Rapid Review of Hematology.1E.2014.PDF - unitedVRG

Rapid Review
of
Hematology
Rapid Review
of
Hematology
Ramadas Nayak MBBS MD

Professor
Department of Pathology
Kasturba Medical College
Manipal University
Mangalore, Karnataka, India
askdr.nayak@gmail.com
Sharada Rai MBBS MD

Associate Professor
Kasturba Medical College
Manipal University
Mangalore, Karnataka, India
askdr.nayak@gmail.com
Foreword
AR Raghupathy
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Rapid Review of Hematology
First Edition: 2014
ISBN 978-93-5090-961-4
Printed at
Dedicated to
Students who inspired us,
patients who provided the knowledge,
our parents and family members who
encouraged and supported us.
Foreword
DEPARTMENT OF PATHOLOGY BANGALORE MEDICAL COLLEGE

VICTORIA HOSPITAL COMPLEX
BENGALURU - 560 002
Phone: 670 1150 Ext.: 314, 315, 316, 317
It gives me great pleasure to write a short foreword for this new book on Rapid Review of Hematology.
This is a well-written concise but precise and student-friendly text that will be highly valuable to medical students. It
will help in revising and reinforcing the fundamental concepts in hematology. It is very well organized with optional and
correct usage of good pictures, schematic diagrams and flow charts. Every essential topic has been discussed giving opt
importance and stress on salient features. Each statement mentioned in the text is well written as it carries the required
essential points.
In short, this book provides within one volume a user-friendly review of the basic essential concepts in hematology.
It will be of great help to not only second year MBBS students, but also for students preparing for entrance examinations,
and students of allied sciences.
This book will certainly serve as a valuable gift and a valuable addition to the students’ library and the user will
definitely appreciate the content and presentation of the information in this book.
In conclusion, I am sure, this book brought out by Dr Ramadas Nayak and Dr Sharada Rai will be a very useful
compendium for second year MBBS students, the students preparing for entrance examinations, and students of allied
health sciences.
I hope the reader of this new book will get as much pleasure and knowledge as I did.
AR Raghupathy MBBS MD PGDHHM (IGNOU)

Professor and Head
Bangalore Medical College and Research Institute
Bengaluru, Karnataka, India
Preface
Hematology is one of the rapidly expanding fields of medicine and emerging as a clinical specialty in its own right.
Hematology is difficult to teach at the undergraduate level, as it is a part of the curriculum in Pathology, during which
undergraduate students do not have enough exposure to diseases of blood. This results in less attention to hematological
diseases at undergraduate level. After many years of teaching undergraduates, we found that undergraduate students
either neglect hematology or find it difficult to understand the subject. It is a nightmare for many students especially
during examinations. There are many hematology textbooks, but undergraduates face difficulties to refresh their
knowledge of hematology during examinations. This encouraged us to write a book to fill the niche, to provide basic
information to an undergraduate in a nutshell. With this view in mind, Rapid Review of Hematology is intended for the
undergraduates from medical, dental and paramedical fields. Most students are fundamentally “visually oriented”. As
the saying “one picture worth thousand words”, it encouraged us to provide many illustrations (e.g. etiopathogenesis,
clinical presentation, complications, peripheral blood smear and other relevant laboratory tests).
Organization
This book is organized into four sections namely disorders of red cells, disorders of white cells, disorders of hemostasis
and clinical scenario.
The final section deals with common clinical scenario encountered during theory examination.
How to use this book

We recommend that this book not to be used as a hematology textbook rather than a supplement to “Essentials in
Hematology and Clinical Pathology” (Authored by Dr Ramadas Nayak, Dr Sharada Rai and Dr Astha Gupta). The
concepts of hematology have been oversimplified in this book, but all the information, the student will ever need to
know, have been provided. The readers are requested to give more emphasis on word in bold letters that represents
the key words to be remembered. The peripheral smear and bone marrow findings have been highlighted in colored
background. Boxes have been provided at the sides of main text. These include some of the key points as well as commonly
expected questions during examinations. This book can serve as a source of rapid review of hematology.
Ramadas Nayak
Sharada Rai
Acknowledgments
•• Our sincere thanks to Ms Prathiba Bhat for her untiring efforts, patience and excellent support in creating many
illustrations for this book.
•• Acknowledgments are also due to Dr Astha Gupta (Consultant Pathologist, New Delhi, India), Dr Rakshatha
(KS Hegde Medical college, Mangalore, Karnataka, India), Ms Rekha Nayak, Ms Rashmitha Nayak, and Mr Ramnath Kini
for their contribution in the preparation of the manuscript.
•• Our sincere thanks to Dr AR Raghupathy, Professor and Head, Department of Pathology, Bangalore Medical College
and Research Institute, Bengaluru, Karnataka, India, for his support and guidance.
•• We are grateful to Dr K Ramnarayan, Vice Chancellor of Manipal University, Manipal, Karnataka, India, and
Dr M Venkatraya Prabhu, Dean, Kasturba Medical College, Mangalore, Manipal University, Karnataka, India, for their
encouragement.
•• We are grateful to all our friends, undergraduate and postgraduate students who have inspired and supported us.
•• We wholeheartedly thank Shri Jitendar P Vij (Group Chairman), Mr Ankit Vij (Managing Director), Mr Tarun Duneja
(Director-Publishing), Ms Chetna Malhotra Vohra (Sr Manager, Business Development) of M/s Jaypee Brothers
Medical Publishers (P) Ltd, New Delhi, India, for publishing the book in the same format as wanted well in time.
•• We acknowledge the wonderful work done by Ms Sunita Katla (Publishing Manager), Ms Samina Khan
(PA to Director), Mr KK Raman (Production Manager), Mr Rajesh Sharma (Production Coordinator), Ms Seema
Dogra (Cover Designer), Mr Sarvesh Kumar Singh (Proofreader), Mr Rajesh Ghurkundi (Graphic Designer), and
Mr Raj Kumar (DTP Operator) of M/s Jaypee Brothers Medical Publishers (P) Ltd, New Delhi, India.
•• We thank especially Mr Venugopal V and Mr Vasudev H of M/s Jaypee Brothers Medical Publishers (P) Ltd,
Bengaluru Branch, Karnataka, India, for taking this book to every corner of Karnataka.
Contents
Section 1: Disorders of Red Cells

1. Anemias of Impaired Red Cell Production 3
Anemia 3
Red cell indices 4
Iron deficiency anemia 5
Megaloblastic anemia 8
Pernicious anemia 11
Aplastic anemia 13
2. Hemolytic Anemias Due to Red Cell Membrane and Enzyme Defects 16
Hemolytic anemia 16
Hereditary spherocytosis 17
Glucose-6-phosphate dehydrogenase deficiency 20
3. Thalassemia Syndrome 22
Classification of hereditary defects in hemoglobin 22
Thalassemia syndrome 22
b-thalassemia 22
b-thalassemia major 23
b-thalassemia minor/trait 27
a-thalassemia 28
4. Sickle Cell Disease 29
Sickle cell disease 29
Sickle cell anemia 29
Sickle cell trait 34
5. Other Anemias 36
Immunohemolytic anemias 36
Hemolytic disease of the newborn 36
Antiglobulin (Coombs) test 39
Autoimmune hemolytic anemia 40
Fragmentation syndrome 41
Paroxysmal nocturnal hemoglobinuria 41
Anemias of blood loss 41
Sideroblastic anemias 42
xiv Rapid Review of Hematology
Section 2: Disorders of White Cells

6. Quantitative and Qualitative Disorders of Leukocytes 45
Normal differential leukocyte count (DLC) 45
Quantitative disorders of leukocytes 45
Qualitative disorders of leukocytes 50
Infectious mononucleosis (Glandular fever) 51
7. Acute Leukemia 52
Acute leukemia 52
Acute lymphoblastic leukemia/lymphoma 55
Acute myelogenous leukemia 57
Myeloid sarcoma 59
8. Myelodysplastic Syndromes 60
Myelodysplastic syndromes 60
9. Myeloproliferative Neoplasms 62
Myeloproliferative neoplasms (MPN) 62
Polycythemia or erythrocytosis 63
Polycythemia vera 63
Essential thrombocythemia 65
Primary myelofibrosis 66
10. Chronic Myelogenous Leukemia 68
Chronic myelogenous leukemia 68
Natural history of chronic myeloid leukemia 70
11. Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma 73
Chronic lymphocytic leukemia 73
Hairy cell leukemia 75
12. Plasma Cell Neoplasms 76
Plasma cell myeloma (multiple myeloma) 76
Plasmacytoma 80
Immunoglobulin deposition disease 80
Monoclonal gammopathy of uncertain significance (MGUS) 80
13. Lymphoid Neoplasms 81
Classification of lymphoid neoplasms 81
Follicular lymphoma (FL) 82
Diffuse large B cell lymphoma (DLBCL) 83
Burkitt lymphoma (BL) 83
Mature T cell and NK cell neoplasms 85
14. Hodgkin Lymphomas 87
Definition 87
Classification 87
Morphology of neoplastic cells 88
Classical Hodgkin lymphoma 88
Nodular lymphocyte predominant Hodgkin lymphoma (NLPHL) 92
Etiology and pathogenesis of Hodgkin lymphoma 93
Contents xv
Laboratory findings 93
Staging of Hodgkin lymphoma 94
Differences between Hodgkin lymphoma and non-Hodgkin lymphoma 94
15. Langerhans Cell Histiocytosis/Histiocytosis X 95
Morphology 95
Laboratory findings 95
Section 3: Disorders of Hemostasis

16. Disorders of Primary Hemostasis 99
Normal hemostasis 99
Classification of hemostatic disorders 99
Bleeding disorders caused by vessel wall abnormalities 99
Bleeding disorders due to abnormalities of platelet 100
Thrombocytopenia 100
Immune thrombocytopenic purpura 102
Thrombocytosis 104
Qualitative platelet disorders 104
17. Bleeding Disorders: Due to Abnormalities of Coagulation/Clotting Factor 105
Classification of coagulation disorders 105
Hereditary coagulation disorders 106
Hemophilia 106
Hemophilia A (Factor VIII deficiency) 106
Hemophilia B (Christmas disease, factor IX deficiency) 108
von Willebrand disease (vWD) 108
Acquired coagulation disorders 109
Disseminated intravascular coagulation 110
18. Thrombotic Disorders: Hypercoagulable State 113
Hypercoagulable state (Thrombophilia) 113
Inherited hypercoagulable states 114
Acquired hypercoagulable states 114
Section 4: Clinical Scenario

19. Clinical Scenario 119
Symptoms and signs that suggest a blood disease 119
Patterns strongly suggestive of a blood disease 120
Appendix 127
Bibliography 133
Index 135
Anemias of Impaired Red Cell Production CHAPTER 1 1
SECTION 1
Disorders of
Red Cells
Anemias of Impaired
Red Cell Production
1
CHAPTER
ANEMIA Q. Define anemia.
Definition
•• Decrease below normal of the hemoglobin concentration (Hb)/RBC count/hematocrit
WHO criteria for anemia:
(packed cell volume). adult males Hb <13 g/dL
•• Reduction of the total circulating red cell mass below normal limits. and adult female Hb <12
•• Decrease in the oxygen-carrying capacity of the blood, which leads to tissue hypoxia. g/dL.
Anemia may be absolute (decreased RBC mass), or relative (associated with a higher plasma
volume). Anemia is conventionally used for absolute anemia. Grading of anemia:
mild (Hb 9.1–10.5 g/dL),
moderate (Hb 6.0–9.0 g/dL)
and severe (Hb < 6.0 g/dL).
Classification of Anemia
1. Morphological classification (Table 1.1): it is based on:
a. Red cell size (normocytic, microcytic, or macrocytic), and
Anemia is characterized by
b. Degree of hemoglobinization (normochromic or hypochromic). decreased oxygen carrying
capacity of blood. Shows
TABLE 1.1: Morphological classification of anemia decreased Hb and PCV.
Type of anemia Microcytic hypochromic Normocytic normochromic Macrocytic

Size of RBCs Smaller than normal Normal Larger than normal Q. Classify anemia.
Central pallor in RBCs More than 1/3 Normal Normal
Mean corpuscular Reduced (< 80 fL) Normal (82–98 fL) Increased (>100 fL)
volume (MCV)
Classification: anemias are
Mean corpuscular Reduced (< 30 g/dL) Normal (31–36 g/dL) Normal (31–36 g/dL) mainly classified based
hemoglobin on 1) morphology and 2)
concentration (MCHC) etiology.
Examples Iron deficiency anemia, During blood loss, anemia of Deficiency of vitamin
thalassemia chronic diseases B12 and folic acid
Morphology of RBC
Spurious anemia is the
term used when RBC
concentration decreases
due to hemodilution as
seen in third semester of
pregnancy.
4 SECTION 1 Disorders of Red Cells
Anemia is the expression 2. Etiological classification: The etiological classification of anemia is listed in Table 1.2.
of underlying disease and
from treatment point, the
cause of anemia must be TABLE 1.2: Etiological classification of anemia
identified. 1. IMPAIRED RED CELL PRODUCTION
Disturbed Proliferation and Maturation of Erythroblasts
• Defective DNA synthesis
Causes of anemia: –– Megaloblastic anemias due to deficiency or impaired utilization of vitamin B12 and folic acid
1. Decreased RBC –– Anemia of renal failure due to deficiency of erythropoietin
production –– Anemia of chronic disease due to iron sequestration and relative erythropoietin deficiency
2. Increased RBC
–– Anemias of endocrine disorders
destruction (hemolysis)
• Defective hemoglobin synthesis
or
–– Defective heme synthesis: iron deficiency, sideroblastic anemia
3. Blood loss.
–– Defective globin synthesis: thalassemias
Marrow Replacement
• Primary hematopoietic neoplasms: acute leukemia, myelodysplastic syndromes
Marrow Infiltration (myelophthisic anemia)
• Metastatic neoplasms
Disturbed Proliferation and Differentiation of Stem Cells
• Aplastic anemia, pure red cell aplasia
Iron deficiency anemia is 2. INCREASED RED CELL DESTRUCTION (HEMOLYTIC ANEMIAS)
the most common anemia.
Intrinsic (Intracorpuscular) Abnormalities
• Hereditary
–– Membrane abnormalities: spherocytosis, elliptocytosis
–– Enzyme deficiencies: glucose-6-phosphate dehydrogenase, pyruvate kinase
–– Disorders of hemoglobin synthesis
◆◆ Deficient globin synthesis: thalassemia syndromes
◆◆ Structurally abnormal globin synthesis (hemoglobinopathies): sickle cell anemia
• Acquired
–– Membrane defects: paroxysmal nocturnal hemoglobinuria
Extrinsic (Extracorpuscular) Abnormalities
• Antibody-mediated
–– Isohemagglutinins: transfusion reactions, Rh disease of the newborn
–– Autoantibodies: idiopathic (primary), drug-associated, systemic lupus erythematosus
• Mechanical trauma to RBCs:
–– Microangiopathic hemolytic anemia: disseminated intravascular coagulation
• Infections: malaria
3. BLOOD LOSS
• Acute: trauma
• Chronic: lesions of gastrointestinal tract (e.g. carcinoma colon), gynecological disturbances
Q. Write short notes on red cell RED CELL INDICES

indices.
Red cell indices are useful in morphological characterization and diagnosis of anemias. They
are either directly measured or automatically calculated by specialized instruments. Red cell
indices include:
Red cell indices: MCV, MCH, 1. Mean Corpuscular Volume (MCV)
MCHC and RDW. •• MCV is indicative of average volume of the RBC and is expressed in femtoliters (fL).
•• It is used for classification and differential diagnosis of anemias.
•• Normal range: 82–98 fL.
Microcytic anemia
have MCV < 80 fL and PCV × 1000
macrocytic anemia have MCV = = 0.45 × 1000/5 = 90 fL
RBC count in millions
MCV> 100 fL.
2. Mean Corpuscular Hemoglobin (MCH) MCH < 26 pg is seen in

•• MCH indicates the amount of Hb (weight) per RBC and is expressed as picograms (1 pg microcytic anemia and
= 10-12 g). MCH > 33 pg is seen in
macrocytic anemia.
•• It is of limited value in differential diagnosis of anemias.
•• Normal range: 27–32 pg
MCH = Hb (in g/L)/RBC (in millions/μL) = 15 × 10/5 = 30 pg
3. Mean Corpuscular Hemoglobin Concentration (MCHC) MCHC<31 g/dL is seen in

•• MCHC denotes the average concentration of hemoglobin in the RBC taking volume into hypochromic RBC such
account. It is expressed as g/dL (earlier it was expressed as %). as IDA and thalassemia.
MCHC >36 g/dL is an
•• It is a better indicator of hypochromasia than MCH.
indication of hyperchromic
•• Normal range: 31–35 g/dL. RBCs.
MCHC = Hb (in g/dL)/PCV = 15/0.45 = 33 g/dL
4. Red Cell Distribution Width (RDW) RDW is useful for

•• RDW is a quantitative measure of anisocytosis. differentiating anemia
•• Normal RDW is 11.5% to 14.5%. due to iron deficiency and
thalassemia.
•• A normal RDW indicates that RBCs are relatively uniform in size. A raised RDW indicates
that red cells are heterogeneous in size and/or shape. In early iron deficiency anemia,
RDW increases along with low MCV while in thalassemia trait, RDW is normal with low
MCV.
RDW = (Standard deviation ÷ mean cell volume) × 100
IRON DEFICIENCY ANEMIA

Iron deficiency anemia (IDA) is the most common nutritional disorder.
Etiology (Table 1.3) Q. Discuss the etiopathogenesis

of iron deficiency anemia.
IDA is due to deficiency of iron causing defective heme synthesis.
TABLE 1.3: Causes of iron deficiency anemia

1. Dietary deficiency/lack Dietary deficiency is the
commonest cause of IDA.
• Milk-fed infants
• Elderly with improper diet and poor dentition
• Low socioeconomical sections In adult men and
• Vegetarians (contains poorly absorbable inorganic iron) postmenopausal women,
deficiency may be due to
2. Impaired absorption
chronic gastrointestinal
• Total/partial gastrectomy blood loss.
• Intestinal absorption is impaired in sprue, other causes of intestinal steatorrhea and chronic diarrhea
• Specific items in the diet, like phytates of cereals, tannates, carbonates, oxalates, phosphates and drugs
can impair iron absorption Iron is absorbed in the
3. Increased demand/requirement duodenum.
• Growing infants, children and adolescents
• Pregnancy and lactation
4. Chronic blood loss: due to bleeding from the Infants who consume large
amounts of cow's milk are
• Gastrointestinal tract (e.g. peptic ulcers, gastric carcinoma, colonic carcinoma, hemorrhoids, hookworm
susceptible to develop IDA.
infestation or nonsteroidal anti-inflammatory drugs)
• Urinary tract (e.g. renal or bladder tumors)
• Genital tract (e.g. menorrhagia, uterine cancer)
• Respiratory tract (e.g. hemoptysis)
Stages of IDA in Pathogenesis of Iron Deficiency Anemia

sequence: absent of
iron stores→decreased It is due to decreased synthesis of heme and can be divided into 3 stages.
serum ferritin→decreased •• Stage 1 (Iron depletion): iron adequate to maintain normal hemoglobin level and only
serum iron→increased serum ferritin decreased.
TIBC → decreased iron
saturation→ microcytic •• Stage 2 (Iron deficient erythropoiesis): lowering of serum iron and transferrin saturation
hypochromic anemia. levels without anemia (Hb, MCV and MCH within normal range). Bone marrow shows iron
deficient erythropoiesis.
•• Stage 3 (Iron deficiency anemia): low serum iron, serum ferritin and transferrin saturation.
Impaired hemoglobin production. Morphologically, first reduction in the size (microcytic)
and later increase in the central pallor (hypochromia) of RBCs.
Q. Discuss the laboratory findings Laboratory Findings

in iron deficiency anemia.
Peripheral Blood
MCV, MCH and MCHC are •• Hemoglobin and hematocrit (PCV): decreased
reduced. RDW is raised. •• Red cell indices:
–– MCV: <80 fL (normal 82–98 fL)
–– MCH: <25 pg (normal 27–32 pg)
–– MCHC: <27 g/dL(31–36 g/dL)
–– RDW: increased and >15%. It is earliest sign of iron deficiency (normal 11.5–14.5%).
Q. Write short notes on peripheral •• Peripheral smear (Figs 1.1 and 1.2):
smear findings in iron deficiency –– RBCs: microcytic (small) and hypochromic (pale). Severe anemia shows ring/pessary cells.
anemia. Moderate anisocytosis and poikilocytosis pencil/cigar-shaped cells.
–– WBCs: normal; eosinophilia in hookworm infestation.
Peripheral smear shows –– Platelets: normal
microcytic hypochromic
RBCs. •• Reticulocyte count: low for the degree of anemia.
Fig. 1.1: Peripheral blood smear showing microcytic Fig. 1.2: Diagrammatic appearance of peripheral blood smear
hypochromic red blood cells with microcytic hypochromic red blood cells
Bone Marrow Bone marrow shows

micronormoblastic
•• Cellularity: moderately hypercellular. eythroid hyperplasia.
•• M:E ratio: varies from 2:1 to 1:2 (normal 2:1 to 4:1). Marrow iron is absent.
•• Erythropoiesis: hyperplasia and micronormoblastic maturation. Prussian blue reaction
negative.
•• Myelopoiesis: normal.
•• Megakaryopoiesis: normal.
•• Absence of bone marrow iron: “Gold standard” test, demonstrated by negative Prussian blue reaction.
Serum Iron Profile (Table 1.4) Reduced: serum iron,

ferritin, % transferrin
saturation.
TABLE 1.4: Serum iron profile in IDA Increased: TIBC, TFR and
red cell protoporphyrin.
Normal range Value in IDA Observation
Serum ferritin 15–300 µg/L <15 µg/L
Serum iron 50–150 µg/dL 10–15 µg/dL
Serum transferrin saturation 30–40% <15%
Total plasma iron-binding capacity (TIBC) 310–340 µg/dL 350–450 µg/dL
Serum transferrin receptor (TFR) 0.57–2.8 µg/L 3.5–7.1 µg/L
Red cell protoporphyrin 30–50 µg/dL >200 µg/dL
Reticulocyte Hemoglobin The earliest laboratory

indicator of IDA is reduced
It is decreased and is an early feature of IDA. reticulocyte hemoglobin.
Clinical Features of IDA Q. Mention the various clinical

features of iron deficiency anemia.
Nonspecific and related to both severity and the cause of the anemia (e.g. gastrointestinal
disease)
•• Onset: insidious.
•• Nonspecific symptoms: fatigue, palpitations, breathlessness, weakness and irritability.
•• Pharyngeal/esophageal webs formed cause dysphagia.
•• Patterson-Kelly or Plummer-Vinson syndrome: Patterson-Kelly or
–– Microcytic hypochromic anemia Plummer-Vinson
syndrome: microcytic
–– Atrophic glossitis
hypochromic anemia,
–– Esophageal webs atrophic glossitis and
•• Congestive heart failure in severe anemia. esophageal webs.
•• Central nervous system: pica-unusual craving for substances with no nutritional value
like clay or chalk. Craving for ice (pagophagia) specific to iron deficiency. Pica may be the
cause rather than effect of IDA.
Physical Findings Koilonychia (spoon nails)

is a physical finding seen
Diminished tissue enzymes cause characteristic epithelial changes of iron deficiency anemia. in iron deficiency. First
•• Angular stomatitis and glossitis fingernails become thin
•• Chronic atrophic gastritis and flat-platonychia, then
brittle and finally spoon
•• Koilonychia (spoon nails) shaped.
Q. Enumerate the causes of Causes of Microcytic Hypochromic Anemia

microcytic hypochromic anemia.
•• Iron deficiency anemia
•• Thalassemia major
•• Anemia of chronic disorders
•• Others: alcohol, lead poisoning and drugs
•• Sideroblastic anemia (rare cause).
Q. Discuss the causes and MEGALOBLASTIC ANEMIA

pathogenesis of megaloblastic
anemia. Anemias characterized by defective/impaired DNA synthesis and distinct megaloblasts in
the bone marrow. Megaloblastic anemias are common among anemias due to impaired red
cell production.
Vitamin B12 is present in
animal products.
Etiology of Megaloblastic Anemia (Table 1.5)

TABLE 1.5: Causes of megaloblastic anemia
VITAMIN B12 DEFICIENCY
1. Decreased Intake: inadequate diet, “pure vegetarians” (vegans)
Deficiency of vitamin B12 2. Impaired Absorption
and folic acid are the main • Gastric: deficiency of gastric acid or pepsin or intrinsic factor
causes of megaloblastic –– Pernicious anemia
anemia. –– Post-gastrectomy
• Intestinal
–– Loss of absorptive surface
◆◆ Malabsorption syndromes
◆◆ Diffuse intestinal disease, e.g. lymphoma, systemic sclerosis
◆◆ Ileal resection, Crohn disease
–– Bacterial or parasitic competition for vitamin B12
◆◆ Bacterial overgrowth in blind loops and diverticula of bowel
◆◆ Fish tapeworm infestation
3. Increased Demand: pregnancy, hyperthyroidism, disseminated cancer
FOLIC ACID DEFICIENCY
Folic acid is absorbed in 1. Decreased Intake: inadequate diet—alcoholism, malnutrition

the jejunum. 2. Impaired Absorption
• Malabsorption states: nontropical and tropical sprue
• Diffuse infiltrative diseases of the small intestine (e.g. lymphoma)
• Drugs: anticonvulsant phenytoin and oral contraceptives
3. Increased Loss: hemodialysis
4. Increased Demand: pregnancy, infancy, disseminated cancer, markedly increased hematopoiesis
5. Impaired Utilization: folic acid antagonists, such as methotrexate
Deficiency of vitamin B12

and folic acid → delayed
Pathogenesis of Megaloblastic Change
nuclear maturation → 1. Impaired DNA synthesis: megaloblastic anemia is commonly due to deficiency of
megaloblast → macrocyte. vitamin B12 (cyanocobalamin) or folic acid. Both are required for the synthesis of DNA.
a. Delayed maturation of nucleus. The nuclear maturation lags behind the cytoplas-
mic maturation and results in abnormally large nucleated erythroid precursors
named as megaloblasts.
b. Cytoplasm matures normally. RBCs are larger than normal → macrocytes.
Ineffective erythropoiesis
c. Affects all rapidly dividing cells of the body (including skin, GI tract, and bone marrow).
and hemolysis are
responsible for anemia. 2. Ineffective erythropoiesis: megaloblast precursors undergo intramedullary destruction.
Laboratory Findings of Megaloblastic Anemia Q. Write short note on

the laboratory findings in
Blood findings in vitamin B12 and/or folic acid deficiency are similar. megaloblastic anemia.
Peripheral Blood
•• Hemoglobin and hematocrit (PCV): reduced
•• Red cell indices
–– MCV: above 100 fL (normal 82–98 fL)
–– MCH (normal 27–32 pg)
–– Normal MCHC (31–36 g/dL)
•• Peripheral smear (Figs 1.3 and 1.4): pancytopenia (decreased RBC, WBCs and platelets). Megaloblastic anemia
–– RBCs: • Pancytopenia
• Macro-ovalocytes
◆◆ Macrocytic and oval (egg-shaped macro-ovalocytes)-diagnostic. • Hypersegmented
◆◆ Most macrocytes lack the central pallor (Figs 1.3 and 1.4). neutrophils
◆◆ Marked variation in the size and shape of red cells (anisopoikilocytosis). • Macropolys.
◆◆ Evidence of dyserythropoiesis: basophilic stippling, Cabot ring and Howell Jolly bodies.
–– WBCs:
◆◆ Decreased WBC count (leukopenia). In megaloblastic anemia
due to vitamin B12
◆◆ Hypersegmented neutrophils (more than five nuclear lobes): first and specific morphological deficiency, reticulocyte
sign of megaloblastic anemia. These neutrophils are also larger than normal (macropolys). count may be normal or
–– Platelets: decreased. low and high reticulocyte
count is seen on 7th day
following vitamin B12
•• Reticulocyte count: normal or low. therapy.
Dimorphic Anemia
•• Combined vitamin B12/folic acid and iron deficiency.
•• Peripheral smear shows two populations of RBCs namely: macro-ovalocytes and microcytic
hypochromic (Fig. 1.5).
Fig. 1.3: Peripheral blood smear showing macro-ovalocytes (arrows) and Fig. 1.4: Diagrammatic peripheral blood smear showing
hypersegmented neutrophil (inset ) macro-ovalocytes (thick arrows) and hypersegmented neu-
trophil (thin arrow )
A mixture of microcytic
hypochromic and
macrocytic RBCs is termed
as dimorphic picture and
occurs in mixed deficiency
of iron and folic acid or
vitamin B12.
Fig. 1.5: Diagrammatic peripheral blood smear of dimorphic

anemia showing macro-ovalocytes and microcytes
Megaloblastic anemia-
bone marrow: Bone Marrow
• Megaloblasts •• Cellularity: moderately to markedly hypercellular.
• Giant metamyelocytes. •• M: E ratio: due to marked erythroid hyperplasia, M: E ratio is reversed ranging from 1:1 to 1:6 (normal
2:1 to 4:1).
Megaloblast are large, •• Erythropoiesis: megaloblastic type (Figs 1.6 and 1.7)
abnormal precursors of –– Megaloblasts: large, abnormal counterparts of normal normoblasts. Megaloblast shows asyn-
RBCs seen in the bone chrony of nuclear and cytoplasmic maturation. The cytoplasm shows normal hemoglobinization.
marrow of patients with –– Ineffective erythropoiesis: developing megaloblasts die in marrow (intramedullary hemolysis).
megaloblastic anemia.
•• Myelopoiesis:
–– Myeloid cells adequate in number.
–– Granulocytic precursors display nuclear-cytoplasmic asynchrony in the form of giant metamyelo-
cytes and band forms.
•• Megakaryopoiesis: normal or increased in number.
•• Bone marrow iron: moderately increased.
The differences between normoblasts and megaloblasts are shown in Table 1.6
Q. List the differences between TABLE 1.6: Differences between normoblast and megaloblast
normoblast and megaloblast. Characteristics Normoblast Megaloblast
Cell size Normal Larger than corresponding normoblast
Nuclear chromatin Normal Open sieve-like
Megaloblasts: Nuclear maturation Normal Lags behind cytoplasmic maturation

• Nuclear maturation lags Mitosis Normal Increased and abnormal
behind cytoplasmic
maturation. Maturation in bone Normal (Late > Increased proportion of more primitive erythroid cells
• Nuclei have open sieve- marrow intermediate > early (Late < intermediate < early megaloblast)
like chromatin. normoblast)
Evidence of Absent Present (irregular nuclei, Howell Jolly bodies)
dyserythropoiesis
Myelopoiesis Normal Shows giant metamyelocytes
Found in Normal bone marrow Bone marrow of megaloblastic anemia
Fig. 1.6: Bone marrow aspirate showing megaloblastic precursors Fig. 1.7: Diagrammatic picture of bone marrow aspirate showing
(arrows) in varying stages of maturation (inset shows early megalo- megaloblastic precursors (thick arrows) in varying stages of maturation
blast)
Biochemical Tests for Megaloblastic Anemia

Common for both vitamin B12 and folic acid deficiency Deoxyuridine suppression
Deoxyuridine suppression test: it is a sensitive measure of deficiency of 5, 10-methylene THF, test is abnormal even
which occurs in both folic acid and vitamin B12 deficiency. before the morphological
changes.
•• Serum homocysteine
•• Serum bilirubin: mild increase causes mild jaundice
•• Serum iron and ferritin
•• Plasma lactate dehydrogenase (LDH)
•• Serum vitamin B12/folate decreased.
Diagnostic tests for vitamin B12 deficiency

•• Serum vitamin B12 levels: decreased
–– Serum methylmalonic acid
–– Urinary excretion of methylmalonic acid
•• Schilling test for vitamin B12 absorption (Refer page 12). Schilling test determines
the cause of vitamin B12
deficiency.
Specific tests for folic acid deficiency
•• Serum folic acid levels: decreased
•• FIGLU in urine: excessively excreted.
PERNICIOUS ANEMIA Q. Discuss the etiopathogenesis

and morphology of pernicious
Pernicious anemia (PA) is an autoimmune disease due to deficiency of intrinsic factor causing anemia.
impaired absorption of vitamin B12 and megaloblastic anemia.
Rare in India. A genetic predisposition is suspected.
Age: older age—fifth to eighth decades of life Vitamin B12 is absorbed
in terminal ileum and
Sex: females are more involved than males (F: M is 1.5: 1). requires IF.
PA: autoimmune disease Etiopathogenesis

• Atrophic gastritis
• IF deficiency •• An autoimmune disease due to destruction of gastric mucosa.
• Autoantibodies. •• Stomach shows damage to parietal cells, dense infiltration by lymphocytes and plasma
cells → chronic atrophic gastritis → failure of production of intrinsic factor.
•• Presence of autoantibodies: two major types of autoantibodies—
–– Anti-intrinsic factor (IF) antibody
◆◆ Type I (blocking) antibody: blocks the binding of vitamin B12 to IF. Present in 50–75%
of the cases.
◆◆ Type II (binding) antibody: attaches to the IF–vitamin B12 complex and prevent its
binding to receptors in the ileum. Present in about 40% of patients.
–– Parietal cell (Type III) antibody: neither specific for PA nor other autoimmune disorders.
It is found in 90% of patients.
Morphology
Alimentary System
Atrophic gastritis may •• Atrophic glossitis: tongue shiny, glazed and beefy.
predispose to carcinoma
stomach. •• Stomach:
–– Diffuse chronic atrophic gastritis and impaired secretion of hydrochloric acid, pepsin
and intrinsic factor.
◆◆ Histologically atrophy of the glands, with loss of both chief cells and parietal cells.
◆◆ Nuclei of mucosal cells look similar to that of megaloblasts.
◆◆ Dense infiltration by lymphocytes and plasma cells.
–– Intestinal metaplasia.
Central Nervous System

