Leukocytes: The Granulocytic and Monocytic Series PDF

 PART THREE Leukocytes...

 PART THREE Leukocytes Leukocytes: The Granulocytic CHAPTER 14 and Monocytic Series OBJECTIVES The granulocytic series Compare the bone marrow maturation of the monocyte with that of Briefly explain the factors related to the development of multipo- the neutrophil. tential progenitor cells into specific leukocyte cell lines. Describe the nuclear and cytoplasmic characteristics of the List each type of immature neutrophil found in the proliferative com- monocyte as it develops. partment of the bone marrow along with the percentage of each Normal values and functional properties of and the approximate time spent in each developmental stage. List each type of neutrophil found in the maturation-storage com- granulocytes and macrophages partment of the bone marrow along with the percentage of each List the normal values for neutrophils, eosinophils, basophils, and and the approximate time spent in this phase. monocytes in normal peripheral blood. Describe the chemical factors and cellular characteristics that permit neu- Explain the activities of the acute inflammatory response. trophils to leave the bone marrow and enter the peripheral circulation. Describe the characteristics of sepsis syndrome. Define the terms marginating and circulating pools and discuss Describe the general characteristics and specific details of phago- the length of time the neutrophils, eosinophils, and basophils spend cytosis. in each pool. Discuss the specialized functions of eosinophils, basophils, and Describe the nuclear and cytoplasmic characteristics of the neutro- monocytes. phils, eosinophils, and basophils throughout the maturation process. Assessment methods Explain the appearance and etiology of the various morphological abnormalities encountered in mature granulocytes. Describe the rationale for each of the methods used in the Describe the abnormalities associated with mature granulocytes in assessment of inflammatory conditions. body fluids. Case study The monocytic-macrophage series Apply laboratory data to the stated case study and discuss the Discuss the differentiation of monocytes and macrophages from implication of this case to the study of hematology. the multipotential stem cell. INTRODUCTION eosinophils, basophils, and the monocytic-macrophage series. The cellular elements of the blood are produced from a common, multipotential hematopoietic cell. This cell, the progenitor cell, undergoes mitotic division. Subsequent mat- THE GRANULOCYTIC SERIES uration of progenitor cells produces the major categories of the cellular elements of the circulating blood: the erythro- Production of Neutrophils, Eosinophils, cytes, leukocytes, and thrombocytes. and Basophils On the basis of function, leukocytes can be divided into the granulocytic, monocytic, and lymphoid series. This chapter Factors that regulate the commitment of a human discusses the granulocytic leukocytes, which can be further hematopoietic progenitor cell to a specific cell line, such as subdivided on the basis of morphology into neutrophils, the granulocytic cell type, and their function are influenced 235 236 PART 3 Leukocytes by the hematopoietic growth factors, the interleukins, and constitutes approximately 3% of the nucleated bone marrow the microenvironment (see Chapter 4). cells. This stage lasts about 24 hours. The myelocyte is the Cells that are committed to differentiation as granulocytes next maturational stage, with approximately 12% of the pro- have been cloned in vitro and have produced a mixture of liferative cells existing in this stage. The stage from myelo- granulocytes and macrophage cells. Further growth of these cyte to metamyelocyte lasts an average of 4.3 days. Once the cells is dependent on colony-stimulating factor (CSF) and metamyelocyte stage has been reached, cells have undergone interleukins. The presence of different CSFs favors interleu- four or five cell divisions and the proliferative phase comes to kins, and the microenvironment of the progenitor cell favors an end. differential development of either the granulocytic (myeloid) Following the proliferative stage, granulocytes enter a mat- series or the macrophage-monocytic series. In addition to uration-storage compartment (Fig. 14.1B). The metamyelocytes the differentiation of granulocytes and monocytes, different and band forms mature into segmented granulocytes in this CSFs stimulate specific differentiation, such as the develop- compartment of the bone marrow. The relative proportions ment of eosinophils. of these cells are approximately 45%, 35%, and 20%, respec- tively. The segmented neutrophils in the maturation-storage Sites of Development and Maturation compartment are frequently referred to as the marrow reserve. This reserve constitutes