Blood Components: Cells of the Blood Foundations 2023 Session 12 Outline • Where does it all begin? (Hematopoiesis) • Red blood cells (RBC) – structure and function • = Erythrocytes • Species differences • White blood cells (WBC) – structure and function • = Leukocytes • Platelets (PLT) – structure and function • May also be called thrombocytes, especially in non-mammalian species A brief overview of hematopoiesis • What: the production of blood cells (“hem-” = blood, “-poiesis” = production) • Where: predominantly within the bone marrow of adult animals • Minor amounts in the spleen (most animals) +/- liver and kidney (amphibians, fish) • Some WBCs (lymphocytes) complete their maturation outside of the marrow in lymphoid tissues • When: begins in the embryo, and continues throughout the life of the animal • Why: all blood cells are essential for maintaining life, but they don’t live forever so continuous production is needed • It is important to understand normal hematopoiesis so that we can identify pathologic alterations in the process • How: starting with the earliest blood cell, a hematopoietic stem cell (HSC), a series of differentiating steps and cell divisions results in the production of mature red blood cells, platelets, and all types of white blood cells Hematopoiesis as a continuous process – why? • Once mature blood cells leave the bone marrow and enter circulation, their time is limited • Normal RBC lifespan in the blood: • Cats: 65-76 days (~2 months) • Dogs: 110-120 days (~4 months) • Cattle: 130-160 days (~6 months) • Horses: 140-150 days (~6 months) • Normal WBC lifespan: • Neutrophils: 5-15 hours in the blood lifespan in tissues can be prolonged by inflammatory stimuli • Monocytes: Hours to 3 days in blood longer in tissues (where they become macrophages) • Eosinophils and basophils: ~1-2 days (may be longer with appropriate stimuli) • Lymphocytes*: Days-years (these are the only WBCs that can go back and forth between blood and (lymphoid) tissues!) • Normal PLT lifespan in the blood: ~5-7 days Red blood cells (RBC), aka erythrocytes • Functions: • Transport oxygen (from lungs to tissues) • Transport carbon dioxide (from tissues to lungs) • Buffer excess hydrogen ions (protons) to maintain acid-base balance • RBCs carry hemoglobin (Hgb), which is the key to all RBC functions This Photo by Unknown Author is licensed under CC BY “Eryth-” = red RBC structure • Mammals • Most mammals have anucleate erythrocytes in the shape of a biconcave disc (discocytes) • Biconcavity is best visualized in 3D (see scanning electron micrograph ) • In two dimensions on a typical blood smear, the central biconcavity appears as a region of central pallor, with the red hemoglobin present in a ring around the periphery of the cell • This is best observed in canine RBCs, which are some of the larger mammalian RBCs • Discocyte shape helps to increase RBC surface area to better enable gas exchange, and to make the cell deformable as it flows through vessels of varying sizes • Depend on anaerobic glycolysis for energy Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey RBC structure: mammals • Cats, ruminants, and horses also have discocyte-shaped RBCs, but they often lack central pallor on blood smears • RBCs from these species are much smaller than canine RBCs • Camelids • Camel, llama, alpaca, vicuña, guanaco • RBCs are anucleate, but elliptical or oval in shape, and very flat RBC structure: non-mammalian species • Elliptical or oval in shape • Nucleated! • Tend to be larger than mammalian erythrocytes Image from Atlas of Veterinary Hematology, 2nd ed., J Harvey Normal life of an erythrocyte • Produced in the bone marrow This Photo by Unknown Author is licensed under CC BY • The hormone erythropoietin (EPO) stimulates RBC production • Takes 3-4 days for a committed erythrocyte progenitor cell to produce mature RBCs • Mature erythrocytes released into the blood • Circulating lifespan: 2-6 months (species dependent) • Aged erythrocytes are removed from the blood by macrophages in the spleen • Premature removal of RBCs from the blood can result in anemia How we assess RBCs clinically – CBC + blood smear! • Measures of erythrocyte mass: • RBC count (RBC) • Hematocrit (HCT) or packed cell volume (PCV) • Hemoglobin concentration (Hgb) Increased RBC mass = erythrocytosis Decreased RBC mass = anemia • Erythrocyte indices: • RBC size – mean cell volume (MCV) • RBC hemoglobin content – mean cell hemoglobin concentration (MCHC) • Erythrocyte production: • Reticulocyte count • Erythrocyte morphology: • Shape changes • Inclusions • Infectious organisms EClinPath White blood cells (WBC), aka leukocytes • Granulocytes “Leuk” = white • Neutrophils • Eosinophils • Basophils • Mononuclear cells • Lymphocytes • Monocytes • Rarely seen in blood of healthy animals • Mast cells • Plasma cells EClinPath Increased total WBC = leukocytosis Photo courtesy of Laurie Holm Decreased total WBC = leukopenia Neutrophil function • Key player in the host’s innate immune system • Primary defense against invading bacteria • Leave the blood and enter tissues in response to cellular and chemical signals of inflammation • Cytokines and chemokines • Phagocytosis (“cell eating”) of bacteria and other pathogens • Ingested bacteria are neutralized by chemical and enzymatic reactions inside the neutrophil cytoplasm (enzymes and chemicals contained in neutrophil granules) • Granule contents and degraded bacteria released into surrounding tissue after digestion • Fun fact: non-mammalian “neutrophils” are called heterophils • Different morphology, similar function! Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey Neutrophil structure and morphology Dog • Cell size: larger than RBCs and lymphocytes; smaller than monocytes • Segmented nucleus with condensed (very darkly staining) chromatin Dog • 2-5 nuclear segments (lobes separated by thin constrictions) are typical of mature neutrophils Cow Horse Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey • Cytoplasm contains numerous granules, which are often colorless (“neutral”), but may be light pink in some animals Normal life of a neutrophil • Produced in the bone marrow • The production and maturation process can be accelerated with inflammation • Stored in the bone marrow • There is a “storage pool” of neutrophils in the bone marrow, which contains mature (segmented) neutrophils and lower numbers of immature neutrophils • With acute inflammation, the storage pool is utilized first while new cells are being made • Mature neutrophils released into the blood • Circulating lifespan very short: 5-15 hours • Migration into tissue • Inflammatory stimuli in the tissues signal to neutrophils to leave the blood • Neutrophils may survive for a few days in tissues, then undergo apoptosis and are cleared by tissue macrophages How we assess neutrophils clinically: CBC + blood smear! • Neutrophil numbers Increased neutrophil numbers = neutrophilia Decreased neutrophil numbers = neutropenia • Relative (%) • Absolute concentration • Segmented neutrophils (“segs”): mature cells that comprise the majority of all WBC in the blood of healthy animals • Neutrophil maturity • Band neutrophils (“bands”): immature neutrophils that may be present in very low numbers in health • Increased numbers of bands in the blood indicate active tissue inflammation • Neutrophil morphology • Certain changes in the neutrophil cytoplasm can indicate an accelerated rate of cell production • Inclusions • Infectious organisms Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey Eosinophil and basophil function • Key players in immediate (type I) hypersensitivity reactions • Allergic diseases • Provide host defense against parasite infections • Helminths (worms) • Insects (fleas) • Some fungi • Cytoplasmic granules contain unique chemicals and enzymes • Defense against invading pathogens • Attractants for other inflammatory and immune system cells • Basophil granules contain histamine • “Worms, wheezes, and weird diseases” Eosinophils – morphology Cat Dog • Cell size: eosinophils generally equal to or slightly larger than a neutrophil • Segmented nuclei with condensed chromatin, similar to neutrophils • Cytoplasm contains distinctly colored granules • Eosinophil granules of most mammalian species are orangepink in color, and tend to be numerous Horse Cow Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey • Extreme species variability in eosinophil granule color and shape! Stacy and Raskin. Reptilian eosinophils. Vet Clin Pathol 44/2 (2015) 177–178 ©2015 American Society for Veterinary Clinical Pathology Basophils - morphology • Cell size: generally equal to or larger than a monocyte • Often the largest WBC in circulation (when present!) Dog Dog • Segmented nuclei with condensed chromatin, similar to neutrophils • Basophil chromatin may occasionally appear slightly less condensed, and the nucleus itself may be “ribbon-like,” lacking distinct constrictions between lobes Cat Cat Cow Horse Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey • Cytoplasm contains distinctly colored granules • Basophil granules of most mammalian species are dark purple in color, but may be very few in number (esp. dogs) • In cats, granules are often light lavendergrey in color How we assess eosinophils and basophils clinically • Cell numbers (CBC) • Relative (%) • Absolute concentrations Increased eosinophil numbers = eosinophilia Decreased eosinophil numbers = eosinopenia* • Cell maturity (blood smear) • Banded eosinophils/basophils indicate active production of these cells • There are no bone marrow storage pools for eosinophils or basophils Increased basophil numbers = basophilia *Decreased numbers of eosinophils or basophils are not clinically significant, and often not detectable. Lymphocyte functions • T-lymphocytes (T-cells) • Cell-mediated immunity • Delayed hypersensitivity reactions (type IV) • Regulation of other immune and inflammatory processes • B-lymphocytes (B-cells) • Production of immunoglobulins (antibodies) • Natural killer (NK) cells • Cytotoxicity • Can kill virus-infected and tumor cells • Regulation of other immune and inflammatory processes Lymphocyte morphology • Cell size: mature lymphocytes are the smallest WBC in the blood – only slightly larger than an erythrocyte • Should be smaller than a neutrophil • Nucleus is small, but takes up most of the cell (high nucleus to cytoplasm ratio) Normal - dog Normal - cow • Round to oval shaped nucleus • Chromatin is highly condensed (very dark staining) • Small amount of pale blue (basophilic) cytoplasm Reactive - cow Granular - cow Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey • Low numbers of lymphocytes may contain a few dark pink granules • Reactive lymphocytes may be larger and have more abundant cytoplasm (seen with antigen stimulation) Lymphocyte production (lymphopoiesis) & circulation • Produced in the bone marrow (low overall %) and to thymus for further lymphoid organs (lymph differentiation and nodes, spleen, thymus) maturation • Unlike other WBCs, lymphocytes are able to reenter lymphoid tissues from the blood • Circulation time in blood may be very short, but some cells (especially memory B-cells) can live for years in lymphoid tissue How we assess (blood) lymphocytes clinically – CBC & blood smear! • Lymphocyte numbers • Relative (%) • Absolute concentration • Increase in lymphocytes = lymphocytosis • Decrease in lymphocytes = lymphopenia • Lymphocyte morphology • Size (small, intermediate, large) • Compared to size of neutrophil • Nuclear and cytoplasmic changes associated with reactivity (inflammation, antigen stimulation) vs. neoplasia (immature, atypical cells) • Not always possible to differentiate reactive from neoplastic lymphocytes via light microscopy alone • Granules, vacuoles, other inclusions in cytoplasm Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey Monocyte function • Phagocytosis of invading pathogens • Presenting antigens to T-lymphocytes to stimulate immune response to pathogens • Regulation of other inflammatory and immune processes • Once monocytes leave the blood and enter tissues, they differentiate into macrophages or dendritic cells • Similar functions, different name Monocyte morphology • Cell size: larger than granulocytes; often the largest WBC present in blood • Nucleus is variably shaped, with less densely-stained chromatin than other WBCs • Round to oval, kidney beanshaped, band-shaped, irregular (“amoeboid”) • Moderate to abundant amounts of pale to moderately blue (basophilic) cytoplasm Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey • Monocyte cytoplasm often contains variable numbers of discrete clear vacuoles How we assess monocytes clinically – CBC + blood smear! • Monocyte numbers • Relative (%) • Absolute concentration • Increased in blood with chronic inflammation • Increase in monocytes = monocytosis • Decrease in monocytes = monocytopenia* • Monocyte morphology • Activated monocytes may have increased numbers of cytoplasmic vacuoles, or be actively phagocytizing cells or microorganisms • Important to differentiate from band neutrophils and intermediate/large lymphocytes Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey *Decreased blood monocyte concentration is not clinically significant Platelet structure and function • In mammals, platelets are not intact cells but fragments of cytoplasm from a bone marrow precursor cell (megakaryocyte) • Granules within the cytoplasm contain molecules needed for clot formation • Platelet aggregation (formation of a “platelet plug”) is one of the first responses to a blood vessel injury • This platelet aggregate forms the foundation for the rest of the clotting cascade Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey Platelet structure and function • In non-mammalian species, thrombocytes are intact, nucleated cells • Morphology is similar to that of lymphocytes • Function is the same as mammalian platelets Thrombocyte aggregate Lymphocyte Thrombocyte Images from Atlas of Veterinary Hematology, 2nd ed., J Harvey How we assess platelets clinically – CBC, blood smear, and more! • Platelet numbers • Platelet count (automated and manual estimate) • Platelet concentration (platelet “crit” or PCT) • ~ HCT for red blood cells • Platelet size • Mean platelet volume (MPV) • Young, recently produced platelets tend to be larger than mature platelets • Certain platelet disorders can result in increased numbers of large platelets • Platelet morphology (blood smear) • Clumping! • Can interfere with platelet count • Large platelets (aka macroplatelets) • Platelet function • Specialized coagulation assays 40x Increased platelets = thrombocytosis Decreased platelets = thrombocytopenia Red blood cells White blood cells Platelets