Found in 75% of cases.
•• Demyelination in the dorsal and lateral tracts: subacute combined degeneration
•• Peripheral neuropathy.
Q. Write short note on laboratory Laboratory Findings (Fig. 1.8)

findings in pernicious anemia.
Blood, bone marrow and biochemical test findings are similar to those described earlier for
megaloblastic anemias (Refer page 9 to 11).
Specific Diagnostic Tests for Pernicious Anemia

Schilling test: diagnostic •• Schilling test for vitamin B12 absorption: abnormal
of PA but now very –– Radioactive vitamin B12 is used to assess the status of intrinsic factor (IF) and vitamin B12.
infrequently performed.
–– Helps in distinguishing megaloblastic anemia due to IF deficiency (pernicious anemia)
from other causes of vitamin B12 deficiency.
•• Serum antibodies to intrinsic factor are highly specific for pernicious anemia
•• Achlorhydria with histamine/pentagastrin stimulation.
•• Severe deficiency of intrinsic factor.
Pernicious anemia
present with features of
megaloblastic anemia due
to vitamin B12 deficiency.
In addition, it may show
features of atrophic
gastritis and achlorhydria.
PA patients sometimes
have a lemon-yellow color
owing to a combination
of pallor and mild
jaundice caused by excess
breakdown of hemoglobin.
Nonmegaloblastic causes
of macrocytic anemia:
1. Alcohol
2. Liver disease
3. Myxedema
4. Cytotoxic drugs
5. Myeloma
6. Aplastic anemia
7. Reticulocytosis
8. Red cell aplasia.
Fig. 1.8: Clinical features and laboratory findings in pernicious anemia
Clinical Features of Megaloblastic Anemia Q. Mention the various clinical

features of megaloblastic anemia.
The clinical features of vitamin B12 deficiency anemia and pernicious anemia are:
•• Onset: insidious and progresses slowly.
•• Classic triad of presentation: weakness, sore throat and paresthesias.
•• Tongue: painful red “beefy” tongue.
•• Neurological manifestations:
Folate deficiency anemia
–– Bilateral peripheral neuropathy: glove and sock distribution of numbness or paresthesia presents with features of
–– Demyelination of spinal cord: subacute combined demyelination/degeneration megaloblastic anemia due
of dorsal and lateral tracts—ataxia, uncoordinated gait, impairment of vibration and to vitamin B12. Unlike with
position sense. vitamin B12 deficiency,
neurological symptoms
•• Atherosclerosis: serum homocysteine level is raised and is a risk factor for atherosclerosis does not occur.
and thrombosis.
APLASTIC ANEMIA Q. Write short notes on aplastic

anemia.
Hematopoietic stem cell (HSC) disorder characterized by:
•• Pancytopenia (anemia, neutropenia and thrombocytopenia)
•• With markedly hypocellular bone marrow (less than 30% cellularity).
Etiology
The most common causes associated with aplastic anemia are shown in Table 1.7.
6 “I” s of the causes of TABLE 1.7: Common causes of aplastic anemia

aplastic anemia:
1. Idiopathic 1. ACQUIRED
2. Ingestion of drugs and
Idiopathic
chemicals
• Acquired defects in stem cell
3. Idiosyncratic
• Immune mediated
4. Irradiation
5. Infections and Secondary
6. Inherited. Chemical Agents
• Cytotoxic drugs: alkylating agents, antimetabolites • Benzene
• Inorganic arsenicals • Chloramphenicol
Idiosyncratic
• Chloramphenicol • Phenylbutazone
• Penicillamine • Carbamazepine
• Gold salts • Organic arsenicals
• Methylphenylethyl hydantoin
Physical Agents: whole-body irradiation
Viral Infections: hepatitis virus, Epstein-Barr virus, cytomegalovirus , herpes zoster (Varicella zoster) , HIV
2. INHERITED: fanconi anemia, telomerase defects
Pathogenesis: Pathogenesis (Fig. 1.9)

• Direct damage to the
hematopoietic stem
cells and progenitor
cells.
• Immune-mediated
destruction.
• Primary stem cell
abnormality—inherited
defect in the stem cells.
Fig. 1.9: Pathogenesis of aplastic anemia
Clinical Features
•• Any age of both sexes
•• Insidious
–– Progressive weakness, pallor and dyspnea due to anemia
–– Frequent (mucocutaneous bacterial infections) or fatal infections due to neutropenia
–– Bleeding manifestations in the form of petechiae, bruises and ecchymoses due to

thrombocytopenia.
Laboratory Findings
Peripheral Blood Reticulocyte count
is markedly low in
•• Hemoglobin aplastic anemia and is
•• PCV characteristic feature.
•• Reticulocyte count: markedly decreased.
•• Peripheral smear: pancytopenia, i.e. decreased red cells, neutrophils and platelets.
–– RBCs: normocytic normochromic anemia
–– WBCs: total leukocyte count decreased. Neutrophils markedly diminished and neutropenia is a
reflection of the severity of aplasia. Initial stages, lymphocytes normal in number as the disease
progresses their count decreases.
–– Platelets: count is decreased.
Bone marrow elements

Bone Marrow are replaced by fat and
•• Marrow aplasia—best appreciated in a bone marrow (trephine) biopsy aspiration usually yields
–– Cellularity: marked hypocellularity. dry tap.
–– Hematopoiesis: paucity of all erythroid, myeloid and megakaryocytic precursors.
–– Other cells: lymphocytes and plasma cells are prominent.
No Splenomegaly Absence of splenomegaly

and in its presence the
Diagnosis: diagnosis is made with peripheral blood and bone marrow biopsy findings. diagnosis of aplastic
anemia should not be
made.
Differential Diagnosis
•• Should be distinguished from other causes of pancytopenia (Table 1.8)
TABLE 1.8: Causes of pancytopenia

Decreased bone marrow function
•• Aplastic anemia
–– Idiopathic
–– Secondary
–– Inherited
•• Myelodysplastic syndromes
•• Bone marrow infiltration with
–– Leukemia
–– Lymphoma
–– Myeloma
–– Tumors (carcinoma)
–– Granulomatous diseases (e.g. tuberculosis, sarcoidosis)
•• Nutritional deficiencies:
–– Megaloblastic anemia (vitamin B12 and folic acid deficiency)
•• Paroxysmal nocturnal hemoglobinuria
•• Myelofibrosis (rare)
•• Hemophagocytic syndrome
Increased peripheral destruction
•• Hypersplenism
Prognosis: unpredictable.
16
2 Hemolytic Anemias Due to

Red Cell Membrane and
CHAPTER Enzyme Defects
Q. Define and classify hemolytic HEMOLYTIC ANEMIA

anemia.
Definition
Normal lifespan of red Hemolytic anemias are due to increase in the rate of red cell destruction (hemolysis).
cell is about 120 days.
In hemolytic anemias
RBC survival time is
considerably shortened. Classification of Hemolytic Anemias (Table 2.1)
Depending on:
•• Location of hemolysis: intravascular and extravascular
•• Source of defect causing hemolysis: intracorpuscular defect and extracorpuscular
defect
•• Mode of onset: hereditary and acquired disorders.
TABLE 2.1: Classification and causes of hemolytic anemia

Intrinsic (intracorpuscular) abnormalities Extrinsic (extracorpuscular) abnormalities
Breakdown of normal RBCs
occurs in the macrophages Hereditary Antibody-mediated
of the bone marrow, liver •• RBC membrane abnormalities •• Isohemagglutinis: Rh disease of the new-born,
and spleen. –– Membrane skeletal abnormalities: transfusion reactions
spherocytosis, elliptocytosis •• Autoantibodies: idiopathic (primary), drug-
–– Membrane lipids: abetalipoproteinemia associated, systemic lupus erythematosus
•• Enzyme deficiencies Mechanical trauma to RBCs
–– Enzymes of hexose monophosphate shunt: •• Microangiopathic hemolytic anemia: disseminated
glucose-6-phosphate dehydrogenase intravascular coagulation
Decreased red cell survival
–– Glycolytic enzymes: pyruvate kinase •• Defective cardiac valves
does not always cause
anemia as there is a •• Disorders of hemoglobin synthesis Infections: malaria
compensatory increase in –– Deficient globin synthesis: thalassemia Drugs, chemicals and toxins
red cell production by the syndromes •• Drugs: oxidant drugs, primaquine, dapsone, etc.
bone marrow. –– Structurally abnormal globin synthesis •• Chemicals: naphthalene, nitrites, nitrates, etc.
(hemoglobinopathies): sickle cell anemia •• Toxins: snake venom, lead poisoning, clostridial
Acquired sepsis
•• Membrane defects: paroxysmal nocturnal
hemoglobinuria
Hemolytic Anemias Due to Red Cell Membrane and Enzyme Defects CHAPTER 2 17
Location of Hemolysis Q. List the differences between

extravascular hemolysis and
It may be intravascular and/or extravascular. The differences between these two types are intravascular hemolysis.
listed in Table 2.2.
TABLE 2.2: Differences between extravascular and intravascular hemolysis In most hemolytic anemias
red cell destruction is
Characteristics Extravascular hemolysis Intravascular hemolysis extravascular.
Site of hemolysis RE system (spleen, bone marrow) Within circulation
Splenomegaly Usual Uncommon
Laboratory findings
•• Serum bilirubin-unconjugated Moderately raised Mildly raised
•• Serum haptoglobin Normal Decreased
•• Hemoglobinemia Not seen Positive
Urine
•• Hemoglobinuria Absent Present
•• Hemosiderinuria Absent Present
Examples Thalassemia, sickle cell anemia G6PD deficiency, PNH
HEREDITARY SPHEROCYTOSIS
Hereditary spherocytosis (HS) is a rare inherited hemolytic anemia resulting from the defect Q. Describe the etiopathogenesis
in the red cell membrane. of hereditary spherocytosis.
Normal structure of RBC membrane is depicted in Figure 2.1.
Etiopathogenesis HS, is due to defect in the

RBC membrane protein.
•• Autosomal dominant disorder The common mutations
•• RBC membrane protein defect caused by various mutations. Most common mutations involve ankyrin, band 3,
involve ankyrin, band 3, spectrin, or band protein 4.2. spectrin or band protein
4.2.
Mechanism of Hemolysis in HS (Fig. 2.2) HS: intrinsic defect of RBC

•• Young HS RBCs are normal in shape. But as they age, they undergo loss of membrane membrane-extravascular
hemolysis.
fragments in the circulation. These small RBCs assume a spherical shape (spherocytes).
•• Spherocytes are rigid, inflexible and less deformable. They get trapped in the spleen
leading to premature destruction of spherocytes.
Fig. 2.1: Structure of the red cell membrane

Fig. 2.2: Mechanism of hemolysis in hereditary spherocytosis
Q. Write short notes on laboratory Laboratory Findings

findings in HS.
Peripheral Blood
In hereditary spherocytosis •• Hemoglobin: decreased and level depends on degree of hemolysis.
MCHC is > 35 g/dL. •• Red cell indices:
–– MCV: reduced (normal 82–98 fL)
–– MCHC: raised and > 35 g/dL (normal 31–36 g/dL).
Spherocytes and •• Peripheral smear: very important for diagnosis (Figs 2.3 and 2.4).
reticulocytosis are –– RBCs:
observed in the peripheral
blood. ◆◆ Spherocytes are most distinctive but not pathognomonic. Spherocytes are small, dark-
staining (hyperchromic) RBCs without any central pallor.
◆◆ Polychromatophilia due to reticulocytosis.
Spherocytes may also
–– WBCs: total leukocyte count (TLC) increased.
be seen in autoimmune
hemolytic anemia and –– Platelets: normal.
burns.
• Reticulocyte count: increased (Fig. 2.5).
Bone marrow shows

erythroid hyperplasia.
Bone Marrow
•• Cellularity: markedly hypercellular
•• Erythropoiesis: erythroid hyperplasia
•• Myelopoiesis: normal
Fig. 2.3: Peripheral blood smear with numerous spherocytes (arrows) Fig. 2.4: Diagrammatic peripheral blood smear
with numerous spherocytes (arrows)
Fig. 2.5: Smear shows reticulocyte with blue filamentous/granular

material (new methylene blue stain) (arrows)
Biochemical Findings
•• Serum bilirubin: mildly raised.
•• Urine urobilinogen: increased.
•• Serum haptoglobin: decreased.
Osmotic Fragility Test HS: osmotic fragility is

increased with a shift of
Osmotic fragility is increased and there is shift of the curve to the right (Fig. 2.6). curve to the right.
Clinical Features Clinical features of

•• Age: anytime from the neonatal period to adulthood. intermittent jaundice,
splenomegaly and
•• Family history: most (75%) are inherited as autosomal dominant trait. spherocytes in the
•• Anemia: mild to moderate. peripheral smear is highly
suggestive of HS.
Fig. 2.6: Osmotic fragility test. Normal curve (blue) and increased
osmotic fragility in hereditary spherocytosis
•• Jaundice: intermittent attacks, precipitated by pregnancy, fatigue, or infection.

•• Splenomegaly: moderate (500 to 1000 g).
•• Gallstones: pigment gallstones.
•• Aplastic crises: may be triggered by an acute parvovirus infection.
GLUCOSE-6-PHOSPHATE
DEHYDROGENASE DEFICIENCY
•• Hemolytic disease due to red cell enzyme defects.
•• In G6PD deficiency, RBCs are susceptible to oxidative injury by free radicals.
•• It is an X-linked recessive disorder and its full expression is seen only in males.
•• There are different subtypes.
G6PD deficiency is an Role of G6PD (Fig. 2.7)

intrinsic defect and
hemolysis is primarily • Reduced glutathione (GSH) in the normal RBCs protects them against oxidant injury by
intravascular. breakdown of compounds such as H2O2 to H2O. The housekeeping enzyme, G6PD is
required for normal GSH.
In G6PD, RBCs exposed Sequence of Events in G6PD Deficiency

to oxidant stress, the
hemoglobin is oxidized In G6PD deficiency, oxidants can cause both intravascular and extravascular hemolysis.
to methemoglobin which •• In G6PD deficiency, there is decreased synthesis of reduced glutathione.
forms Heinz bodies in the
•• RBCs when exposed to oxidant stress (during infections, exposure to drugs or chemical,
cytoplasm of RBCs.
fava beans) accumulate H2O2. It damages RBC membrane causing hemolysis.
•• Hemolyzed red cells liberate hemoglobin.
•• The hemoglobin is oxidized by oxidants leading to formation of methemoglobin, which
forms Heinz bodies (Fig. 2.8) in the cytoplasm of RBCs.
Fig. 2.7: Role of G6PD against injury by oxidants

Fig. 2.8: Peripheral blood smear in G6PD deficiency with “bite cells”
(arrows). Inset shows Heinz bodies (supravital stain)
•• Heinz bodies removed from RBC membrane by macrophages in the spleen and produce G6PD deficiency has a
bite cells. These bite cells are removed via erythrophagocytosis in the spleen. protective effect against
Plasmodium falciparum
malaria.
Clinical Presentation
G6PD deficiency manifests in several distinct clinical patterns. Usually present as acute self-
limited acute intravascular hemolytic anemia following exposure to oxidative stress.
Laboratory Findings
Peripheral Blood
•• Hemoglobin: decreased.
•• Reticulocyte count: increased.
G6PD deficiency–oxidant
damage to RBC
•• Peripheral smear: • Bite cells
–– RBCs: moderate anisopoikilocytosis with polychromatophilia, microspherocytes and bite cells • Heinz bodies.
(Fig. 2.8). Heinz bodies identified with a supravital stain and are best seen during active hemolysis.
–– WBCs: mild leukocytosis.
–– Platelets: normal.
•• Self-limited hemolysis: primarily the old red cells are hemolyzed, hence hemolysis is self-limited.
Urine
G6PD: enzyme analysis–
Hemoglobinuria will be found during hemolysis and may last for about 1–6 days. confirmatory test.
RBC Enzyme Analysis

Tests for G6PD deficiency are positive and should be assessed a few weeks after the acute
hemolytic episode.
22
3 Thalassemia Syndrome
CHAPTER
Q. Classify hereditary disorders of CLASSIFICATION OF HEREDITARY

hemoglobin.
DEFECTS IN HEMOGLOBIN
The term Hemoglobin defects may be quantitative (reduced production of normal hemoglobin) or
hemoglobinopathy qualitative (production of abnormal hemoglobin).
is usually used for a •• Quantitative defect: genetic mutations in the globin loci (e.g. thalassemia) may quan-
qualitative hereditary
disorder of hemoglobin. titatively reduce the synthesis of a-globin or b-globin chain. It leads to net reduction of
hemoglobin.
•• Qualitative defect: genetic mutations in the a-globin or b-globin locus may produce
abnormal hemoglobin (e.g. sickle cell anemia). The abnormal hemoglobin may be func-
tionally normal, but its physical or physiologic properties differ from normal hemoglobin.
Q. Classify thalassemia

syndromes.
THALASSEMIA SYNDROME
•• These are group of inherited disorders due to abnormality of globin production.
•• It is characterized by decreased or absence of synthesis of either a or b-globin chain of
adult hemoglobin, HbA (a2b2).
In β-Thalassemia, there Classification

is decreased/absence of
synthesis of β-chains. They are mainly classified as:
•• b-Thalassemia syndromes: impaired synthesis of b-chains of globin.
In α-Thalassemia, there
is reduced/absence of
•• a-Thalassemia syndromes: impaired synthesis of a-chains of globin.
synthesis of α-chains of •• Miscellaneous thalassemia syndromes.
globin.
b-THALASSEMIA
•• Autosomal recessive hereditary disorder
•• Diminished synthesis of b-globin chains and normal synthesis of a-chains.
Thalassemia Syndrome CHAPTER 3 23
Molecular Pathology Point mutations leading

to aberrant RNA splicing is
•• b-globin chains are encoded by a single gene. the most common cause of
•• The molecular errors over 200 genetic defects leading to b-thalassemia have been identified. β-thalassemia.
•• Different types of mutations in b-globin gene can occur but mainly point mutations
rather than gene deletions (unlike in a-thalassemia). The mutations result in defects in
transcription, RNA splicing and modification, translation via frame shifts and nonsense
codons. Mutations leading to aberrant RNA splicing are the most common cause.
Clinical and Genetic Classification (Table 3.1)

TABLE 3.1: Clinical and genetic classification of b-thalassemias • β0 = Total absence of
β-globin synthesis;
Clinical syndromes Genotype Clinical features
• β+ = Markedly reduced
b-thalassemia major Homozygous (b0/b0,b+/b+) Severe form, severe anemia or diminished β-globin
or double heterozygous ( b0/b+) and transfusion dependent synthesis;
High level of HbF in the blood • β = normal β-globin
synthesis.
b-thalassemia intermedia Variable (b /b , b /b , b /b, b /b)
0 + + + 0 +
Moderately severe and not
transfusion dependent
b-thalassemia minor/b-thalassemia trait Heterozygous (b0/b, b+/b) Mild anemia and asymptomatic
b-THALASSEMIA MAJOR β-thalassemia is the

commonest quantitative
•• It is a hereditary hemolytic anemia due to absence of synthesis of b-globin chain of disorder of hemoglobin.
hemoglobin. The synthesis of a-globin chain is not affected.
•• Homozygous form of b0/b0 or b+/b+ or double heterozygous b0/b+ (Table 3.1) β-thalassemia major also
•• Most common in Mediterranean countries, parts of Africa and South East Asia. called Mediterranean or
Cooley’s anemia.
•• Hemolytic anemia is of severe degree.
Pathophysiology of b-thalassemia Major (Fig. 3.1) Q. Describe the pathophysiology/

pathogenesis of β-thalassemia
Consequence of Defective or Absent b-chains major.
•• Severe hemolytic anemia due to:
1. Absence of b-globin chain: results in absence of synthesis of HbA (a2b2). This produces β-thalassemia major
RBCs that are poorly hemoglobinized (hypochromic) and small in size (microcytic). • Absence of synthesis of
2. Ineffective erythropoiesis: unpaired and excess a-chains aggregate into insoluble HbA produces severe
precipitates, which bind to and damage the membrane of erythroid precursors. These anemia
erythroid precursors fail to mature and undergo apoptosis in the marrow. • Increased synthesis of
3. Extravascular hemolysis: RBCs with a-chain inclusions are removed by macrophages of HbF.
spleen (extravascular hemolysis).
•• Synthesis of fetal hemoglobin (HbF): the ϒ-globin chain synthesis continues even 6 months
after birth and combines with a-globin leading to increased levels of HbF (a2ϒ2). The level
of HbF varies from 30% to 90%.
Consequences of Ineffective Erythropoiesis β-thalassemia major

• Thalassemic facies
•• Changes in bone marrow: marked erythroid hyperplasia.
• Crew cut appearance on
•• Changes in bone: skull x-ray
–– Skull X-ray: hair on end (“crew-cut”) appearance (Fig. 3.2) • Splenomegaly.
–– Typical facies: thalassemic facies (Fig. 3.3)—prominent forehead, cheekbones and
upper jaw.
Fig. 3.1: Pathogenesis of β-thalassemia major and its consequence
•• Extramedullary hematopoiesis: in liver and spleen → consequent hepatosplenomegaly.

•• Cachexia: develops in untreated patients.
β-thalassemia major Iron Overload and its Consequences

• Iron overload damgaes
parenchymal organs
•• Causes of iron overload:
due to hemosiderosis 1. Increased absorption of dietary iron from duodenum
and secondary 2. Hemolysis
hemochromatosis.
3. Repeated transfusions (usual mode of treatment).
•• Consequences: iron overload produces hemosiderosis and secondary hemochromatosis
and damages to parenchyma of organs (e.g. heart, liver and pancreas).
Failure to thrive, retarded Clinical Features

growth, monogoloid face,
and hepatosplenomegaly •• Age: infants develop moderate to severe anemia 6–9 months after birth.
are clinical features of •• Growth and development: untreated/untransfused children fail to thrive and die within
β-thalassemia major. 4–5 years of age.
•• Bone changes: those who survive longer develop distortion of skull and facial bones. X-ray
skull shows hair on end appearance (Fig. 3.2) and face shows a characteristic thalassemic
facies (Fig. 3.3).
•• Marked splenomegaly: up to 1500 grams due to hyperplasia and extramedullary
hematopoiesis.
•• Extramedullary hemopoiesis: liver and lymph nodes may show extramedullary
hematopoiesis.
Fig. 3.2: X-ray appearance of skull in β-thalassemia showing hair-on- Fig. 3.3: Appearance of typical thalassemic facies
end appearance (Courtesy: Dr Nuthan Kamath) (Courtesy: Dr Nuthan Kamath)
•• Iron overload: multiple blood transfusions may lead to iron overload and result in
hemosiderosis and secondary hemochromatosis (heart, liver and pancreas).
Laboratory Findings Q. Mention the laboratory

findings in b-thalassemia major.
Peripheral Blood
•• Hemoglobin (ranges from 3 to 8 g/dL) and hematocrit (ranges from 8 to 23%): markedly
reduced
•• RBC count increased/normal (in contrast to iron deficiency anemia).
•• Reticulocyte count increased and in the range of 5 to 15%.
•• Red cell indices:
–– MCV decreased and in the range of 45–70 fL (normal range 82–98 fL).
–– MCHC decreased and in the range of 22–30 g/dL (normal range 31–35 g/dL). RDW normal
–– MCH decreased and in the range of 20–28 pg (normal range 27–32 pg). MCV, MCH and MCHC
–– RDW-within normal limits (in contrast to iron deficiency anemia where it is increased). decreased.
•• Peripheral smear: Q. Write short note on peripheral

–– RBCs: smear findings in b-thalassemia
◆◆ Microcytic hypochromic anemia major.
◆◆ Moderate to marked anisocytosis and poikilocytosis
◆◆ Many target cells (Figs 3.4 and 3.5) β- thalassemia major: RDW
◆◆ Basophilic stippling normal. The peripheral
blood smear shows
◆◆ Nucleated red cell precursors (normoblasts) in variable numbers (5–40%).
–– WBCs: leukocytosis with mild left shift. anemia, target cells and
–– Platelets: normal. anisopoikilocytosis.
Bone marrow in β-
Bone Marrow thalassemia major shows
•• Cellularity: markedly hypercellular. marked normoblastic
•• M: E ratio: reversed to 1:1 to 1:5 depending upon the degree of erythroid hyperplasia. erythroid hyperplasia.
•• Erythropoiesis: normoblastic with marked erythroid hyperplasia. Marrow iron is markedly
increased.
•• Bone marrow iron: markedly increased due to increased dietary absorption and hemolysis.
Fig. 3.4: Peripheral blood smear in β-thalassemia showing target Fig. 3.5: Diagrammatic appearance of peripheral blood smear in β-thalassemia
cells (arrows) showing target cells (short arrows) and nucleated red cells (long arrows)
•• Bilirubin: increased—mainly of unconjugated type.
•• Urine urobilinogen: increased
•• Serum haptoglobin: markedly reduced.
•• Serum iron status:
–– Serum iron, serum ferritin and transferrin saturation are markedly increased
–– Total iron binding capacity (TIBC): reduced.
Reduced/absence of Special Tests

synthesis of β-chains; the
•• Fetal hemoglobin (HbF): increased to 30% to 90% (normal range 0 – 1%).
excess α-chains combine
with γ-chains leading to •• Hemoglobin electrophoresis (Table 3.2):
increased HbF. –– b+ thalassemia (b+/b+ or b0/b+ genotypes): demonstrates bands of both HbA and HbF.
–– bo thalassemia (b0/b0 genotype): since no b-chains are formed, there is no HbA. Major
hemoglobin is HbF with normal or low HbA2.
•• High performance chromatography (HPLC): HbF is increased (30–90%). HPLC measures
various fractions of hemoglobin (Hb) and is used for confirmation of diagnosis.
•• Prenatal diagnosis by molecular analysis of DNA.
•• Estimation of globin chains: normally a: b ratio is 1:1. Lack of b chain alter this ratio to
5–30:1
Note: normal adult cell TABLE 3.2: Hemoglobin F and A2 percentage in thalassemia syndromes
contains 96% HbA (α2β2),
Type HbF HbA2
3% HbA22(α2d2) and 1%
HbF(α2γ2). β -Thalassemia major (homozygous) 30–90% < 3.5%
β -Thalassemia intermedia (double heterozygous) 10–30% < 3.5%
β -Thalassemia minor/trait (heterozygous) 0–5% 3.6–8%
Differences between Iron Deficiency Anemia and

b-Thalassemia Major (Table 3.3)
TABLE 3.3: Differences between iron deficiency anemia and b-thalassemia major
Character Iron deficiency anemia b-thalassemia major β-thalassemia major
should be differentiated
Etiology Deficiency of iron Reduced synthesis of β chain
from iron deficiency
Laboratory findings anemia. Treatment with
iron in β-thalassemia major
•• RBC count Decreased (< 5 million/cu mm) Increased (> 5 million/cu mm)
worsens the iron load and
•• Peripheral smear its consequences.
–– Type of RBCs Microcytic hypochromic Microcytic hypochromic
–– Anisopoikilocytosis Mild to moderate Severe
–– Target cells Absent Present
•• Bone marrow iron Absent Markedly increased
•• Serum iron profile
–– Serum ferritin Reduced < 15 µg/L Increased (300 – 1000 µg/L)
–– Serum iron Reduced Increased
–– TIBC Increased Normal
•• Fetal hemoglobin (HbF) Normal (0–1%) Markedly increased (30–90%)
•• RDW Increased Normal
Clinical features
•• Age Any age Presented < 2 years of age β-thalassemia intermedia:
it is a clinical entity
•• Growth and development Normal Retarded
intermediate between
•• Hepatosplenomegaly Absent Present thalassemia trait and
thalassemia major.
X-ray findings Nil Hair on end appearance
Abbreviations: RDW, red cell distribution width; TIBC, total iron-binding capacity.
b-THALASSEMIA MINOR/TRAIT
•• More common than b-thalassemia major.
•• Most patients are heterozygous for thalassemic gene.
•• Usually asymptomatic and anemia is mild.
Laboratory Findings in b-Thalassemia Minor

•• Peripheral blood: microcytosis, hypochromia, basophilic stippling and target cells.
•• Bone marrow: mild erythroid hyperplasia.
•• Hemoglobin electrophoresis: increase in HbA2 (a2δ2) to 4 to 8% of the total hemoglobin
(normal 2.5 ± 0.3%). HbF levels may be normal or slightly increased.
•• NESTROF test (Naked eye single tube red cell osmotic fragility test): positive. NESTROF test positive
because the microcytic
–– In this test, 0.02 mL of patient’s blood is added to 5 mL of 0.35% saline in a test tube.
hypochromic RBCs of
–– After half an hour white paper with a dark black line is held behind the tube. β-thalassemia minor are
–– The microcytic hypochromic RBCs of thalassemia minor are resistant to lysis than resistant to lysis than
normocytic normochromic RBCs. normocytic normochromic
RBCs.
–– Hence, the black line on the paper is not clearly visible through the test tube compared
to normal cells.
•• Estimation of HbA2: HPLC is used for accurate estimation. HbA2 estimation is diagnostic
and level ranges from 4% to 8%.
β-thalassemia trait should TABLE 3.4: Differences between iron deficiency anemia and b-thalassemia minor/trait
be differentiated from iron
deficiency (Table 3.4). Character Iron deficiency anemia b-thalassemia minor
Etiology Deficiency of iron Reduced synthesis of b chain
Laboratory findings
•• Peripheral smear - RBCs Microcytic hypochromic Microcytic hypochromic
•• Serum iron profile
–– Serum ferritin Reduced < 15 µg/L Normal /slightly incresaed
–– Serum iron Reduced Normal
–– TIBC Increased Normal
•• HbA2 level Normal or decreased (2.5 + 0.3 %) Increased (4–8 %)
•• RBC count < 5 million/cu mm >5 million/cumm
•• RDW Increased Normal
α-Thalassemia:
anemia due to— a-THALASSEMIA
• Lack of adequate •• Inherited disorders characterized by reduced or absent synthesis of a-globin chains.
hemoglobin
• Effect of excess
•• Autosomal recessive disorder.
unpaired non-α-chains
(β, γ, δ).
Molecular Pathology
In contrast to a single gene coding b-globin chain, each a-globin chain are encoded by two
genes. Deletion of a-gene is the most common cause of reduced a-chain synthesis.
α-thalassemia is one of Clinical Syndromes

the cause of non-immune
hydrops fetalis. Four genes control a-chain synthesis. Severity of a-thalassemia varies greatly depending on
the number of a-globin genes deleted (Table 3.5). Each of the four a-globin genes normally
contributes 25% of the total a-globin chains.
Immune hydrops fetalis TABLE 3.5: Clinical syndromes associated with a-thalassemia disorders
is a hemolytic disease
caused by blood group Clinical syndrome No. of Clinicopathological features
incompatibility between a-globin
mother and fetus. deleted
Silent carrier state 1 Asymptomatic
a-Thalassemia trait 2 Usually asymptomatic. Normal hemoglobin level or minimal anemia
Hemoglobin H disease 3 Moderate microcytic hypochromic anemia
Hydrops fetalis (Hb Barts) 4 Severe form, fatal and usually results in intrauterine death
Sickle Cell Disease 4
CHAPTER
SICKLE CELL DISEASE Sickle cell diseases are

hemoglobinopathies
Definition characterized by
qualitative defect in
Sickle cell disease (SCD) is a group of hereditary disorders of hemoglobin characterized by hemoglobin synthesis.
production of defective hemoglobin called sickle hemoglobin (HbS). On low oxygen tension
or deoxygenation, HbS imparts sickle shape to RBCs. HbS is produced due to qualitative
defect in hemoglobin production caused by mutation in β-globin gene.
Classification of Sickle Cell Disease (Table 4.1) Sickle cell anemia is a

homozygous state in
which both β-globin
TABLE 4.1: Classification of sickle cell disease chains are abnormal.
Sickle cell anemia (SS) Sickle cell trait (AS)
•• Homozygous state—both the β-globin chains are •• Heterozygous state—one gene is defective Sickle cell trait: one
β-globin chain is abnormal
abnormal/defective (for HbS) and while the other gene is
and other β-globin chain is
normal (for HbA)
normal.
Other sickling syndromes (Compound heterozygous)
•• If both the β-globin chains have different abnormalities, (e.g. Hb SC, Hb S-β-thalassemia)—termed as
compound heterozygous
SICKLE CELL ANEMIA Sickle cell anemia:

autosomal recessive
Characteristic Features disorder with extravascular
hemolysis.
•• Autosomal recessive disorder manifests early in life.
•• Homozygous state (SS) caused by a mutation in the β-globin gene.
•• HbS constitutes more than 70% of hemoglobin in their RBCs with no HbA. HbS provides protection
against falciparum malaria.
Etiopathogenesis Q. Discuss the etiopathogenesis

of sickle cell anemia.
•• Production of abnormal hemoglobin called sickle hemoglobin (HbS).
Fig. 4.1: Replacement of glutamic acid with valine in the sixth position of b-globin
Replacement of the •• Missense point mutation: in HbS, there is substitution of glutamic acid by valine in the
glutamic acid residue by 6th position the β-globin chain of hemoglobin (Fig. 4.1). It alters the solubility or stability
valine in 6th position of of the hemoglobin and produces hemolytic anemia.
β-globin chain.
•• HbS is responsible for the characteristics of the disease.
Molecular Basis of Sickling (Fig. 4.2)