a 4- to 8-day supply of neutrophils. It The development, distribution, and destruction of neutro- is estimated that neutrophilic granulocytes normally remain phils, eosinophils, and basophils are collectively referred to in the maturation-storage phase for 7 to 10 days. Eosinophils as granulocytic kinetics. The neutrophil, basophil, and eosino- remain for about 2.5 days, and basophils remain in this phase phil each begin as a multipotential cell in the bone marrow. for the shortest period, approximately 12 hours. Throughout the normal processes of differentiation, mul- tiplication, and maturation, these cells remain in the bone Distribution of Neutrophils, Eosinophils, marrow. After developing into either band or segmented and Basophils forms, mature cells enter into the blood circulation. The release of neutrophils from the bone marrow into the Development and Proliferation of Neutrophils, circulatory system is a complex process. Certain charac- Eosinophils, and Basophils teristics (Fig. 14.2) and physiological regulators promote movement of the granulocytes through the sinusoid wall of When the colony-forming-unit-granulocyte-erythrocyte- the bone marrow, which is normally an anatomical barrier. megakaryocyte (CFU-GEMM) progenitor cell differenti- Some of the factors that influence cellular release include ates into the colony-forming-unit-granulocyte-macrophage (CFU-GM) progenitor cell, the cell line becomes committed to developing into a myeloblast; the maturational develop- Immature cell Mature cell ment from the myeloblast through the myelocyte stage and mitotic division take place in what is referred to as the bone marrow’s proliferative compartment (Fig. 14.1A). This is also called the mitotic pool and includes cells capable of DNA Decreased overall synthesis. cell size The myeloblast is the first identifiable cell in the granu- locytic series. Myeloblasts constitute approximately 1% of the total nucleated bone marrow cells. This stage lasts approximately 15 hours. The next stage, the promyelocyte, Decreased nuclear- cytoplasmic ratio Progenitor cells 0.1 - 0.2 % Myeloblasts 1% 15 hours Promyelocytes Metamyelocytes 24 hours 45% 3% Myelocytes 4.3 days 12% Increased flexibility Band form Segmented neutrophils neutrophils and mobility 35% 20% Metamyelocytes FIGURE 14.2 Comparative maturational characteristics. As A B cells mature, they are able to move through the sinusoids of the FIGURE 14.1 Bone marrow compartments. A: Proliferative. bone marrow because of a decreased overall cell size, a decreased B: Maturation-storage. nuclear-cytoplasmic ratio, and increased flexibility and mobility.  CHAPTER 14 Leukocytes: The Granulocytic and Monocytic Series 237 the interleukins. Cellular characteristics include an overall Myeloblast reduction in cell size and a smaller nuclear-cytoplasmic ratio. The greater flexibility and mobility of mature cells enhance the migration of cells through the marrow sinusoids into the peripheral blood pool. Promyelocyte The peripheral blood circulation is subsequently divided into two pools of equal size: the circulating and the marginating pools. The marginating granulocytes adhere to the endothe- lium of the blood vessels. Some granulocytes are additionally Neutrophilic Myelocyte found in the spleen. Mature granulocytes in the peripheral blood are only in transit to their potential sites of action in the tissues. The movement of granulocytes from the circulat- ing pool to the peripheral tissues occurs by a process called Neutrophilic Metamyelocyte diapedesis. Once in the peripheral tissues, granulocytes, par- ticularly the neutrophils, are able to carry out their function of phagocytosis. Neutrophilic Band The average life span of a segmented neutrophilic granu- locyte in the circulating blood is approximately 7 to 10 hours. Once mature cells have migrated into the tissues, their life span is considered to be several days, unless the cells encoun- ter antigens, toxins, or microorganisms. Eosinophils are in Segmented Neutrophil the peripheral blood for a few hours and are believed to (Polymorphonuclear Neutrophil) reside in the tissues for several days. Basophils have an aver- age circulation time of about 8.5 hours. If excessive numbers FIGURE 14.3 Neutrophilic series. (Reprinted with permission from Anderson SA, Poulsen KB. Anderson’s Atlas of Hematology, of eosinophils are present because of a disease state, dam- Philadelphia, PA: Lippincott Williams & Wilkins, 2003.) aged or degenerated eosinophils give rise to Charcot-Leyden crystals found in body secretions, such as the sputum and stool. If cells are not prematurely destroyed while defending Normal Maturational Characteristics of the body, they are sloughed off with various body secretions, Neutrophils, Eosinophils, and Basophils such as the urine, saliva, or