HEMA1 - White Blood Cells Morphology and Disorders

Innate Immunity

Naturally occurring, non-specific first-line defense mediated primarily by phagocytic myeloid cells (Neutrophils, Monocytes, Eosinophils, Basophils)

Adaptive Immunity

Acquired, specific defense mediated by immunocytes (T lymphocytes, B lymphocytes → plasma cells)

Leukocyte Classification — 3 Schemes

1. By Progenitor: Myeloid (Neutrophils, Eos, Basos, Monos) vs. Lymphoid (T, B, NK lymphocytes)
   2. By Function: Phagocytes (Neutrophils, Eos, Basos, Monos/Macs) vs. Immunocytes (Lymphocytes)
   3. By Physical Attributes: Mononuclear/Agranulocytes (Monos, Lymphs) vs. Segmented/Granulocytes (Neutrophils, Eos, Basos)

Diapedesis

The outward passage of blood cells through intact vessel walls as cells migrate between endothelial cells lining capillaries. Leukocytes are transient in blood and transfer to peripheral tissues via diapedesis.

Leukocyte Lifespans

• Neutrophils: 7 hours in blood; variable in tissues
• Eosinophils: 18 hours in blood (longer in eosinophilia); at least 6 days in tissues
• Basophils: 60 hours in blood; variable in tissues
• Monocytes: 3 days in blood; variable (months as macrophages) in tissues
• Lymphocytes: Days (B cells), Months to years (T cells) in blood; long-lived/recirculating

Chemotaxis

Directional cellular movement along a chemical concentration gradient

Positive Chemotaxis

Movement toward the stimulus (e.g., neutrophil migration toward bacterial chemokines)

Negative Chemotaxis

Movement away (repulsion) from a chemical stimulus

Opsonization

The process by which antibodies (IgG) or complement fragments (C3b) attach to foreign antigen or microorganisms to facilitate recognition and binding by phagocyte membrane receptors.

Pinocytosis

Cellular ingestion of small soluble particles or fluid droplets ('cell drinking')

Phagocytosis

Cellular ingestion of large insoluble particles or live microorganisms ('cell eating')

Phagocytosis — 4 Sequential Stages

1. Recognition and Attachment: Opsonized particles bind to phagocyte Fc and complement receptors
   2. Ingestion: Pseudopods enclose particle in a phagosome; actin/myosin pulls it centrally
   3. Killing and Digestion: Oxygen-dependent (respiratory burst → superoxide, H₂O₂, HOCl) and Oxygen-independent (lysosomal enzymes)
   4. Formation of NETs / Exocytosis: Extrusion of DNA traps or debris

Neutrophil Extracellular Traps (NETs / NETosis)

A specialized cell death pathway where nuclear/organelle membranes dissolve, activated enzymes attach to DNA, and the plasma membrane ruptures to release a web of chromatin and antimicrobial enzymes that ensnares and digests extracellular bacteria.

Four Criteria in Leukocyte Identification

1. Cell Size (µm)
   2. Nuclear-Cytoplasmic (N:C) Ratio
   3. Cytoplasmic Characteristics (background color, presence/absence of granules, granule color and size)
   4. Nuclear Characteristics (shape, color, chromatin pattern, presence/absence of nucleoli)

Normal Segmented Neutrophil (PMN) — Morphology & Function

• Size: 10–15 µm
• Nucleus: 2 to 5 lobes connected by thin filaments; coarse clumped chromatin
• Cytoplasm: Fine lilac-pink specific granules distributed diffusely (do not lie over nucleus)
• Function: Phagocytosis of bacterial pathogens and acute inflammatory response

Normal Band Neutrophil — Morphology & Function

• Size: 10–15 µm
• Nucleus: Curved, elongated sausage or C, S, Z shaped nucleus without filament constriction; coarse clumped chromatin
• Cytoplasm: Same as PMN (lilac-pink granules)
• Function: Phagocytosis of bacterial pathogens

Normal Eosinophil — Morphology & Function

• Size: 10–15 µm
• Nucleus: Segmented into 2 lobes (bilobed); coarse clumped chromatin
• Cytoplasm: Large, spherical, reddish-orange granules that do not lie over the nucleus
• Function: Immune defense against parasitic helminths and modulation/regulation of allergic responses