•• During low O2 tension or deoxygenation, HbS molecules undergo aggregation and
polymerization.
During low oxygen tension

or deoxygenation RBCs
assume sickle shape and
predisposes to vessel
occlusion.
RBCs in sickle cell anemia

have shorter lifespan and
causes hemolytic anemia.
Fig. 4.2: Pathogenesis of sickle cell anemia

Sickle Cell Disease CHAPTER 4 31
•• If deoxygenation continues, the aggregated HbS molecules form long needle-like fibers
(or pseudocrystalline structures known as tactoids) within RBCs.
•• The tactoids grow in length beyond the diameter of RBCs and distort RBC shape.
•• RBC become elongated and assumes a shape like sickle (or crescent moon or holly-leaf or
boat) and predisposes to stasis and vascular occlusion.
•• When the oxygen tension returns to normal, the sickled red cell returns to normal
shape.
•• Recurrent sickling causes red cell membrane damage and these RBCs become irreversibly
sickled cells (ISC).
Factors Affecting Sickling (Table 4.2)
TABLE 4.2: Factors affecting sickling

Factors Favors sickling Hinders sickling
Type of other associated
hemoglobins - HbA
- HbF
HbC - In sickle cell anemia, HbF
hinders sickling.
Transit time in microvasculature Slowing of bloodstream -
MCHC Increased MCHC Decreased MCHC
Intracellular pH Decreased pH -
°
Other factors Temperature above 37 C -
Infections -
Abbreviation: MCHC, mean corpuscular hemoglobin concentration.
Mechanism of Red Cell Damage With repeated sickling the

RBCs become irreversibly
•• HbS polymerization: when HbS polymerizes, it grows beyond the RBC membrane and sickled cells (ISC) and leads
project through it. to RBC membrane damage
•• Dehydration: repeated episodes of sickling leads to increased dehydration of RBCs. These and hemolysis.
RBCs become more rigid and nondefromable (irreversible sickled cells).
•• Percentage of ISC: degree of the hemolysis correlates with the percentage of irreversibly
sickled cells.
•• Impaired cation homeostasis: structural changes in the RBC membrane causes the influx
of Ca+ ions, which activate an ion channel resulting in the efflux of K+ and H2O.
Pathogenesis of the Microvascular Occlusions

Most serious clinical features are due to occlusion of microvasculature.
•• Deformability: sickle cells are rigid and tend to aggregate. The aggregated sickle cells block
the small blood vessels.
•• Factors that slow the blood flow: RBC cytoskeletal damage slow the movement of RBCs Most serious clinical
through microvascular beds. features of sickle cell
anemia are due to
•• Higher expression of adhesion molecules: sickle cells express higher levels of adhesion microvascular occlusion.
molecules and thus become abnormally sticky to the endothilium.
•• Inactivation of nitric oxide: lysed sickle cells liberate free hemoglobin, which binds and
inactivates nitric oxide (NO). This narrows the vessels and produces microvascular stasis
and sickling.
Clinical Features (Fig. 4.3)

•• Presence of HbF in the first 6 months of life has a protective role.
The cardinal clinical •• Symptoms appear after 6 months of age as the HbF disappears.
features are due to •• Infants and children present with acute problems like severe infection, acute chest
chronic hemolytic anemia,
crises (recurrent painful syndrome, splenic sequestration and stroke.
episodes), infections and •• Chronic hypoxia in children is responsible for generalized impairment of growth and
chronic organ damage. development. Adults manifest with chronic organ damage.
Chronic Hemolytic Anemia

•• Lifelong hemolysis (mainly extravascular) and causes chronic hemolytic anemia, which
is of moderate degree. This produces raised unconjugated (indirect) bilirubin, and
predisposes to pigment bilirubin gallstones (cholelithiasis) and cholecystitis.
Four crises encountered Crises

in sickle cell anemia:
sickling crisis, hemolytic Four types of crises are encountered. These are:
crisis, aplastic crisis and
sequestration crisis. 1. Sickling crisis (vaso-occlusive/pain/painful/infarctive crisis)
•• Most common
•• Blockage of microcirculation by sickled red cells produces hypoxic injury and infarction.
•• Bone: manifest as the hand-foot syndrome, dactylitis of the bones of the hands or feet or both.
Recurrent splenic
infarction due to •• Lung: acute chest syndrome (dangerous).
sickling crisis lead to •• Spleen: acute abdominal pain due to infarcts of abdominal viscera caused by occlusion
autosplenectomy. of vessels. Recurrent splenic infarction results in autosplenectomy.
Infants most commonly

present with dactylitis.
Most common cause of

death in adults is acute
chest syndrome.
Fig. 4.3: Various effects of vascular occlusion and hemolysis in sickle cell anemia
2. Hemolytic crisis
•• Rare type and presents with marked increase in hemolysis.
3. Aplastic crisis Reticulocytopenia is

•• Associated with parvovirus B19. seen in aplastic crisis
•• Reticulocytopenia. and reticulocytosis in
sequestration crisis.
4. Sequestration crisis
•• Usually occurs in children.
•• Sudden trapping of blood in spleen or liver causes rapid enlargement of the organ and
drop in hematocrit leading to hypovolemic shock.
Other crises encountered rarely are hypoplastic crisis and megaloblastic crisis (due to
inadequate folate).
Increased Susceptibility to Infections Susceptible to acute

•• Common infections are pneumonia due to Pneumococcus, meningitis due to S. infections with
encapsulated organisms.
pneumoniae and osteomyelitis due to Salmonella. Increased frequency of osteomyelitis is
due to bone infarcts, which act as a nidus for infection.
•• Septicemia and meningitis are the most common causes of death in children.
Causes of susceptibility to infections:
•• Hypofunction of spleen:
In children: due to congestion and poor blood flow. Common pathogens:
In adults: due to multiple infarcts and resultant autosplenectomy. S. pneumonia,
•• Defects in the alternative complement pathway. Salmonella and
Impairs opsonization of encapsulated bacteria such as pneumococci and Haemophilus influenzae. Pneumococcus.
Chronic Organ Damage SCA: severe hemolytic

anemia
Particularly seen in the spleen, bones, kidneys, heart, lungs, brain and skin. Sickling crisis
•• Spleen Autosplenectomy.
– Children after 6 months of life present with splenomegaly (up to 500 g).
– After 5–6 years of age, the spleen gets fibrosed and gradually reduces in the size due to
multiple infarcts.
– Gradual loss of splenic function secondary to infarcts results in autosplenectomy.
•• Bone: osteomyelitis, particularly with Salmonella typhimurium
•• Extremities: skin ulcers over the lower extremities
Laboratory Findings in Sickle Cell Anemia Q. Laboratory findings in sickle

cell anemia.
Peripheral Blood
•• Hematocrit (PCV): decreased. Sickle cell anemia: ESR is
•• ESR: reduced. reduced because sickle
•• Reticulocyte count: increased and range from 3% to 10%. cells do not form rouleaux.
•• Peripheral smear
–– RBCs:
◆◆ Normocytic normochromic to mildly hypochromic.
◆◆ Moderate to severe degree of anisopoikilocytosis.
◆◆ Characteristic cell is the sickle cell—appear as long, curved cells with pointed ends (Figs 4.4
and 4.5); may also show target cells (due to red cell dehydration) and ovalocytes.
◆◆ Polychromatophilia due to reticulocytosis. Peripheral smear shows
–– WBCs: mildly increased with shift to left. characteristic sickle cells
–– Platelets: mildly increased. number of which varies.
Fig. 4.4: Peripheral blood smear with sickle cells (arrows) Fig. 4.5: Diagrammatic peripheral blood smear with sickle cells (arrows)
In severe cases, skull

bone shows crew-
Bone Marrow
cut appearance in •• Cellularity: hypercellular.
roentgenograms. •• Erythropoiesis: compensatory normoblastic erythroid hyperplasia, which expands the marrow and
causes resorption of bone and secondary new bone formation.
•• Iron stores: usually increased.
Extramedullary Serum Findings

hematopoiesis can also
•• Serum bilirubin: raised and predisposes to pigment gallstones.
develop as a compensatory
mechanism. •• Iron status: raised serum iron, serum ferritin and transferrin saturation.
•• Serum haptoglobin: reduced.
•• Urine Urobilinogen: increased.
Diagnostic/Confirmatory Tests
•• Sickling test:
–– Sickling is induced by adding a reducing (oxygen-consuming) agent like 2% sodium
metabisulfite or sodium dithionite to blood sample.
–– Red cells with HbS show sickled (Fig. 4.6) and holly leaf appearance.
–– It is diagnostic of sickle cell anemia.
•• Hemoglobin electrophoresis: HbS is a slow moving compared to HbA and HbF.
Sickle cell anemia:HbS •• Estimation of HbF: in homozygous state constitutes about 10–30% of hemoglobin.
70–90%, HbF 10–30%, •• HPLC: useful for confirmation of diagnosis.
no HbA.
•• Prenatal diagnosis: by analysis of fetal DNA obtained by amniocentesis or chorionic villous
biopsy, to detect the point mutations.
SICKLE CELL TRAIT

Heterozygous state for the hemoglobin S mutation and shows both HbA and HbS (HbAS). One
defective gene (from one parent with HbS) and while the other gene is normal.
Sickling test is a diagnostic

test for sickle cell anemia.
Fig. 4.6: Sickling test. Sickled red cells (arrows) induced by reducing agent
(2% sodium metabisulfite)
Pathogenesis Sickle cell trait:

• Usually no anemia
In sickle cell trait, the hemoglobin A in RBCs prevents hemoglobin S polymerization. However, • No significant clinical
RBCs may sickle under extreme conditions (e.g. flight at high altitude in unpressurized aircraft, features
deep sea diving). • Amount of HbS varies
from 25% to 40%
• Hb A in RBCs prevents
polymerization of Hb S.
Clinical Features
Usually asymptomatic. Normal growth and development, lifespan and life expectancy.
Laboratory Findings
Peripheral Blood
•• Hemoglobin: normal or mildly decreased.
•• Peripheral smear:
–– RBCs: normocytic normochromic picture with very few target cells and mild degree of anisopoi-
kilocytosis.
–– WBCs: normal.
Bone Marrow
Hypercellular because of a compensatory normoblastic erythroid hyperplasia.
Diagnostic Tests In sickle cell trait: HbS

•• Hb electrophoresis: demonstrates two bands of HbS and HbA. 40–45% and HbA 55–60%.
•• Sickling test: sickling test is positive.
•• High-performance liquid chromatography (HPLC): useful for confirmation of diagnosis.
5 Other Anemias
CHAPTER
Immunohemolytic
anemias are characterized
IMMUNOHEMOLYTIC ANEMIAS
by the destruction of Anemias due to premature RBC destruction (hemolysis) mediated by antibodies that bind to
RBCs by either allo or auto RBCs. The antibodies may be either allo or auto type.
antibodies.
Immunohemolytic Classification of Immunohemolytic Anemias (Table 5.1)

anemias are mainly
classified as:
1. Alloimmune and TABLE 5.1: Classification of immunohemolytic anemias
2. Autoimmune hemolytic
Alloimmune hemolytic anemia
anemia.
•• Hemolytic disease of the newborn
•• Hemolytic transfusion reactions: mismatched blood transfusion
Autoimmune hemolytic anemia
•• Warm antibody type (IgG antibodies active at 37°C)
Hemolytic transfusion
reactions are due to ABO –– Primary (Idiopathic)
mismatch. The antibodies –– Secondary: autoimmune disorders (systemic lupus erythematosus), drugs, lymphomas
present in the recipient’s •• Cold agglutinin type (IgM antibodies active at 4°C–18°C)
serum coat donor’s RBCs –– Acute: mycoplasmal infection, infectious mononucleosis
and lead to intravascular –– Chronic: idiopathic, lymphomas
hemolysis. •• Cold hemolysin type (Donath-Landsteiner antibodies)
Alloimmune Hemolytic Anemia

•• Production of antibody against foreign antigen not present on individual’s red blood cell.
•• Allo-antibodies are present either in the serum or bound to red cells.
Q. Write short notes on hemolytic

disease of newborn.
HEMOLYTIC DISEASE OF THE NEWBORN
•• It is an allo-immune hemolytic anemia developing in the fetus and newborn baby.
•• Hemolysis is extravascular.
•• HDN develops when the IgG antibodies against blood group of fetus passes from mother to
fetus through the placenta.
Other Anemias CHAPTER 5 37
•• Occurs in two forms: HDN may be either due to

–– Rh incompatibility in which mother is Rh negative and fetus is Rh positive. The anti-D Rh or ABO incompatibility
antibodies are responsible for the hemolytic anemia. between mother and fetal
RBCs.
–– ABO incompatibility in which mother’s blood group is O and fetus is either of A or B
blood group. Either anti-A or anti-B antibodies cause hemolysis.
Rh Hemolytic Disease of the Newborn (Fig. 5.1)

Rh hemolytic disease of the newborn is more important than due to ABO incompatibility.
Pathogenesis
•• Occurs when mother is Rh (D antigen) negative and fetus is Rh positive. HDN usually does not
manifest during first
•• Sensitization occurs when fetal Rh positive RBCs enter into Rh negative mothers. Rh pregnancy. Sensitization
negative mother develops anti-Rh antibodies. develops during delivery
•• Sensitization occurs only at the time of delivery or during miscarriage. So, it does not or miscarriage.
manifest in the first pregnancy.
Rh HDN develops when

mother is Rh-ve and fetus
is Rh+ve.
Fig. 5.1: Pathogenesis of Rh hemolytic disease of the newborn

Hydrops fetalis is fatal •• In subsequent pregnancy, anti-Rh antibodies from mother cross placenta and coat the
condition, characterized Rh positive fetal red cells. These antibodies cause immune destruction of fetal red cells
by left and right-sided results in severe hemolytic anemia leading to jaundice of the newborn.
heart failure producing
•• Fetus may develop cardiac failure—hydrops fetalis (immune type).
generalized edema and
may result in death.
Clinicopathological Features
In Rh HDN, high levels of
•• Infants may have jaundice at birth.
unconjugated bilirubin can •• When the disease is severe, the levels of unconjugated bilirubin in the blood are high and
cross blood brain barrier bilirubin can pass the blood brain barrier.
causing kernicterus and
death of infant.
•• Bilirubin is deposited in the central nervous system (especially the basal ganglia) producing
neurological damage and is known as kernicterus (yellow coloration of cerebellum and
basal ganglia due to bilirubin deposition). It can cause death of the infant.
Prevention of Rh HDN: by the prophylactic removal of fetal cells entering the maternal
circulation before sensitization develops, by injecting anti-D into the Rh D negative mother.
Laboratory Findings
Peripheral blood
•• Reticulocyte count: increased.
Peripheral smear: •• Peripheral smear:

normocytic
–– RBCs: normocytic normochromic anemia with numerous nucleated RBCs, polychromatophils
normochromic anemia
with nucleated RBCs and and occasional spherocytes.
polychromatophils. –– WBCs: normal.
•• Antiglobulin test (Coombs test): antibodies in the mother and baby are detected by indirect
and direct Coombs test respectively (Fig. 5.2).
In direct antiglobulin
test, patient’s RBCs are
used where as in indirect
antiglobulin test patient’s
serum is used for the test.
Fig. 5.2: Direct and indirect methods of antiglobulin test (Coombs test)
Serum findings Antiglobulin test is useful

•• Serum bilirubin: increased. for diagnosis of HDN.
•• Lactate hydrogenase (LDH): increased.
•• Haptoglobin: decreased.
ABO Hemolytic Disease of the Newborn ABO HDN is more common

but less severe. It may be
•• It is less severe. seen in first pregnancy.
•• The fetus may be affected in the first pregnancy of a mother with blood group O.
•• The IgG antibodies to A or B from maternal blood cross placenta and enter the fetal
circulation. These anti-A or anti- B antibodies react with A and B antigenic determinants
present in fetal fluids and tissues.
•• This results in consumption of major portion of the maternal IgG and the small portion,
which is left combines with fetal red cells causing only mild hemolysis.
ANTIGLOBULIN (COOMBS) TEST Q. Write short notes on Coombs

(antiglobulin) test.
It is useful to detect the presence of incomplete antibody (IgG) and/or complement on the
RBC membrane.
Principle
•• RBCs coated with incomplete antibody (IgG) or C3 complement does not cause aggluti-
nation of RBCs.
•• Coombs reagent contains antibodies (antiglobulins) against human IgG/IgM/complement.
•• If the RBCs coated by incomplete antibody or complement, are treated with Coombs
reagent, the antiglobulins in the reagent will induce agglutination of such RBCs.
Types of Antiglobulin Test (Fig. 5.2) There are 2 types of

antiglobulin test: direct
•• Direct (Coombs) antiglobulin test (DAT) and indirect.
•• Indirect (Coombs) antiglobulin test (IAT)
Direct Antiglobulin Test (Fig. 5.2) Patient’s red cells are used
in direct antiglobulin test.
Direct antiglobulin test (DAT) (direct Coombs test) detects antibodies (IgG) and/or comple-
ment coated on the surface of patient’s RBC membrane.
•• Patient’s RBCs are taken in a test tube and washed three times in normal saline.
•• Coombs (anti-globulin) reagent is added and observed for agglutination.
•• Agglutination indicates the presence of antibody on the RBC membrane and interprets as
positive DAT.
Uses of Direct Antiglobulin Test

•• Hemolytic disease of the newborn (HDN), in which direct Coombs test is performed on the
newborn baby’s red cells from the cord blood.
•• Autoimmune hemolytic anemia: to demonstrate in vivo attachment of antibodies to red
cells.
•• Drug induced red cell sensitization.
•• Investigation of hemolytic transfusion reaction.
•• In HDN, newborn baby’s RBCs from cord blood is used for direct antiglobulin test, which
will be positive.
Patient’s serum is used for Indirect Antiglobulin Test (Fig. 5.2)

indirect antiglobulin test.
Indirect antiglobulin test (IAT) (indirect Coombs test) detects the presence of incomplete
(IgG) antibodies and/or complement in the patient’s serum.
•• In this test, patient’s serum is taken and “O” Rh positive cell suspension of any normal
individual is added.
•• “O” Rh positive RBCs are coated with (lgG) anti-Rh antibodies (if present) in the patient’s serum.
•• Add Coombs (antiglobulin) reagent and examine for agglutination.
•• Agglutination of RBCs indicates the presence of antibodies in the patient’s serum and test is
reported as positive for indirect antiglobulin test.
Patient’s serum + O Rh positive RBC suspension + Coombs reagent → Agglutination (test positive).
Uses of Indirect Antiglobulin Test

•• Hemolytic disease of newborn: mother’s serum is tested to detect anti-Rh antibody.
•• Cross-matching for blood transfusion: to detect incompatibility of recipient’s serum with
donor’s cells.
The type of antibody

causing autoimmune
AUTOIMMUNE HEMOLYTIC ANEMIA
hemolytic anemia may •• Antibodies against self-antigens on the RBC membrane cause premature destruction of
be warm antibody or RBCs.
cold agglutinin or cold •• Anti-RBC antibodies can be divided into three general categories (Table 5.1). Interaction of
hemolysin. the autoantibody with the red cell antigen is dependent on the temperature, i.e. warm or
cold type.
Warm AIHA: mediated Warm Antibody Type

by IgG autoantibody-
optimally active at 37°C. •• Most common type (50–70%).
•• Idiopathic (primary) or secondary to drug exposure or predisposing disease.
•• IgG type antibodies combine with RBC antigen at 37°C—warm antibody.
•• Direct antiglobulin test: DAT (Coombs test) positive in 90–95% cases.
•• LE cell test: positive in SLE with secondary autoimmune hemolytic anemia (AIHA).
Cold Agglutinin Type

•• Caused by cold agglutinins.
•• Mediated by IgM antibodies optimally active below 30°C.
•• Occurs as a complication of infections (e.g. infectious mononucleosis, Mycoplasma
infections) and lymphoid neoplasms.
Cold Hemolysins Type

(Donath-Landsteiner Antibodies)
•• Autoantibodies directed against the P antigen system on red cells.
•• Responsible for a rare disorder known as paroxysmal cold hemoglobinuria.
•• Direct antiglobulin test is usually negative.
FRAGMENTATION SYNDROME
The RBCs subjected to trauma (physical or mechanical) in the circulation can undergo
fragmentation and result in intravascular hemolysis leading to hemolytic anemias. These are
known as fragmentation syndrome.
Classification
According to the site of hemolysis it is classified as:
•• Macroangiopathic (large vessels) hemolytic anemia: red cell trauma from an abnormal
vascular surface (e.g. prosthetic heart valve, synthetic vascular graft).
•• Microangiopathic hemolytic anemia (MAHA): it occurs in capillaries due to abnormal
narrowing of the lumen (e.g. disseminated intravascular coagulation).
PAROXYSMAL NOCTURNAL PNH is an acquired

disorder in which there is
HEMOGLOBINURIA deficiency of GPI linked
proteins, which normally
It is a rare and is the only hemolytic anemia acquired mutation in the hematopoietic stem cell. protect the red cells
against complement
mediated lysis.
Etiology and Pathogenesis
•• Acquired mutations in the phosphatidylinositol glycan-group A (PIGA) gene in the
hematopoietic stem cell.
•• PIGA gene mutation causes deficient synthesis of GPI-linked proteins in blood cells
and loss of anchor for decay-accelerating factor (DAF). Normally, DAF responsible for In PNH, RBCs are very
complement degradation. sensitive to complement-
•• RBCs are abnormally sensitive to complement-mediated intravascular hemolysis. mediated hemolysis.
Clinical Features
•• Intravascular hemolysis: hemoglobin in acidic urine is converted into acid hematin and
results in dark brown urine.
•• Thrombosis: in the hepatic, portal or cerebral veins.
Laboratory Findings
•• Ham’s acidified serum test and sucrose hemolysis test: patient’s RBCs undergo lysis when PNH: Ham’s acidified serum
incubated with acidified serum (Ham test) or sugar (sucrose hemolysis test). test and sucrose hemolysis
•• Flow cytometry: detects RBC deficient in GPI-linked proteins (CD55 and CD59) and is test +ve.
useful for diagnosis of PNH.
ANEMIAS OF BLOOD LOSS

Acute Blood Loss (Hemorrhage) During recovery phase
of acute blood loss,
•• Causes loss of intravascular volume and if massive can lead to hypovolemic shock and peripheral smear show
death. reticulocytosis.
•• Bleeding may be external (e.g. open fracture, knife wound) or internal (e.g. ruptured
spleen, ruptured abdominal aneurysm).
–– RBCs: normocytic normochromic anemia. Polychromasia during the recovery phase due to
increased reticulocytes.
–– WBCs: leukocytosis.
–– Platelets: increased in number (thrombocytosis) during recovery phase.
Chronic Blood Loss

Produces anemia when the rate of blood loss exceeds the regenerative capacity of the bone
marrow or when iron reserves are depleted and results in iron deficiency anemia.
Sideroblastic anemias are SIDEROBLASTIC ANEMIAS

rare refractory anemias
which may be hereditary Rare heterogeneous group of refractory anemias characterized by:
or acquired. •• Ring sideroblasts in the bone marrow aspirate (Fig. 5.3).
•• Dimorphic peripheral blood picture: microcytic hypochromic red cells in hereditary form
and macrocytic in the acquired forms of the disease mixed with normochromic cells.
•• Iron-containing inclusions (Pappenheimer bodies) in the
RBCs.
•• Increased serum iron concentration and markedly
increased storage iron.
•• Ineffective erythropoiesis.
It is classified as:
1. Hereditary sideroblastic anemia Fig. 5.3: Ring sideroblasts with par-
2. Acquired sideroblastic anemia: idiopathic or secondary. tial perinuclear ring of iron granules
SECTION 2
Disorders of
White Cells
Quantitative and
Qualitative Disorders of 6
Leukocytes CHAPTER
NORMAL DIFFERENTIAL LEUKOCYTE COUNT (DLC) Differential leukocyte

count (DLC) is one of
The normal range of DLC in an adult is presented in Table 6.1. the routine, useful and
important investigations.
TABLE 6.1: Normal range of different leukocytes in an adult

Type of white blood cell Normal range
Neutrophils 40–70% (2.0–7.0 × 109/L)
Lymphocytes 20–40% (1.0–3.0 × 109/L)
Monocytes 2–10% (0.2–1.0 × 109/L)
Eosinophils 1–6% (0.02–0.5 × 109/L)
Basophils Less than 1% (0.02–0.1 × 109/L)
QUANTITATIVE DISORDERS OF LEUKOCYTES

Leukocytosis Q. Define leukocytosis and list
An increase in the total number of leukocytes in the blood more than 11,000/cu mm (11 × 10 /L). its causes.
9
Causes: common causes of leukocytosis are shown in Table 6.2.

Leukocytosis is usually
due to increase in the
TABLE 6.2: Common causes of leukocytosis neutrophils, but may
also be due to increased
•• Infections lymphocytes (or
–– Bacterial rarely monocytes and
–– Viral infections (e.g. infectious mononucleosis) eosinophils).
•• Leukemia
–– Acute
–– Chronic: chronic lymphocytic leukemia and chronic myeloid leukemia
•• Leukemoid reactions
•• Physiological
–– Pregnancy
–– Exercise
46 SECTION 2 Disorders of White Cells
Leukopenia is the decrease Leukopenia

in the WBC count below
4,000/cumm. Total leukocyte count is less than 4,000/cu mm (4 × 109/L).
Causes: common causes of leukopenia are shown in Table 6.3.
TABLE 6.3: Common causes of leukopenia

The causes of leukopenia •• Typhoid and paratyphoid
include typhoid and
•• Anemia
paratyphoid fever and
–– Aplastic anemia
aplastic anemia.
–– Megaloblastic anemia
•• Hypersplenism
•• Drugs including cytotoxic drugs
•• Radiation
•• Rarely leukemia
Q. Define neutrophilia and Disorders of Neutrophils

mention its causes.
Neutrophilia (Fig. 6.1)
Neutrophilia: absolute
neutrophil count more
An absolute neutrophil count of more than 8000/cu mm (8 × 109/L). Differential count shows
than 8000 cells/mm. more than 70% neutrophils and is usually accompanied by leukocytosis (15–30 × 109/L).
Common causes of Causes of neutrophilia: major causes of neutrophilia are shown in Table 6.4.
neutrophilia are infections,
inflammatory conditions
and tissue necrosis.
TABLE 6.4: Major causes of neutrophilia
1. Pathological:
–– Acute bacterial and fungal infections:
◆◆ Localized: pyogenic microorganisms causing infections, e.g. pneumonias, pyogenic meningitis,
cellulitis, diphtheria, abscess, tonsillitis, etc.
◆◆ Generalized: septicemia, acute rheumatic fever
–– Acute inflammatory processes: inflammatory conditions (acute appendicitis), vasculitis
–– Tissue necrosis: burns, myocardial infarction, gangrene, neoplasms (tumor necrosis)
–– Acute stress or hypoxic states: following hemorrhage, hemolysis and surgery
–– Myeloproliferative neoplasms: chronic myeloid leukemia, polycythemia vera
–– Metabolic: uremia, acidosis, gout
–– Miscellaneous: eclampsia, steroid therapy
2. Physiological:
–– Exercise (shift from marginating pool to circulating pool), newborns, extremes of temperature, pain,
emotional stress and during obstetric labor
Fig. 6.1: Peripheral smear showing neutrophilia

Quantitative and Qualitative Disorders of Leukocytes CHAPTER 6 47
Leukemoid Reaction Leukemoid reaction:

benign exaggerated
Benign leukocytic proliferation characterized by a total leukocyte count of more than 25 × leukocyte proliferation
109/L with immature white cells (like band forms, metamyelocytes and myelocytes). to be differentiated from
leukemia.
It is different from chronic myelocytic/myeloid leukemia (Table 6.5).
TABLE 6.5: Differences between leukemoid reaction and chronic myeloid leukemia Q. Tabulate the differences
between leukemia and leukemoid
Leukemoid reaction Chronic myeloid leukemia
reaction.
Clinical features Features of causative disease Splenomegaly, and bone pain
are common
Peripheral blood findings Neutrophils in bacterial
infections show toxic
WBC granules.
Total WBC count Moderately increased, rarely exceeds Markedly increased and usually
50 × 109/L 50 × 109/L
Differential leukocyte count Shift to the left with few immature Shift to the left with numerous Dohle bodies are small
forms. Toxic granulation seen immature forms. Myelocyte and round to oval structures
neutrophil peak seen in the cytoplasm
can also be observed in
Eosinophilia and basophilia Variable Present
bacterial infections.
Leukocyte alkaline phosphatase Increased Decreased
(LAP)
RBC
Anemia Usually minimal or absent Severe and progressive
Platelets
Number Variable Normal or increased In leukemoid reaction
LAP score is raised and
Extramedullary myeloid tumors Absent Present neutrophils may show
Philadelphia chromosome Absent Present toxic granulation.
Neutropenia (Agranulocytosis) Neutropenia: absolute

neutrophil count below
Reduction in the absolute neutrophil count (total WBC × % segmented neutrophils and 1500 cells/cu mm.
band forms) below 1.5 × 109/L (1500/cu mm).
Etiology: the causes of neutropenia are presented in Table 6.6.
Eosinophilia (Fig. 6.2)

Eosinophilia: eosinophil
Eosinophil count of more than 450/cu mm (0.45 × 109/L). count more than 450 cells/
Causes of eosinophilia are presented in Table 6.7. cu mm.
Fig. 6.2: Peripheral smear showing eosinophilia

TABLE 6.6: Causes of neutropenia

1. Inadequate production:
–– Suppression of stem cells: in these disorders granulocytopenia represents a component of
pancytopenia
◆◆ Aplastic anemia
Agranulocytosis: ◆◆ Marrow infiltration
neutrophil count below ◆◆ Metastatic tumors
0.5 × 109/L. The patients ◆◆ Granulomatous disorders
are highly susceptible –– Suppression of committed granulocytic precursors
to bacterial and fungal ◆◆ Drugs and chemicals (e.g. sulfonamides, analgesics, arsenicals)
infections. ◆◆ Ionizing radiation
–– Diseases associated with ineffective hematopoiesis
◆◆ Megaloblastic anemias: vitamin B12 or folate deficiency
◆◆ Myelodysplastic syndromes
–– Congenital: Kostmann syndrome (rare)
–– Severe infections
◆◆ Bacterial (e.g. typhoid, paratyphoid, septicemia)
◆◆ Viral (e.g. influenza, infectious mononucleosis, hepatitis, measles)
◆◆ Rickettsial (e.g. scrub typhus)
◆◆ Protozoal (e.g. malaria, kala-azar)
2. Increased destruction of neutrophils:
–– Immunologically mediated destruction
◆◆ Idiopathic
◆◆ Secondary
◊ Drugs
◊ Autoimmune disorders, e.g. systemic lupus erythematosus
–– Splenic sequestration may be associated with pancytopenia
3. Shift from the circulating pool to marginating pool:
–– Hemodialysis and cardiopulmonary bypass
4. Idiopathic: mechanism not known
–– Hodgkin and non-Hodgkin lymphoma
–– Chronic lymphocytic leukemia
–– Viral infections (HIV, hepatitis)
–– Cyclic neutropenia
Q. Define eosinophilia and list TABLE 6.7: Causes of eosinophilia

its causes. 1. Allergic/atopic conditions
– Asthma – Urticaria
– Hay fever – Drug reactions
– Allergic rhinitis
2. Parasitic infestations (with tissue invasion)
– Roundworm infestation – Hookworm infestation
– Filariasis
Eosinophilia is seen in 3. Fungal infections (e.g. coccidioidomycosis)
allergic reactions and
4. Skin diseases
parasitic infestations with
tissue invasion. – Dermatitis (eczema) – Pemphigus
– Scabies – Dermatitis herpetiformis
5. Hematological diseases
– Chronic myeloid leukemia – Polycythemia
– Hodgkin lymphoma – Acute myelomonocytic leukemia
– Eosinophilic leukemia
6. Miscellaneous
– Tropical eosinophilia – Pulmonary eosinophilia
– Löeffler’s syndrome – Hypereosinophilic syndrome
– Eosinophilic granuloma
Basophilia
Normally basophils (Fig. 6.3) are less than 1% of WBCs in peripheral blood.
Causes of basophilia include chronic myeloid leukemia, immediate hypersensitivity reactions,
mastocytosis, etc.
Monocytosis (Table 6.8)

More than 10% of differential count or an absolute monocyte (Fig. 6.4) count exceeding 500/ Fig. 6.3: Diagrammatic
cu mm (0.5 × 109/L). appearance of basophil
TABLE 6.8: Causes of monocytosis

