 8.
• As gastric emptying occurs, pancreatic
secretion raises the pH
and vitamin B12 released from the diet
switches from the R protein to intrinsic factor.

27. VIT B 12 ABSORPTION STEP 3
• The vitamin B12 intrinsic factor complex binds
to specific receptors in the terminal ileum
and vitamin B12 is actively transported by the
enterocytes to plasma, where it binds to
transcobalamin II, a transport protein
produced by the liver,
which carries it to the tissues for utilisation.

28. Absorption vit B12

29. Absorption vit B12
The liver stores enough vitamin B12 for 3
years
Bile also contains vitamin B12 which is available
for reabsorption in the intestine.
ENTERO HEPATIC CICULATION due to these
vitamin B12 deficiency takes years to become
manifest even if all dietary intake is stopped.

30. CAUSES VIT B12 DEFICIENCY
Dietary deficiency
This only occurs in strict vegans
The breastfed offspring of vegan mothers
are at risk of developing nutritional vitamin B12 deficiency.
Less strict vegetarians often have slightly low vitamin
B12 levels
but are not tissue vitamin B12-deficient.

31. Causes vit B12 Deficiency
Gastric factors
Normal gastric acid and enzyme secretion is
required for the release of vitamin B12 from the
food.
Hypochlorhydria in elderly patients or following
gastric surgery can impair the release of
vitamin B12 from food. Total gastrectomy
invariably results in vitamin B12 deficiency
within 5 years,

32. B12 DEFICIENCY contd
Pernicious anaemia
• This is an autoimmune disorder in which
the gastric mucosa is atrophic with loss of
parietal cells causing intrinsic factor
deficiency.
• In the absence of intrinsic factor less than
1% of dietary vitamin B12 is absorbed

33. B12 DEFICIENCY
• It is more common in individuals with other
autoimmune disease
• (Hashimoto's thyroiditis, Graves' disease,
vitiligo, hypoparathyroidism or Addison's
disease)
• or a family history of these
or pernicious anaemia.

34. ANTIBODIES 2 types
1 --Anti-parietal cell antibodies
are present in over 90%
2-- Antibodies to intrisic factor
are found in the serum of 60%

35. Pernicious anaemia
• Anti-parietal cell antibodies are present in
over 90% of cases but are also present in 20%
of normal females over the age of 60 years.
• A negative result makes pernicious anaemia
less likely but a positive result is not
diagnostic.
• Antibodies to intrisic factor are found in the
serum of 60% of patients with pernicious
anaemia and, if present, are diagnostic.

36. VIT B 12 DEFICIENCY contd
Small bowel
factors
One-third of all patients with
pancreatic insufficiency
fail to transfer dietary vitamin B12
from R protein to intrinsic factor.

37. VIT B 12
Motility disorders
or hypogammaglobulinaemia
can result in bacterial overgrowth
and
the resulting competition for free vitamin B12
can result in deficiency.

38. Vit B12 deficiency
A small number of people heavily infected
with the fish tapeworm develop vitamin B12
deficiency.
Inflammatory disease of the terminal ileum,
such as Crohn's disease, may impair the
interaction of the vitamin B12-intrinsic factor
complex with its receptor AND
surgery on this part of
the bowel will both result in malabsorption of
Vit B12.

39. Diagnostic workup
• The evaluation of suspected cobalamin
deficiency generally proceeds in two stages:
• documenting the presence of the vitamin
deficiency
• and determining its cause (e.g., pernicious
anemia, malabsorption, dietary lack).

40. Diagnostic workup
• The diagnosis can often be established by
• 1) measurement of the serum cobalamin
concentration,
• 2) evaluation of specific metabolites,
and
• 3) use of the Schilling test to establish
malabsorption of cobalamin.

41. Peripheral blood smear
• The peripheral smear shows
• macro-ovalocytes, fish-tailed red blood cells,
hypersegmented neutrophils, and,
occasionally, nucleated red blood cells
Examination of the bone marrow reveals
megaloblastic erythroid hyperplasia
and giant metamyelocytes.

42. macro-ovalocytes, and bizarre-
shaped red cell forms.

43. SCHILLING,S TEST part 1
• Give 1 mg radio-labelled B12 orally+1 mg
ordinary B12 intramuscularily to block
transcobalamin binding sites to ensure any radio-
labelled B12absorbed is excreted by kidneys.
If normal gut & normal intrinsic factor
B12-----Normally absorbed
>30% excreated in urine in next 24 hours
If excretion < 30%--malabsorbtion/perncious
anaemia---part 2 of test to differentiate

44. SCHILLING,S TEST PART 2
• Give 1 mg radiolabelled B12 + intrinsic factor
orally + 1 mg ordinary B12 intramuscularily.
If gut disease---diminished absorbtion
<30% urinary excretion
In pernitious anaemia
addition intrinsic factor normalises absorbtion
> 30 % extreted in urine.

