Leukocytes

Granulocytes

• neutrophils, eosinophils, basophils

  • heterophils (neutrophil-like) in avian, reptilian, & small mammal species

• segmented nuclei + cytoplasmic granules (vary by color + shape)

CBC leukocyte parameters

• leukogram: part of CBC involving number/type/morphology of WBCs

  • total leukocyte count

  • leukocyte differential: % of each leukocyte type

  • absolute number of each leukocyte type**

Total leukocyte count

• equals total nucleated cell count

  • increased nRBCs (>5%) needs correction factor

• determined by automated counting, manual counting w/hemocytometer + microscope, or visual estimation w/blood smear (least accuratee)

  • manual count good for non-mammalian samples

Leukocytosis

• increased total WBC count above reference interval

• often due to neutrophilia

• may be pathologic (inflammation, leukemia) or non-pathologic (stress, physiologic)

Leukopenia

• decreased WBCs below lower reference interval

• usually due to neutropenia

• always pathologic (increased utilization, decreased production, redistribution)

Panleukopenia

• decrease in all individual WBC types

Errors in total leukocyte counts

• false decrease: clumped/fragile

• false increase: clumped platelets or RBCs, excessive Heinz bodies dissociated from RBCs

Leukocyte differential

• microscope exam is gold standard for determination

  • count 100-200 cells on 100X oil in monolayer & classify

• multiply by total WBC to obtain absolute count

Absolute numbers

• best evaluation of leukogram

• reflects proportion of each WBC type in relation to count

Species variation in WBCs

• predominant neutrophils, small lymphocytes in carnivores

• equal neutrophils (more segmented) & lymphocytes in horses

  • often get neutropenia from inflammation

• predominant large lymphocytes w/low storage pool of neutrophils in ruminants/rodents

• increased eosinophils, more neutrophils than other ruminants in pseudoruminants

Mature segmented neutrophil

• most frequently seen WBC

• segmented nucleus, coarse + clumped chromatin

• numerous small cytoplasmic granules (pink)

• live in blood and tissues, function as host immunity

  • move into tissue inflammation sites, phagocytic, degrade via enzymatic digestion

Neutrophil granules

• packets that compartmentalize cellular constituents, multiple types w/different functions

  • fuse to phagosomes to kill/digest phagocytized material

  • fuse to cell membrane, exocytosis of adhesion factors/digestive enzymes

• primary, secondary, & tertiary at different stages of development

Neutrophil maturation

• proliferation & maturation: myeloblast → progranulocyte → myelocyte

• maturation only: metamyelocyte → band → segmented

• 7-10 days

Neutrophil pools

• bone marrow: proliferation + maturation & storage pools

• blood: circulating (blood flow) & marginal (pulled to side of vessel) pools

Band neutrophils

• U or S shaped nucleus (no pinched areas)

• precursor to segmented neutrophils, very low numbers in the blood during health

Left shift

• increased band neutrophils in the blood

• indicate inflammation requiring high levels of neutrophil response

  • increased severity with increased severity of inflammation

• appropriate (regenerative): segmented neutrophils > immature/bands

• inappropriate (degenerative): immature/bands > segmented neutrophils (more severe)

Neutrophil toxic changes

• morphologic features seen in blood caused by accelerated granulopoiesis (most common in bands)

• occur with moderate-severe inflammation

• cytoplasmic basophilia (blue cytoplasm): retained ribosomes

• Dohle bodies (blue dots): aggregates of rough ER

• toxic granulation: retained primary granules

• cytoplasmic vacuolation (clear spots): cell not fully developed

Degenerate neutrophils

• seen in tissues

• nuclear morphology changes

Hyper-segmented nucleus

• indicates longer time in circulation

• normal in horses

Neutrophil inclusions

• often caused by infectious agents

• Erlichia: obligate intracellular bacteria appear as morulae w/pancytopenia

• canine distemper: seen in acute phase of infection w/Diff-quik stain

Pelger Huet anomaly

• autosomal dominant inheritance causing hyposegmentation of granulocyte nuclei

• chromatin has mature pattern, function is normal

• automated analyzer will read results as left shift

Birman Cat neutrophil granulation anomaly

• autosomal recessive inherited condition

• neutrophils have fine eosinophilic - magneta granules

• neutrophil function is normal

Mucopolysaccharidoses

• neutrophils contain numerous distinct dark purple - magenta cytoplasmic granules