1. Infections
–– Bacterial: tuberculosis, bacterial endocarditis, brucellosis
–– Protozoal: malaria, kala-azar
–– Spirochetal: syphilis
–– Rickettsial: typhus, rocky mountain fever
–– Recovery phase of neutropenia and acute infections
2. Inflammatory diseases
–– Inflammatory bowel disease: ulcerative colitis, Crohn disease
–– Autoimmune diseases: systemic lupus erythematosus, rheumatoid arthritis Fig. 6.4: Diagrammatic
–– Sarcoidosis appearance of monocyte
3. Hematologic malignancies
–– Acute monocytic, myelomonocytic and myelocytic leukemias
–– Chronic myelomonocytic leukemia
–– Hodgkin lymphoma
–– Multiple myeloma
Lymphocytosis Lymphocytosis:
lymphocyte count more
Lymphocyte (Fig. 6.5) count more than 4,000/cu mm (4 × 109/L) in adults and more than than 4,000/cu mm in
8,000/cumm (8 × 109/L) in child. adults and more than
8,000/cumm (8 × 109/L)
Common causes of lymphocytosis are given in the Table 6.9 in child.
Dengue fever is caused

by flavi virus transmitted
TABLE 6.9: Causes of lymphocytosis by freshwater mosquito
1. Acute infections (Aedes egypti). Peripheral
smear shows transformed
–– Viral infections: infectious mononucleosis, mumps, measles, chickenpox, infectious hepatitis
lymphocytes and
–– Toxoplasmosis
thrombocytopenia.
2. Chronic infections/inflammatory diseases
–– Tuberculosis
–– Syphilis
–– Brucellosis
–– Inflammatory bowel disease: Crohn disease and ulcerative colitis
3. Hematologic malignancies
–– Acute lymphoblastic leukemia
–– Chronic lymphocytic leukemia
–– Non-Hodgkin lymphoma with spill over
–– Adult T cell leukemia/lymphoma
–– Hairy cell leukemia
Fig. 6.5: Diagrammatic
appearance of lymphocyte
Lymphocytopenia
Lymphocyte count below 1,500/cu mm (1.5 × 109/L) in adults and below 3000/cu mm
(3 × 109/L) in children.
Some of the important causes of lymphocytopenia are listed in the Table 6.10.
TABLE 6.10: Causes of lymphocytopenia

1. Increased destruction
–– Corticosteroids
–– Cytotoxic drugs
–– Radiation
2. Decreased production
–– Aplastic anemia
–– Advanced malignancy: Hodgkin lymphoma
–– Infections: AIDS, miliary tuberculosis
3. Increased loss via GI tract
–– Obstruction to intestinal lymphatic drainage (e.g. tumor)
–– Congestive heart failure
QUALITATIVE DISORDERS OF LEUKOCYTES

Qualitative disorders of leukocytes are rare familial disorders that manifest as morphologic
changes in the leukocytes (Fig. 6.6).
Chediak-Higashi anomaly
is associated with
increased susceptibility to
pyogenic infections.
CGD is associated with

impaired phagocytosis and
killing of organisms.
Fig. 6.6: Various quantitative disorders of leukocytes

INFECTIOUS MONONUCLEOSIS
(GLANDULAR FEVER)
Acute, benign, self-limiting lymphoproliferative disorder caused by Epstein-Barr virus (EBV). EBV infects B cells but the
•• Incubation period: 4 to 8 weeks. peripheral blood shows
CD8 + T cells, which appear
•• Mode of transmission: oropharyngeal secretions (kissing), hence the nickname kissing
as atypical lymphocytes.
disease.
Pathogenesis
•• EBV infects B lymphocytes by binding to CD21 (CR2) receptor.
•• Viral infection begins in the submucosal lymphoid tissues of nasopharynx and oropharynx.
•• Virus remains dormant inside the B cells.
•• B cells are “immortalized” and are capable of proliferation indefinitely. Lesions caused by EBV:
1. Infectious
mononucleosis
Clinical Features 2. Burkitt lymphoma
3. Nasopharyngeal
•• Age: young adults among upper socioeconomic classes in developed nations and children carcinoma
of low socioeconomic status. 4. Hodgkin lymphoma
5. X-linked
•• Signs and symptoms: classical triad lymphoproliferataive
–– Fever disorders, and
–– Pharyngitis (sore throat) 6. Body cavity lymphoma.
–– Lymphadenopathy.
Laboratory Findings Q. Mention the laboratory

findings in infectious
•• Total leukocytes count increased (12,000 to 25,000 cells/cu mm): absolute lymphocytosis.
mononucleosis.
•• Atypical lymphocytosis (mononuclear cells): these are CD8 + subset (cytotoxic) of T cells
and not the virus-infected B cells.
•• Serological tests
–– Demonstration of heterophile antibodies
◆◆ Paul Bunnell test is characteristically positive.
◆◆ Monospot test is a sensitive slide test. Demonstration of specific
antibodies to EBV is the
–– Demonstration specific antibodies against EBV antigens: most specific test for
◆◆ Antibody against viral capsid antigens (anti-VCA). infectious mononucleosis.
◆◆ Antibodies to Epstein-Barr nuclear antigen (EBNA).
52
7 Acute Leukemia
CHAPTER
ACUTE LEUKEMIA
Q. Define and classify leukemia. Definition
Acute leukemia is a malignant disease of the bone marrow stem cell and its characteristic
features are:
Normally blast cells are less •• Bone marrow: diffuse replacement with proliferating neoplastic blast cells that fail to
than 5% of nucleated cells mature. Blast cells more than 20% (WHO criteria) of the nucleated cells in the marrow.
in the marrow. •• Peripheral blood: abnormal numbers and forms of immature white blood cells.
Aleukemic/subleukemic leukemia is characterized by very few/no blasts in the peripheral
blood.
Leukemia: malignant Acute leukemia are mainly divided into two groups namely acute lymphoblastic leukemia
disease of bone marrow (ALL) and acute myeloblastic leukemia (AML).
stem cell, arises in the
marrow and spreads.
Risk Factors: risk factors (Table 7.1) may cause mutations in the proto-oncogenes and tumor
suppressor genes.
TABLE 7.1: Risk factors for acute leukemia

ENVIRONMENTAL FACTORS
•• Ionizing radiation
•• Drugs:
–– Alkylating agents—nitrogen mustard, chlorambucil, etc.
–– AML occurs in myeloma patients treated with melphalan
–– Leukemia follows chemotherapy of lung and ovarian cancer
•• Chemicals: benzene (used in paint industry, plastic glues, etc.)
GENETIC DISORDERS
Example: Down syndrome (ALL or AML), Fanconi anemia (AML), ataxia telangiectasia (ALL, NHL)
ACQUIRED DISORDERS
•• PNH and aplastic anemia may transform into acute leukemia
•• AML may develop de novo or secondary to myelodysplastic syndrome (MDS)
Acute Leukemia CHAPTER 7 53
Classification Q. Classify acute leukemia.
Traditional classification depending on microscopic appearance of the involved cell and the
course of leukemias is presented in Table 7.2.
TABLE 7.2: Traditional classification of leukemia

•• Acute leukemia
–– Acute myelogenous/myeloblastic/myelocytic/myeloid leukemia (AML)
–– Acute lymphoblastic/lymphocytic leukemia (ALL)
•• Chronic leukemia
–– Chronic myeloid leukemia (CML)
–– Chronic lymphocytic leukemia (CLL)
FAB Classification of Acute Leukemias FAB criteria for the

•• First French, American and British (FAB) classification (1976) was based on the diagnosis of acute
1. morphological and 2. cytochemical characteristics of blast cells. leukemia: bone marrow
•• Revised FAB classification (Table 7.3): it includes should show a blast count
of 30% or more.
1. M: Morphology and cytochemistry of blast cells
2. I: Immunophenotyping
3. C: Cytogenetics
4. M: Molecular genetics.
TABLE 7.3: Revised French, American and British (FAB) classification of acute leukemias
Acute Lymphoid Leukemia
L1 Small homogenous cells with inconspicuous nucleoli
L2 Large cells with variable size and 1–2 nucleoli
L3 Large, homogeneous cells with finely stippled chromatin and prominent nucleoli. Cytoplasm is
basophilic and vacuolated
Acute Myeloid Leukemia
M0 Minimally differentiated AML
M1 AML without maturation
M2 AML with maturation
M3 Promyelocytic leukemia
M4 Myelomonocytic leukemia
M5 Monocytic leukemia
M6 Erythroleukemia
M7 Megakaryocytic leukemia
WHO Classification (2008) of Acute Leukemia (Table 7.4) WHO classification of

ALL: two main categories
TABLE 7.4: WHO classification (2008) of acute lymphoblastic and myeloid leukemia namely,
1. Precursor-B
A. Acute Lymphoblastic Leukemia lymphoblastic
I. Precursor-B lymphoblastic leukemia/lymphoma leukemia/lymphoma,
II. Precursor -T lymphoblastic leukemia/lymphoma and
2. Precursor T
B. Acute Myeloid Leukemia lymphoblastic
I. AML with recurrent genetic abnormalities leukemia/lymphoma.
◆◆ AML with t(8;21)(q22;q22); RUNX1-RUNX1T1
◆◆ AML with inv(16)(p13;1q22); CBFB-MYH11
Contd...
Contd...
WHO classification ◆◆ APL with t(15;17)(q22;q12); PML-RARA
of AML: based on ◆◆ AML with t(9;11)(p22;q23); MLLT3-MLL
clinical, morphological, II. AML with MDS-related changes
immunophenotypic and III. Therapy-related myeloid neoplasms
genetic features. IV. AML not otherwise specified
◆◆ AML minimally differentiated
◆◆ AML without maturation
Minimum blast cells in
◆◆ AML with maturation
bone marrow should be
more than 20%. ◆◆ Acute myelomonocytic leukemia
◆◆ Acute monoblastic and monocytic leukemia
◆◆ Acute erythroid leukemia
◆◆ Acute megakaryoblastic leukemia
V. Myeloid sarcoma
VI. Myeloid proliferation related to Down syndrome
Q. List the differences between Abbreviations: AML, Acute myeloid leukemia; APL, Acute promyelocytic leukemia; MDS, Myelodysplastic syndrome
myeloblast and lymphoblast.
It is important to
differentiate between
Differences between Myeloblast and
lymphoblast and Lymphoblast (Table 7.5)
myeloblast because of
difference in treatment
and prognosis of AML and TABLE 7.5: Differences between myeloblast and lymphoblast based on morphology and
ALL. cytochemistry
Lymphoblast (Figs 7.1 and 7.3) Myeloblast (Figs 7.2, 7.4 and 7.5)
Myeloblast: comparison Size 2–3 times the size of lymphocyte 3–5 times the size of lymphocyte
with lymphoblast has 4 Ms Cytoplasmic characters
M: more in size
Amount Scanty (less cytoplasm than myeloblast) Scanty to moderate (more cytoplasm
M: more nucleoli (3–5)
than lymphoblast)
M: moderate cytoplasm
M: myeloperoxidase +ve Color Blue Gray
Auer rod :+. Cytoplasmic granules Agranular May have cytoplasmic granules
Auer rod Negative Positive
Nuclear characters
Nuclear chromatin Uniform, coarse Uniform, fine
Nucleoli Inconspicuous or 1 to 2 3 to 5, prominent
N:C ratio High High
Accompanying cells Lymphocytes Promyelocytes, myelocytes, meta-
myelocytes, band forms and neutrophils
Cytochemistry
Myeloperoxidase Negative Positive
Sudan Black Negative Positive
Fig. 7.1: Diagrammatic PAS Block positivity Negative
appearance of lymphoblast Nonspecific esterase Negative Positive in M4 and M5
Fig. 7.2: Diagrammatic Fig. 7.3: Periodic acid Fig. 7.4: Myeloblast Fig. 7.5: Myeloblast
appearance of myeloblast Schiff (PAS) stain showing stained positively with stained positively with
lymphoblast with block myeloperoxidase (MOP) Sudan Black
positivity
ACUTE LYMPHOBLASTIC Differentiating

malignant pre-B and
LEUKEMIA/LYMPHOMA pre-T lymphoblasts on
morphology is difficult.
•• Acute Lymphoblastic Leukemia/Lymphoma (ALL) is a group of neoplasms consisting of
lymphoblasts.
•• Lymphoblast is immature, precursor B (pre-B) or T (pre-T) lymphocyte.
•• WHO classification (Table 7.4):
–– Precursor B cells ALL (about 85%) seen in childhood and present as acute leukemias.
–– Precursor T cells ALL (15%) present in adolescent males as lymphomas, often with
involvement of mediastinum (thymus).
Molecular Pathogenesis Requires

immunophenotyping for
•• Chromosomal abnormalities are found in about 90% of ALLs. subclassification of ALL.
–– Numerical abnormality: hyperploidy (>50 chromosomes) and hypoploidy.
–– Structural abnormality: balanced chromosomal translocations (e.g. Philadelphia
chromosome).
◆◆ Most T-ALLs have mutations in NOTCH1 gene.
◆◆ Most B-ALLs have mutations in genes PAX5, E2A and EBF or a balanced translocation T-ALL has worse prognosis
compared to B-ALL.
t (12; 21) involving the genes TEL and AML1.
Classification of Acute Lymphoblastic

Leukemia (Tables 7.3, 7.4 and 7.6)
TABLE 7.6: Characteristics of FAB subtypes of acute lymphoid leukemias (ALL)
FAB type L1 L2 L3 Morphologically, as per
the FAB classification
Cell size Small cell size Large heterogeneous Large, homogeneous
lymphoblast are classified
cell population cell population as L1, L2 and L3.
Nuclear characteristics
Shape Regular Irregular, clefting and Regular, oval or round
indentation common
Chromatin Condensed Dispersed chromatin Finely stippled ALL-L1 has better prognosis
than ALL-L3.
Nucleolus Small and Visible, 1–2 in number Usually prominent
inconspicuous
Cytoplasmic characteristics
ALL-L3 is a leukemic
Amount Scanty Variable, often Moderately abundant counterpart of Burkitt
abundant lymphoma.
Cytoplasmic basophilia Slight to moderate Variable Strong
Cytoplasmic vacuolation Absent Variable Prominent and oil red O
stain positive
Clinical Features
Age: most common hematological malignancy of children. Most common between 1 and 5 ALL is the most common
years of age and between 30 and 40 years. leukemia in children and
is usually associated with
Sex: slight male preponderance. lymphadenopathy.
Onset: abrupt.
Symptoms are due to bone Symptoms:

marrow infiltration by •• Bone marrow failure:
blasts. –– Anemia: causes fatigue, weakness.
Bone marrow failure –– Neutropenia: infections by bacteria or opportunistic fungi. Develop sore throat and
• anemia respiratory infections.
• neutropenia –– Thrombocytopenia: bleeding into the skin and mucosa in the form of purpura or
• thrombocytopenia.
ecchymoses.
–– Bone pain and sternal tenderness.
•• Extramedullary infiltration:
–– Lymphadenopathy: 75% of patients, usually involve cervical lymph nodes.
–– Bone pain and tenderness.
–– Hepatosplenomegaly: splenomegaly is more common than hepatomegaly.
–– Mediastinal thymic mass: more common in T-ALL.
•• CNS involvement: spread into the meninges causes leukemic meningitis ALL (pre-B).
•• Testicular involvement (ALL).
Q. Write short note on laboratory/ Laboratory Findings

peripheral smear findings in acute
lymphoblastic leukemia. Peripheral Blood
•• Total WBC count: markedly raised ranging from 20 × 109/L to 200 × 109/L
Subleukemic leukemia: •• Platelet count: reduced (thrombocytopenia).
total WBC count lower than •• Hemoglobin: decreased and may be as low as 3 g/dL.
4 × 109/L and peripheral
blood shows very few
blasts. •• Peripheral smear (Figs 7.6 and 7.7):
–– RBCs: normocytic normochromic anemia.
Aleukemic leukemia: total –– WBCs: total count markedly increased and 20% or more lymphoblasts.
white cell count is low ◆◆ Morphology of lymphoblasts:
(< 4 × 109/L) with no blasts
◊ Larger than small lymphocyte
in the peripheral blood.
◊ High N:C ratio
◊ Nucleus with condensed chromatin and nucleoli are either absent or inconspicuous
Lymphoblasts should
be differentiated from ◊ Scant to moderate agranular basophilic cytoplasm.
myeloblasts (see Table 8.3). –– Platelets: thrombocytopenia.
Fig. 7.6: Peripheral blood smears in acute lymphoblastic leukemia Fig. 7.7: Diagrammatic peripheral blood smear in acute
showing lymphoblasts (arrows). Inset shows lymphoblast with block lymphoblastic leukemia showing lymphoblasts (arrows)
positivity with PAS stain
Cytochemistry of Lymphoblasts Lymphoblast: cytoplasm

shows block positivity with
•• PAS: cytoplasmic aggregates of PAS positive (Figs 7.3 and 7.6) material (block positivity). PAS stain.
•• Myeloperoxidase (MPO) negative.
•• Sudan black B negative.
Bone Marrow
•• Cellularity: markedly hypercellular due to proliferation of blasts.
•• Erythropoiesis and myelopoiesis: reduced.
•• Megakaryopoiesis: megakaryocytes gradually decrease.
•• Blasts: constitute 20–100% of the marrow cells.
Immunophenotyping
Terminal-deoxynucleotidyl-transferase (TdT) + in pre-B and pre-T lymphoblasts.
•• Immature B cells + positive for pan B cell marker CD19 and CD10 (CALLA—common ALL Distinction between
antigen). precursor B and T cell ALL
•• Precursor T ALL cells are positive for CD2, CD5 and CD8. requires lineage-specific
markers.
•• Serum uric acid: raised due to destruction of leukemic cells during chemotherapy leading
to hyperuricemia.
•• LDH: raised, because of increased turnover of leukemic cells.
CSF Examination
To know/rule out CNS involvement.
Prognosis: prognostic features of ALL are presented in Table 7.7. Presence of Philadelphia
chromosome in ALL:
prognosis unfavorable.
TABLE 7.7: Prognostic factors in ALL
Unfavorable prognosis Favorable prognosis
Age Below 2 years and above 10 years Between 2 to 10 years
(adolescence or adulthood)
Prognosis is far better in
Sex Males Females ALL than AML.
Total WBC count High (more than 50,000 cells/cu mm) Low
95% of children develop
Meningeal involvement Present Absent complete remission.
Cytogenetic t(9;22) (the Philadelphia chromosome) Hyperploidy, trisomy of chromosomes 4, 7 75 to 85% are cured with
abnormalities and 10 and t(12;21) current chemotherapy.
Time required for clearing More than 1 week Less than 1 week
blasts from blood
ACUTE MYELOGENOUS LEUKEMIA

Definition: neoplasm of hematopoietic progenitors characterized by proliferation resulting in
accumulation of immature myeloblasts in the marrow.
Classification of acute myelogenous leukemia (AML): refer Tables 7.3 and 7.4.
AML synonyms: acute Molecular Pathogenesis

myeloid/myeloblastic/
myelocytic leukemia. •• Many recurrent genetic abnormalities can disrupt genes encoding transcription factors
involved in normal myeloid differentiation.
•• Mutated tyrosine kinase activation is a common.
Clinical Features
AML: develop at any age. Age: AML may develop at any age, but is more common in adults.
Usually 15–60 years of age. Onset: acute leukemias are abrupt in onset.
Symptoms: related to depressed marrow function.
•• Bone marrow failure:
Symptoms are due to
anemia, neutropenia and –– Anemia: fatigue and weakness.
thrombocytopenia. –– Neutropenia: life-threatening infections by bacteria or opportunistic fungi.
–– Thrombocytopenia: bleeding, patient may also develop disseminated intravascular
coagulation (DIC) in AML M3 and primary fibrinolysis.
Acute promyelocytic
–– Bone pain and tenderness
leukemia (AML-M3)
may be associated with •• Extramedullary infiltration
widespread bleeding due –– Gingival hypertrophy (M4 and M5) and infiltration of skin (leukemia cutis).
to DIC. –– Hepatosplenomegaly: usually more than in ALL.
Laboratory Findings
Q. Write short note on laboratory/ Peripheral Blood
peripheral smear findings in AML. •• Total WBC Count: markedly raised ranging from 20 × 109/L to 100 × 109/L.
•• Hemoglobin: decreased and ranges from 5 to 9 g/dL.
Subleukemic leukemia: •• Peripheral smear (Figs 7.8 and 7.9):

total WBC count lower than –– RBCs: normocytic normochromic type of anemia.
4 × 109/L and peripheral
–– WBCs: total WBC count markedly increased.
blood shows very few
blasts.
◆◆ Differential count: more than 20% myeloid blasts. May show more than one type of blast or
blasts with hybrid features.
◆◆ Morphology of myeloblasts
Aleukemic leukemia: total ◊ 3 to 5 times larger than the diameter of a small lymphocyte.
white cell count is low ◊ High N:C ratio.
(< 4 × 109/L) with no blasts ◊ Fine nuclear chromatin with 2-4 variably prominent nucleoli.
in the peripheral blood. ◊ More cytoplasm than lymphoblasts—azurophilic, peroxidase-positive granules.
◊ Presence of Auer rods is definitive evidence of myeloid differentiation.
AML: Auer rods in the ◆◆ Auer rods are azurophilic needle-like peroxidase-positive structures in the cytosol of myelo-
cytoplasm of myeloblasts, blasts (M2 and M3 subtype).
seen in AML; not in CML. –– Platelets: moderate to severe thrombocytopenia and causes bleeding from skin and mucosa.
Myeloblasts stain positively Cytochemistry of Myeloblasts (Figs 7.10 and 7.11)

with myeloperoxidase and •• Stain positively with myeloperoxidase (MPO) and Sudan black B.
Sudan black B.
•• Monoblasts stain with nonspecific esterases.
Both in subleukemic and

aleukemic leukemia bone
Bone Marrow
marrow contains blasts •• Cellularity: markedly hypercellular.
more than 20%. •• Erythropoiesis: markedly suppressed.
•• Myelopoiesis: suppression of myeloid maturation and myeloblasts constitute more than 20% of
marrow cells.
•• Megakaryopoiesis: gradually decreased.
Fig. 7.8: Peripheral smear in AML with myeloblasts. Inset shows Fig. 7.9: Diagrammatic peripheral blood smear in AML with
myeloblast with Auer rod myeloblasts. One myeloblast with two Auer rods (arrow)
Fig. 7.10: Myeloblast stained positively with myeloperoxidase Fig. 7.11: Myeloblast stained positively with Sudan black
Immunophenotyping AML prognosis:

• Fulminant course and
Diagnosis of AML is confirmed by using stains for myeloid specific antigens. has worse prognosis
than ALL.
• Cytogenetic markers are
major determinants of
Cytogenetics prognosis.
Very important in the WHO classification of AML (Table 7.4).
Myeloid sarcoma synonym:
Extramedullary myeloid
MYELOID SARCOMA tumor/granulocytic
sarcoma/chloroma.
Tumor mass consisting of myeloid blasts with or without maturation occurring at extra–
medullary sites. Myeloid sarcoma is
•• On sectioning: tumor is green (hence the term chloroma) frequent in skin, lymph
•• Microscopically myeloblasts with or without features of promyelocytic or neutrophilic node, GI tract, bone, soft
maturation. tissue and testis.
8 Myelodysplastic
Syndromes
CHAPTER
MYELODYSPLASTIC SYNDROMES
Myelodysplastic Syndromes (MDS) are a heterogeneous group of acquired clonal stem cell
disorders affecting stem cells.
MDS: cytopenias with MDS is characterized by:
hypercellular bone •• Progressive cytopenias
marrow.
About 30% progress to
•• Dysplasia in one or more cell lines
AML. •• Ineffective hematopoiesis
•• Risk of development of AML.
Classification
•• Idiopathic or primary MDS
•• Secondary/therapy-related MDS (t-MDS): complication of previous cytotoxic drug or
radiation therapy.
WHO classification of myelodysplastic syndromes is presented in Table 8.1.
Clinical Features
•• Elderly above 60 years
•• Slightly more common in males
•• Symptoms are due to cytopenias
•• About 10% to 40% of MDS patients progress to AML.
Laboratory Findings
•• Peripheral smear: cytopenias in the peripheral blood
–– RBCs: mild to moderate degree of macrocytic or dimorphic anemia.
–– WBCs: normal or low total leukocyte count.
–– Platelets: variable thrombocytopenia, large hypogranular or giant platelets.
Myelodysplastic Syndromes CHAPTER 8 61
TABLE 8.1: WHO classification of myelodysplastic syndromes

Disease Peripheral blood picture Bone marrow features
1. Refractory cytopenia with Unicytopenia or bicytopenia 1 Unilineage dysplasia in > 10% of
unilineage dysplasia (RCUD): the cells in one myeloid lineage
refractory anemia (RA), refractory < 5% blasts
neutropenia (RN), refractory < 15% ring sideroblasts
thrombocytopenia (RT)
2. Refractory anemia with ring Anemia Dyserythropoiesis >15% ring
sideroblasts (RARS) No blasts sideroblasts <5% blasts
3. Refractory cytopenia with Bi/pancytopenia Dysplasia in > 10% of cells in 2
multilineage dysplasia (RCMD) Rare blast myeloid lineages (neutrophil
No Auer rods and/or erythroid and/or
<1 × 109/L monocytes megakaryocytes) < 5% blasts
No Auer rods ± 15% ring
sideroblasts
4. Refractory anemia with excess < 5% blasts2 5–9% blasts
blasts-1 (RAEB-1) Bi/pancytopenia Unilineage or multilineage
No Auer rods dysplasia
<1 × 109/L monocytes No Auer rods
5. Refractory anemia with excess 5–19% blasts 10–19% blasts
blsts-2 (RAEB-2) Cytopenia Unilineage or multilineage
Auer rods ± 3 dysplasia
<1 × 109/L monocytes Auer rods ± 3
6. MDS unclassified (MDS-U) < 1% blasts < 5% blasts
Cytopenia only Unequivocal dysplasia in less
than 10% of cells in one or
more myeloid cell lines when
accompanied by cytogenetic
abnormalities considered as
presumptive evidence for
diagnosis of MDS
7. MDS with isolated del (5q) No or rare blasts < 5% blasts
Anemia Increased to normal
Platelets increased or normal megakaryocytes with
hypolobated nuclei. Isolated 5q
deletion. No Auer rods
1. Cases with pancytopenia should be classified as MDS-U
2. If marrow blasts < 5% with 2–4% myeloblasts in blood = RAEB-1. Cases with RCUD and RCMD with 1% myeloblasts in blood
should be classified as MDS-U
3. Cases with Auer rods and < 5% myeloblasts in blood and 10% in marrow = RAEB 2
Bone Marrow
Dysplasia of all non-lymphoid lineages (erythroid, granulocytic, monocytic and megakaryocytic)
associated with cytopenias.
Bone marrow in MDS:
•• Cellularity: hypercellular.
pawn ball megakaryocytes,
•• Erythropoiesis: dysplastic changes in erythroid precursors with megaloblastoid change and presence dysgranulopoiesis,
of ringed sideroblasts in iron stain. erythroid precursors with
•• Myelopoiesis: hyperplasia with dysgranulopoiesis. megaloblastoid change
•• Megakaryopoiesis: dysmegakaryopoiesis—pawn ball megakaryocytes. and presence of ringed
•• Iron stores: increased with ring sideroblasts. sideroblasts.
Ineffective hematopoiesis
Bone Marrow Trephine Biopsy

Abnormal localization of immature precursors (ALIP) in (refractory anemia with excess blasts
(RAEB).
9 Myeloproliferative
Neoplasms
CHAPTER
MYELOPROLIFERATIVE NEOPLASMS (MPN)

MPN peaks in the 5th to Definition: clonal hematopoietic stem cell disorders characterized by proliferation of one or
7th decade. more of the myeloid lineages (erythroid, granulocytic, megakaryocytic and mast cells).
All MPN show spleno- •• Splenomegaly and hepatomegaly due to sequestration of excess hematopoietic cells or
megaly. proliferation of abnormal hematopoietic cells.
WHO Classification of MPN

It is presented in Table 9.1.
TABLE 9.1: WHO (2008) classification of myeloproliferative neoplasm (MPN)

WHO (2008) Myeloproliferative neoplasms
Chronic myelogenous leukemia, BCR-ABL-1 positive
Chronic neutrophilic leukemia
Polycythemia vera—JAK2 V617F or exon 12 mutation
Primary myelofibrosis—JAK2 or MPL mutation
Essential thrombocythemia
•• Platelet count > 450 × 109/L
•• JAK2 mutation
Chronic eosinophilic leukemia, NOS
•• No BCR-ABL1, PDGFRA, PDGFRB or FGFR1 translocation
Mastocytosis—KIT mutation
Myeloproliferative neoplasm, unclassifiable
Pathogenesis
Presence of mutated, constitutively activated tyrosine kinases leads to proliferation of
hematopoietic stem cells and results in hypercellular marrow.
Myeloproliferative Neoplasms CHAPTER 9 63
POLYCYTHEMIA OR ERYTHROCYTOSIS
Polycythemia is characterized by increase in the RBC mass, usually with a corresponding
increase in hemoglobin level.
Pathophysiologic classification of polycythemia is given in Table 9.2.
TABLE 9.2: Pathophysiologic classification of polycythemia

ABSOLUTE
Primary (low erythropoietin level)
•• Polycythemia vera
Increase in red cells can be
Secondary (high erythropoietin level)
absolute or relative.
•• Physiologically appropriate
–– Compensatory
–– Lung disease
–– Living in high-altitude
–– Cyanotic heart disease (Tetralogy of Fallot)
•• Physiologically inappropriate (with increased erythropoietin)
–– Paraneoplastic: erythropoietin-secreting tumors (e.g. renal cell carcinoma, uterine leiomyoma,
hepatocellular carcinoma)
RELATIVE
Reduced plasma volume
•• Hemoconcentration (dehydration due to diarrhea, vomiting)
•• Gaisböck’s syndrome (spurious polycythemia)
POLYCYTHEMIA VERA Q. Write short notes on polycy-

themia vera.
Definition: polycythemia vera (PV) is an acquired myeloproliferative neoplasm arising from
malignant transformation of hematopoietic stem cell.
•• It is characterized by trilineage (erythroid, granulocytic, and megakaryocytic) hyperplasia
in the bone marrow.
•• It leads to uncontrolled production of red cells, granulocytes and platelets (panmyelosis)
and leads to erythrocytosis (polycythemia) and or granulocytosis and thrombocytosis.
Molecular Pathogenesis (Figs 9.1 and 9.2)

•• Normally, a tyrosine kinase protein called JAK2 (Janus 2 kinase gene), is activated following JAK2 mutation is
binding of the growth hormone erythropoietin. diagnostic of polycythemia
•• JAK2 then activates a signaling pathway causing cells to replicate. vera.
•• This process is strictly regulated by various feedback pathways.

•• Polycythemia vera (PV) is due to mutation in tyrosine kinase JAK 2 V617F, which causes PV: erythropoietin is
proliferation of not only erythroid lineage but also granulocytic and megakaryocytic lineage. decreased.
Clinical Features
1. Insidious.
2. Late middle age (median age at onset is 60 years).
3. Plethora and cyanosis, headache, dizziness and visual problems result from vascular PV: most symptoms are
disturbances in the brain and retina. due to the increased red
4. Thrombotic episodes: e.g. deep venous thrombosis, myocardial infarction, thrombosis of cell mass and hematocrit.
hepatic veins (producing Budd-Chiari syndrome).
Fig. 9.1: Normal signaling by JAK2 Fig. 9.2: In polycythemia vera, the presence of a mutant
version of JAK2 results in dysregulated downstream
signaling in the absence of erythropoietin
Phases
There are three phases of Polycythemia vera
PV develops into acute •• Proliferative phase: erythroid proliferation and increased red cell mass.
myelogenous leukemia in •• Spent phase: in 10%, excessive proliferation of erythroid cells ceases with stable or
2% to 5%.
decreased RBC mass.
•• Myelofibrosis: about 10% progress to myelofibrosis.
Laboratory Findings
Peripheral Blood (Fig. 9.3)
•• Hemoglobin: increased and are more than 18.5 g/dL in men and 16.5 g/dL in women.
•• Hematocrit: increased and about 60%.
Polycythemia vera is a •• Red cell count: increased and usually about 6 million/cu mm (6 × 1012/L).
chronic myeloproliferative •• White cell count: normal or increased.
neoplasm with RBC count •• Platelet count: normal or increased.
of more than 6 million/
cu mm.
–– RBCs: show normocytic normochromic picture.
–– WBCs:
◆◆ Mild to moderate leukocytosis
◆◆ Neutrophils are morphologically normal
◆◆ Basophils often increased
◆◆ NAP (LAP) score is increased to 150–300 (Normal 40–100).
–– Platelets: abnormally large and functionally defective.
PV: hematocrit is increased

and > 60%.
Fig. 9.3: Normal hematocrit in comparison with anemia and

polycythemia vera
Bone Marrow
•• Hypercellular due to hyperplasia of all elements (trilineage hyperplasia/panmyelosis) namely
erythroid, myeloid and megakaryocytic series with prominence of erythroid precursors in the bone
marrow.
Bone Marrow Biopsy

Shows increased reticulin fibers and fibrosis as the disease progresses.
Other Findings In PV arterial oxygen

saturation (pO2) is normal
•• Extramedullary hematopoiesis in the liver and spleen that causes hepatosplenomegaly.
(>92%) whereas in
•• Arterial oxygen saturation (pO2): normal (75–100 mm Hg) and is useful for differentiating secondary polycythemia it
it from secondary polycythemia. is <90%.
•• Erythropoietin levels: decreased in contrast to secondary polycythemia.
•• Serum vitamin B12 and uric acid: increased indicating increased cell turnover.
•• JAK2 V617F mutation: it can be demonstrated.
ESSENTIAL THROMBOCYTHEMIA ET synonym: primary

(essential/idiopathic)
Definition: chronic myeloproliferative neoplasm (MPN) primarily of megakaryocytic thrombocytosis.
lineage. It is characterized by increased megakaryopoiesis and thrombocytosis (more than
450 × 109/L).
Etiology ET: mutation of JAK2 gene

Thrombocytosis with a
•• Most due to point mutations in JAK2 gene and constitutive activation of JAK2, and count of > 450 × 109/L.
thrombopoietin-independent proliferation of megakaryocytes.
ET: throbbing and burning Clinical Features

sensation of hands and
feet due to blocking of •• Age: 50–60 years
arterioles by aggregates •• Thrombosis and hemorrhage
of platelets is known as •• Erythromelalgia: one of the characteristic features.
erythromelalgia.
Laboratory Findings
–– RBCs: normocytic normochromic.
–– WBCs: mild leukocytosis.
–– Platelets:
◆◆ Increased number (thrombocytosis)> 600,000/cu mm.
◆◆ Variation in size and shape-abnormally large platelets are common.

