Anemia

1. MEGALOBLASTIC
ANEMIA

2. MEGALOBLASTIC ANEMIA
 Megaloblastic anemia are a heterogenous group
of disorders caused by impaired DNA synthesis
and characterized by the presence of
megaloblasts, the morphologic hallmark of this
group of anemia.
 2nd most common type of anemia
 Usually due to vitamin B12/ Folic acid deficiency
 Multisystem involvement- All the organs with
increased cell turnover are affected.

3. Approach to Macrocytic Anemias
 Divided into 2 groups
– Megaloblastic anemias
– non-Megaloblastic anemias
two most valuable findings for differentiating
megaloblastic from non- megaloblastic anemias
are
– Hpersegmentation of neutrophil
– oval macrocytes
 neutrophil hypersegmentation – neutrophils with 6 or
more lobes (1 or more) or presence of at least 4 – 5 %
of neutrophils with 5 lobes

4. CLASSIFICATION OF MEGALOBLASTIC
ANEMIA
 COBALAMIN DEFICIENCY:
I. Inadequate intake – vegetarians
II. Malabsorption
a) Defective release of cobalamin from food
 Gastric achlorhydria
 Partial gastrectomy
 Drugs that block acid secretion
b) Inadequate production of intrinsic factor
 Pernicious anemia
 Total gastrectomy
 Congenital absence or functional abnormality of IF.
c) Disorders of terminal ileum
 Tropical sprue
 Non tropical sprue
 Regional enteritis
 Intestinal resection
 Neoplasm and granulomatous disorders

5. d) Competition for cobalamin
 Fish tape worm
 Blind loop syndrome
e) Drugs
 Para amino salicylic acid
 Colchicin
 Neomycin

6.  FOLIC ACID DEFICIENCY
I. Inadequate intake – unbalanced diet (common in
alcoholics, teenagers, infants)
II. Increased requirements
a) Pregnancy
b) Infancy
c) Malignancy
d) Increased haematopoiesis (chronic hemolytic anemia)
e) Chronic exfoliative skin disorders
f) Haemodialysis
III. Malabsorption
a) Tropical sprue
b) Non tropical sprue
c) Drugs – phenytoin, barbiturates, ethanol
IV. Impaired metabolism
a) Inhibitors of DHFR – methotrexate, pyrimethamine,
triamterene, pentamidine, trimethoprim
b) Alcohol
c) Rare enzyme deficiencies - DHFR

7.  OTHER CAUSES
I. Drugs that impair DNA metabolism
a) Purine antagonist – 6 mercaptopurine, azathioprine
b) Pyrimidine antagonist – 5 fluorouracil, cytosine arabinoside.
c) Others – hydroxyurea, procarbazine, acylovir, zidovudine
II. Metabolic disorders
a) Hereditary orotic aciduria
b) Lesch Nyhan syndrome

8. PATHOGENESIS OF MEGALOBLASTIC
ANEMIA
1. Unbalanced cell growth:
 There is decreased DNA synthesis while RNA
synthesis is normal. As the cell differentiates the
chromatin condenses more slowly than normal to
give the nucleus a characteristic fenestrated
appearance. (sieve like)
 As the cytoplasm acquires hemoglobin, it’s
growing maturity contrasts with the immature
looking nucleus, a featured termed NUCLEAR
CYTOPLASMIC ASYNCHRONY.
 This results in impaired cell division of erythroid
precursors leading to anemia.

9. 2. Ineffective erythropoiesis: Though
erythropoiesis is hyperplastic, yet RBCs
formed are few because of intramedullary
death of intermediate and late normoblast

10. VITAMIN B12 AND FOLIC ACID-
PHYSIOLOGIC CONSIDERATIONS
Vitamin B12 Folic acid
Sources meat, fish green
vegetables, yeast
Daily requirement 2-5 ug 50-100 ug
Body stores 3-5 mg (liver) 10-12mg(liver)
Places of absorption Ileum Duodenum
and proximal
segment of small
intestine

11. VITAMIN B-12 METABOLISM

12. FOLIC ACID ABSORPTION
Polyglutamates (food)
Monoglutamates
Dihydrofolate
Tetrahydrofolate
Methyl tetrahydrofolate
Absorbed from jejunum and enters circulation.
Carried by plasma to the cells where DNA synthesis takes place
Intestinal conjugases

13. METABOLIC EFFECTS OF VIT B-12 AND FOLIC ACID
Directly or indirectly both folic acid and vitamin B-12 are involved in DNA
synthesis
FH2
5,10 Methylene Polyglutamate
FH4
N5 – Methyl FH4
Vit. B12
Methionine
Homocysteine
Vit. B12
Methionine Synthase
d UMP
d TMP
DNA

14. METHYL FOLATE TRAP
HYPOTHESIS
 In vitamin B12 deficiency, N5
Methyl FH4 is not converted
into FH4, which is further not
converted into DNA.
 This N5 Methyl FH4 gets
accumulated into the cell.
 This is the methyl folate trap
hypothesis.
 This hypothesis explains why
tissue folate stores in
cobalamin deficiency are
substantially reduced with
disproportionate reduction in
conjugated folates, despite
normal or supra normal
serum folate levels.

