in this ppt you will learn about aplastic anemia and its lab diangnosis.

1. APLASTIC ANEMIA &
LABORATORY DIAGNOSIS
GUIDED BY:-
RESP. MR. S.K BOSE SIR
RESP. MR. ISHWAR SIR
PRESENTED BY:-
BALRAM KRISHAN
B.SC MLT PART III

2. History of Aplastic anaemia
• Paul Ehrlich (1854 -1915) described the first case of aplastic
anaemia in a pregnant woman who died of marrow failure
in1888.
• The term “aplastic anaemia” first used by Anatole Chauffard in
1904.
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3. Epidemiology
• incidence -2/ 1 million people
•Acquired AA-First peak 15-25 years(maximum) & ≥60
years- second peak.
•Inherited AA- first decade of life, associated with
physical anomalies.
•
Aplastic anemia is defined by-
•Pancytopenia.
•Hypocellular bone marrow where normal
haemopoietic cells are replaced by fat.
•Absence of abnormal or malignant cells .

4. 1. Acquired aplastic anaemia- 80%.
2. Inherited aplastic anaemia- 20%.
CATEGORIES

5. Aplastic Anemia
•Congenital/inherited (20%)
• Patients usually have dysmorphic features or physical stigmata.
Occasionally, marrow failure may be the initial presenting
feature.
• Fanconi anemia
• Dyskeratosis congenita
• Diamond blackfan anemia
Acquired: (80%)
• Drugs
- Cytotoxic drugs e.g. Vincristine, andriamycin
- Antibiotics e.g. chloramphenicol, cotrimoxazole, nitrofurantioin.
- - Anti-inflammatory e.g. Phenylbutazone, oxyphenbutazone, diclofenac
- Antithyroid drugs e.g. carbimazole
Chemicals e.g. Benzene
- 2- 6 months usually between exposure and the development of aplastic
anemia. 5

6. •Viruses- EBV, Hepatitis Non-A,B,C,D,E,G
viruses, HIV and Parvo virus B19.
•Autoimmune/ Connective tissue disorders-
Eosinophilic fasciitis, SLE.
•Iatrogenic- Radiation, Cytotoxic therapy.

7. PATHOGENESIS
• Aplastic anemia is due to stem cell defect and the stem cells
fail to proliferate and differentiate.
• Mechanisms :
1. AUTOIMMUNITY
• Cytotoxic T cells produce interferon γ & TNFα
inhibits FAS on CD34 cell
Hematopoeisis apoptosis of progenitor
Stem cells undergo apopotosis and hence stem cells gradually
diminish in marrow resulting in aplasia of marrow.
2. STEM CELL
• Marked reduction in number

8. CLINICAL FEATURES
Clinical features of patients are due to
thrombocytopenia, anemia, and neutropenia.
•Pallor, weakness, fatigue, dyspnea
•petechiae, bruising, epistaxis(nose bleeding) ,
unexpected bleeding at other sites.
•Fever, chills, pharyngitis.
•Infection because of neutropenia

9. LAB DIAGNOSIS
A. Full blood count –
i. Hemoglobin ≤ 10gm/dl
ii. TLC ↓
iii. Platelet count↓
Peripheral blood smear – RBC -normocytic/ normochromic
• no abnormal cells/blasts.
• In initial stages lymphocyte count is normal but as the
disrease progresses lymophocyte count also falls.

10. 2. Reticulocyte count (decrease)
• Normal value - 0.5-2.5%
(i). Manual count
• New methylene blue or Brilliant crystal blue
(ii) automated counting
3.Corrected reticulocyte count
⁼Retic(%)× {PCV of patient (%)÷45}

11. 2. BONE MARROW EXAMINATION
Aspirate –
1. Morphology-
2. Cytogenetics &Immunophenotyping to differentiate MDS
3. Iron staining.
4. DNA/antigen-based viral tests
Trephine biopsy
• Assess cellularity
• Infiltrative ,malignant disease or storage disease
• Fibrosis or granuloma.

13. 3. PNH work up- Ham’s test and Sensitive multi-color flow-
cytometry.
4. PCR for Hepatitis viruses and Parvovirus B19.
5 Liver function tests.
6. Auto-antibody screen.
7. Vertebral MRI- Uniformly fatty in AA
(Myelodysplastic syndromes)

14. Laboratory Evaluation for Aplastic Anemia
Complete blood cell count and differential  Define severity of aplasia
1. Morphology  Define abnormal cells (e.g., blasts or storage cells)
Vitamin B 12 deficiency
2. Reticulocyte count  Define severity Differentiate production vs.
destruction
Bone marrow biopsy  To assess cellularity

15. Features AA Hypoplastic MDS
•
• Cytopenia Present Present
• BM
1. cellularity Aplastic or hypo cellular Hypo cellular
2. Erythropoiesis Reduced Present
3. Myelopoiesis decreased Present
4. Megakaryopoiesis Decreased or absent Present
5. Dysplasia Normal Possible
6. Blasts Absent Variable
7. PNH clone Frequent Unusual

16. AA and PNH
•≥50 % patients with aplastic anemia may have a PNH
cell population.
•PNH – defect in glycosylphosphatidylinositol (GPI)-
linked proteins (CD 55 &59)
•Due to PIG-A mutation
•The probability of patients with aplastic anemia
developing PNH is 10 to 20%
•Diagnosis - Flow cytometry
•Ham’s test

17. Inherited aplastic anaemia
1. Fanconi anaemia
2. Dyskeratosis Congenita
3. Shwachman-Diamond syndrome
4. Congenital Amegakaryocytic Thrombocytopenia

18. FANCONI ANAEMIA
• most common
• Autosomal recessive.
PATHOGENESIS
• Cells from FA show
chromosomal breakage and
hypersensitivity to the effects
diepoxybutane (DEB) and
mitomycin-C (MMC).
•

19. Clinical features
•Growth retardation
•Short stature
•Skeletal anomalies- Absent, misshapen, or
supernumerary thumbs, dysplastic radii, hip and
vertebral abnormalities
•heart defects, eye abnormalities,fused kidneys,
•mental retardation.

20. LAB. DIAGNOSIS
• chromatid breaks seen in
myeloid cells, lymphocytes.
• This chromosome damage is
intensified after exposure to
DNA cross-linking agents such as
mitomycin C or diepoxybutane.

21. Dyskeratosis congenita
• Cutaneous and mucous membrane abnormalities.
• Progressive marrow insufficiency.
• Predisposition to malignant transformation.
• Inheritance- X- linked recessive.

22. Diagnosis
• Phenotypic findings.
• Cytopenias.
• Genetic analysis for telomerase complex gene mutations.
• Shortened telomere length in leukocytes also can be
assessed by flow cytometric fluorescence in situ
hybridization studies.

23. Shwachman Diamond syndrome
• Once in every 100,000 births.
• Exocrine pancreatic insufficiency with secondary steatorrhea.
• Blood cell deficiencies.
• Skeletal abnormalities.

24. Some imp points regarding lab
• ESR is increased in case of pancytopenia.
• Roulex formation is also occur because of high sedimentation rate .
• Toxic granules occasionally occur in aplastic anaemia
• Bone marrow failure resulting in aplastic anaemia may manifest
first and them may develop PNH also because of being stem cell
disorder .
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25. Thankyou