Megakaryocytic
hyperplasia and abnormal
Bone Marrow
(giant) platelets are •• Cellularity: mild to marked hypercellularity.
characteristic features. •• Erythropoiesis: normal or mild hyperplasia.
•• Myelopoiesis: normal or mild hyperplasia.
•• Megakaryopoiesis: markedly increased in number with abnormally large megakaryocytes (giant
ET course: indolent. megakaryocytes).
Extramedullary hematopoiesis: mild hepatosplenomegaly.
PRIMARY MYELOFIBROSIS
Myelofibrosis: mutation in Clonal MPN characterized by a proliferation of predominantly megakaryocytes and
JAK2 gene. granulocytes in the bone marrow.
Fully developed disease results in reactive marrow fibrosis and replaces hematopoietic
cells leading to cytopenias and extensive extramedullary hematopoiesis.
Molecular Pathogenesis
Most show JAK2 mutations.
Massive splenomegaly Clinical Features

due to extramedullary
hemopoiesis. •• Age: above 60 years of age.
•• Progressive anemia.
•• Splenomegaly.
Laboratory Findings
•• Peripheral smears: Primary myelofibrosis:
–– RBCs: moderate to severe degree of normochromic normocytic anemia accompanied by peripheral smear shows
leukoerythroblastosis and
leukoerythroblastosis. Tear drop-shaped red cells (dacryocytes), probably due to damage in the
tear drop cells.
fibrotic marrow can also be found.
–– WBCs: total white cell count is usually normal or reduced, but can be markedly elevated 80 to 100
× 109/L in early stages of the disease.
–– Platelets: they may be abnormally large. The platelet count is usually normal or elevated, but as
the disease progresses the count decreases.
Bone Marrow Primary myelofibrosis:

bone marrow fibrosis leads
•• Cellularity: in early stages, it is often hypercellular due to increase in maturing cells of all lineages. In to cytopenias.
later stages, it is replaced by fibrosis and becomes hypocellular and diffusely fibrotic resulting in a dry
tap.
•• Erythroid and granulocytic precursors: these are morphologically normal.
•• Megakaryocytes: these are large, dysplastic and abnormally clustered.
Bone Marrow Biopsy Bone marrow biopsy is

essential for the diagnosis
Stages: two stages have been recognized. of myelofibrosis as aspirate
1. Prefibrotic (cellular) stage: hypercellular bone marrow. Megakaryocytes increased and results in a dry tap late in
the course of the disease.
markedly abnormal.
2. Fibrotic stage: fibrosis distorts the marrow and prematurely releases nucleated erythroid
and early granulocyte progenitors (leukoerythroblastosis).
Reticulin stain demonstrates the increase in reticulin fibers (fibrosis).
Extramedullary hematopoiesis in spleen and liver produces hepatosplenomegaly.
Course: variable.
10 Chronic Myelogenous
Leukemia
CHAPTER
CML synonyms: chronic

myelocytic/myeloid/
CHRONIC MYELOGENOUS LEUKEMIA
granulocytic leukemia.
Definition
CML is an acquired MPN of
pluripotent hematopoietic Chronic myelogenous leukemia (CML) is one of the myeloproliferative neoplasm (MPN) of
stem cell. pluripotent hematopoietic stem cell characterized by overproduction of cells of the myeloid
series which results in marked splenomegaly and leukocytosis.
Distinguished from other myeloproliferative neoplasms by the presence of:
1. Chimeric fusion BCR-ABL gene.
2. Philadelphia (Ph) chromosome in more than 90% of cases.

Risk factor: exposure to ionizing radiation and benzene.
Q. Write short notes on

Philadelphia chromosome. Molecular Pathogenesis
Philadelphia (Ph) Chromosome (Fig. 10.1)
•• Acquired chromosomal abnormality in all proliferating hematopoietic stem cells
Philadelphia (Ph)
chromosome is a
(erythroid, myeloid, monocytic and megakaryocytic precursors).
shortened chromosome •• Balanced reciprocal translocation between long arm of chromosome 9 and 22, i.e. t (9; 22)
22 and is due to balanced (q 34; q 11.2). It increases the length of chromosome 9 and shortening of 22. This shortened
reciprocal translocation chromosome 22 is known as Philadelphia chromosome (Fig. 10.1).
between chromosome 9
and 22-t (9; 22).
BCR-ABL Fusion Gene (Fig. 10.2)

•• ABL proto-oncogene from chromosome 9 joins the BCR on chromosome 22.
Translocation results in
a BCR-ABL fusion gene, •• It produces a new chimeric (fusion) gene called BCR-ABL, thus converting ABL proto-
which produces neoplastic oncogene into oncogene. The product of the fusion gene plays a central role in the
proliferation. development of CML
Chronic Myelogenous Leukemia CHAPTER 10 69
CML: translocation results

in the head-to-tail fusion
of the breakpoint cluster
region (BCR) gene on
chromosome 22 with
the ABL (named after the
abelson murine leukemia
virus) gene located on
chromosome 9.
Fig. 10.1: Balanced reciprocal translocation between long arm of chromosome 9 and chromosome 22 resulting in
shortened chromosome 22 known as Philadelphia chromosome
Fig. 10.2: Fusion of ABL gene from chromosome 9 with BCR on chromosome 22 and its consequences
•• The product of this oncogene i.e., oncoprotein (e.g. p210) causes cell division and
inhibition of apoptosis.
Clinical Features
•• Age: usually occurs between 40 to 60 years of age. CML: usually occurs
between 40 and 60 years
•• Sex: males slightly more affected than females. of age.
•• Onset: insidious.
Symptoms:
•• Nonspecific symptoms: fatigue, weakness, weight loss, anorexia.
CML: moderate to massive •• Fullness of abdomen due to splenomegaly (caused by leukemic infiltration and extra
splenomegaly. medullary hematopoiesis). Splenomegaly is moderate to severe and is characteristic feature
in majority (80–90%) of patients.
•• Hepatomegaly: mild or moderate seen in 60–70% of cases.
NATURAL HISTORY OF CHRONIC

CML has three phases: MYELOID LEUKEMIA
chronic stable, accelerated
and blast phase. Three different phases: 1. chronic phase, 2. accelerated phase and 3. blastic phase.
Chronic/Stable/Indolent Phase (CP)

•• Most are diagnosed in this phase.
•• Lasts for 2 to 6 years.
•• If not treated, progresses gradually to accelerated phase or abruptly to blastic phase.
Q. Write short notes on laboratory
findings /peripheral smear in CML.
Laboratory Findings
Peripheral blood
CML: neutrophilia with the •• Hemoglobin: usually less than 11 g/dL
whole spectrum of mature
myeloid precursors. •• Peripheral smear:
–– RBCs: normocytic normochromic anemia
CML is characterized –– WBCs:
by anemia, extreme ◆◆ Marked leukocytosis (12–600 × 109/L) total leukocyte count usually exceeds 100 × 109/L
leukocytosis, granulocytic (1,00,000/cu mm).
immaturity, basophilia, ◆◆ Shift to left (shift to immaturity)—granulocytes at all stages of development (neutrophils,
thrombocytosis. metamyelocytes, myelocytes, promyelocytes and an occasional myeloblasts).
◆◆ Predominant cells are neutrophils and myelocytes.
The preponderance of ◆◆ Blasts are usually less than 10% of the circulating WBCs (Figs 10.3 and 10.4).
myelocyte is called as
◆◆ Basophilia and eosinophilia.
myelocyte bulge.
◆◆ Decreased NAP/LAP score: NAP score in CML is decreased below 20 (normal score range is
40–100). Helpful in differentiating CML from leukemoid reaction (see Table 7.5).
In CML LAP (NAP) is mark- –– Platelets: platelets range from normal (150–450 × 109/L) to greater than 1000 × 109/L. Up to 50%
edly reduced. have thrombocytosis.
Bone Marrow
•• Cellularity: markedly hypercellular due to myeloid hyperplasia.
•• M: E ratio: often exceeds 20:1.
•• Erythropoiesis: diminished erythropoiesis as disease progresses.
•• Myelopoiesis: marked hyperplasia. Blast cells usually less than 10%. Basophils, eosinophils and their
precursors are usually found.
•• Megakaryopoiesis: megakaryocytes are either normal or increased. Dwarf megakaryocytes.
•• Sea-blue histiocytes (Gaucher-like cells/pseudo Gaucher cells) are seen.
Biochemical findings:
•• Serum uric acid raised
•• Serum LDH raised.
Philadelphia chromosome and BCR-ABL fusion gene: demonstrated either by chromosomal
analysis or fluorescent in situ hybridization (FISH) or PCR based tests.
Chronic Myelogenous Leukemia CHAPTER 10 71
Fig. 10.3: Peripheral blood picture in chronic/stable phase of chronic Fig. 10.4: Diagrammatic peripheral blood picture in
myeloid leukemia chronic/stable phase of chronic myeloid leukemia
Accelerated Phase (AP) (Figs 10.5 and 10.6) CML: accelerated phase
is more aggressive and
•• More aggressive and lasts for few months. myeloblasts range from
•• Myeloblasts: 10–19% in the blood or bone marrow. 10% to 19%.
•• Striking basophilia (20% or more).
•• Persistent thrombocytopenia (less than 100 × 109/L) unrelated to therapy or persistent
thrombocytosis (more than 1000 × 109/L) uncontrolled by therapy.
•• Megakaryocyte proliferation in sheets or clusters in association with fibrosis.
•• Persistent or increasing splenomegaly unresponsive to therapy.
Fig. 10.5: Peripheral blood picture in accelerated phase of chronic myeloid Fig. 10.6: Diagrammatic peripheral blood picture in accelerated
leukemia showing numerous blasts (10–19%) and striking basophilia phase of chronic myeloid leukemia showing numerous blasts
(10–19%) and striking basophilia
Blast Phase/Crisis (BP)

CML blast crisis: blasts 20% Blood picture resembles acute leukemia and has poor prognosis.
or more, myeloblast (no
Auer rods) or lymphoblasts. •• Peripheral smear (Figs 10.7 and 10.8):
–– Blasts 20% or more. May be either myeloblast (70% cases) or lymphoblast (30% cases).
Prognosis: poor with Myeloblast does not contain Auer rods.
accelerated phase or blast
–– Thrombocytopenia causes bleeding episodes.
crisis.
Fig. 10.7: Peripheral blood picture in blast phase of chronic myeloid Fig. 10.8: Diagrammatic peripheral blood picture in blast phase of
leukemia showing numerous blasts (20% or more) and striking basophilia chronic myeloid leukemia showing numerous blasts (20% or more)
and striking basophilia
Chronic Lymphocytic
Leukemia/Small 11
Lymphocytic Lymphoma CHAPTER
CHRONIC LYMPHOCYTIC LEUKEMIA

Definition Q. Write short notes on chronic
lymphocytic leukemia.
Chronic lymphocytic leukemia (CLL)/small lymphocytic lymphoma (SLL) is a tumor
composed of monomorphic small B lymphocytes in the peripheral blood, bone marrow and
lymphoid organs (spleen and lymph nodes).
•• Both CLL and SLL is a single entity with different presentations.
•• Small lymphocytic lymphoma (SLL) is tissue equivalent of chronic lymphocytic leukemia
CLL/SLL are tumors derived
(CLL).
from B lymphocytes.
•• CLL/SLL tumor cells coexpress CD5 and CD23.

•• Environmental factors: suggested but none proved.
•• Hereditary factors: families with higher risk of CLL or other lymphoid neoplasms.
Cytogenetic Abnormalities
Common mutations are deletions of 13q14.3, 11q22-23, and 17p13. About 20% of CLL show
trisomy 12.
Clinical Features CLL patients may

be asymptomatic or
•• Age: between 50–60 years of age. present with generalized
lymphadenopathy.
•• Sex: more in males than in females (2:1).
•• Symptoms:
–– Asymptomatic in about 25–30%
–– Nonspecific symptoms: fatigue, loss of weight and anorexia
–– Generalized lymphadenopathy
–– Immunological defects either as immune deficiency or autoimmunity.
Laboratory Findings
CLL: absolute lymphocyte Peripheral Blood
count is more than 5 ×
10 /L. It is the characteristic •• Hemoglobin: decreased and usually below 13 g/dL.
9
feature. •• Total leukocyte count is increased (20–50 × 109/L).
CLL: lymphocytosis with •• Peripheral smear (Figs 11.1 and 11.2):

smudge cells in the
–– RBCs: normocytic normochromic anemia.
peripheral smear. Smudge
cells are fragile leukemic –– WBCs:
cells produced due to ◆◆ Differential leukocyte count shows lymphocytosis and constitutes more than 50% of the white cells.
rupture while making the ◆◆ Lymphocytes mature type—small with scant cytoplasm, nuclei round with clumped coarse
peripheral smear. chromatin (”soccer ball”/block-type chromatin). Nucleoli absent.
◆◆ Smudge cells or basket cells (fragile leukemic cells).
–– Platelets: initially normal count and later may be decreased.
Lymphocytes constitute
more than 30% of the
Bone Marrow
nucleated cells of the bone •• Cellularity: hypercellular marrow due to infiltration by mature lymphocytes.
marrow cells—diagnostic •• Erythropoiesis: normal.
feature of CLL. •• Myelopoiesis: normal.
•• Lymphocytic infiltrate: as the disease advances neoplastic lymphocytes replace the normal erythroid, myeloid
and megakaryocytic series in the bone marrow resulting in anemia, neutropenia and thrombocytopenia.
Immunophenotype
Tumor cells express the pan-B cell markers CD19 and CD20. CD5+ and CD23+ are distinctly
positive in CLL.
Lymph Node
•• Show loss of normal architecture
•• Diffuse infiltration by monomorphic, small, round lymphocytes
Fig. 11.1: Peripheral blood smears in chronic lymphocytic leukemia showing Fig. 11.2: Diagrammatic peripheral blood smears in chronic
numerous small lymphocytes (long arrows) and few smudge cells (short arrow) lymphocytic leukemia showing numerous small lymphocytes (long
arrows) and few smudge cells (short arrows)
Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma CHAPTER 11 75
•• Lymphocytes have nuclei with coarse chromatin and scanty cytoplasm CLL/SLL: lymph node with
•• Small, nodular aggregates of medium to large-sized lymphocytes known as proliferation centers proliferation centers are
or pseudo-follicles or growth centers and when found are pathognomonic for CLL/SLL. pathognomonic.
Course and prognosis: median survival rate is 4 to 6 years. They may progress to B cell
prolymphocytic transformation or into diffuse large B cell lymphoma (Richter syndrome).
HAIRY CELL LEUKEMIA

Definition Hairy cells have hair-like
cytoplasmic projections.
Uncommon neoplasm of small mature B cells having abundant cytoplasm with fine hair-
like cytoplasmic projections (hence the name hairy cell leukemia) when viewed under the
phase-contrast microscope.
Laboratory Findings HCL: it is B cell neoplasm

and involves peripheral
Peripheral Blood blood, bone marrow,
spleen and liver and
•• Hemoglobin: decreased. usually seen in old age.
•• Total leukocyte count: decreased (leukopenia).
•• Platelet count: decreased (less than 50 × 109/L)
•• Peripheral smear: pancytopenia

–– RBCs: normocytic normochromic.
–– WBCs: leukopenia with few hairy cells (Fig. 11.3).
–– Platelets: reduced.
Bone Marrow Aspiration

•• Dry tap
•• Hairy cells may be seen in the marrow Fig. 11.3: Hairy cell
•• Moderate to marked reduction in myeloid, erythroid and megakaryocytic cell lines.
Bone marrow trephine biopsy: neoplastic cells have “fried egg” or “honeycomb” appearance. HCL: bone marrow biopsy-
Reticulin stain shows marked increase of thin reticulin fibers surrounding neoplastic cells. hairy cells have fried egg
appearance.
Spleen
•• Enlarged due to leukemic infiltrate
•• Sinuses lined by hairy cells and grossly impart a beefy red appearance.
Immunophenotype and Molecular Characteristics Tartrate resistant acid

phosphatase (TRAP)
Express the CD20, CD22, CD11c and CD25 (the IL-2 receptor α-chain) positivity. Annexin A 1 positivity in the cytoplasm
is the most specific marker of hairy cell leukemia. is a characteristic feature
of HCL.
Clinical Features HCL: only leukemia

without lymphadenopathy.
•• Affects middle-aged to elderly men.
•• Male-to-female ratio of 5:1. HCL prognosis: indolent
•• Massive splenomegaly. course and prognosis is
•• Pancytopenia. excellent.
12 Plasma Cell Neoplasms
CHAPTER
Plasma cell neoplasms are

of B cell origin in which
INTRODUCTION
single clone of plasma cells
proliferate.
Definition
Plasma cell neoplasms are group of B cell neoplasms associated with the proliferation of single
clone (monoclonal) of immunoglobulin-secreting plasma cells (also known as dyscrasias).
Plasma cell neoplasms: Characteristics of Plasma Cell Neoplasms

tumor cells secrete
single type of complete Monoclonal neoplastic plasma cells secrete complete single type of immunoglobulin (Ig) or
or fragment of Ig fragment. Hence, are known as monoclonal gammopathies.
immunoglobulins. •• Serum: single Ig proteins detected as monoclonal spike [M protein (M for myeloma)] on
electrophoresis.
•• Urine: excess of free light chains is excreted in the urine as Bence-Jones (BJ) proteins.
Classification of Plasma Cell Neoplasms (Table 12.1)

TABLE 12.1: Classification of plasma cell neoplasms (WHO 2008)
•• Plasma cell myeloma
•• Plasmacytoma
•• Immunoglobulin deposition diseases
•• Monoclonal gammopathy of undetermined significance (MGUS)
•• Osteosclerotic myeloma (POEMS syndrome)
PLASMA CELL MYELOMA (MULTIPLE MYELOMA)

Definition
Multiple myeloma is a Plasma cell myeloma is a malignant, multifocal plasma cell neoplasm of the bone marrow
multifocal malignant associated with M-protein in the serum and/or urine.
tumor of plasma cell and
arises in the bone marrow.
•• Most common monoclonal gammopathy.
•• Presents as multiple tumor masses throughout the skeletal system.
Plasma Cell Neoplasms CHAPTER 12 77
Etiology Plasma cell neoplasms

arise from post-germinal
Risk Factors center B cells.
•• Genetic predisposition
•• Exposure to ionizing radiation
•• Chronic antigenic stimulation associated with chronic infections (HIV and chronic
osteomyelitis) and chronic inflammatory disorders (e.g. rheumatoid arthritis)
•• Exposure to chemicals like benzene, herbicides and insecticides.
Laboratory Findings Q. Write short notes on the

laboratory diagnosis of multiple
Peripheral Blood myeloma.
•• Hemoglobin: decreased and ranges from 6 to 10 g/dL.
•• Peripheral smear: MM: hypergammaglo–

–– RBCs: normocytic normochromic anemia, red blood cells show rouleaux formation (Fig. 12.1) due bulinemia is responsible
to increased immunoglobulins. for high ESR and rouleaux
formation seen in peri–
–– WBCs: normal. pheral smear.
•• ESR: high and is due to high gamma globulin (immunoglobulin) and rouleaux formation.
•• Bleeding time: increased.
Bone Marrow
•• Cellularity: hypercellular due to myeloma plasma (myeloma) cells (neoplastic plasma cells) (Figs 12.2 Bone marrow in MM:
and 12.3). hypercellular, and contains
•• Myeloma plasma cells: more than 30% are diagnostic. more than 30% neoplastic
–– Myeloma plasma cells are neoplastic plasma cells (Fig. 12.2), which are large oval cells having plasma cell.
abundant pale blue cytoplasm.
–– The nucleus is round to oval, eccentric and shows perinuclear clearing/hof. Myeloma plasma cells
–– The nuclear chromatin appears like a clock-face/spoke wheel. are commonly called as
–– These cells are usually uninucleated or may show binucleation. myeloma cells.
–– Other cells can also be seen in myeloma (Fig. 12.4).
•• Erythropoiesis: diminished and is normoblastic.
Fig. 12.1: RBCs showing rouleaux formation in the peripheral Fig. 12.2: Bone marrow aspirate in multiple myeloma. With numerous myeloma
blood plasma cells. Inset shows flame cell (left lower corner) and mott cell (right upper corner)
Serum Findings
•• Serum β2 microglobulin: prognostic marker and
high values signify poor prognosis.
•• Hypercalcemia
•• Renal function tests: blood urea, serum creatinine
and uric acid levels are raised with renal involvement.
•• Serum albumin: decreases in advance stages of the
disease.
Electrophoretic Studies on Serum and

Urine (Figs 12.5 and 12.6)
•• Monoclonal spikes in 80% to 90% of cases.
•• Raised monoclonal immunoglobulins in the blood.
Immunoglobulin may be IgG (most common)/IgD/
IgA/IgE type.
•• Light chains or Bence Jones (BJ) proteins in the urine
Fig. 12.3: Diagrammatic appearance of bone marrow in multiple
myeloma showing plasmablasts (short arrow) and plasma cells (long may be seen in 60% to 80% of cases. BJ protein may be
arrow) of κ or λ type of light chain.
Fig. 12.5: Serum electrophoresis showing normal pattern
Fig. 12.4: Diagrammatic appearance of the various cells that can be Fig. 12.6: Serum electrophoresis showing monoclonal
seen in bone marrow in multiple myeloma immunoglobulin (“M Band”) in multiple myeloma
Plasma Cell Neoplasms CHAPTER 12 79
Morphology of Organs Involved MM: IgG is the most

common immunoglobulin
•• Bone: destructive -punched-out lytic lesions (Fig. 12.7). secreted.
•• Renal lesions:
–– Myeloma kidney: light-chain cast of BJ protein damages renal tubules.
–– Amyloidosis of the AL type and leads to nephrotic syndrome.
◆◆ Hypercalcemia leads to nephrocalcinosis
◆◆ Prone to acute and chronic pyelonephritis
◆◆ Renal failure.
MM:
• Monoclonal
gammopathy peak 50
Clinical Manifestations (Fig. 12.8) to 60 years
• Multiple lytic lesions in
Onset: insidious. bones
Age and sex: affects old age between 50 and 60 years with slight male preponderance. • Hypercalcemia.
MM: involved bone shows

multiple punched out lytic
lesions.
Fig. 12.7: Skull X-ray showing multiple punched out lytic lesions
Fig. 12.8: Clinical features and laboratory findings in myeloma

MM: renal failure and The clinical features of multiple myeloma are
sepsis are common causes 1. Due to tumor cells causing bone lesions:
of death.
•• Resorption of bone: this results in pathologic fractures, chronic bone pain and
tenderness.
•• Compression: lesion in the vertebra may compress the spinal cord nerve root.
MM: higher levels of •• Hypercalcemia.
serum β2 microglobulin •• Pallor: due to anemia and result in weakness and fatigue.
are associated with poor
prognosis. 2. Production of M-proteins (increased immunoglobulins):
•• Bleeding tendency
•• Coagulation abnormalities
•• Amyloidosis of the AL type.
MM: prognosis— 3. Humoral immune deficiency: humoral immune deficiency predisposes to recurrent
progressive course with bacterial infections.
poor prognosis. 4. Renal disease: renal insufficiency, infections or nephrotic syndrome.
Extra-osseous
plasmacytoma is usually
PLASMACYTOMA
found in the upper Localized proliferation forms a single discrete plasma cell tumor in bone (usually) or soft
respiratory tract, especially tissue.
in the nasal cavity and
sinuses, nasopharynx and
•• Solitary plasmacytoma of bone (osseous plasmacytoma)
larynx. •• Extra-osseous (extramedullary) plasmacytoma
IMMUNOGLOBULIN DEPOSITION DISEASE

Primary Amyloidosis
Plasma cell neoplasm secretes abnormal immunoglobulin light chains, which may get
deposited in tissues and form a β-pleated sheet structure (AL amyloid).
MONOCLONAL GAMMOPATHY OF
UNCERTAIN SIGNIFICANCE (MGUS)
•• Presence of serum M protein concentration lower than 3 g/dL.
MGUS: prognosis—most of •• Bone marrow clonal plasma cells less than 10% in an asymptomatic patient.
the patients remain stable. •• Etiology: may represent an early stage of myeloma development.
•• Clinical manifestations: asymptomatic.
Lymphoid Neoplasms 13
CHAPTER
CLASSIFICATION OF LYMPHOID
NEOPLASMS (TABLE 13.1)
TABLE 13.1: WHO classification of the lymphoid neoplasms (2008) Majority (80 to 85%) of
lymphoid neoplasms are of
I. PRECURSOR LYMPHOID NEOPLASMS
B cell origin and remaining
B lymphoblastic leukemia/lymphoma of T cell/NK cell type.
T lymphoblastic leukemia/lymphoma
II. MATURE B CELL NEOPLASMS
Chronic lymphocytic leukemia/small lymphocytic lymphoma Lymphoid neoplasms:
B cell prolymphocytic leukemia most resemble some stage
Splenic B cell marginal zone lymphoma of B or T cell differentiation.
Hairy cell leukemia
Lymphoplasmacytic lymphoma
Heavy chain disease
Plasma cell neoplasm
Follicular lymphoma
Mantle cell lymphoma
Diffuse large B cell lymphoma
Burkitt lymphoma
Lymphoid neoplasms:
III. MATURE T AND NK CELL NEOPLASMS second most common
malignant tumor in HIV.
T cell prolymphocytic leukemia
T cell large granular lymphocytic leukemia
Mycosis fungoides
Sézary syndrome
Peripheral T cell lymphoma, NOS
Angioimmunoblastic T cell lymphoma
Anaplastic large cell lymphoma
Adult T cell leukemia/lymphoma
Extranodal NK/T cell lymphoma, nasal type Lymphoid neoplasms:
about 1/3rd arise from
IV. HODGKIN LYMPHOMA extranodal sites.
Classical Hodgkin lymphoma
–– Nodular sclerosis classical Hodgkin lymphoma
–– Mixed cellularity classical Hodgkin lymphoma T-cell lymphoblastic
–– Lymphocyte-rich classical Hodgkin lymphoma lymphoma or Burkitt
–– Lymphocyte depleted classical Hodgkin lymphoma lymphoma usually seen in
Nodular lymphocyte predominance Hodgkin lymphoma childhood.
WHO classification of lymphoid neoplasms depends on clinicopathological and immuno-

logical profile (Table 13.2) and has clinical and therapeutic importance.
TABLE 13.2: Cell type and its antigens detected by monoclonal antibodies
Cell type Antigen detected
T cell CD1, CD3, CD4, CD5, CD8
B cell CD10, CD19, CD20, CD21, CD23, CD79a
Monocyte or macrophage CD11c, CD13, CD14, CD15, CD33, CD64
NK cell CD16, CD56
Stem cell and progenitor cell CD34
All leukocytes CD45 (LCA)
Abbreviations: CD, cluster designation; NK, natural killer; LCA, leukocyte common antigen
Q. Write short notes on follicular FOLLICULAR LYMPHOMA (FL)

lymphoma.
Composed of follicle center (germinal center) B cells of lymphoid follicles (centrocytes and
centroblasts).
Morphology
FL: arises from follicle
center B cells. Gross
•• Involves lymph nodes, spleen and bone marrow.
•• Architecture of lymph node is lost; frequently infiltrate the perinodal tissue (Fig. 13.1).
Microscopy
FL: centrocytes and •• Follicular (nodular) growth pattern, neoplastic follicles are poorly defined (Fig. 13.2).
centroblasts form poorly
defined follicles. •• Two types of B cells.
–– Centrocytes (small cleaved cells)
◆◆ Cleaved nuclei
◆◆ Inconspicuous nucleoli.
–– Centroblasts (large non-cleaved cells)
FL: grade ranges from 1 to ◆◆ Round or oval nuclei with open nuclear (vesicular) chromatin
3. Grade 1 with less than 5
centroblasts/hpf and grade ◆◆ Multiple (1 to 3) nucleoli.
3 with more than 15/hpf. ◆◆ Usually 3 times the size of lymphocyte.
Fig. 13.1: Diagrammatic appearance of follicular lym- Fig. 13.2: Follicular lymphoma shows nodular
phoma. Neoplastic follicles are seen in both the cortex aggregates of malignant lymphoid cells
and medulla and infiltration of the perinodal tissue
Lymphoid Neoplasms CHAPTER 13 83
Immunophenotype: expresses CD19, CD20 (pan-B cell markers), CD10 (CALLA), surface FL: peripheral blood smear
immunoglobulin and BCL2 protein. may show lymphocytosis
[less than 20 × 109/L
Cytogenetics and molecular genetics: t (14; 18) (q32:q21), with IgH and BCL2 as partner (20,000/cu mm)].
genes and leads to constitutive overexpression of BCL2 protein.
FL: bone marrow involved
in 85%.
Clinical Features
•• Peak in sixth and seventh decades.
FL: prognosis indolent.
•• Generalized lymphadenopathy.
DIFFUSE LARGE B CELL LYMPHOMA (DLBCL) DLBCL: aggressive, diffuse

large B cell neoplasm.
Heterogeneous group of aggressive, neoplasm of large B cell with diffuse growth pattern.
Constitutes about 20 to 30% of NHL and 60% to 70% of aggressive lymphoid neoplasms.
Microscopy DLBCL may involve lymph

nodes or extranodal sites.
•• Loss of lymph node architecture with diffuse growth pattern.
•• Neoplastic cells:
–– Large round or oval cells, 4 to 5 times of a small lymphocyte.
–– Moderate pale or basophilic cytoplasm
–– Nucleus equals or larger than the nucleus of a macrophage with different appearances.
Immunophenotype
•• Express pan-B cell markers such as CD19, CD20, CD22 and CD79a.
•• Also express germinal center markers like CD10 and BCL6.
•• Negative for TdT.
Cytogenetics and Molecular Profile

•• Translocation of BCL2 gene: t (14; 18) translocation
•• Mutations of the BCL6 gene.
Clinical Features DLBCL: prognosis

aggressive and rapidly fatal
•• More common between 65 and 70 years of age. if untreated.
•• Slight male preponderance.
•• Rapidly enlarging mass at a single or multiple nodal or extranodal sites.
BURKITT LYMPHOMA (BL) Q. Write short notes on Burkitt

lymphoma.
•• Highly aggressive B cell neoplasm, often presents as extranodal lymphoma or as an acute
leukemia.
•• Composed of medium-sized, monomorphic lymphoid cells with basophilic vacuolated
cytoplasm.
Clinical Variants
•• Endemic (African) Burkitt lymphoma (BL):
–– Occurs in Africa, affects children and adolescents.
–– Associated with Epstein-Barr virus infection and malaria.
BL: aggressive B cell –– Usually involves the jaw and present as a mandibular mass.
lymphoma, 3 clinical
variants. Endemic: involves •• Sporadic (nonendemic) BL:
jaw and associated with –– Occurs in children or young adults.
EBV. –– Abdominal mass and involves ileocecum and peritoneum.
•• Immunodeficiency-associated (HIV) BL:
–– Involves lymph nodes and bone marrow.
BL: medium sized B cells. Microscopy

Starry sky pattern.
•• Burkitt lymphomas, irrespective of the categories, are histologically similar.
•• Lymph node shows loss of architecture.
•• Involved tissues show diffuse infiltrate of monotonous medium-sized lymphoid cells (Figs
13.3 and 13.4).
•• Appearance of neoplastic lymphoid cells:
–– Medium-sized cells.
–– Round or oval nuclei having clumped coarse chromatin with several (2 to 5) nucleoli.
–– Moderate amount of deeply basophilic cytoplasm, multiple, small, round lipoid (clear)
vacuoles which stain positive with oil red O.
–– Numerous mitotic figures.
•• Starry sky pattern: tumor cells undergo apoptosis and nuclear remnants of these apoptotic
cells are phagocytosed and cleared by benign macrophages. These macrophages in the
background of lymphoid cells creates “starry sky” appearance (Figs 13.3 and 13.4).
Fig. 13.3: Burkitt lymphoma composed of medium-sized lymphoid cells admixed Fig. 13.4: Diagrammatic appearance of Burkitt lymphoma
with benign macrophages giving a “starry sky” appearance composed of medium-sized lymphoid cells admixed with
benign macrophages giving a “starry sky” appearance
Lymphoid Neoplasms CHAPTER 13 85
Immunophenotype
•• Express surface IgM, monotypic κ or λ light chain.
•• Positive for common B cell antigens (CD19, CD20, and CD22).
•• Positive for CD10 and BCL6.
•• BCL2 negative.
Cytogenetic and Molecular

Genetic Features (Fig. 13.5)
Translocations of c-MYCgene BL: translocation of c-MYC
gene.
•• MYC (c-MYC) is a proto-oncogene-on chromosome 8.
•• Most common translocation t (8:14) (q24; q32).
•• Translocations of c-MYC gene, converts proto-oncogene into MYC oncogene, which
leads to overexpression of MYC protein (oncoprotein). This causes uncontrolled cell BL: prognosis—very
proliferation and stimulation of apoptosis. aggressive but responds
•• Mutations inactivate p53. well chemotherapy.
•• Poor prognostic factors:

–– Involvement of blood, bone marrow and central nervous system.
–– Bulk of the disease-unresected tumor of more than 10 cm in diameter.
–– High serum LDH levels.
–– Presence of residual disease after excision.
Fig. 13.5: Chromosomal translocation and activated MYC oncogene in Burkitt lymphoma
MATURE T CELL AND NK CELL NEOPLASMS Peripheral T cell tumors

constitute less than 15% of
Peripheral T Cell Lymphoma (PTCL), NOS non-Hodgkin lymphomas.
NK cell tumors very rare.
Mainly involves lymph node.
PTCL: clinical features Microscopy

• Fifth to seventh decade.
• Generalized
•• Lymph node with effacement of the normal architecture.
lymphadenopathy. •• Paracortical or diffuse infiltration by neoplastic T cells.
•• Neoplastic T cells
–– Small, intermediate to large cells with sparse or abundant; clear, eosinophilic or
basophilic.
–– Vesicular or hyperchromatic nuclei, prominent nucleoli.
PTCL: prognosis—highly Immunophenotype

aggressive with a poor
response to therapy.
•• Lack TdT (expressed by immature T cells).
•• Express pan-T cell-CD2, C3, CD5 and either α β or γ δ T cell receptors (TCR).
Mycosis fungoides and Mycosis Fungoides

Sézary syndrome: T cell
neoplasms with skin •• Cutaneous T cell lymphoma
involvement. •• Lymphoid cells with irregular nuclear outlines
•• Limited to skin.
Mycosis fungoides has
three stages:
Age: most are adults or elderly.
1. Patch stage
2. Plaque stage
3. Tumor stage. Microscopy
Immunophenotype: •• Epidermis (epidermotropism) and upper dermis is infiltrated by neoplastic T cells.
Express pan-T-CD2+, CD3+ •• Groups of neoplastic cells in the epidermis—Pautrier’s microabscess.
and CD5. •• Tumor cells have convoluted (cerebriform) nuclear contours.
Sézary cells are neoplastic Sézary Syndrome

T cells with cerebriform
nuclei. Rare disease and is defined by the triad namely:
1. Widespread exfoliative erythroderma
Immunophenotype: tumor
2. Generalized lymphadenopathy
cells express-CD2+, CD3+
and CD5+. 3. Presence of characteristic Sézary cells in the skin, lymph nodes and peripheral blood.
Prognosis: aggressive disease and most die of opportunistic infections.