15. MMCoA
Mutase
Methylmalonyl CoA
Succinyl malonyl CoA
Vit B12
METABOLIC EFFECT OF Vit B12 DEFICIENCY OF Vit B12
MM CoA
accumulates
Formation of abn
fatty acids
Neurologic complications
of Vit B12
Predisposes to
myelin breakdown
Excreted in urine
as MMA

16. MEGALOBLASTIC ANEMIAS
clinical features
1.Signs & symptoms of anemia – weakness, fatigue, light
headedness, pallor, slight
jaundice.
.
2. Symptoms associated with deficiency
 Neurologic manifestations (exclusivly in vit. B12
deficiency)
- megaloblastic madness or psychosis,
- unsteadiness of gait (spastic or scissor gait)
- reduced vibration sense, esp. at higher frequency
- impaired position sense.
These symptoms occur due to subacute combined
demyelination of posterolateral columns of the spinal cord.

17. Peripheral Neuropathy
 Most of the patients complain of bilaterally
symmetrical tingling (PINS & NEEDLES
SENSATION), numbness or coldness of
extremities, beginning from toes and extending
proximally.
Later on patients develop STOCKING AND
GLOVE distribution of numbness in all the four
limbs.
Gastrointestinal complaints
(vit.B12 and folic acid deficiency)
 - loss of appetite
 - glossitis (red, sore, smooth tongue)
 - diarrhoea or constipation .

18. Less common neurological manifestations
 Atony of the bladder
 Impotence
 Loss of senses of taste & smell
The degree of nervous system involvement
does not correlate with the degree of anemia.
If untreated, the neurologic disease is progressive
and the severity of manifestations is strongly
related to the duration of symptoms.

19. Investigations of Megaloblastic
anemia
 Hematologic findings
 Biochemical profile
HEMATOLOGIC FINDINGS:
 hemoglobin – decreased
 hematocrit – decreased
 MCV – increased (>110 fl)
 MCH – increased
 MCHC – Normal

20. –Peripheral smear findings
 RBC sparse distribution
 pancytopenia
 Mod. to marked degree anisopoikilocytosis
 macro-ovalocytes
 hypersegmented neutrophil
 nRBC with megaloblastoid open nuclear
chromatin
 Evidences Of Dyserythropoiesis
–Basophilic stippling
–Cabot ring
–Howell Jolly bodies

21. – Leucocytes
 marked leucopenia with relative
lymphocytosis
 Hypersegmented neutrophils
– Platelets
 variable degree of thrombocytopenia

22. Normal Blood Cells

23. PBS In Megaloblastic anemia

24. Megaloblastic Anemia :

25. ANISOPOIKILOCYTOSIS

26. Basophilic Stippling

27. BASOPHILIC STIPPLING (Punctate basophilia)
 Randomly distributed, fine to coarse granular blue
black inclusions seen in RBCs.
 These are precipitated ribosomal RNA.

28. Howell- Jolly bodies
 These inclusions are nuclear remnants
(aggregates of chromatin material) seen in
red cells, intermediate / late normoblast.

29. Cabot ring

30. Bone marrow picture
 Cellularity – moderately to markedly hyper
cellular
 Marked erythroid hyperplasia
 Megaloblastic erythropoiesis – Megaloblasts are
larger than normoblasts and have open seive
like nuclear chromatin.

31.  Myelopoiesis- Though myeloid cells appear adequate
in number, yet patients with severe anemia manifest
neutropenia.
 Larger myeloid cells esp.GIANT METAMYELOCYTES
seen.
 Megakaryopoiesis-
 Reduction in the number of megakaryocytes
 Nuclear abnormalities like: hypersegmentation of the
nucleus
 Open nuclear chromatin network which may result in
thrombocytopenia.

32.  B. M. Iron-
- Moderately increased.
- Megaloblasts show coarse iron granules
in the cytoplasm (ABNORMAL
SIDEROBLASTS).

33. Normoblast Megaloblast
Size Normal Larger for corresponding
normoblast
Nuclear Chromatin Normal More open – sieve like
Evidence of
dyserythropoisis
None Present – irregular nuclei
- Howell jolly bodies
Maturation Late > intermediate >
early normoblasts
Late < intermediate < early
megaloblasts
Mitosis Normal Increased, abnormal
Nuclear maturation Normal Lags behind the cytoplasmic
maturation
Late normoblasts Pyknotic nuclei Nuclei have open chromatin
Myelopoiesis Normal Giant metamyelocytes +

36. Biochemical changes in
megaloblastic anemia
Test Normal values Vit.B12 def. F.A. def.
S.Vit. B12 200 – 800 pg/ml ↓ N/↓
S. Folate 3 – 16 ng/ml N/↑ ↓
Red Cell Folate 150 – 450 ng/ml ↓/N ↓
S. Methylmalonic
acid
<280 nmol/l ↑ N
Urinary MMA 0 – 3.4 mg/day ↑ N
FIGLU in urine N ↑

37. Schilling Test
 Detects deficiency of intrinsic factor vs nutritional
deficiency of vit b12.

38. PERNICIOUS ANEMIA
 It is a chronic disease resulting from deficiency
of INTRINSIC FACTOR, leading to impaired
absorption of vitamin B12 and eventually
megaloblastic anemia.

39. SEQUENCE OF EVENTS IN PERNICIOUS ANEMIA
T-Cell mediated mucosal cell injury (stomach) & formation of antibodies (Abs).
Antibodies further damage the mucosal cells.
Parietal cells damaged + mononuclear cell infiltration.
Atrophic gastritis.
Parietal cell mass diminishes.
IF + HCl secretion falls.
Reduced absorption of Vit. B12
B12 Deficiency
Megaloblastic Anemia.

40. Non megaloblastic macrocytic
anemias
 RBCs of newborn : physiological
 Alcoholism
 Liver disease
 Hypothyroidism
 Reticulocytosis
 Aplastic anemia
 Myelodysplastic syndrome