• heritable lysosomal storage disorder

• usually seen in young animals, progressive

• accompanied by neuro signs, ortho signs, facial dysmorphia, & corneal clouding ± lymphocyte abnormalities

Chediak Higashi

• autosomal recessive inherited condition

• breed/species specific

• neutrophils have large fused lysosomes that stain light pink/eosinophilic, some w/prominent cytoplasmic inclusions

• slight bleeding tendency from abnormal platelets, abnormal neutrophil function

Bovine leukocyte adhesion deficiency

• lethal autosomal recessive disorder

• mutation in surface adhesion molecule needed for neutrophil entry to tissue (predisposed to bacterial infections)

• shows up at neutrophilia

Heterophils

• similar function to neutrophils in non-mammals & some small mammals

• large red seed-shaped granules

Lymphocytes

• circulate between blood and tissue, most common WBC in cows/rodents

• small lymphocytes are most common (nucleus smaller in diameter than neutrophil) except for cows

• round to oval nucleus with dense clumped chromatin, scant cytoplasm

Reactive lymphocytes

• large with intensely basophilic cytoplasm, indented/irregular nucleus, & clumped chromatin

• seen in juveniles or with antigenic stimulation

Granular lymphocytes

• small number of pink-purple cytoplasmic granules, intermediate in size

• seen in normal ruminant blood

• often caused by antigenic stimulation

Lymphocyte vacuolation

• caused by ingestion of toxic plants or lysosomal storage disorder

Lymphocytosis

• caused by: epinephrine (physiologic), age (young animals, antigenic stimulation), chronic inflammatory disease, hypoadrenocorticism, neoplasia

Physiologic lymphocytosis

• epinephrine release causes increased blood flow rates, moving lymphocytes from marginal to circulating pool

  • cats + young horses get splenic contraction

• transient & mild

Chronic inflammatory lymphocytosis

• increased lymphocyte production due to chronic antigenic/cytokine stimulation

• caused by certain bacterial, fungal, parasitic, & viral infections + IMHA

• usually mild to moderate, often occurs with neutrophilia or monocytosis

BLV induced persistent lymphocytosis

• virus prevents apoptosis of virus-infected B lymphocytes & promotes proliferation of non-infected B-lymphocytes

• persistent, can develop into neoplasia

Hypoadrenocorticsm lymphocytosis

• decreased glucocorticoids increase circulating lymphocytes

Lymphoid neoplasia

• degree of lymphocytosis varies by type

Lymphopenia

• causes: corticosteroids, acute inflammation/systemic infections (marginating), loss (often viral), decreased production

Lymphopenia of acute inflammation

• caused by acute bacterial or viral infections

• lymphocytes marginate into inflamed + lymphoid tissues, decreased recirculation

• lymphocyte apoptosis associated with changes in cytokines

• partially due to corticosteroid stress response

Lymphocyte depletion

• GI loss (protein losing enteropathy) or neoplasia

• lymphangeictasia (rupture of intestinal lacteals)

• chylothrorax/abdomen from ruptured lymph vessels

Severe combined immunodeficiency

• seen in Arabian foals, autosomal recessive

• lack B & T lymphocytes

Monocytes

• produced in bone marrow, common ancestor with neutrophil

• hard to identify, nucleus not consistent shape

  • larger than neutrophils, less dense chromatin

• no storage pool in bone marrow, circulate briefly in blood

• migrate to tissues to differentiate into macrophages - phagocytic, antigen presenters, recycle old RBCs

Monocytosis

• causes: inflammatory/infectious disease, corticosteroids, leukemia, paraneoplastic response (uncommon)

Inflammatory/infectious monocytosis

• acute or chronic granulomatous inflammation (fungi, mycobacteria, protozoa)

• increases production of monocytes

Monocytopenia

• usually not clinically significant

• rarely may be due to bone marrow injury

Eosinophils

• defend against parasites, cause allergic inflammation & immune-complex reactions

• red - orange granules, segmented nuclei

Eosinophilia

• worms: parasites (inconsistent finding, not seen with organisms that infect blood cells)

• wheezes: hypersensitivity/allergy, asthma, anaphylaxis (inconsistent finding)

• weird diseases: hypoadrenocorticism, fungal infections, neoplasms via IL-5

Hypereosinophilic syndrome

• idiopathic marked eosinophilia due to increased bone marrow production

• increase of mature, segmented, morphologically normal eosinophils in the blood (>15000/uL) for 6+ months

• eosinophils infiltrate tissues/organs, cause organ damage via cytotoxic substance release