Hodgkin
Lymphomas
14
CHAPTER
DEFINITION Hodgkin
lymphoma synonym:
HL: Malignant lymphoid neoplasms with following characteristics: Hodgkin disease.
• Minority (1–3%) of specific neoplastic cells (Hodgkin cells and Reed-Sternberg cells).
• Majority background of reactive non-neoplastic cells.
• Usually involves lymph nodes.
• Majority occurs in young adults.
CLASSIFICATION (TABLE 14.1) Q. Classify Hodgkin lymphoma.
Hodgkin lymphoma (HL) is broadly divided into two types, which differ in clinical features,
behavior, morphology and immunophenotype.
Cell of Origin and Immunophenotype

• Classical Hodgkin lymphoma
– Cell of origin: germinal center or post-germinal center B cell
– Immunophenotype: CD15 and CD 30 positive
HL is mainly divided into:
• Nodular lymphocyte predominant Hodgkin lymphoma 1. Classical and
– Cell of origin: germinal center B cell at the centroblastic stage of differentiation. 2. Nodular lymphocytic
– Immunophenotype: CD15 and CD30 negative. Hodgkin lymphoma.
TABLE 14.1: WHO classification (2008) of Hodgkin lymphoma Classical HL: CD15 + and
CD30 +,
• Classical Hodgkin lymphoma (CHL)
NLPHL: CD15-,CD 30-,
– Nodular sclerosis classical Hodgkin lymphoma (NSCHL) CD20+, and CD 45+.
– Mixed cellularity classical Hodgkin lymphoma (MCCHL)
– Lymphocyte-rich classical Hodgkin lymphoma (LRCHL)
– Lymphocyte depleted classical Hodgkin lymphoma (LDCHL)
• Nodular lymphocyte predominant Hodgkin lymphoma (NLPHL)
Q. Write short note on RS cell and MORPHOLOGY OF NEOPLASTIC CELLS

its variants.
Reed-Sternberg (RS) Cells are neoplastic cells (Fig. 14.1) pathognomonic of Hodgkin
lymphoma.
Appearance and description of diagnostic Reed-Sternberg cells and its variants are shown
in Figure 14.2.
Various types of cells found in Hodgkin lymphoma are listed in Table 14.2.
TABLE 14.2: Types of cells found in Hodgkin lymphoma

HL: majority are non- Non-neoplastic cells Neoplastic cells
neoplastic cells and
minority are neoplastic • Reactive lymphocytes • Reed-Sternberg cells (classical)
cells. • Macrophages/histiocytes • Variants
– Granulocytes – Mononuclear
– Eosinophils – Lacunar
– Neutrophils – Mummified
• Plasma cells – Anaplastic/pleomorphic
– Lymphocyte predominant (LP) cell/popcorn
CLASSICAL HODGKIN LYMPHOMA

Classical Hodgkin lymphoma (CHL) account for 95% of Hodgkin lymphomas and has 4 subtypes.
Q. Write short note on nodular Nodular Sclerosis Classical

sclerosis HL.
Hodgkin Lymphoma (NSCHL)
Nodular sclerosis is the Subtype of CHL characterized by collagen bands that surround nodules and have lacunar cell
most common subtype variant of Reed-Sternberg cells.
of CHL. Lacunar cells are • Most common: 40% to 70% of cases.
commonly seen.
• Most between 20 and 30 years of age with equal frequency in males and females.
• Rarely associated with EBV.
• Involves mediastinal lymph nodes.
Fig. 14.1: Microscopic appearance of Hodgkin lymphoma showing RS

cells (short arrow and inset) and Hodgkin cells (long arrow) within the
background of mixed population of reactive cells
Hodgkin Lymphomas CHAPTER 14 89
Classical RS cell is
binucleated with owl-eyed
nuclei having mirror image
appearance.
Lacunar cell has clear

cytoplasm and seen in
nodular sclerosis CHL.
LP/popcorn cell is seen

in nodular lymphocyte
predominant HL.
Fig. 14.2: Diagrammatic appearances and characteristic features of Reed-Sternberg cells and its variants
Microscopy of NSCHL (Fig. 14.3) NSCHL

• Nodules separated by
• Loss of lymph node architecture. broad bands of collagen
• Sclerosis and nodules: broad collagen bands (sclerosis) divide the lymphoid tissue into • CD15+,CD30+, EBV-ve
nodules of varying sizes and shapes. and CD 45-ve.
• Presence of lacunar cell.
• Background: small T lymphocytes, eosinophils, plasma cells, and macrophages.
Immunophenotype NSCHL Prognosis: better

than other types of CHL,
• RS cells are CD15+ and CD30+; CD45- and T cell markers negative. with a cure rate of 80% to
• EBV negative. 85%.
Fig. 14.3: Nodular sclerosis classical Hodgkin lymphoma with nodules Fig. 14.4: Mixed cellularity classical Hodgkin lymphoma with classical
separated by bands of collagen. Also seen are lacunar cells and RS cells RS cells, Hodgkin cells in the background of mixed cellular population
in each nodule within the background of lymphocytes, eosinophils, consisting of lymphocytes, eosinophils, plasma cells and macrophages
plasma cells and macrophages
Q. Write short note on Mixed Mixed Cellularity Classical Hodgkin Lymphoma (MCCHL)
cellularity HL.
• Second common subtype: 20% to 25% of cases
• More common in males
• Strongly associated with EBV
• Older age, with systemic symptoms (such as night sweats and weight loss) and advanced
tumor stage
MCCHL: scattered classical
RS cells and mixed • Involves peripheral lymph nodes.
inflammatory background,
CD15+, CD30+ and EBV+.
Microscopy of MCCHL (Fig. 14.4)
• Lymph node architecture obliterated
• Plenty of Reed-Sternberg cells and Hodgkin cells
MCCHL: prognosis—very • Back ground: small lymphocytes, eosinophils (sometimes numerous), neutrophils,
good.
plasma cells and benign macrophages (histiocytes).
Immunophenotype: RS cells are CD15+, CD30+ and EBV+ (about 70%).
Lymphocyte-rich Classical Hodgkin Lymphoma (LRCHL)

• Subtype of classical Hodgkin lymphoma with scattered Hodgkin and RS cells.
• Uncommon—about 5% of classical HL.
• More in elderly patients, associated with EBV in 40% of cases.
• Involves peripheral lymph nodes.
Microscopy of LRCHL (Fig. 14.5) LRCHL:

• Uncommon
• Growth patterns: may show two patterns • Few RS cells
– Nodular—common • Abundant lymphocytes
– Diffuse—rare • CD15+, CD30+, CD45-
and CD20-.
• Only few Reed-Sternberg cells and Hodgkin cells
• Background: abundant reactive small lymphocytes.
Immunophenotype: CD45–, CD20–, CD15+ and CD30+. LRCHL: prognosis—good

to excellent prognosis.
Lymphocyte-depleted Classical
Hodgkin Lymphoma (LDCHL)
Subtype of classical Hodgkin lymphoma rich in Hodgkin and RS cells in a background LDCHL:
depleted in non-neoplastic lymphocytes. • Rarest
• Rarest—less than 5% of cases • Paucity of lymphocytes
• Plenty of RS cells
• Predominantly in older, HIV-positive patients, often EBV-associated (over 90%) • CD15+, CD30+; majority
• Predominantly retroperitoneal lymph nodes, abdominal organs and bone marrow. are EBV+.
Microscopy of LDCHL (Fig. 14.6)

• Paucity of lymphocytes.
• Plenty of RS cells or their anaplastic/pleomorphic variants.
• Histological types
– Reticular: numerous Hodgkin and RS cells with depletion of lymphocytes.
– Diffuse sclerosis/fibrosis: hypocellular infiltrate containing bizarre RS cells with fine
fibrosis. LDCHL: prognosis—
outcome less favorable
Immunophenotype: RS cells are CD15+, CD30+; majority are EBV+. than with other subtypes.
Fig. 14.5: Lymphocyte-rich classical Hodgkin lymphoma. One RS cell is Fig. 14.6: Lymphocyte-depleted classical Hodgkin lymphoma with
seen in a background of many small lymphocytes and few histiocytes the pleomorphic variant of RS cells surrounded by fibrous tissue
NODULAR LYMPHOCYTE PREDOMINANT

HODGKIN LYMPHOMA (NLPHL)
• Uncommon—5% of all Hodgkin lymphomas.
• Not associated with EBV.
• Majority males, usually 30–50 year of age.
• Involves mainly cervical or axillary lymph nodes.
NLPHL: Microscopy of NLPHL (Fig. 14.7)

• Uncommon
• Loss of lymph node architecture.
• Abundant lymphocytes
• LP cells • Nodular and/or diffuse infiltrate of abundant small lymphocytes with histiocytes and
• No Hodgkin/RS cells scattered LP cells.
• CD20+, CD 45+ and
• Lymphocyte predominant cells (LP cells)/"popcorn" cells (Fig. 14.2):
CD15 -, C30- and
EB -ve. – Specific to NLPHL.
– Large with relatively abundant, pale cytoplasm.
– Single large delicate multilobulated nucleus or folded nuclei resembling bubbly outlines
of popcorn kernels.
– One or more inconspicuous nucleoli.
• Hodgkin and RS cells are not found.
Immunophenotype: LP cell are CD20+, CD 45+ and CD15–, C30– and EBV–ve. Express
BCL6.
NLPHL: prognosis—more
likely to recur than the
classical subtypes, but the
prognosis is very good.
Fig. 14.7: Nodular lymphocyte predominant Hodgkin lymphoma with ‘popcorn’

cells in a background of reactive lymphocytes and few macrophages
ETIOLOGY AND PATHOGENESIS OF

HODGKIN LYMPHOMA
• EBV: previous EBV infection (infectious mononucleosis) ↑ risk of HL.
• Genetic factors: HLA-B18 higher in HL.
• Immune status: HL more frequent in immunocompromised patients and autoimmune
diseases (e.g. rheumatoid arthritis).
Pathogenesis (Fig. 14.8)

• EBV and HL: HL is associated with EBV infection.
• Activation of nuclear factor (NF-κB) common event in classical HL → rescue germinal
center B cells from apoptosis → produces Reed-Sternberg cells.
• Accumulation of reactive cells in response to cytokines (such as IL-5, IL-6 and TGF-β) and
chemokines secreted by Reed-Sternberg cells.
LABORATORY FINDINGS
• Peripheral smear: ESR: raised.
– RBCs: normocytic normochromic anemia.
– WBCs: leukocytosis occurs in 1/3rd of the patients. Eosinophilia is frequent. Bone marrow: involved in
– Platelets: normal or increased. the later stages.
Fine Needle Aspiration Cytology (FNAC)

RS cells/its variants against a background of inflammatory cells (depending on the subtype).
Clinical features:
• Painless enlargement of
lymph nodes.
• Systemic/constitutional
symptoms: fever, night
sweats and weight loss.
HL: Pel-Ebstein fever

is characterized by
Fig. 14.8: Pathogenetic mechanism and interaction of various cell types in Hodgkin lymphoma alternating bouts of fever
followed by remissions.
Spread
• Mainly by contiguity
• First nodal disease → then splenic disease, hepatic disease → and finally marrow
involvement and extranodal disease.
STAGING OF HODGKIN LYMPHOMA (TABLE 14.3)

TABLE 14.3: Clinical staging of Hodgkin lymphomas (Cotswold revision of Ann Arbor staging
classification)
Stage Definition
I Involvement of a single lymph node region or lymphoid structure (e.g. spleen, Waldeyer ring, thymus)
II Involvement of two or more lymph node regions on the same side of the diaphragm (the mediastinum
is a single site; hilar lymph nodes are lateralized); the number of anatomic sites should be indicated by
suffix (e.g. II3)
III Involvement of lymph node regions or structures on both sides of the diaphragm
III1 With or without splenic, hilar, celiac or portal nodes
III2 With para-aortic, iliac or mesenteric nodes
IV Involvement of extranodal site(s) beyond those designated E
E—involvement of a single extranodal site, or contiguous or proximal to known nodal site of disease
DIFFERENCES BETWEEN HODGKIN

LYMPHOMA AND NON-HODGKIN LYMPHOMA
HL differs from NHL in several respects and their main differences are shown in Table 14.4.
Q. List the differences between TABLE 14.4: Differences between Hodgkin and non-Hodgkin lymphomas
HL and NHL. Sl. No. Characteristics Hodgkin lymphoma Non-Hodgkin lymphoma
1. Site of involvement Arises in a single node or Mainly involves multiple
chain of nodes (cervical, peripheral nodes
mediastinal, para-aortic)
2. Pattern of spread Orderly spread by contiguity Noncontiguous spread in an
HL: extranodal in a predictable fashion unpredictable fashion
involvement uncommon. 3. Mesenteric nodes and Waldeyer ring Rarely involved Commonly involved
4. Extranodal involvement Uncommon Common
5. Characteristic of neoplastic cells Neoplastic cells—Hodgkin Neoplastic cells form the
or Reed-Sternberg cells form major tumor cell mass
minor tumor cell mass (1–5%)
Langerhans Cell
Histiocytosis/ 15
Histiocytosis X CHAPTER
INTRODUCTION
•• Histiocytic and dendritic cell neoplasms
•• Clonal proliferative disorder arising from Langerhans cells
•• Langerhans cell histiocytosis (LCH) spectrum ranges from unifocal to multifocal and
unisystem to multisystem disease.
MORPHOLOGY
•• Light microscopy: the characteristic feature is proliferations of Langerhans cells.
–– These are large cells 10–15 μm in diameter, moderate slightly eosinophilic cytoplasm
folded, indented, grooved or lobulated nucleus having fine chromatin.
–– Background: mixed background of eosinophils, histiocytes (mononuclear and Langerhans cell contains
multinuclear), neutrophils and small lymphocytes. pathognomonic Birbeck
•• Electron microscopy: Langerhans cell contains pathognomonic Birbeck granules—tennis granules.
racket-like shape, with a zipper-like appearance.
•• Immunological markers: express CD1a, langerin and S-100 protein.
LABORATORY FINDINGS
•• Peripheral blood: pancytopenia (anemia, neutropenia and thrombocytopenia).
•• Bone marrow: extensive infiltration by histiocytes.
Prognosis: depends on the age at presentation, extent of disease and rate of progression.
Groups: depending on the site involved and distribution of lesion, LCH can be divided into
three groups (Table 15.1).
TABLE 15.1: Types of Langerhans cell histiocytosis

Terminology/site Involved tissue/ organ Clinical features
Eosinophilic granuloma Bone and adjacent soft issue (skull, Usually seen in older children or
Localized to a single site/solitary femur, vertebra, pelvic bones and adults. Presents with lytic bone
(unifocal) ribs). Less commonly lymph nodes lesion
Hand-Schüller-Christian disease Usually bone and soft issue Usually seen in young children
Multiple sites within a single Multiple destructive bone lesions
system (multifocal unisystem) with adjacent soft tissue masses
Letterer-Siwe disease Skin, bone, liver, spleen and bone Usually seen in infants.
Disseminated and multisystemic marrow Present with fever, cytopenias,
disease (multifocal multisystem skin and bone lesions and
disease) hepatosplenomegaly
SECTION 3
Disorders of
Hemostasis
Disorders of
Primary Hemostasis
16
CHAPTER
NORMAL HEMOSTASIS Platelet sequence in

hemostasis: platelet
•• Hemostasis is the body’s response to vascular damage/injury. adhesion → release of
•• Includes several sequences of events at the site of vascular injury. They are as follows: granule contents →
platelet aggregation
→ primary (temporary)
hemostatic plug →
Primary Hemostatic Plug activation of coagulation
system → fibrin →
Platelet adhere to subendothelial structures at the site of injury. The platelets change their secondary (permanent)
shape and release granule contents. The released contents cause platelet aggregation and hemostatic plug.
form primary hemostatic plug.
Secondary Hemostatic Plug

Exposure of tissue factor at the site of vascular injury activates the extrinsic coagulation
system. The fibrin formed develops into a secondary hemostatic plug.
CLASSIFICATION OF HEMOSTATIC Q. Classify bleeding disorders.
DISORDERS (TABLE 16.1)

1. Bleeding disorders (hemorrhagic disorders/hemorrhagic diathesis): bleeding disorders
have an abnormal tendency to bleed (hemorrhage) due to failure of hemostasis.
2. Thrombotic disorders: they cause thrombus formation.
BLEEDING DISORDERS CAUSED BY

VESSEL WALL ABNORMALITIES
Vascular purpura (nonthrombocytopenic purpura) is group of disorders of blood vessels
that results in bleeding. They should be distinguished from bleeding disorders due to
abnormalities of platelets.
100 SECTION 3 Disorders of Hemostasis
Hemostatic disorders TABLE 16.1: Classification of disorders of hemostasis

are broadly classified as
1. Bleeding disorders
bleeding disorders and
thrombotic disorders. –– Disorders of primary hemostasis
◆ Vessel wall abnormalities
◊ Congenital, e.g. Ehlers-Danlos syndrome
Bleeding disorders may be
◊ Acquired, e.g. Henoch-Schönlein purpura
due to
• Diseases of blood ◆ Platelet abnormalities
vessels ◊ Quantitative: thrombocytopenia (e.g. ITP, drug-induced, congenital)
• Platelet disorders ◊ Qualitative: platelet function disorders
• Coagulation disorders. -- Inherited, e.g. Glanzmann thrombasthenia, Wiskott-Aldrich syndrome, Bernard Soulier
syndrome
-- Acquired, e.g. Uremia, drugs
Vascular purpuras –– Disorders of coagulation system (disorders of secondary hemostasis)
are also known as ◆ Congenital: hemophilia A, B; von Willebrand disease; other coagulation factor deficiencies [XI, VII, II, V, X]
nonthrombocytopenic ◆ Acquired: vitamin K deficiency, liver disease, disseminated intravascular coagulation
purpuras.
2. Thrombotic disorders
–– Inherited
◆ Deficiency of antithrombotic factors: antithrombin III deficiency, protein C deficiency, protein S
deficiency
◆ Increased prothrombotic factors: activated protein C (APC) resistance (Factor V mutation/factor
V Leiden)
◆ Prothrombin (G20210A mutation)
–– Acquired: fibrinolytic system defects
Classification of bleeding disorders caused by vessel wall abnormalities are presented in

Table 16.2.
TABLE 16.2: Classification of bleeding disorders caused by vessel wall abnormalities

Acquired disorders
Senile purpura is due to 1. Due to decreased amount of connective tissue

vessel instability. –– Senile purpura
–– Scurvy
–– Cushing syndrome and steroid therapy
2. Due to vasculitis
Henoch-Schönlein purpura
–– Henoch-Schönlein purpura
is characterized by
–– Infections
hypersensitivity vasculitis
–– Drug reactions
and palpable purpura.
3. Associated with plasma cell neoplasms
–– Amyloidosis
4. Miscellaneous
–– Simple easy bruising
Congenital/inherited disorders
–– Hereditary hemorrhagic telangiectasia
–– Ehlers-Danlos syndrome
–– Marfan syndrome
BLEEDING DISORDERS DUE TO

ABNORMALITIES OF PLATELET
Classification of Platelet Disorders (Table 16.3)
THROMBOCYTOPENIA
•• Decrease in the platelet count below the lower limit of 150,000/cu mm (150 × 109/L).
Disorders of Primary Hemostasis CHAPTER 16 101
TABLE 16.3: Classification of platelet disorders

Quantitative platelet disorders
•• Thrombocytopenia
– Increased destruction – Decreased production
– Sequestration – Dilutional
•• Thrombocytosis
Qualitative platelet disorders
•• Hereditary
– Defective adhesion of platelets – Disorders of platelet secretion
– Defective platelet aggregation
•• Acquired
Clinical Features of Thrombocytopenia

•• Cutaneous bleeding appears as pinpoint hemorrhages (petechiae) and ecchymoses. Petechiae are pinpoint
•• Mucosal bleeding. hemorrhages seen only
•• Intracranial bleed (subarachnoid and intracerebral hemorrhage) rare but serious. with thrombocytopenia.
Severity of Bleeding Intracranial bleeding

•• Post-traumatic bleeding—when the platelet count is 20,000 to 50,000/cu mm occurs-when platelet
count is <10,000/cu mm.
•• Spontaneous bleeding—when the platelet count falls below 20,000/cu mm
•• Intracranial bleeding—when platelet count is <10,000/cu mm.
Causes of Thrombocytopenia (Table 16.4)

TABLE 16.4: Causes of thrombocytopenia
1. Increased platelet destruction
–– Immune mediated
◆◆ Autoimmune
◊ Primary: immune thrombocytopenic purpura (acute and chronic)
◊ Secondary: systemic lupus erythematosus, B cell lymphoid neoplasms ITP is the most common
◆◆ Alloimmune: post-transfusion or pregnancy form of thrombocytopenia.
◆◆ Drug-induced: quinidine, heparin, sulfa compounds
◆◆ Infections: HIV infection, infectious mononucleosis, cytomegalovirus
–– Non-immune mediated
◆◆ Disseminated intravascular coagulation
◆◆ Thrombotic thrombocytopenic purpura, hemolytic uremic syndrome
◆◆ Mechanical destruction: prosthetic heart valves, malignant hypertension
◆◆ Microangiopathic hemolytic anemias
2. Decreased production of platelets
–– Generalized primary diseases of bone marrow: aplastic anemia (congenital and acquired)
–– Bone marrow invasion/infiltration: leukemia, disseminated cancer
–– Selective impairment of platelet production
◆◆ Drug-induced: alcohol, thiazides, cytotoxic drugs
◆◆ Infections: measles, human immunodeficiency virus (HIV)
–– Ineffective megakaryopoiesis
◆◆ Megaloblastic anemia
◆◆ Myelodysplastic syndromes
3. Sequestration
–– Hypersplenism
4. Dilutional
IMMUNE THROMBOCYTOPENIC PURPURA

•• Most common form of thrombocytopenia.
•• Due to increased destruction of platelets by immune mechanisms—mainly autoimmune
mechanism.
Types of Immune Thrombocytopenic Purpura (ITP)

Acute Immune Thrombocytopenic Purpura
Acute ITP is seen mainly •• Self-limited disease.
in children between 2 to •• Children: 2 to 4 years and seen equally in both sexes.
4 years.
•• Presents 1 to 3 weeks after viral (measles, rubella, EBV)
infection.
Acute ITP: autoimmune •• Platelet destruction by antiplatelet autoantibodies.
disease, sudden onset, •• Platelet count is decreased, sometimes even below
shorter duration and 10,000/cu mm (10 × 109/L).
usually resolves within 6
months.
Clinical Features
•• Sudden onset.
•• Petechiae, gum bleeding, epistaxis and mild fever.
•• Usually resolve spontaneously within 6 months.
•• Excellent prognosis.
Chronic ITP: autoimmune Chronic Immune Thrombocytopenic Purpura

disease and the antibodies
•• Persistent thrombocytopenia for more than 6 to 12
are directed against
glycoprotein IIb/III a of months
platelets. •• Indolent, females are more affected than males (F:M=3:1).
•• More common and usually seen in adults (20 to 40
years).
Pathogenesis of ITP (Fig. 16.1)

•• Autoimmune disorder characterized by formation of
antiplatelet antibodies, directed against membrane
glycoproteins (most often IIb-IIIa or Ib-IX of platelets).
•• Antiplatelet antibodies in about 80% of patients and are
of the IgG type.
•• Antiplatelet antibodies act as opsonins and are
recognized by IgG Fc receptors present on mononuclear
Spleen is the major site of phagocytes of RE system (mainly spleen) and are
destruction of platelets destroyed there resulting in thrombocytopenia.
and important site of •• Splenectomy causes marked improvement in 75% to
autoantibody synthesis.
80% of patients.
Clinical Features
ITP: splenomegaly and •• More common in females (F:M ratio is 3:1).
lymphadenopathy are
uncommon and in their •• Age between 20 and 40 years.
presence one should •• Clinical features are due to thrombocytopenia: skin
consider the diagnosis bleeding, mucosal bleeding, menorrhagia in females, Fig. 16.1: Pathogenesis of idiopathic
other than ITP. etc. thrombocytopenic purpura
Disorders of Primary Hemostasis CHAPTER 16 103
Laboratory Findings Q. Write short notes on laboratory

findings in ITP.
Peripheral Blood
•• Platelet count: markedly reduced—below 80,000/cu mm (80 × 109/L).
•• Hemoglobin: ranges from 7 to 12 g/dL.
•• Peripheral smear ITP: platelets markedly

–– Platelets: markedly reduced (thrombocytopenia) and abnormally large sized platelets (mega- reduced below 80,000/
cu mm.
thrombocytes/giant platelets).
–– RBCs: chronic blood loss (e.g. menorrhagia) due to ITP may lead to microcytic hypochromic
anemia.
–– WBCs: normal.
Bone marrow in chronic

Bone Marrow ITP shows megakaryocytic
•• Cellularity: hypercellular. hyperplasia with immature
•• Megakaryopoiesis: megakaryocytes.
–– Moderate increase in number (Fig. 16.2) of both immature and mature forms of megakaryocytes.
–– Immature megakaryocytes predominate-large nonlobulated single nuclei and basophilic
cytoplasm (Fig. 16.3).
•• Erythropoiesis:
ITP: bone marrow—
–– Prolonged bleeding may cause anemia leading to normoblastic erythroid hyperplasia. decreased megakaryo-
–– Constant bleeding leads to iron deficiency and micronormoblastic erythroid hyperplasia. cytes—against the
•• Myelopoiesis: normal. diagnosis of ITP.
•• Storage iron: severe and chronic bleeding causes iron deficiency with reduced iron stores.
•• Bleeding time (BT): prolonged, but PT and PTT are normal. ITP: bleeding time
prolonged
•• Tourniquet test: positive. PT and APTT normal.
•• Clotting time (CT): normal.
•• Tests for platelet autoantibodies: may be positive.
•• Spleen: normal size.
1 2
3
Fig. 16.2: Bone marrow in ITP showing moderate increase in number of Fig. 16.3: Diagrammatic appearance of megakaryocytes in
megakaryocytes (arrows) different stages of maturation (1-immature to 3-mature)
THROMBOCYTOSIS
Platelet count more than 4,50,000/cu mm is known as thrombocytosis.
Causes: various causes of thrombocytosis are shown in Table 16.5.
TABLE 16.5: Causes of thrombocytosis

Idiopathic/primary (autonomous production)
•• Essential thrombocytosis
•• Polycythemia vera
•• Chronic myeloid leukemia
Secondary (reactive thrombocytosis)
•• Iron deficiency
•• Malignancy
•• Following hemorrhage
•• Following splenectomy
QUALITATIVE PLATELET DISORDERS

Classification of platelet functional (qualitative) disorders are presented in Figure 16.4 and
Table 16.6.
Fig. 16.4: Functional disorders of platelet
TABLE 16.6: Classification of platelet functional (qualitative) disorders

A. Hereditary
Aspirin blocks the cyclo- 1. Disorders of platelet adhesion: Bernard-Soulier syndrome
oxygenase enzyme of 2. Disorders of platelet secretion: storage pool deficiency
platelets and prevents 3. Disorders of platelet aggregation: Glanzmann thrombasthenia
aggregation of platelets.
B. Acquired
1. Drugs: aspirin, nonsteroidal anti-inflammatory drugs (NSAIDs), dipyridamole, sulfinpyrazone
2. Renal failure: uremia
3. Hematologic malignancies: myeloproliferative neoplasms and myelodysplastic syndromes
17
Bleeding Disorders:
Due to Abnormalities of
Coagulation/Clotting
Factor CHAPTER
INTRODUCTION
Bleeding due to coagulation disorders must be distinguished from those due to platelet/
vascular disorders (Table 17.1).
TABLE 17.1: Distinguishing patterns of bleeding in platelet/vascular and coagulation disorders

Characteristics Platelet/vascular disorders Coagulation disorders
Onset Spontaneous and develops Delayed bleeding after trauma/
immediately after trauma/surgery surgery
Type of lesion Petechiae, ecchymoses Hematomas
Sites Skin, mucous membrane Deep tissues
•• Mucous membrane Common from nose, mouth, Uncommon except from
gastrointestinal and genitourinary gastrointestinal or genitourinary tract
tracts
•• Into the joint Absent Common in severe factor deficiencies
•• Into the muscle Following trauma Spontaneous
CLASSIFICATION OF COAGULATION
DISORDERS (TABLE 17.2)
TABLE 17.2: Classification of coagulation disorders
A. Hereditary coagulation disorders
1. Hemophilia A 2. Hemophilia B
3. von Willebrand disease 4. Others
B. Acquired (secondary) coagulation disorders
1. Vitamin K deficiency 2. Liver disease
3. Others
HEREDITARY COAGULATION DISORDERS

Usually, due to deficiency of single coagulation factor.
vWF is synthesized by Factor VIII-vWF Complex

endothelial cells and
megakaryocytes. •• Factor VIII-vWF complex has two components:
–– Plasma factor VIII
–– von Willebrand factor.
vWF may be located in the •• vWF protects factor VIII and important for its stability. Subendothelial vWF promotes
plasma and subendothelial platelet adhesion.
tissue.
•• Whenever there is vascular endothelial injury, plasma vWF gets adsorbed to exposed
subendothelial matrix and augments adhesion of platelets.
Three common hereditary

disorders are:
HEMOPHILIA
1. Hemophilia A •• Hemophilia A and B are similar in both clinical and pathological features, the difference
(deficiency of factor VIII) being in the deficient factor.
2. Hemophilia B
•• Both are sex-linked recessive disorders resulting in inherited deficiency of the clotting
(deficiency of factor IX)
3. von Willebrand disease factor or synthesis of a defective clotting factor.
(deficiency of vWF). •• Males are affected and females are carriers.
HEMOPHILIA A (FACTOR VIII DEFICIENCY)

•• Common hereditary X-linked recessive disease.
•• About 30% of hemophiliacs may be due to acquired mutations.
•• Reduced amount or activity of factor VIII is associated with life-threatening bleeding
•• Bleeding is due to both inadequate coagulation and inappropriate clot removal (fibrinolysis).
Mode of Inheritance (Fig. 17.1)