gastrointestinal secretions. An alternative route for the removal of granulocytes from the Myeloblast circulation is phagocytosis by the mononuclear phagocyte In the maturational sequence (Fig. 14.3; Table 14.1), the cells of the spleen. earliest morphologically identifiable granulocytic precursor TABLE 14.1 Maturational Characteristics of Neutrophilic Granulocytes Myeloblast Promyelocyte Myelocyte Metamyelocyte Band Segmented Size (mm) 10–18 14–20 12–18 10–18 10–16 10–16 N:C ratio 4:1 3:1 2:1–1:1 1:1 1:1 1:1 Nucleus Shape Oval or Oval or round Oval or Indented Elongated, Distinct lobes round indented curved (2–5) Nucleoli 1–5 1–5 Variable None None None Chromatin Reticular Smooth Slightly Clumped Very clumped Densely packed clumped Cytoplasm Inclusions Auer rods None None None None None Granules None Heavy Fine Fine Fine Fine Nonspecific Specific Specific Specific Specific Amount Scanty Slightly Moderate Moderate Abundant Abundant increased Color Medium blue Moderate blue Blue-pink Pink Pink Pink 238 PART 3 Leukocytes FIGURE 14.4 Type I (left cell) and II (right cell) myeloblasts. FIGURE 14.6 Photomicrograph of a bone marrow aspirate smear (Reprinted with permission from McClatchey KD. Clinical Labo- shows erythroid and granulocytic maturation. MB, myeloblast; ratory Medicine, 2nd ed, Philadelphia, PA: Lippincott Williams & PM, promyelocyte; MY, myelocyte; ME, metamyelocyte; MO, Wilkins, 2002.) monocyte; PN, pronormoblast; BN, basophilic normoblast; PCN, polychromatophilic normoblast; ON, orthochromatic normoblast; PC, plasma cell. (Reprinted with permission from McClatchey KD. is the myeloblast (Fig. 14.4). This cell has an average overall Clinical Laboratory Medicine, 2nd ed, Philadelphia, PA: Lippincott diameter of 10 to 18 mm. The nuclear chromatin is finely Williams & Wilkins, 2002.) reticular, with one to five light-staining nucleoli. The cyto- plasm appears as a small rim of basophilic cytoplasm that The promyelocyte is larger than the blast stage, with an aver- lacks granules. Auer rods (Fig. 14.5), which are aggregates age diameter of 14 to 20 mm. The N:C ratio is lower in the of fused lysosomes, may appear as red, needle-like crystal- promyelocyte than in the myeloblast. The nuclear chromatin line cytoplasmic inclusions. These inclusions may appear is more condensed than in the blast, and nucleoli are present. alone or in groups. Auer rods are pathological, not normal, The cytoplasm is a pale grayish blue. inclusions. Myelocyte Promyelocyte The myelocyte (Fig. 14.7) is the third maturational stage. The promyelocyte (Fig. 14.6) represents the second matura- This cell is characterized by the recognizable appearance of tional stage seen in granulocytes. The outstanding feature of secondary or specific cytoplasmic granulation. The sepa- this cell is the presence of prominent granulation that may rate cell types—neutrophils, eosinophils, and basophils— actually obscure the other morphological features of the cell. become visibly recognizable at this stage. Neutrophilic These granules are primarily azurophilic granules and are rich granules are fine and stain a blue-pink color with Wright in the enzymes myeloperoxidase and chloroacetate esterase. stain. Eosinophilic granules are larger than neutrophilic granules. These round or oval-shaped granules are orange and have a glassy or semiopaque texture. Basophilic gran- ules have a dark blue-black color and a dense appearance. The myelocyte has an average diameter of 12 to 18 mm. The Promyelocyte Myelocyte Myeloblast FIGURE 14.5 Auer rods. (Reprinted with permission from FIGURE 14.7 Myeloblast, myelocyte, promyelocyte. (Reprinted with Anderson S, Poulsen K. Anderson’s Atlas of Hematology, Philadelphia, permission from Anderson S, Poulsen K. Anderson’s Atlas of Hema- PA: Lippincott Williams & Wilkins, 2003.) tology, Philadelphia, PA: Lippincott Williams & Wilkins, 2003.)  CHAPTER 14 Leukocytes: The Granulocytic and Monocytic Series 239 Myeloblast Promyelocyte Eosinophilic Myelocyte FIGURE 14.8 Two neutrophil myelocytes (one large and one small), a neutrophil metamyelocyte, and a juvenile neutrophil (stab form) from a normal marrow smear. (Reprinted with permission from Mills SE. Histology For Pathologists, 3rd ed, Philadelphia, PA: Eosinophilic Lippincott Williams & Wilkins, 2007.) Metamyelocyte N:C ratio continues to decrease. The nucleus has a more oval appearance than in previous stages, nucleoli are no longer visible, and the chromatin is much more clumped Eosinophilic than in previous stages. Band Metamyelocyte The metamyelocyte (Fig. 14.8) is the fourth maturational stage. Its most characteristic feature is that the nucleus begins to assume an indented or kidney bean shape, which Eosinophil will continue to elongate as the cell matures through this phase. The chromatin continues to become more condensed