medullary myeloblastosis ¾ Aplastic anemia ¾ Pancytopenia with hyperplastic bone marrow ¾ Bone marrow infiltration or replacement ¾ Hypersplenism ¾ Megaloblastic macrocytic anemia ¾ Systemic lupus erythematosus

 

Clinical Hematology 257

In b Thalassemia

Level of HbA, A2 and F are equally depressed leading on

to microcytic, hypochromic anemia. In the absence of

normal chains, b or γ chains increase and form HbH (b4)

or Hb Bart’s (γ4).

Laboratory Diagnosis of b Thalassemia Minor

Blood Picture

Hemoglobin is normal, MCV and MCH. MCHC is normal.

Peripheral Smear

¾ Microcytic, hypochromic cells; target cells

¾ Basophilic stippling, reticulocytosis (up to 6%)

¾ Osmotic fragility is decreased.

¾ Evidence of hemolysis

¾ HbA2 increased, slight increase in HbF.

Laboratory Diagnosis of b Thalassemia Major

Blood Picture

(Resembles iron deficiency anemia)

¾ Marked microcytosis with hypochromia

¾ Moderate degree of anisopoikilocytosis, teardrop cells

¾ Target cells are prominent

¾ All; MCV, MCH and MCHC are diminished

¾ Normoblasts (intermediate/late) in peripheral smear.

¾ Granular inclusions in cytoplasm, represent a-chain

aggregates which can be shown by methyl violet

¾ Polychromasia and moderate degree of punctate

basophilia

¾ Reticulocytosis (10%)

¾ Leukocytosis (5,000–40,000) with shift to the left

(immature forms seen)

¾ Platelet count normal but decreases with splenomegaly

¾ Osmotic fragility decreased, the curve has a tail because

of a few very fragile cells

¾ Evidences of intravascular hemolysis.

Bone Marrow

¾ Erythroid hyperplasia

¾ Micronormoblastic reaction

¾ Increase in early and intermediate normoblasts

¾ Methyl violet positive inclusion bodies and PAS positive

glycogen (PAS-periodic acid Schiff’s reagent)

¾ Sideroblastosis, often ring sideroblasts.

Hb Pattern on Electrophoresis

¾ HbF (10–98%) estimated also by acid elution and alkali

denaturation tests

¾ HbA (very little or absent)

¾ HbA2 (variable).

Laboratory Diagnosis of HbH Disease

Blood Picture

¾ Marked microcytosis with hypochromia

¾ Target cells, fragmented cells and normoblasts in

peripheral blood

¾ Reticulocytosis, basophilic stippling

¾ Numerous HbH inclusions.

Hb Pattern (Electrophoresis)

¾ HbH = 5–25%

¾ Remainder is HbA, A2 and F.

Demonstration of HbH Inclusions

With a redox dye, e.g. brilliant cresyl blue (BCB) or new

methylene blue (NMB); HbH being relatively unstable,

precipitates and the red cells appear pitted with numerous

inclusions (golfball appearance).

Hemoglobin Electrophoresis

At pH 8.6, both HbH and Hb Bart’s are fast moving, they

migrate in front of HbA towards anode.

Hb Bart’s—Hydrops Fetalis

Blood Picture

¾ Marked anisopoikilocytosis with hypochromia

¾ Polychromasia

¾ Target cells and normoblasts.

Hb pattern

¾ Hb Bart’s 80–90%

¾ Some HbH and Hb Portland

¾ No HbA, A2 or F.

Laboratory Diagnosis of Aplastic Anemia

Blood Picture

¾ RBC morphology is usually normal (polychromasia,

normoblasts and stippling usually not seen)

¾ Sometimes macrocytosis, mild anisopoikilocytosis

¾ Hb and PCV are diminished to about 7 g% and 20%

respectively

¾ Red cell osmotic fragility is normal

¾ Leukopenia—neutropenia, polymorphs are qualitatively

and quantitatively abnormal, show coarse granules

• Absolute lymphocytopenia but relative lymphocytosis

• Neutrophil alkaline phosphatase count is increased

258 Concise Book of Medical Laboratory Technology: Methods and Interpretations ¾ Thrombocytopenia—bleeding time is increased

¾ Clot retraction time is increased

¾ Coagulation parameters are abnormal

¾ Serum iron is increased

¾ Radio-iron bone marrow uptake is reduced

¾ Red cell life may be decreased

¾ Plasma erythropoietin level is raised.