Hemophilia A: X-linked •• X-linked recessive disease. Genes for factor VIII are located on the long arm of the
recessive disorder. X-chromosome.
•• Does not manifest when there is a normal copy of X-chromosome.
•• Males with a defective/mutant factor VIII gene (hemophiliac gene) on their single X
chromosome (XH) suffer from hemophilia.
•• Heterozygous females are carriers and do not express the full clinical disease because of
the paired normal X-chromosome.
•• However, females with two copies of the defective XH chromosome may rarely suffer from
hemophilia.
Molecular Genetics
Causative mutations include deletions, inversions, point mutations and insertions.
Clinical Features
Clinical severity depends on the level of factor VIII activity with normal range expressed as
percentage (Table 17.3). Severe cases have less than 1% residual factor VIII activity.
Bleeding Disorders: Due to Abnormalities of Coagulation/Clotting Factor CHAPTER 17 107
Hemophilia A: males are

suffers and females are
carriers.
Fig. 17.1: Mode of inheritance in hemophilia
TABLE 17.3: Factor VIII level and clinical severity in hemophilia A

Clinical severity Level of factor VIII activity in Clinical features Normal range for factor
percentage VIII: 45–158 IU/dL.
Mild More than 6 In the mildest form, it may be

unnoticed. Bleeding develops after
trauma only
Moderate 2–5 Bleeding after trauma, including
dental and other surgical trauma
Easy bruising
Severe Less than 1 Frequent and spontaneous
hemorrhage into joints
(hemarthrosis) and soft tissues
Common clinical presentations include: Hemophilia A: percentage

•• Frequent and spontaneous hemorrhage into the joints-hemarthrosis. of level of factor VIII
•• Hemorrhage into soft tissues. activity correlates with
severity of disease.
•• Prolonged bleeding following trauma
Hemophilia A: common
presentation
Laboratory Findings • Hemarthrosis
• Hemorrhage into soft
•• Bleeding time: normal
tissues.
•• Clotting time: prolonged, but is not a sensitive test
•• Platelet count: normal
•• Prothrombin time: normal
•• Activated partial thromboplastin time (APTT): increased (normal 30–40 seconds)
Hemophilia A: •• Factor VIII assay: essential for the diagnosis and to assess the levels and severity of disease
decreased: factor VIII •• Fibrinogen assay: normal
Increased: APTT and
clotting time. •• FDP: negative
•• Detection of carriers: by DNA markers
–– To detect female carriers
–– Prenatal diagnosis of affected fetuses.
Complications
Causes of death Due to Hemophilia
• Intracranial hemorrhage •• Deforming arthritis and contractures: this is due to repeated bleeding into the joints.
• Prolonged bleeding.
Organization and fibrosis of intramuscular hematomas → contractures of involved muscles.
•• Anemia: excessive, spontaneous or repeated bleeding leads to anemia.
Treatment Due to Therapy

• Factor VIII concentrate •• Viral hepatitis: hepatitis B, C and D in patients who received multiple transfusions of
• Recombinant factor VIII.
FFP/cryoprecipitate.
•• AIDS: in individuals who received fresh frozen plasma (FFP) or cryoprecipitate, when
screening tests for HIV were not available.
•• Factor VIII inhibitors: makes further management difficult.
Hemophilia B:
• X-linked recessive
HEMOPHILIA B (CHRISTMAS DISEASE,
disorder FACTOR IX DEFICIENCY)
• Mutation in factor IX
• Deficiency of factor IX. •• Clinically indistinguishable from hemophilia A
•• X-linked recessive disorder
•• Variable clinical severity
•• Assay of factor IX should be done to diagnose Christmas disease (named after the first
patient).
Hemophilia B: clinical
features
Laboratory Findings
• Usually milder than Similar to hemophilia A.
hemophilia A. •• Bleeding time: normal
• Hemarthrosis is the
common presentation. •• Clotting time: prolonged
•• Platelet count: normal
Hemophilia B:
decreased: factor IX
increased: APTT and •• Activated partial thromboplastin time (APTT): increased (normal 30–40 seconds)
clotting time. •• Factor IX assay: factor IX is decreased.
vWF: causes platelet

adhesion and prevents
degradation of Factor
VON WILLEBRAND DISEASE (vWD)
VIII in plasma. Platelet •• Most common inherited bleeding disorders
adehsion molecule •• Most cases are autosomal dominant disorders
is synthesized in the
Weibel-Palade bodies in •• Variable clinical picture with more than 20 variants.
endothelial cells.
Categories
Grouped into two major categories:
•• Quantitative deficiency of vWF: decreased circulating vWF vWD: autosomal dominant
disorders caused by
–– Type 1- autosomal dominant, mild disorder and form about 75% of all cases mutations in vWF.
–– Type 3-autosomal recessive, severe disorder and least common type.
•• Qualitative defects in vWF:
–– Type 2-autosomal dominant, accounts for 25% with several subtypes.
Clinical Features
•• Most cases are of mild bleeding
•• Common symptoms
–– Spontaneous bleeding from mucous membranes (e.g. epistaxis)
–– Excessive bleeding from wounds or menorrhagia
•• In severe cases, similar to hemophilia A.
Laboratory Findings vWD: increased

•• Platelet count: normal bleeding time, clotting
time and prolonged
•• Bleeding time: prolonged APTT. Plasma vWF is
•• Clotting time: prolonged decreased. Defective
•• Tourniquet test (Hess test): positive due to defect in platelet adhesion ristocetin induced platelet
aggregation test is
•• APTT: prolonged APTT diagnostic.
•• PT: normal
•• vWF assay: plasma level of active vWF is decreased
•• Platelet function test: defective ristocetin induced platelet aggregation test is diagnostic
of vWF.
Laboratory tests in hereditary disorders are summarized in Table 17.4.
TABLE 17.4: Summary of laboratory tests in hereditary coagulation disorders

Hemophilia A Hemophilia B von Willebrand disease Hemophilia A, B and
vWD: prothrombin time,
Bleeding time N N Increased
thrombin time and platelet
APTT Increased Increased Increased count are normal. APTT
Factor VIII Decreased N Low or Normal increased in all the three.
Factor IX N Decreased N
vWF N N Decreased
Abbreviation: N, normal
ACQUIRED COAGULATION DISORDERS Vitamin K dependent

coagulation factors: II, VII,
Coagulation Factor Abnormalities IX and X.
Usually characterized by multiple clotting abnormalities

•• Vitamin K deficiency: in neonates, low levels of vitamin K levels may produce life-threaten-
ing hemorrhage during the first week of life known as hemorrhagic disease of the newborn.
•• Liver disease: liver synthesizes all the clotting factors and severe liver disease is associated
with a hemorrhagic diathesis.
•• Other causes: disseminated intravascular coagulation that involves deficiency of several
coagulation factors.
DISSEMINATED INTRAVASCULAR
COAGULATION
Widespread disorder with combination of thrombosis and hemorrhage.
Etiology
Develops as a secondary complication of wide variety of disorders (Table 17.5).
DIC: TABLE 17.5: Major disorders associated with disseminated intravascular coagulation
• Sepsis, major trauma,
obstetric complications Infections
and certain cancers are • Gram-negative bacterial sepsis • Meningococcemia and other bacteria
the common triggers. • Fungi, viruses, Rocky Mountain
spotted fever, malaria
Obstetric Complications
• Retained dead fetus • Septic abortion
• Abruptio placentae • Amniotic fluid embolism
• Toxemia and pre-eclampsia
Neoplasms
•• Carcinomas of pancreas, prostate, lung and stomach
•• Acute promyelocytic leukemia
Massive Tissue Injury
• Traumatic • Burns
• Fat embolism • Surgery
Vascular Disorders
•• Aortic aneurysm, giant hemangioma
Immunologic Reactions
• Transfusion reactions • Transplant rejection
Respiratory Distress Syndrome
Miscellaneous
•• Snakebite, liver disease, acute intravascular hemolysis, shock, heat stroke, hypersensitivity, vasculitis
DIC: widespread thrombo- Pathogenesis (Fig. 17.2)

hemorrhagic disorder
secondary to wide variety Disseminated intravascular coagulation (DIC) is a disorder that shows combination of 1.
of disorders. thrombosis and 2. hemorrhage.
1. Thrombi/Clot Formation
Mechanism of thrombi formation:
A. Initiation of thrombotic process: two major mechanisms initiate the thrombotic process
of DIC namely entry of thromboplastic (procoagulant) substances into the circulation
and widespread endothelial injury.
•• Entry of thromboplastic (procoagulant) substances into the circulation: source of
thromboplastic/procoagulant substance in majority is tissue factor, which activates.
•• Widespread endothelial injury: endothelial injuries expose the thrombogenic sub
endothelial matrix.
Fig. 17.2: Pathogenesis of thrombosis, ischemic tissue necrosis and bleeding in disseminated intravascular coagulation
B. Development of thrombi: DIC:

•• Both procoagulant substances (tissue factor) and endothelial injury activate coagulation • Consumption of
system resulting in fibrin-platelet thrombi formation in the microvasculature. coagulation factors
• Widespread thrombosis
•• During this process there is consumption of clotting factors, fibrin and platelets. Hence,
in small blood vessels.
it is also referred to as consumptive coagulopathy or defibrination syndrome.
C. Consequences of thrombi formation: widespread deposition of fibrin-thrombi within
the microcirculation leads to:
•• Ischemic necrosis: microvascular thrombi produces micro-infarcts or large areas of
infarction and multiorgan failure.
•• Microangiopathic hemolytic anemia: RBCs trapped in the intravascular fibrin-
thrombi deposits undergo fragmentation. These RBCs appear as schistocytes in blood
smears; but, frank hemolytic anemia is unusual in DIC.
2. Hemorrhagic Diathesis
A. Causes of hemorrhagic/bleeding diathesis:
•• Consumption of platelets
•• Consumption of coagulation factors
•• Activation of fibrinolytic system.
B. Mechanism of hemorrhagic diathesis fibrin-thrombi activate secondary fibrinolytic
system and generate plasmin. The plasmin cleaves fibrinogen and fibrin and generates
fibrin split products (FSPs) [or fibrin degradation products (FDP)]. FSPs are potent
anticoagulant and antiplatelet effect and produces hemorrhagic diathesis.
Prognosis
• Depends on the
underlying disorder.
Clinical Features • Mortality is high in
•• Serious, often fatal, clinical condition severe cases.
•• Signs and symptoms are related to:
Treatment
–– Hemorrhagic diathesis/bleeding: most common, manifest as ecchymoses, petechiae or • Removal of the
bleeding from mucous membranes or at the sites of venipuncture. underlying cause
–– Microvascular thrombi: tissue hypoxia and infarction of the organ leading to multiorgan • Replacement of clotting
failure. factors and platelets.
Laboratory Findings in DIC

DIC laboratory findings: Screening Assays
• Increased: APTT, PT, BT, •• Coagulation abnormalities
D-dimer
• Decreased: platelets, –– APTT: increased as a result of consumption and inhibition of the function of clotting
fibrinogen. factors.
–– Prothrombin time: increased.
–– Thrombin time (TT): increased because of decreased fibrinogen.
–– Fibrinogen: decreased.
•• Bleeding time: increased due to decreased platelet count.
•• Platelet count: decreased due to utilization of platelets in microthrombi.
•• Peripheral smear: microangiopathic hemolytic anemia with schistocytes.
Confirmatory Tests
•• Fibrinolysis abnormalities
•• FDP (fibrin degradation/split products): secondary fibrinolysis results in generation of
DIC: D-dimer test is specific FDPs, which can be measured by latex agglutination
diagnostic test. •• D-dimer test: it is specific for diagnosing DIC.
Thrombotic Disorders:
Hypercoagulable State
18
CHAPTER
HYPERCOAGULABLE STATE (THROMBOPHILIA)

Group of inherited or acquired conditions that are associated with increased tendency or risk
to develop thrombosis.
Causes of Hypercoagulability State (Table 18.1)

TABLE 18.1: Major causes of hypercoagulable state
A. Inherited (genetic/primary)
Deficiency of antithrombotic (anticoagulant) factors
1. Antithrombin III deficiency 2. Protein C deficiency
3. Protein S deficiency
Increased prothrombotic factors
1. Activated protein C (APC) resistance (Factor V mutation/ factor Va/ factor V Leiden)
2. Excessive levels of prothrombin (prothrombin G20210A mutation)
3. High levels of factors VII, XI, IX, VIII; von Willebrand factor; fibrinogen
B. Secondary (acquired)
1. Antiphospholipid antibody syndrome
2. Venous stasis
◆ Prolonged immobilization ◆ Congestive cardiac failure
◆ Prolonged bed rest
3. Increased platelet activation
◆◆ Hematological disorders: myeloproliferative neoplasms (polycythemia vera, essential
thrombocythemia, myelofibrosis), paroxysmal nocturnal hemoglobinuria
◆◆ Cancer
◆ Nephrotic syndrome ◆ Atrial fibrillation
◆ Myocardial infarction ◆ Heparin-associated thrombocytopenia
◆ Thrombotic thrombocytopenic purpura (TTP)
4. Increased hepatic synthesis of coagulation factors and reduced anticoagulant synthesis
◆ Oral contraceptive pill ◆ Hyperestrogenic states
(pregnancy and postpartum)
5. Release of procoagulant substances: disseminated cancers
6. Tissue injury: surgery, fracture, extensive burns
7. Reduced endothelial PGI2: old age
8. Endothelial injury: homocysteinemia
9. Fibrinolytic system defects
10. Unknown: smoking, obesity, sickle cell anemia
INHERITED HYPERCOAGULABLE STATES

Clinical Presentation
•• Thrombosis develops at young age (less than 45 years)
•• Recurrent thromboembolism
•• Family history of thromboembolic episodes
•• Thrombosis develops in the venous system and at unusual anatomical sites like visceral
veins.
Deficiency of Antithrombotic Factors

Antithrombin (AT) III Deficiency
•• Autosomal dominant disorder
•• Deficiency of antithrombin—either quantitative or qualitative
•• Risk of a thrombosis—20% to 80%.
Protein C and S Deficiency

•• Normally, activated proteins C (APC) and protein S act as a complex, which degrades
activated factors V and VIIl.
•• When there is deficiency of these proteins, the activated factor V and VIII are not neutralized.
This leads to activation of the clotting system and formation of thrombus.
Increased Prothrombotic Factors

Factor V Leiden/Leiden Activated Protein C (APC) Resistance (Factor V Leiden)
mutation is characterized
by factor V variant. •• Most common genetic disorder associated with familial thrombophilia.
•• Activated proteins C (APC) and protein S complex inhibits activated factor normal V and
VIII. The variant clotting factors cannot be degraded.
Factor V Leiden is resistant •• Point mutation in the factor V gene synthesis of a factor V variant. This variant is known as
to inhibition by activated
protein C (APC). It is
factor V Leiden/Leiden mutation.
associated with familial •• Factor V variant has normal procoagulant activity but is resistant to inhibition by activated
thrombophilia. protein C (APC).
ACQUIRED HYPERCOAGULABLE STATES

Causes
Causes of the acquired hypercoagulable states are listed in Table 18.1.
Antiphospholipid Antibody Syndrome (APLA/APS)

•• Presence antiphospholipid antibodies (APAs) in the plasma are associated with hyper
coagulable state.
•• Antiphospholipid antibody reacts with plasma proteins, which are bound to phospholipids
(Fig. 18.1).
•• Two important antiphospholipid antibodies: lupus anticoagulant antibody and anti-β2
glycoprotein antibody.
1. Lupus anticoagulant antibody: prolongs the phospholipid-dependent coagulation
tests in vitro (e.g. prolongation of APTT).
Thrombotic Disorders: Hypercoagulable State CHAPTER 18 115
2. Antibodies against the phospholipid–β2-glycoprotein com Antiphospholipid

plex: it also bind to cardiolipin antigen used in the serological antibodies includes lupus
test for syphilis. anticoagulant antibody
and anti-β2 glycoprotein
antibody.
Types
•• Primary antiphospholipid syndrome: no predisposing cause.
•• Secondary antiphospholipid syndrome: association with
autoimmune diseases, like systemic lupus erythematosus, hence
known as lupus anticoagulant syndrome.
Clinical Features Fig. 18.1: Antiphospholipid anti

body against plasma proteins
•• Hypercoagulable state: commonest acquired hematologic bound to phospholipids Triad of thrombosis,
cause of recurrent thromboembolic events. recurrent spontaneous
•• Repeated spontaneous abortions: normally, tissue plasminogen activator (t-PA) is neces- abortions and immune
thrombocytopenia
sary for the invasion of uterine blood vessels by placental trophoblastic tissue. Recurrent may be the presenting
spontaneous abortions develop due to antibody-mediated inhibition of t-PA activity. clinical features of
•• Immune thrombocytopenia. antiphospholipid
syndrome.
Laboratory Tests
Coagulation tests
•• APTT: prolonged
•• Factor VIII levels: normal
•• Thrombin time: normal
•• Fibrinogen level: normal.
Confirmatory test
•• Test for lupus anticoagulant:
–– Dilute Russell’s viper venom test (DRVVT): Russell’s viper venom (RVV) activates factor
X leading to fibrin clot. Lupus anticoagulant prolongs clotting time by binding to RVV
and preventing the action of RVV.
•• Antibodies against the phospholipid–β2-glycoprotein complex:
–– Detected by enzyme-linked immunosorbent assay (ELISA) and radioimmunoassay
(RIA).
SECTION 4
Clinical
Scenario
Clinical Scenario 19
CHAPTER
INTRODUCTION
Medical undergraduates in pathology are frequently required to analyze clinical-oriented
cases. These cases are provided with symptoms and signs and laboratory findings. Students
are expected to often diagnose these cases with history alone. Some of the classical scenarios
in hematology, which are frequently asked in pathology examination, are presented in this
chapter. First, symptoms, signs and general characteristics of blood diseases are given. Next
common patterns of clinical features observed in hematology are discussed, which will help in
suggesting the diagnosis. This is followed by clinical scenarios and their interpretations.
SYMPTOMS AND SIGNS THAT SUGGEST A

BLOOD DISEASE (TABLE 19.1)
TABLE 19.1: Symptoms and signs that favor a blood disease
Symptoms and signs Suggestive of blood disease During pregnancy
anemia may develop due
•• Tiredness, weakness, malaise, lightheadedness to combination of iron
and easy fatigability deficiency and/or folate
•• Dyspnea (breathlessness) on mild exertion deficiency.
relieved by lying flat Anemia
Pallor: appreciated better
•• Pallor in the conjunctiva, mucous
•• Tachycardia, palpitation and systolic murmur membrane of tongue and
nail beds.
•• Pica (craving for ice or soil)
Pica is a craving for certain
•• Spoon-shaped nails (koilonychia) substances with no
Iron deficiency anemia
•• Cheilosis (fissures at the corners of the mouth) nutritional value like clay
or chalk.
•• Dysphagia
Vitamin B12 -peripheral
•• Painful red “beefy” tongue (glossitis)
neuropathy: glove and
•• Pain or weakness in legs sock distribution of
Vitamin B12 deficiency tingling and numbness or
•• Peripheral neuropathy
paresthesia in the fingers
•• Psychosis and toes.
Contd...
120 SECTION 4 Clinical Scenario
Contd...
•• Passage of dark urine (? red cells, ? hemoglobin) Hemolytic anemia—intravascular hemolysis
•• Jaundice/recurrent jaundice Hemolytic anemia, megaloblastic anemia (lemon-
yellow jaundice)
•• Enlarged spleen Chronic hemolytic anemias, malaria, leukemia,
lymphoma
•• Splenic infarction producing severe abdominal
pain Sickle cell anemia
•• Leg ulcers
Massive splenomegaly
•• Frontal bossing—thalassemic facies Chronic hemolytic anemia, e.g. thalassemia major
causes abdominal
discomfort. •• Massive splenomegaly (greater than 20 cm in size) Chronic myelocytic leukemia, myelofibrosis, kala-azar
PATTERNS STRONGLY SUGGESTIVE OF

A BLOOD DISEASE
Various patterns that favor a blood disease are mentioned below.
Pattern 1: Iron Deficiency Anemia

1. History strongly indicative of iron deficiency (IDA): a pale patient with tiredness,
Craving for ice
weakness, malaise, a sore tongue (glossitis), pica (craving for ice) and spoon-shaped nails
(pagophagia), is believed
to be the most specific to (koilonychia). Signs and symptoms are usually due to both anemia and the underlying
iron deficiency. cause of anemia.
Cheilosis and are 2. Diagnosis of IDA would be strengthened in a patient who has underlying cause of anemia
koilonychia (spoon such as gastrointestinal (e.g. carcinoma colon, hemorrhoids) or gynecologic disease
shaped or flattened)
are characteristically
(e.g. menorrhagia/excessive menstrual blood flow), malnutrition, pregnancy, and
and specifically seen in malabsorption.
advanced cases of IDA. 3. IDA: diagnosis is typically based on laboratory results. Microcytic hypochromic anemia,
low MCV, MCH, MCHC, reduced serum ferritin and other serum profile observed in IDA
(refer Table 1.4).
Inference (case No. 1): Case No. 1

spoon shaped nails and
pica indicates IDA. History: A 28-year-pregnant lady comes to hospital with complains of weakness, easy fatigability and
breathlessness of 4 months duration. She is a laborer of low economic status and used to eat clay. On
examination, she is pale and had spoon shaped (koilonychia) nails.
Inference (case No. 2): Case No. 2

spoon shaped nails and
History: A 20-year-old girl complains of weakness, easy fatigability and breathlessness of 6 months
cheilosis indicates IDA.
Menorrhagia is the cause. duration. She also complains of heavy menstrual bleeding every month. On examination, she is pale
and had spoon shaped (flattened) brittle nails and cheilosis.
Inference (case No. 3):

cause of IDA is bleeding
Case No. 3
from GI tract. History: A 73-year-old male complains of increasing weakness, malaise, and easy fatigability for the past
10 months. On examination, he is pale without hepatosplenomegaly. He also complains of black tarry
stool (Stool for occult blood was positive).
Clinical Scenario CHAPTER 19 121
Laboratory findings for cases 1, 2, 3: Hb 9.1 g/dL, hematocrit 27.3%, RBC count 3 million/cumm, WBC Laboratory findings for
count 6,700/cumm and platelet count 4,20,000/cumm. MCV 70 fL, MCH 26 pg, MCHC 28 g/dL and RDW cases 1 to 3: The decreased
18. hemoglobin, RBC count,
MCV, MCH and MCHC
favors iron deficiency
Cause of IDA: the causes for iron deficiency: anemia.
•• Case 1 is probably due to inadequate intake (due to low socioeconomic status) or increased
demand of iron during pregnancy.
Increased RDW helps in
•• Case 2 is caused by chronic blood loss, due to excessive menstrual flow. differentiating IDA from
•• Case 3 is due to chronic blood loss through gastrointestinal tract. The black tarry stool thalassemia (refer Table
indicates that the patient has occult bleeding probably from GI tract malignancy. 3.3).
Pattern 2: Megaloblastic Anemia Unlike with B12 deficiency,

folic acid deficiency is not
•• History strongly indicative of megaloblastic anemia (vitamin B12 deficiency and/or associated with significant
pernicious anemia): middle-aged, pale patient with prematurely gray hair, severe glossitis neuropathy.
and stomatitis and a smooth painful tongue, significant neuropathy and difficulty in
walking, uncoordinated gait, impairment of vibration sense and position sense with
delirium/dementia.
•• Patients with megaloblastic anemia due to folic acid/folate deficiency present with
symptoms of anemia or of the underlying cause.
•• Peripheral smear with macro-ovalocytes, hypersegmented neutrophils, raised MCV,
bone marrow with megaloblasts are features of megaloblastic anemia.
Inference (case No. 4): in a

Case No. 4 pure vegetarian, tingling
History: A 34-year-male, pure vegetarian, executive complains of anorexia, premature graying of hair, and numbness, glossitis,
weight loss and tingling and numbness in fingers and toes. On examination, he is pale and anemic. The raised MCV point towards
tongue is shiny and has glazed appearance (glossitis). vitamin B12 deficiency.
Laboratory findings: Hb 9 g/dL, WBC count 3,800/cumm, platelet count 60,000/cumm, RBC count 2.5
million/cumm. MCV is 104 fL, MCH 32 pg and MCHC is 32 g/dL.
Case No. 5 Inference (case No. 5):

raised MCV, history of
History: A 45-year-old female complains of tiredness, mild breathlessness while climbing steps and tingling and numbness,
tingling and numbness. On examination, she is pale, has raspberry-red tongue, and shows numbness raspberry tongue point
and paresthesia. Biochemical investigation reveals normal serum iron profile. to megaloblastic anemia
due to vitamin B12
deficiency. The evidence of
Laboratory findings: Hb 8.2 g/dL, WBC count 4,800/cumm, platelet count 1,20,000/cumm, MCV is 108 fL, atrophic gastritis points to
MCH 33 pg and MCHC is 32 g/dL. Peripheral smear and bone marrow examination confirms the clinical pernicious anemia.
diagnosis. Patient is further subjected to gastric biopsy and shows atrophic gastritis. Other test including
serological tests is done.

Case No. 6 characteristic history
History: A 50-year-old male complains of loss of appetite, increasing fatigue tingling and numbness of with increased MCV,
both lower limbs and difficulty in walking for the past 10 months. On physical examination, he is pale macro-ovalocytes
and tongue was beefy red. and hypersegmented
neutrophils in the
peripheral smear points to
Laboratory findings: Hb 9.2 g/dL, hematocrit 27.9%, MCV 132 fL, platelet count 242,000/cumm, and megaloblastic anemia.
WBC count 7590/cumm. Peripheral smear shows macro-ovalocytes and hypersegmented neutrophils.
HS may present during Pattern 3: Hereditary Spherocytosis

anytime from the neonatal
period to adulthood. When •• History strongly indicative of chronic extravascular hemolysis, usually but not always
there is a family history, it hereditary spherocytosis: normal patient presenting with pallor, mild jaundice, an enlarged
is usually easy to suspect spleen (classic triad) and a family history of a relative who had been ‘‘cured’’ of the same
the diagnosis. problem by splenectomy. Also family history of pigment gallstones in a young person.
•• Peripheral smear with spherocytes and raised osmotic fragility test are features of HS.
Case No. 7
Inference (cases No. 7 History: A 24-year-women complains of chronic fatigue since childhood and mild icterus. On examination
and 8): mild jaundice, she is anemic, jaundiced and has moderate splenomegaly. Her mother had similar complaints and has
anemia and splenomegaly multiple pigment gallstones.
constitute a classical
triad, and along with
family history/pigment
gallstones favor hereditary Case No. 8
spherocytosis. History: A 15-year-old girl presents with weakness and fatigue. On examination, her sclera shows mild
jaundice and mild splenomegaly. Family history indicated about her father having recurrent gallstones.
Positive osmotic test Laboratory findings for cases 7 and 8: Hb 11.1 g/dL, RBC count 3.6 million/cumm. WBC count 6,800/cumm
fragility confirms the and platelet count 1,75,000/cumm. MCV is 78 fL, MCH is 32 pg and MCHC is 38 g/dL. In a laboratory test,
diagnosis of HS. her RBCs lyse at a higher concentration of saline (osmotic fragility test) compared to normal patients.
Pattern 4: Thalassemia Major

•• History strongly indicative of thalassemia major: short stature, failure to thrive, severe
pallor, jaundice, maxillary over-growth (bossing), frontal bossing, sallow complexion and
splenomegaly.
•• Laboratory finding of microcytic hypochromic anemia with many target cells, reduced
MCV,MCH and MCHC, increased serum iron normal RDW are features of thalassemia
major.

failure to thrive, severe Case No. 9
pallor, jaundice, History: An 11-month-old male child was brought to the pediatric outpatient by his parents and
splenomegaly, microcytic complained that the child was failing to thrive. On examination, jaundice, pallor and a palpable spleen
hypochromic anemia
was detected.
with target cells, reduced
MCV and normal RDW
points to the diagnosis Laboratory findings: Hb 7.8 g/dL, hematocrit 23.4%, MCV 66 fL, platelet count 1,75,000/cumm, and
of β-thalassemia major. WBC count 8,200/cumm. His serum ferritin was 3250 ng/mL. Peripheral examination showed microcytic
Increased iron stores and hypochromic anemia with many target cells. A bone marrow aspiration performed and reveals a
raised serum ferritin is myeloid: erythroid ratio of 1:4, and increased iron stores.
against the diagnosis of
IDA (refer Table 3.3).
Pattern 5: Sickle Cell Anemia

•• History strongly indicative of sickle cell anemia: recurrent episodes of vaso-occlusion in
Inference (case No. 10): the connective and musculoskeletal structures anywhere in the body producing ischemia and
recurrent episodes of pain manifesting as acute pain, tenderness and fever. Child may show generalized impairment
abdomen and backache of growth and development. Adults manifest with chronic organ damage.
points to a vaso-occlusive
crisis in sickle cell anemia.
•• Presence of sickle cells in the peripheral smear, positive sickling test, along with
demonstration of HbS by hemoglobin electrophoresis and HPLC is diagnostic of sickle
cell anemia.
Presence of sickle cells in

Case No. 10 the peripheral smear and
History: A 10-year-old boy complains of severe pain in the chest, abdomen and bones. On enquiry, his presence of more than
70% Hb S on hemoglobin
mother reveals that he had several episodes of severe abdominal and back pain since early childhood.
electrophoresis is
Physical examination shows anemia, jaundice and leg ulcer. confirmative.
Laboratory findings: Hb 11.0 g/dL, RBC count 3.2 million/cumm, WBC count 8,800/cumm and platelet
count 1,95,000/cumm. Peripheral smear examination, hemoglobin electrophoresis and HPLC confirms
the diagnosis.
Pattern 6: Immune Thrombocytopenic Purpura (ITP) The severity/nature of

bleeding depends on the
•• History strongly indicative of acute immune thrombocytopenic purpura: sudden platelet count.
development of bruising, petechiae and muco-cutaneous bleeding in a child following
a bacterial or nonspecific viral (upper respiratory or gastrointestinal) infection without
features of acute leukemia (such as anemia and bone tenderness).
•• Laboratory findings of markedly reduced platelet count and megakaryocytic hyperplasia
with immature megakaryocytes. In the bone marrow favors the diagnosis of acute immune
thrombocytopenic purpura.
petechial rashes of short
Case No. 11 duration in a child, low
platelet count is due to
History: A 14-year-male comes to the OPD with petechial rashes of 2 days duration. He has no history of
acute idiopathic thrombo-
bleeding in past. He had an attack of viral infection 3 weeks before this complaint. cytopenic purpura.
Laboratory findings: Hb 14.5 g/dL, hematocrit 45%, RBC count 5.1 million/cumm. WBC count 7,200/
cumm and platelet count 75,000/cumm. MCV is 85 fL, MCH is 32 pg and MCHC is 31 g/dL and RDW is 14.
Pattern 7: Hemophilia
•• History strongly indicative of one of the hemophilias (A or B): a male child presenting with
a serious bleeding disorder, particularly with hemarthrosis.
•• Decreased factor VIII/IX and increased APTT is diagnostic of one of the hemophilias
(A or B).
Case No. 12
History: An 18-year-male complains of knee joint swelling after minor trauma. On examination, the joint
Inference (cases No. 12 and
appears tense, red and swollen. Family history revealed similar complaints by his uncle. 13): the development of
ecchymosis, family history
with disease affecting
Case No. 13 males, hemarthrosis,
normal platelet count and
History: A 6-year-old boy gives a history of easy bruising and episodes of passing blood in urine since
prolonged APTT favor
infancy. On examination, many ecchymoses are noted in the skin of lower limbs. Family history reveals
hemophilia A/B.
similar complaints in members of the family involving only males and history of hemarthrosis in few of
them.
Laboratory findings for cases 11 and 12: Hb 13 g/dL, hematocrit 36%, RBC count 4.4 million/cumm, WBC
count 8,200/cumm and platelet count of 2,35,000/cumm. MCV is 84 fL, MCH 32 g and MCHC is 33 g/dL.
Bleeding time, clotting time and prothrombin time are within normal limits. APTT is prolonged.
ALL: pallor, fatigue, Pattern 8: Acute Lymphoblastic Leukemia (ALL)

bleeding, fever, and
infection are due •• History strongly indicative acute lymphoblastic leukemia: child (1 to 6 years of age)/adult
to peripheral blood (30 to 40 years) presenting with rapid onset (few weeks) of pallor, fatigue, bleeding, fever,
cytopenias resulting from and infection. Presence of lymphadenopathy, hepato- or splenomegaly, CNS disease,
bone marrow failure. testicular enlargement.
•• Laboratory findings of marked raised WBC count, presence of more than 20% lymphoblasts
in the blood and bone marrow, blasts + with PAS (block positivity) and TdT + are features
of acute lymphoblastic leukemia.
Inference (cases No. 14 to

16): presence of anemia, Case No. 14
thrombocytopenia and History: A 6-year-old boy presents with fatigue, bone pain, weakness and low-grade fever of 1-week
leukocytosis with presence duration. Physical examination shows anemia, sternal bone tenderness and lymphadenopathy.
of PAS positive blasts in the
peripheral smear and bone
marrow in a child is in favor
of acute lymphoblastic
Case No. 15
leukemia. History: A 5-year-old boy complains of sudden onset of fever, tiredness and pallor. On examination,
there are many enlarged cervical lymph nodes, hepatosplenomegaly, bone tenderness, and petechial
hemorrhages on the skin.
Bone pain is due to

expansion of the marrow
Case No. 16
caused by proliferation of History: A 4-year-old boy is becoming increasingly lethargic for the past 1 month. On examination, he is
blasts. having fever, ecchymoses on the skin of his lower legs and shoulder, generalized lymphadenopathy and
tenderness on palpation of long bones.
Laboratory findings of cases 14 to 16: Hb 8 g/dL, hematocrit 24%, WBC 18,200 cells/cumm and platelet
count of 90,000/cumm. Serum LDH is markedly increased. Peripheral blood smear shows abnormal
cells, which are PAS positive. A bone marrow examination shows 100% cellularity with replacement
by primitive cells. These abnormal primitive cells have scanty cytoplasm and large nuclei with indistinct
nucleoli.
Following blood smear examination, the child is admitted to the medical oncology ward.
Pattern 9: Acute Myeloblastic Leukemia (AML)

•• History strongly indicative acute myeloblastic leukemia: adult presenting with rapid onset
(few weeks) of pallor, fatigue, bleeding, fever, and infection.
•• Laboratory findings of marked raised WBC count, presence of more than 20% myeloblasts
in the blood and bone marrow, presence of Auer rods (not seen in all subtypes), blasts +
with MPO and Sudan Black B are features of acute myeloblastic leukemia.