45. Treatment vit B12 deficiency
Vitamin B12 deficiency
Vitamin B12 deficiency is treated with
hydroxycobalamin 1000 μg i.m.
in five doses 2 or 3 days apart
followed by maintenance therapy of 1000 μg every 3
months for life.
The reticulocyte count will peak by the 5th-10th day
after therapy and may be as high as 50%.
The haemoglobin will rise by 10 g/l (1 gm%) every
week.

46. treatment
Vitamin B12 deficiency is treated with
hydroxycobalamin 1000 μg i.m.
in five doses 2 or 3 days apart followed by
maintenance therapy of 1000 μg every 3
months for life.
The reticulocyte count will peak by the 5th-
10th day after therapy and may be as high as
50%.
The haemoglobin will rise by 10 g/l
(1 gm %)every week.

47. Treatment vit B12 deficiency
The response of the marrow is associated with
a fall in plasma potassium levels and rapid
depletion of iron stores.
If an initial response is not maintained and the
blood film is dimorphic, the patient may need
additional iron therapy.
A sensory neuropathy may take 6-12 months to
correct; long-standing neurological damage
may not improve.

48. FOLATE DEFICIENCY
Diet Poor intake of vegetables
• Malabsorption e.g. Coeliac disease
• Increased demand Pregnancy
• Cell proliferation, e.g. haemolysis
• Drugs* Certain anticonvulsants (e.g.
phenytoin)
• Contraceptive pill
• Certain cytotoxic drugs eg Methotrexate

49. Clinical features
• Clinical features IN FOLATE DEFICIENCY
are similar to vit B12 deficiency
EXCEPT THAT
NEUROLOGICAL FEATURES ARE
NOT PRESENT IN
FOLATE DEFICIENCY

50. Diagnosis Folate deficiency
• Low serum folate levels (fasting blood
sample)
• Red cell folate levels low (but may be normal
if folate deficiency is of very recent onset)
• Corroborative findings
• Macrocytic dysplastic blood picture
• Megaloblastic marrow

51. DIAGNOSIS
• A serum folic acid level
less than 2 ng/ml is consistent with folic acid
deficiency,
as is a red blood cell folic acid level
less than 150 ng/ml.

52. Folate absorbtion
• Folates are produced by plants and bacteria;
hence dietary leafy vegetables (spinach,
broccoli, lettuce), fruits (bananas, melons) and
animal protein (liver, kidney) are a rich source.
• An average Western diet contains more than
the minimum daily intake of 50 μg but excess
cooking for longer than 15 minutes destroys
folates.

53. Folate absorption
• Most dietary folate is present as
polyglutamates; these are converted to
monoglutamate in the upper small bowel and
actively transported into plasma.
• Plasma folate is loosely bound to plasma
proteins such as albumin and there is an
enterohepatic circulation.
• Total body stores of folate are small and
deficiency can occur in a matter of weeks.

54. Causes of Folic Acid Deficiency
• inadequate intake
• Alcoholism ,Nutritional deficiencies
• resulting from increased folic acid
requirements
Pregnancy ,Severe hemolysis ,Chronic
hemodialysis or peritoneal dialysis

55. Causes of Folic Acid Deficiency
• Inadequate absorption
• Tropical sprue
• Gluten-sensitive enteropathy (nontropical
sprue)
• Crohn disease
• Lymphoma or amyloidosis of small bowel
Diabetic enteropathy
• Intestinal resections or diversions

56. Causes of Folic Acid Deficiency
Drug-induced interference with folic acid
metabolism
Action of dihydrofolate reductase blocked by
methotrexate, trimethoprim, pyrimethamine
• Reduced folate absorption and tissue folate
depletion caused by sulfasalazine
Interference of unknown mechanism caused
by phenytoin, ethanol, antituberculosis
drugs, ?oral contraceptives

57. Treatment Folate deficiency
Oral folic acid 5 mg daily for 3 weeks will
treat acute deficiency
and 5 mg once weekly is adequate
maintenance therapy.
Prophylactic folic acid in pregnancy will
prevent megaloblastosis in women at risk.

58. Treatment folate deficiency
• Folic acid supplementation may reduce the
risk of neural tube defects, and in some
countries all pregnant women receive routine
folic acid supplementation.
• Prophylactic supplementation is also given in
chronic haematological disease associated
with reduced red cell lifespan (e.g.
autoimmune haemolytic anaemia or
haemoglobinopathies).

59. thanks