Bone marrow may be:

¾ Aplastic

¾ Hypoplastic

¾ Normal or

¾ Patchily hypercellular.

Aspiration

¾ Blood tap or dry tap

¾ Is hypoplastic (there is both myeloid and erythroid

hypoplasia)

¾ Fat cells are increased

¾ Plasma cells and reticulum cells are prominent

¾ Megakaryocytes are diminished in number

¾ WBCs may show maturation arrest

¾ Developing granulocytes are abnormal and may show:

• Abnormal granulation or.

• Vacuolization.

¾ Bone marrow iron may be normal or increased.

Hence, the diagnosis is based upon:

¾ Pancytopenia

¾ Rapid ESR

¾ No immature cells in peripheral smear

¾ Bone marrow is aplastic/hypoplastic (if 2 consecutive

aspirations have been unsuccessful do a bone marrow

trephine biopsy).

Classification and Causes of Aplastic Anemia

1. Primary

• Idiopathic (cause unknown).

2. Secondary

• Effects of chemical/physical agents on bone

marrow.

3. Miscellaneous

• Familial hypoplastic anemia (Fanconi’s syndrome)

• Aplastic anemia associated with

– Infective hepatitis (1–2%) (usually post hepatitis A)

– Pancreatic insufficiency

– Paroxysmal nocturnal hemoglobinuria (PNH).

4. Pure red cell aplasia

• Congenital (Diamond Blackfan type)

• Acquired

– With thymoma

– Without thymoma. (In both primary and secondary aplastic anemia clinical and hematological

features are similar)

– Primary type is less common.

 Secondary type occurs due to:

• Chemicals

– Pancreatic

– Industry

– Domestic

– Drugs.

• Physical

• Ionizing radiation.

Drugs

Especially those which have an amino group near a

benzene ring.

Bone marrow depression depends upon:

¾ Dosage and period of treatment

¾ Idiosyncrasy, susceptibility and hypersensitivity to the

drug.

Drugs which regularly cause aplastic anemia:

¾ Anticancer drugs.

Drugs which occasionally cause bone marrow depression:

(Either because of idiosyncrasy or hypersensitivity)

¾ Antiepileptics

• Phenylhydantoin

• Mesantoin

• Paradione.

¾ Antibacterials

• Chloramphenicol

• Sulfas

• Streptomycin

• Chlortetracycline

• INH.

¾ Tranquillizers

• Chlordiazepoxide

• Chlorpromazine.

¾ Antidiabetics

• Tolbutamide

• Chlorpropamide.

¾ Antirheumatics

• Oxyphenylbutazone

• Phenylbutazone

• Indomethacin

• Gold salts

• Aspirin

• Colchicine. (Commonest drugs—phenyl and oxyphenylbutazone and chloramphenicol).

Clinical Hematology 259

Chemicals

• Benzene

• Lindane

• TNT

• DDT.

Physical Agents

¾ Ionizing radiation is most dangerous (e.g. X-ray, γ rays,

neutrons).

¾ Particles have a limited range and hence cause effects

only on entering the body.

Pancytopenia

Reduction in number of the three blood cells types RBCs,

WBCs and platelets.

Blood Picture

Hb < 13.5 g% in males

< 11.5 g% in females

TLC < 4,000 cells/cu mm,

Platelet count < 1.5 lakh/cu mm.

Causes

¾ Subleukemic leukemia: Refractory anemia with

medullary myeloblastosis

¾ Aplastic anemia

¾ Pancytopenia with hyperplastic bone marrow

¾ Bone marrow infiltration or replacement

¾ Hypersplenism

¾ Megaloblastic macrocytic anemia

¾ Systemic lupus erythematosus

¾ Disseminated tuberculosis.

NORMAL WHITE CELL VALUES AND

PHYSIOLOGICAL VARIATIONS

Normal total leukocyte count = 4,000–11,000 cells/cu mm.

Total leukocyte count (TLC) undergoes minor physiological and diurnal variations. It increases slightly in

the afternoon ‘afternoon tide’. Various stimuli that may