Normal Basophil — Morphology & Function

• Size: 10–14 µm
• Nucleus: Indistinct, unsegmented or bilobed
• Cytoplasm: Unevenly distributed, variable-sized, water-soluble dark bluish-black granules that lie over and obscure the nucleus
• Function: Initiator of allergic, hypersensitivity, and immediate anaphylactic reactions

Normal Monocyte — Morphology & Function

• Size: 10–30 µm (largest cell in normal peripheral blood)
• Nucleus: Indented, kidney-bean shaped or folded; delicate raked/lacy chromatin
• Cytoplasm: Blue to gray, ground-glass appearance with minute dust-like azurophilic granules and frequent vacuoles
• Function: Phagocytosis of foreign material, debris removal, and antigen presentation (APC)

Typical Lymphocyte

6–9 µm; compact round nucleus almost occupying entire cell; scanty sky-blue cytoplasm devoid of granules

Atypical Lymphocyte

12–15 µm; immature-looking nucleus; abundant deeply basophilic to pale blue cytoplasm; antigen-reactive sensitive cell

Lymphocyte Function

B cells produce antibodies and fight viruses; T cells mediate adaptive cellular immunity

Neutrophilia — Diagnostic Cut-Off & Etiologies

• Cut-Off: Absolute neutrophil count > 7.0 × 10⁹/L (or > 8.5 × 10⁹/L)
• Pathological: Bacterial infections, appendicitis, RA, acute tissue necrosis (pancreatitis, colitis, MI, severe hemolysis, burns, trauma), parasites (malaria, liver flukes), lithium, lead, mercury, digitalis, phenacetin, corticosteroids, myelogenous leukemia, venom, Actinomyces
• Physiologic: Exercise, extreme temperatures, nausea, vomiting, pregnancy, labor, panic, stress

Pseudoneutrophilia (Physiologic Neutrophilia)

Transient redistribution of neutrophils from the marginal granulocyte pool to the circulating granulocyte pool induced by exercise, temperature extremes, nausea, vomiting, pregnancy, labor, panic, rage, or acute emotional stress without increased bone marrow production.

Neutropenia & Agranulocytosis — Diagnostic Cut-Offs & Causes

• Neutropenia: Neutrophil count < 1.5 × 10⁹/L (or < 1.2 × 10⁹/L); caused by chemical toxicity (benzene), marrow replacement, nutritional deficiency (B12/folate), cytotoxic drugs, chemotherapy, myelokathexis
• Agranulocytosis: Absolute neutrophil count < 0.5 × 10⁹/L; seen in response to amidopyrine and cephalosporin toxicity

Myelokathexis

A congenital disorder of granulocytes characterized by severe chronic neutropenia and bone marrow hyperplasia, resulting from the intrinsic inability of the bone marrow to release mature granulocytes into the peripheral blood.

Eosinophilia — Diagnostic Cut-Off & Causes

• Cut-Off: Absolute eosinophil count > 0.35 × 10⁹/L
• Causes: Active allergic disorders (asthma, hay fever, eczema), parasitic helminth infections, psoriasis, scarlet fever, eosinophilic leukemia (not characteristic of protozoan infections)

Eosinopenia — Causes & Mechanism

Decreased eosinophil count caused by ACTH administration (which stimulates tissue diapedesis of eosinophils), acute glucocorticoid therapy, or acute systemic stress/infection.

Basophilia — Diagnostic Cut-Off & Causes

• Cut-Off: Absolute basophil count > 0.2 × 10⁹/L (> 0.075 × 10⁹/L per Turgeon)
• Causes: Immediate hypersensitivity reactions, hypothyroidism, ulcerative colitis, chronic urticaria, estrogen therapy, myeloproliferative disorders

Basopenia — Causes

Decreased basophil count associated with elevated corticotropin (ACTH), progesterone administration, ovulation, acute infections, stress, hyperthyroidism (thyrotoxicosis), and increased glucocorticoids.