Case No. 17 the high WBC count with
History: A 45-year-old man presents with fatigue, episodes of epistaxis, bleeding from gums, and low the blasts and Auer rods
grade fever of 1 week duration. Physical examination reveals temperature of 37.4°C and hepatospleno- are very characteristic
megaly. for acute myelogenous
leukemia.
Laboratory findings: Hb 7 g/dL, WBC count 51,000/cumm and platelet count 80,000/cumm. Peripheral
blood smear shows abnormal large cells with Auer rods.
Pattern 10: Chronic Myelogenous Leukemia (CML)

•• History strongly indicative of chronic myelogenous leukemia: patient in fifth and sixth
decades of life presenting with gradual development of fatigue, weakness, weight loss,
anorexia and fullness of abdomen, early satiety and left upper quadrant pain or mass (due
to splenomegaly).
•• Laboratory finding of very high WBC count with myeloid precursors, neutrophils and
myelocyte peak, low LAP and presence of Ph chromosome is diagnostic of chronic
myelogenous leukemia.

Case No. 18 massive splenomegaly
History: A 50-year-old male comes to the hospital with complaints of generalized weakness, weight loss, with very high WBC
easy fatigability, abdominal discomfort and dragging sensation in the left hypochondrium for the last count, peripheral smear
8 months. On examination, he is pale and has marked splenomegaly. with myeloid precursors,
neutrophils and myelocyte
peak t (9:22) positivity and
Laboratory findings: Hb 11.9 g/dL, hematocrit 36%, WBC count 1,20,000/cumm, platelet count 4,12,000/ low LAP score are features
cumm. Peripheral smear examination shows characteristic blood picture with myeloid precursors, of chronic myelogenous
neutrophils and myelocyte peak confirms the clinical diagnosis. The leukocyte alkaline phosphatase leukemia (chronic phase).
(LAP) score is low. Chromosomal analysis shows t(9:22) positivity. He is admitted to the oncology
department for treatment.
Pattern 11: Chronic Lymphocytic Leukemia (CLL)

•• History strongly indicative of chronic lymphocytic leukemia: marked splenomegaly,
generalized lymphadenopathy (any nodes can be involved), involvement of spleen in an
elderly patient with fatigue, loss of weight and anorexia.
•• Peripheral blood with lymphocytosis which constitute more than 50% of the white cells
and smudge cells; bone marrow with more than 30% lymphocytes of the bone marrow
cells is diagnostic of chronic lymphocytic leukemia.

Case No. 19 older patient,
History: A 60-year-old male complains of increasing fatigue and shortness of breath with minimal abdominal discomfort
exercise for the last 6 months. He has noted some abdominal discomfort over the past month. On (splenomegaly) non-
examination, he has non-tender cervical lymphadenopathy. The liver is enlarged, smooth and palpable tender lymphadenopathy,
just below right costal margin. The spleen is palpated 3 cm below left costal margin. hepatosplenomegaly,
high WBC count with
lymphocytosis and
smudge cells in the
peripheral smear are
features of chronic
lymphocytic leukemia.
Laboratory findings: WBC count 22,000/ cumm with 78% lymphocytes on DLC. Hb 10.1 g/dL, hematocrit
33%, platelet count 2,32,000/cumm. Peripheral smear shows many smudge cells.
Pattern 12: Multiple Myeloma

•• History strongly indicative of multiple myeloma: Patient over 50 years of age presenting
with severe bone pain, pallor, and renal failure.
•• Laboratory findings shows nonspecific findings such as raised ESR, rouleaux formation of
RBC in the peripheral smear and lytic lesions in the bone. Bone marrow with more than
30% myeloma cells and M spike on electrophoresis confirms the diagnosis of multiple
myeloma.

patient over 50 years
Case No. 20
of age with bone pain, History: A 61-year-old man has dull, constant back pain for 3 months. On physical examination, he is
pallor, lytic lesions in bone, pale. A plain film radiograph of the spine and skull reveals several 1 to 2 cm lytic lesions of the vertebral
peripheral smear with bodies.
marked rouleaux formation
and markedly raised ESR
suggest multiple myeloma. Laboratory findings: Peripheral smear shows marked rouleaux formation, ESR is 110 mm at 1st hour.
Bone marrow aspiration, urine and serum electrophoresis showed characteristic findings.
Pattern 13: Hodgkin Lymphoma (HL)

•• History strongly indicative of Hodgkin lymphoma: weight loss, fever, night sweats,
lassitude, pruritis, and enlarged cervical nodes. Sometimes accompanied by pain in one
or more sites almost immediately after ingestion of alcohol. Extranodal involvement is very
rare.
•• Histologically/cytologically presence of RS cells is diagnostic of Hodgkin lymphoma.

non-tender
Case No. 21
lympadenoapthy, low- History: A 33-year-old female complains of low-grade fevers, 6 kg weight loss, night sweats, and
grade fever, weight generalized malaise for the past 3 months. On physical examination, she has non-tender right cervical
loss, night sweats and and supraclavicular lymphadenopathy.
presence of characteristic
Reed-Sternberg cells is
diagnostic of Hodgkin Investigation: Fine needle aspiration followed by a biopsy of cervical lymph node is performed. On
lymphoma. microscopic examination, the lymph node showed loss of architecture and characteristic cell that
confirmed the clinical diagnosis. He was admitted to the oncology ward for further management.
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11. Kumar V, Abbas AK, Aster JC. Robbins Basic Pathology, 9th edn. Philadelphia, WB Saunders. 2013.
12. Lewis SM, Bain BJ, Bates I. Dacie and Lewis Practical Hematology, 10th edn. London, Churchill Livingstone. 2006.
13. Lichtman MA, Kipps TJ, Kaushansky K, Beutler E, Seligsohn U, Prchal JT. Williams Hematology, 7th edn. USA,
McGraw-Hill Companies. 2006.
14. Longo DL. Harrison’s Hematology and Oncology. USA, McGraw-Hill Companies. 2010.
15. Longo DL, Kasper DL, Jameson JL, Fauci AS, Hauser SL, Loscalzo J. Harrison’s Principles of Internal Medicine, 18th
edn. USA, McGraw-Hill Companies. 2012.
16. McPherson RA, Pincus MR. Henry’s Clinical Diagnosis and Management by Laboratory Methods, 22nd edn.
Philadelphia, WB Saunders. 2011.
17. Rodak BF, Fritsma GA, Doig K. Hematology: Clinical Principles and Applications, 3rd edn. Philadelphia, WB
Saunders. 2007.
18. Rubin R, Strayer DS. Rubin’s Pathology: Clinicopathologic Foundation of Medicine, 6th edn. Philadelphia, Lippincott
Williams and Wilkins. 2012.
19. Stevens A, Lowe J. Pathology, 2nd edn. Edinburgh, Mosby. 2000.
20. Swerdlow SH, Campo E, Harris NL, Jaffe ES, Pileri SA, Stein H, et al., WHO Classification of Tumours of Haematopoietic
and Lymphoid Tissues. IARC, Lyon. 2008.
21. Underwood JCE, Cross SS. General and Systemic Pathology, 5th edn. Edinburgh, Churchill, Livingstone. 2009.
Appendix
Different stages of hematopoiesis.

Abbreviation: BFU, bursa forming unit, CFU, colony forming unit
128 Rapid Review of Hematology
Stages of erythropoiesis
Variations in color of RBCs and associated conditions

Appendix 129
Variations in shape of RBCs and associated conditions

Inclusions in RBCs and associated conditions
Rouleaux formation and associated conditions

Appendix 131
Variations in size of RBCs and associated conditions
Stages of megakaryopoiesis
Hemoparasites
1. Malarial parasites:
Most common are
Plasmodium vivax and
falciparum
2. Microfilaria
3. Trypanosomes Stages of myelopoiesis
4. Leishmania donovani
5. Babesiosis.
HEMOPARASITES
A B C D
Peripheral blood smear Peripheral blood smear A. Plasmodium vivax ring form, B and C Plasmodium vivax schizont and
showing microfilaria D. Plasmodium falciparum gametocyte
Index
Page numbers followed by f refer to figure and t refer to table
A promyelocytic leukemia 54 Autosomal

Abdominal discomfort 125 Adult T cell dominant
Abnormal localization of immature leukemia 49, 81 disorder 17, 108
precursors 61 lymphoma 49, 81 trait 19
Abnormalities of Agranulocytosis 47 recessive hereditary disorder 22
coagulation/clotting factor 105 Aleukemic leukemia 52 Autosplenectomy 33
globin production 22 Alimentary system 12
platelet 100 Allergic rhinitis 48 B
Abnormally large megakaryocytes 66 Alloimmune hemolytic anemia 36 B cell
ABO Amyloidosis 79, 80 neoplasms 76
hemolytic disease of newborn 39 Anaplastic large cell lymphoma 81 prolymphocytic leukemia 81
incompatibility 37 Anemia 3, 19, 36, 65f Basophilia 49
Achlorhydria 12 of blood loss 41 Bence Jones proteins 76, 78
Acquired of chronic disorders 8 Benign leukocytic proliferation 47
clonal stem cell disorders 60 of impaired red cell production 3 Bernard-Soulier syndrome 100, 104
coagulation disorders 105, 109 Angioimmunoblastic T cell lymphoma 81 Biochemical tests for megaloblastic
disorders 52, 100 Angular stomatitis and glossitis 7 anemia 11
hypercoagulable states 114 Anisopoikilocytosis 33 Black tarry stool 120
mutations 41 Antiglobulin test 38, 39 Bleeding
myeloproliferative neoplasm 63 disorders 99, 100, 105
Anti-intrinsic factor antibody 12
Activated tendency 80
Antiphospholipid antibody syndrome 114
partial thromboplastin time 107 time 103, 107
Antiplatelet autoantibodies 102
protein C resistance 114 Blockage of microcirculation 32
Antithrombin III deficiency 114
Acute Blood
Aplastic
abdominal pain 32 brain barrier 38
anemia 13, 15, 46, 48, 50
bacterial and fungal infections 46 loss 4
crises 20, 33 Bone 79
blood loss 41
Appearance of marrow 7, 10, 15, 18, 25, 27, 34, 35, 52,
chest syndrome 32
neoplastic lymphoid cells 84 57, 65, 66, 67, 77, 95, 103
erythroid leukemia 54
typical thalassemic facies 25f aspiration 75
immune thrombocytopenic purpura
Arterial oxygen saturation 65 biopsy 65, 67
102, 123
Asthma 48 failure 56, 58
leukemia 52, 52t, 53, 55, 72
lymphoblastic Asynchrony of nuclear and cytoplasmic iron 27
maturation 10 trephine biopsy 61
leukemia 49, 52, 53, 55, 56f, 124
Atherosclerosis 13 pain 56
lymphoma 55
lymphoid leukemias 55t Atrophic and tenderness 56
gastritis 121 Brucellosis 49
megakaryoblastic leukemia 54
glossitis 7, 12 Budd-Chiari syndrome 63
monoblastic leukemia 54
Atrophy of glands 12 Burkitt lymphoma 81, 83, 84f, 85f
monocytic leukemia 49, 54
myeloblastic leukemia 52, 53, 124 Atypical lymphocytosis 51
myelocytic leukemia 49, 53 Autoimmune C
myelogenous leukemia 53, 57 diseases 49 Carcinoma 15
myeloid leukemia 53, 54 disorder 102 Causes of
myelomonocytic leukemia 48, 49, 54 hemolytic anemia 36, 39, 40 aplastic anemia 14t
eosinophilia 48t lymphoid neoplasms 81 Direct antiglobulin test 39, 40

hemolytic anemia 16t plasma cell neoplasms 76, 76t Disorders of
hemorrhagic/bleeding diathesis 111 platelet neutrophils 46
hypercoagulability state 113 disorders 100, 101t platelet
iron functional disorders 104t adhesion 104
deficiency anemia 5t sickle cell disease 29, 29t aggregation 104
overload 24 Clonal secretion 104
leukocytosis 45t hematopoietic stem cell disorders 62 primary hemostasis 99, 100
leukopenia 46t proliferative disorder 95 white cells 43
lymphocytopenia 50t Combined vitamin B12 9 Disseminated intravascular coagulation
lymphocytosis 49t Common hereditary X-linked recessive 110, 110t, 111f
megaloblastic anemia 8t disease 106 Distinct megaloblasts in bone marrow 8
microcytic hypochromic anemia 8 Complement-mediated intravascular Donath-Landsteiner antibodies 40
monocytosis 49t hemolysis 41 Down syndrome 54
neutropenia 48t Confirmatory test 112, 115 Dry tap 75
neutrophilia 46 Congestive heart failure 7, 50 Dwarf megakaryocytes 70
pancytopenia 15t Consequences of Dysmegakaryopoiesis 61
thrombocytopenia 101, 101t ineffective erythropoiesis 23 Dysplasia 60
thrombocytosis 104t thrombi formation 111
Central nervous system 7, 12, 38 Coombs test 38, 38f, 39 E
Cervical lymph nodes 56 Crohn disease 49
Eczema 48
Chickenpox 49 Cutaneous T cell lymphoma 86
Ehlers-Danlos syndrome 100
Christmas disease 108 Cyclic neutropenia 48
Endemic Burkitt lymphoma 84
Chronic Cytochemistry of
Endothelial injury 110
antigenic stimulation 77 lymphoblasts 57
Enlarged cervical lymph nodes 124
atrophic gastritis 7, 12 myeloblasts 58
Eosinophilia 47
blood loss 5, 42, 121 Eosinophilic
eosinophilic leukemia 62 D
granuloma 48, 96
hemolytic anemia 32 Defective leukemia 48
hypoxia 32 DNA synthesis 4 Epstein-Barr virus 51
immune thrombocytopenic purpura heme synthesis 5 Erythrocytosis 63
102 hemoglobin synthesis 4 Erythroleukemia 53
infections 49 Deficiency of Erythromelalgia 66
leukemia 53 antithrombotic factors 113, 114 Erythrophagocytosis 21
lymphocytic leukemia 45, 48, 49, 53, intrinsic factor 11
Erythropoiesis 18, 66, 77
73, 74f, 81, 125 Degeneration of dorsal and lateral tracts
Esophageal webs 7
myelogenous leukemia 62, 68, 125 13
Extramedullary
myeloid leukemia 45, 47t, 48, 53, 70, Degree of hemoglobinization 3
hematopoiesis 24, 65-67
72f, 104 Dehydration 31
myelomonocytic leukemia 49 hemopoiesis 24
Delayed maturation of nucleus 8
myeloproliferative neoplasm 65 infiltration 56, 58
Demonstration of heterophile antibodies
neutrophilic leukemia 62 51 myeloid tumors 47
organ damage 33 Demyelination of spinal cord 13 Extravascular hemolysis 23
Classical Hodgkin lymphoma 81, 87, 88 Deoxyuridine suppression test 11
Classification of Dermatitis 48
F
acute myelogenous leukemia 57 herpetiformis 48 Fab classification of acute leukemias 53
anemia 3, 4t Development of thrombi 111 Fetal hemoglobin 26, 27
bleeding disorders 100, 100t Diffuse Fever 51
coagulation disorders 105, 105t chronic atrophic gastritis 12 Fibrinolysis abnormalities 112
disorders of hemostasis 100t large B cell lymphoma 81, 83 Fibrotic stage 67
hemolytic anemias 16 Dilute Russell’s viper venom test 115 Filariasis 48
hemostatic disorders 99 Dimorphic Fine needle aspiration cytology 93
hereditary defects in hemoglobin 22 anemia 9 Fluorescent in situ hybridization 70
immunohemolytic anemias 36, 36t peripheral blood picture 42 Folate deficiency 48
Index 137
Folic acid 8 Hereditary J

and iron deficiency 9 coagulation disorders 105, 106, 109t Jaundice 20
deficiency 8, 11, 15 disorders 29
Follicular lymphoma 81, 82, 82f hemolytic anemia 23 K
Fragmentation syndrome 41 spherocytosis 17, 122
Koilonychia 7
Functional disorders of platelet 104f High-performance liquid
nails 120
Fungal infections 48 chromatography 35
Kostmann syndrome 48
Hodgkin
G cells 87, 90, 91
L
Gallstones 20 lymphoma 48-50, 81, 87, 88t, 90f, 93,
93f, 94, 94t, 126 Lacunar cell. 89
Gaucher-like cells 70
Hookworm infestation 48 Langerhans cell 95
Giant
Humoral immune deficiency 80 histiocytosis 95
megakaryocytes 66 LE cell test 40
Hydrops fetalis 38
metamyelocytes 10 Letterer-Siwe disease 96
Hypercoagulable state 113, 115
Glandular fever 51 Leukemia 15, 55
Hypereosinophilic syndrome 48
Glanzmann thrombasthenia 100, 104 Leukemic meningitis 56
Hypersegmented neutrophil 9f
Glucose-6-phosphate dehydrogenase Leukemoid reaction 47
Hypocellular bone marrow 13
deficiency 20
Hypochromia 6 Leukocyte
Gold standard test 7 alkaline phosphatase 47
Hypofunction of spleen 33
Granulomatous common antigen 82
Hypovolemic shock 33, 41
diseases 15 Leukocytosis 45, 66
disorders 48 I Leukoerythroblastosis 67
Gum bleeding 102 Leukopenia 46
Immune
thrombocytopenia 115 Life-threatening bleeding 106
H Location of hemolysis 16, 17
thrombocytopenic purpura 102
Haemophilus influenzae 33 Löeffler’s syndrome 48
Immunoglobulin deposition disease
Hairy cell 75 76, 80 Loss of
leukemia 49, 75, 81 Immunohemolytic anemias 36 architecture 84
Hand-Schüller-Christian disease 96 Impaired intravascular volume 41
Hay fever 48 absorption 5, 8 lymph node architecture 89
Heavy DNA synthesis 8 membrane fragments in circulation 17
chain disease 81 red cell production 4 Lupus anticoagulant antibody 114
menstrual bleeding 120 Inactivates nitric oxide 31 Lymph nodes 74, 82, 86, 87
Heinz bodies 21, 21f Inactivation of nitric oxide 31 Lymphoblast 54
Hematologic malignancies 49, 104 Incubation period 51 Lymphocyte
Hematopoietic stem cell 63 Indirect antiglobulin test 39, 40 depleted classical Hodgkin lymphoma
Hemoglobin 6, 9, 21, 25, 58 Infectious 81, 87, 91
concentration 3 hepatitis 49 predominant cells 92
electrophoresis 26, 27, 34 mononucleosis 45, 49 rich classical Hodgkin
Hemolytic anemia 4, 16, 23 Inflammatory lymphoma 81, 87, 90
Hemolytic bowel disease 49 Lymphocytopenia 50
crisis 33 diseases 49 Lymphocytosis 49
disease of newborn 36, 39 Inherited hypercoagulable states 114 Lymphoid neoplasms 81
Hemophagocytic syndrome 15 Intestinal metaplasia 12 Lymphoma 15, 55
Hemophilia 106 Intravascular hemolysis 41 Lymphoplasmacytic lymphoma 81
A 106 Iron
B 108 deficiency 104, 120
M
Hemorrhage 41 anemia 5, 6, 8, 27, 28t, 42, 120 Macroangiopathic hemolytic anemia 41
Hemorrhagic deficient erythropoiesis 6 Malignant
bleeding 111 depletion 6 disease of bone marrow stem cell 52
diathesis 99, 111 overload 24 lymphoid
disorders 99 Irreversibly sickled cells 31 cells 82f
Henoch-Schönlein purpura 100 Ischemic necrosis 111 neoplasms 87
Mantle cell lymphoma 81 Mononuclear cells 51 O

Marrow Morphological classification of anemia 3t Osmotic fragility test 19, 19f, 122
aplasia 15 Morphology of
Osteosclerotic myeloma 76
infiltration 4, 48 myeloblasts 58
Massive splenomegaly 75 neoplastic cells 88
P
Mature Mucosal bleeding 102
Multiple Pancytopenia 13, 75
B cell neoplasms 81
myeloma 49, 76, 78f, 125 Pappenheimer bodies 42
T and NK cell neoplasms 81
tumor masses 76 Parietal cell antibody 12
T cell and NK cell neoplasms 85
Mumps 49 Paroxysmal
Mean corpuscular
Mutated tyrosine kinase activation 58 cold hemoglobinuria 40
hemoglobin 5
Mycosis fungoides 81, 86 nocturnal hemoglobinuria 15, 41
concentration 5, 31
Myelodysplastic syndromes 15, 48, 54, Pathogenesis of
volume 4
60, 104 iron deficiency anemia 6
Measles 49
Myelofibrosis 15, 64 megaloblastic change 8
Mediastinal Myeloid Rh hemolytic disease of newborn 37f
lymph nodes 88 hyperplasia 70
thymic mass 56 sickle cell anemia 30f
proliferation 54 thrombosis 111f
Medium-sized cells 84 sarcoma 54, 59
Megakaryocytic Pathognomonic
Myeloma 15
hyperplasia 123 Birbeck granules 95
kidney 79
leukemia 53 of Hodgkin lymphoma 88
plasma cells 77
Megakaryopoiesis 18, 66, 77 Pathophysiologic classification of
Myelomonocytic leukemia 53
polycythemia 63t
Megaloblastic Myelophthisic anemia 4
anemia 8, 9, 11, 13, 15, 46, 48, 121 Myelopoiesis 18, 66, 77 Patterson-Kelly syndrome 7
precursors 11f Myeloproliferative neoplasms 62, 68, 104 Paul Bunnell test 51
Menorrhagia 102 Pemphigus 48
Metastatic tumors 48 N Periodic acid Schiff stain 54f
Microangiopathic hemolytic anemia 41, Peripheral
Neoplastic
111 cells 88 blood smear 6f, 9f, 56
Microcytic hypochromic follicles 82 lymph nodes 90
anemia 7, 25, 120 T cells 86 T cell lymphoma 81, 85
red blood cells 6f Nephrotic syndrome 79 Pernicious anemia 11, 12
Micronormoblastic maturation 7 Neutropenia 47 Persistent thrombocytopenia 71
Microscopically myeloblasts 59 Neutrophilia 46 Pharyngitis 51
Microvascular thrombi 111 Nodular Philadelphia chromosome 47, 68, 69f, 70
Mild erythroid hyperplasia 27 lymphocyte predominant Hodgkin Pigment gallstones 122
Miliary tuberculosis 50 lymphoma 87, 92 Plasma
Missense point mutation 30 sclerosis classical Hodgkin lymphoma cell 89
81, 87, 88 myeloma 76
Mixed cellularity classical Hodgkin
lymphoma 81, 87, 90 Non-Hodgkin lymphoma 48, 49, 94, 94t neoplasm 76, 81
Non-neoplastic cells 87, 88 lactate dehydrogenase 11
Mode of
Non-steroidal anti-inflammatory drugs
inheritance 106 Plasmacytoma 76, 80
104
onset 16 Plasmodium falciparum gametocyte 132
Non-tender cervical lymphadenopathy
transmission 51 Platelet
125
Monoclonal Nonthrombocytopenic purpura 99 count 64, 107
gammopathy of Normochromic normocytic anemia 67 function disorders 100
uncertain significance 80 Normocytic normochromic anemia Plummer-Vinson syndrome 7
undetermined significance 76 38, 42, 70 Poems syndrome 76
spikes 78 Nucleated red cell precursors 25 Polychromatophilia 21, 33
Monocytic leukemia 53 Nucleus of macrophage 83 Polycythemia 48, 63
Monocytosis 49 Numbness of both lower limbs 121 vera 62, 63, 64f, 65f, 104
Monomorphic lymphoid cells 83 Nutritional deficiencies 15 Precursor lymphoid neoplasms 81
Index 139
Premature Reticulocyte Sickling

destruction of spherocytes 17 count 6, 25, 15, 18, 21, 33, 38 crisis 32
RBC destruction 36 hemoglobin 7 test 34, 35, 35f
Primary Rh Sideroblastic anemia 8, 42
amyloidosis 80 hemolytic disease of newborn 37 Skin
antiphospholipid syndrome 115 incompatibility 37 bleeding 102
hemostatic plug 99 Rheumatoid arthritis 49 diseases 48
myelofibrosis 62, 66 Ring sideroblasts 42 Small
Produces hemolytic anemia 30 Rouleaux formation 131 lymphocytic lymphoma 73, 81
Progressive cytopenias 60 Roundworm infestation 48 T lymphocytes 89
Proliferating hematopoietic stem cells 68 Russell’s viper venom 115 Sore throat 51
Promyelocytic leukemia 53 Spherocytes and raised osmotic
Protein C and S deficiency 114 S fragility test 122
Prothrombin time 107 Salmonella typhimurium 33 Splenic B cell marginal zone
Pseudo Gaucher cells 70 Sarcoidosis 15, 49 lymphoma 81
Pulmonary eosinophilia 48 Scabies 48 Spoon shaped nails 120
Schilling test 11, 12 Stages of
Q Secondary erythropoiesis 128
Qualitative antiphospholipid syndrome 115 Hodgkin lymphomas 94t
disorders of leukocytes 50 hemostatic plug 99 megakaryopoiesis 129
platelet disorders 101, 104 Sequestration crisis 33 myelopoiesis 132
Quantitative Serum Starry sky appearance 84f
and qualitative disorders of albumin 78 Striking basophilia 71
leukocytes 45 bilirubin 11, 34 Structure of red cell membrane 17f
deficiency of vWF 109 ferritin 7, 26 Subacute combined demyelination 13
disorders of leukocytes 45 folic acid levels 11 Subleukemic leukemia 52
platelet disorders 101 haptoglobin 26, 34 Substitution of glutamic acid 30
homocysteine 11 Sudden trapping of blood 33
R iron 7, 26 Suppression of stem cells 48
Raspberry-red tongue 121 and ferritin 11 Synthesis of fetal hemoglobin 23
RBC enzyme analysis 21 status 26 Syphilis 49
Reactive marrow fibrosis 66 transferrin Systemic lupus erythematosus 49
Recurrent splenic infarction 32 receptor 7
Red cell saturation 7
T
count 64 vitamin B12 11 T cell
distribution width 5 and uric acid 65 large granular lymphocytic
indices 43, 6, 9, 18, 25 Severe leukemia 81
protoporphyrin 7 anemia 7 prolymphocytic leukemia 81
size 3 hemolytic anemia 23, 38 T lymphoblastic leukemia/lymphoma 81
Reed-Sternberg cells 87, 88, 89f, 90, 91 infections 48 Target cells 122
Refractory Severity of bleeding 101 Thalassemia syndrome 22
anemia with Sex-linked recessive disorders 106 Thrombocytopenia 100, 101
excess blasts 61 Sézary Thrombocytosis 101, 104
ring sideroblasts 61 cells 86 Thrombotic disorders 99, 100, 113
cytopenia 61 syndrome 81, 86 Total
with multilineage dysplasia 61 Sickle iron binding capacity 26
Renal cell leukocyte count 46, 47, 51, 70
disease 80 anemia 29, 32f, 33, 122 plasma iron-binding capacity 7
failure 79, 104 disease 29 WBC count 58
function tests 78 trait 34 Tourniquet test 103
Repeated spontaneous abortions 115 hemoglobin 29 Toxoplasmosis 49
Respiratory distress syndrome 110 Sickled red cells 35f Traditional classification of leukemia 53t
Tropical eosinophilia 48 V WHO classification of

Tuberculosis 15, 49 acute
Vascular
Tumors 15 leukemia 53
disorders 110
Types of lymphoblastic and myeloid
purpura 99
antiglobulin test 39 leukemia 53t
Vessel wall abnormalities 99
immune thrombocytopenic Hodgkin lymphoma 87t
Viral
purpura 102 lymphoid neoplasms 81t
Langerhans cell histiocytosis 96t hepatitis 108
MPN 62
white blood cell 45 infections 45, 48, 49
myelodysplastic syndromes 61t
Vitamin
myeloproliferative neoplasm 62t
U B12 8, 15, 48
Widespread exfoliative erythroderma 86
absorption 12
Ulcerative colitis 49 Wiskott-Aldrich syndrome 100
deficiency 8, 11
Unconjugated bilirubin 38
K deficiency 105
Urine urobilinogen 26, 34
von Willebrand disease 105, 108
Urticaria 48
Uses of
W
direct antiglobulin test 39
indirect antiglobulin test 40 White cell count 64

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Rapid Review

Ramadas Nayak MBBS MD

Sharada Rai MBBS MD

JAYPEE BROTHERS MEDICAL PUBLISHERS (P) LTD

Jaypee Medical Inc Jaypee Brothers Medical Publishers (P) Ltd

Jaypee Brothers Medical Publishers (P) Ltd

© 2014, Jaypee Brothers Medical Publishers

Inquiries for bulk sales may be solicited at: jaypee@jaypeebrothers.com

Rapid Review of Hematology

First Edition: 2014

DEPARTMENT OF PATHOLOGY BANGALORE MEDICAL COLLEGE

AR Raghupathy MBBS MD PGDHHM (IGNOU)

How to use this book

Section 1: Disorders of Red Cells

Section 2: Disorders of White Cells

Section 3: Disorders of Hemostasis

Section 4: Clinical Scenario

ANEMIA Q. Define anemia.

Type of anemia Microcytic hypochromic Normocytic normochromic Macrocytic

Q. Write short notes on red cell RED CELL INDICES

2. Mean Corpuscular Hemoglobin (MCH) MCH < 26 pg is seen in

3. Mean Corpuscular Hemoglobin Concentration (MCHC) MCHC<31 g/dL is seen in

4. Red Cell Distribution Width (RDW) RDW is useful for

IRON DEFICIENCY ANEMIA

Etiology (Table 1.3) Q. Discuss the etiopathogenesis

TABLE 1.3: Causes of iron deficiency anemia

Stages of IDA in Pathogenesis of Iron Deficiency Anemia

Q. Discuss the laboratory findings Laboratory Findings

Bone Marrow Bone marrow shows

Serum Iron Profile (Table 1.4) Reduced: serum iron,

Reticulocyte Hemoglobin The earliest laboratory

Clinical Features of IDA Q. Mention the various clinical

Physical Findings Koilonychia (spoon nails)

Q. Enumerate the causes of Causes of Microcytic Hypochromic Anemia

Q. Discuss the causes and MEGALOBLASTIC ANEMIA

Etiology of Megaloblastic Anemia (Table 1.5)

Folic acid is absorbed in 1. Decreased Intake: inadequate diet—alcoholism, malnutrition

Deficiency of vitamin B12

Laboratory Findings of Megaloblastic Anemia Q. Write short note on

Fig. 1.5: Diagrammatic peripheral blood smear of dimorphic

Megaloblasts: Nuclear maturation Normal Lags behind cytoplasmic maturation

Biochemical Tests for Megaloblastic Anemia

Diagnostic tests for vitamin B12 deficiency

PERNICIOUS ANEMIA Q. Discuss the etiopathogenesis

PA: autoimmune disease Etiopathogenesis

Central Nervous System

Q. Write short note on laboratory Laboratory Findings (Fig. 1.8)

Specific Diagnostic Tests for Pernicious Anemia

Fig. 1.8: Clinical features and laboratory findings in pernicious anemia

Clinical Features of Megaloblastic Anemia Q. Mention the various clinical

APLASTIC ANEMIA Q. Write short notes on aplastic

6 “I” s of the causes of TABLE 1.7: Common causes of aplastic anemia

Pathogenesis: Pathogenesis (Fig. 1.9)

Fig. 1.9: Pathogenesis of aplastic anemia

–– Bleeding manifestations in the form of petechiae, bruises and ecchymoses due to

Bone marrow elements

No Splenomegaly Absence of splenomegaly

TABLE 1.8: Causes of pancytopenia

2 Hemolytic Anemias Due to

Q. Define and classify hemolytic HEMOLYTIC ANEMIA

TABLE 2.1: Classification and causes of hemolytic anemia

Location of Hemolysis Q. List the differences between

Etiopathogenesis HS, is due to defect in the