• diagnosis of exclusion

Eosinopenia

• difficult to detect, lower reference interval often 0

• usually of limited clinical significance

• can be caused by corticosteroids (stress leukogram), increased margination, decreased marrow release, or acute inflammation

Basophils

• very low number in blood in health

• function unknown, associated with parasitic infections & allergic inflammation

• larger than neutrophils, segmented nuclei

• cytoplasmic granules vary based on species

Basophilia

• rare, often concurrent with eosinophilia

• caused by diseases that increase IgE production

• most common cause is heartworm (in dogs)

  • also seen with allergic reactions or neoplasia

Basopenia

• not clinically significant

Neutrophilia

• increased segmented neutrophils

• may be from epinephrine response (physiologic) or corticosteroids (stress)

• pathologic causes are inflammation or neoplasia

• most common cause of leukocytosis

Stress leukogram

• may have normal or mildly elevated total WBC count, no neutrophil left shift or toxic changes

• caused by increased corticosteroids (endogenous or exogenous)

• lymphopenia, eosinopenia, monocytosis, neutrophilia

  • not all changes present in every patient, most commonly lymphopenia & neutrophilia

• may see hyperglycemia and/or increased ALP (dog)

Causes of stress leukogram signs

• neutrophilia: increased neutrophil release from storage pool, decreased neutrophils in marginal pool & movement into tissues - not due to increased production

• lymphopenia: increased margination & tissue sequestration, decreased production/activation/proliferation

• eosinopenia: increased margination, tissue sequestration, & apoptosis, decreased bone marrow production & release

• monocytosis: increased mobilization of marginal pool to circulating pool

Physiologic leukocytosis

• caused by epinephrine (exercise, excitment)

• most common in cats & young horses

• neutrophilia + lymphocytosis

  • no left shift or toxic changes, normal monocytes, eosinophils, and basophils

• may see hyperglycemia

Causes of physiologic leukocytosis signs

• neutrophilia: increased blood flow moves marginal pool neutrophils into circulation

• lymphocytosis: release from the spleen, transient

Inflammatory leukogram

• increased total WBCs

• usually neutrophilia (increased production), ± left shift and toxic changes

  • can see neutropenia depending on time frame, severity, & species

• might have monocytosis

• caused by any condition associated with increased tissue demand

Inflammatory leukogram by species

• dogs + pigs: marked neutrophilia

• cats + horses: intermediate neutrophilia

• cows: neutropenia

• depends on amount of bone marrow storage

Acute inflammation

• releases cytokines that signal bone marrow to produce leukocytes

• increased margination + tissue migration triggers proliferative pool & storage pool

Chronic inflammation

• 1+ weeks of inflammation

• band neutrophils decrease, bone marrow can release segmented neutrophils again

Neoplastic neutrophilia

• hematopoietic neoplasia (leukemia): causes extreme neutrophilia/leukocytosis

• non-hematopoietic neoplasms: inflammatory cytokines/growth factors from tumor cause paraneoplastic neutrophilia

Adhesion molecule disorders

• cause marked neutrophilia & chronic recurrent infections

Prognostic indicators of neutrophilia

• magnitude of neutrophilia

• degree of left shift (more band neutrophils, worse prognosis)

• presence/severity of toxic changes

Leukemoid response

• marked leukocytosis with marked segmented neutrophila (>50)

• rare, granulopoiesis triggered by infection, inflammation, or neoplasia

• often marked left shift + toxic changes

  • cells still more mature than in leukemia

• may have anemia of inflammatory disease

Neutropenia

• causes risk for secondary infection

• always pathologic, caused by increased consumption (inflammation) or margination (endotoxemia), peripheral destruction, decreased production

Inflammatory neutropenia

• increased consumption not met by bone marrow production

• often occurs with left shift + toxic changes

• more common in cattle, indicates severe inflammation with other species

Margination neutropenia

• caused by endotoxemia (Gram - bacteria)

• occurs within first few days, bone marrow increases production after 3-5 days

Peripheral destruction neutropenia

• primary immune-mediated destruction

• uncommon

• myeloid hyperplasia from bone marrow

Decreased production neutropenia

• persistent with no left shift

• due to bone marrow damage or suppression (infectious, neoplastic, drug-mediated, primary disease)

Canine cyclic hematopoiesis

• very rare, seen in grey collies

• autosomal recessive stem cell disorder

• arrest of hematopoiesis at 11-14 day intervals

• susceptible to infections due to neutropenia

• can be cured with bone marrow transplant