Monocytosis — Diagnostic Cut-Off & Causes

• Cut-Off: Absolute monocyte count > 1.0 × 10⁹/L
• Causes: Tuberculosis, subacute bacterial endocarditis (SBE), syphilis, protozoan and rickettsial infections (malaria, typhus), brucellosis, typhoid fever, Gaucher disease, Hodgkin disease, collagen vascular disorders (SLE), GI diseases, surgical trauma, recovery from neutropenia

Monocytopenia — Diagnostic Cut-Off & Causes

• Cut-Off: Absolute monocyte count < 0.2 × 10⁹/L
• Causes: Prednisone/corticosteroid therapy, Hairy Cell Leukemia, aplastic anemia, overwhelming infections, hemodialysis, Epstein-Barr virus infection

Lymphocytosis — Diagnostic Cut-Off & Causes

• Cut-Off: Absolute lymphocyte count > 4.0 × 10⁹/L (> 8.8–9.0 × 10⁹/L in infants/children)
• Causes: Normal in infants/children < 10 yrs; viral infections (infectious mononucleosis, CMV, acute viral hepatitis), Bordetella pertussis (whooping cough), acute infectious lymphocytosis, Brucellosis, Toxoplasmosis

Lymphocytopenia (Lymphopenia) — Diagnostic Cut-Off & Causes

• Cut-Off: Absolute lymphocyte count < 1.0 × 10⁹/L (< 2.0 × 10⁹/L in children)
• Causes: HIV/AIDS infection, cytotoxic/immunosuppressive chemotherapy, systemic debilitation, congenital immunodeficiency, abnormal lymphatic circulation

Toxic Granulations — Morphology & Mechanism

• Morphology: Prominent dark blue/purple azurophilic primary granules in band/segmented neutrophils and monocytes; peroxidase-positive; graded 1+ to 4+
• Mechanism: Precipitation of ribosomal RNA caused by metabolic toxicity during accelerated granulopoiesis
• Clinical: Severe bacterial infections, sepsis, toxemia, burns

Döhle (Inclusion) Bodies — Morphology & Composition

• Morphology: Single or multiple light blue-staining round/oval cytoplasmic inclusions in neutrophil periphery
• Composition: Aggregates of rough endoplasmic reticulum (rRNA)
• Clinical: Severe infections, burns, trauma, scarlet fever, pregnancy; also seen in May-Hegglin anomaly

Hypersegmentation of Neutrophils

• Definition: Segmented neutrophils with > 5 nuclear lobes
• Clinical Significance: Pathognomonic feature of Vitamin B12 or folic acid deficiency (megaloblastic anemia); accompanied by macroovalocytes

May-Hegglin Anomaly — Genetics & Triad

• Genetics: Autosomal dominant (MYH9 gene mutation)
• Triad: Large, pale blue Döhle body-like inclusions in all leukocytes + giant, poorly granulated platelets + thrombocytopenia
• Clinical: Benign condition with mild bleeding tendency from low platelet count

Alder-Reilly Anomaly — Genetics & Composition

• Genetics: Autosomal recessive
• Morphology: Heavy, coarse, purple-red azurophilic granules in cytoplasm of neutrophils, eosinophils, basophils, and monocytes (resembles coarse toxic granulation)
• Composition: Precipitated mucopolysaccharides in lysosomes
• Clinical: Mucopolysaccharidoses (Hurler, Hunter, Maroteaux-Lamy syndromes); leukocyte function is normal

Pelger-Huët Anomaly — Genetics & Morphology

• Genetics: Autosomal dominant (LBR gene mutation)
• Morphology: Hyposegmentation of mature neutrophils into bilobed dumbbell, pince-nez, or spectacle shapes (heterozygotes) or round/oval unsegmented nuclei (homozygotes) with mature, coarse clumped chromatin
• Clinical: Benign; cell function is normal; must differentiate from acquired pseudo-Pelger-Huët (MDS, AML)

Chédiak-Higashi Syndrome — Genetics, Morphology & Pathophysiology

• Genetics: Autosomal recessive (LYST gene defect)
• Morphology: Giant, peroxidase-positive lysosomal granules in neutrophils, monocytes, lymphocytes, and melanocytes
• Pathophysiology: Defective vesicle trafficking/fusion → impaired chemotaxis and delayed bactericidal degranulation
• Clinical: Partial oculocutaneous albinism, recurrent pyogenic infections, peripheral neuropathy, early death

Chronic Granulomatous Disease (CGD) — Defect & Presentation

• Defect: Mutations in NADPH oxidase enzyme complex subunits (predominantly X-linked)
• Pathophysiology: Failure to generate superoxide (O₂⁻) and H₂O₂ (defective respiratory burst)
• Clinical: Recurrent severe infections with catalase-positive organisms (S. aureus, Aspergillus, Burkholderia); normal phagocytosis and degranulation

Myeloperoxidase (MPO) Deficiency

• Defect: Congenital or secondary (MPD, diabetes, drugs, pregnancy) deficiency of MPO enzyme
• Pathophysiology: Failure to convert H₂O₂ + Cl⁻ into hypochlorous acid (HOCl)
• Clinical: Mildly delayed intracellular bactericidal killing; usually compensated and clinically mild or asymptomatic

Specific Granule Deficiency (SGD)

• Defect: Rare congenital absence of neutrophil secondary (specific) granules
• Morphology: Atypical bilobed nuclei in neutrophils
• Clinical: Recurrent severe bacterial infections and defective neutrophil chemotaxis

Leukocyte Adhesion Deficiency (LAD)

• Defect: Mutations in the β₂-integrin subunit (CD18/CD11 complex)
• Pathophysiology: Leukocytes cannot adhere to vascular endothelial cells and fail to undergo diapedesis into tissues
• Clinical: Extreme peripheral blood leukocytosis with absence of pus formation at infection sites; delayed umbilical cord separation

Normal Lymphocyte Differential & CD4/CD8 Ratio in Adults

• Relative Differential: 20% to 40% of total circulating leukocytes
• Absolute Count: 1.5 to 4.0 × 10⁹/L
• Normal CD4/CD8 T-cell Ratio: 1.0 to 3.4

Acute Infectious Lymphocytosis

• Etiology: Coxsackievirus (A, B6), Echovirus, Adenovirus Type 12
• Epidemiology: Contagious childhood disease
• Diagnostic Features: Marked absolute lymphocytosis (20 to 50 × 10⁹/L) with mature, normal-appearing small typical lymphocytes

Whooping Cough (Bordetella pertussis) — Hematologic Findings

• Etiology: Bordetella pertussis bacterial infection, most severe in infants
• Diagnostic Features: Marked absolute lymphocytosis (> 30 × 10⁹/L) with characteristic cleaved or clefted lymphocyte nuclei

Persistent Polyclonal B-Cell Lymphocytosis

• Epidemiology: Rare condition primarily affecting young female smokers and post-splenectomy patients
• Diagnostic Features: Binucleated lymphocytes on peripheral smear; polyclonal serum IgM elevation; immunophenotype IgM+ / IgD+ / CD27+ memory B cells

Infectious Mononucleosis (IM) — Etiology & Smear Features

• Etiology: Epstein-Barr Virus (EBV)
• Clinical Triad: Fever, sore throat (pharyngitis), lymphadenopathy + splenomegaly
• Diagnostic Features: Presence of ≥ 10% reactive lymphocytes (Downey cells, which are cytotoxic CD8+ T cells reacting to EBV-infected B cells) and positive heterophile antibody test

Cytomegalovirus (CMV) Infection — Comparison with IM

• Clinical Presentation: IM-like syndrome with reactive lymphocytosis, malaise, and pharyngitis
• Key Differentials from IM: Heterophile antibody test is negative; characteristically accompanied by an increase in cold agglutinins

Toxoplasmosis (Toxoplasma gondii) — Histopathology

• Etiology: Toxoplasma gondii (protozoan infection)
• Clinical Presentation: Resembles IM; only a small number of immunocompetent patients are symptomatic
• Histopathology: Lymph node biopsy demonstrates reactive follicular hyperplasia with epithelioid histiocytes that characteristically blur the germinal center and mantle zone borders