or clumped. The color of the specific granulation continues FIGURE 14.10 Eosinophilic series. (Reprinted with permission to become a major distinguishing feature. from Anderson SA, Poulsen KB. Anderson’s Atlas of Hematology, Philadelphia, PA: Lippincott Williams & Wilkins, 2003.) Mature Forms Two stages of granulocytes are observed in the circulating A mature, segmented neutrophil has a characteristic multilobed blood: the band form and the segmented form (Fig. 14.9). The nucleus. The separate lobes are attached to each other by a fine band form has a typical elongated nucleus. thread-like filament. The filament, between separate lobes may be hidden. The nucleus of the basophil can be difficult to see because it is usually obscured by dark, large granules. The band form of neutrophils, eosinophils, and basophils and, in the final stage of maturation, the segmented neutro- phils, eosinophils (Fig. 14.10), and basophils (Fig. 14.11) are the cell forms normally found in the circulating blood. Mast cells (tissue basophils) are not observed in the blood of healthy persons. These cells have an appearance similar to that of the blood basophil. Mast cells have a round or oval nucleus. The granules of the mast cell do not overlie the nucleus as they do in basophils. Granulation in Mature Forms FIGURE 14.9 Peripheral blood smear from a normal individual. Segmented neutrophil. (Reprinted with permission from Although all granules are commonly produced by the McClatchey KD. Clinical Laboratory Medicine, 2nd ed, Philadelphia, rough endoplasmic reticulum and transported to the Golgi PA: Lippincott Williams & Wilkins, 2002.) apparatus for packaging, the granules of each cell type stain 240 PART 3 Leukocytes Basophilic granules contain heparin and histamine. Mast cells have granules that have an enzyme content similar to Myeloblast those of the blood basophil. THE MONOCYTIC-MACROPHAGE SERIES Cells of the mononuclear phagocyte system include the Promyelocyte monocytes and macrophages. Macrophages (Fig. 14.12) have a variety of names, including histiocytes in the loose connective tissues, Kupffer cells in the sinusoids of the liver, osteoclasts in bone, and microglial cells in the nervous sys- tem. The name of the cell changes with the location of the Basophilic cell; however, mature macrophages are distributed through- Myelocyte out the body. These cells, along with the reticular cells of the spleen, thymus, and lymphoid tissues, are collectively referred to as the mononuclear phagocyte (Fig. 14.13). The predominant phagocytic cell, the segmented neutrophil, is confined to the circulating blood unless it is recruited into Basophilic the tissues. Metamyelocyte Production and Development of Monocytes and Macrophages Basophilic Cells of the macrophage system are formed from the pro- Band genitor cells in the bone marrow. These cells are derived from the CFU-GM, which can differentiate into either the megakaryocyte-colony-forming- unit (CFU-M) and develop into a monocyte or macrophage or the granulocyte-colony- forming-unit (CFU-G) and develop into a segmented neu- Basophil trophil. A monocyte is influenced by hematopoietic growth factors to transform into a macrophage in the tissues. Func- tionally, monocytes and macrophages have phagocytosis FIGURE 14.11 Basophilic series. (Reprinted with permission from as their major role, although they also have regulatory and Anderson SA, Poulsen KB. Anderson’s Atlas of Hematology, Phila- secretory functions. delphia, PA: Lippincott Williams & Wilkins, 2003.) In contrast to the granulocytic leukocytes, the promono- cyte will undergo two or three mitotic divisions in approx- imately 2 to 2.5 days. Monocytes are released into the differently because their contents vary. The characteristics of circulating blood within 12 to 24 hours after their precursors these granules are as follows. The granules of segmented neutrophils are rich in various antibacterial substances, including lysosomal hydrolases, lysozyme, and myeloperoxidase. Some of these granules are typical lysosomes. Eosinophilic granules differ from neutrophilic granules in that they lack lysozyme. These granules are of two types: 1. Smaller round granules, which have been identified as not containing crystalloids. These granules exist in small quantities in the mature eosinophil and are rich in acid phosphatase. 2. Larger crystalline granules, which are more numerous. These crystalline granules are elliptical, are larger than the granules of the neutrophil, and have an amorphous matrix surrounding an internal crystalline structure. The crystals are thought to represent the enzyme peroxidase (not the same as the myeloperoxidase found in neutro- FIGURE 14.12 Macrophage. (Reprinted with permission from phils), and the matrix contains acid phosphatase. Anderson SA, Poulsen KB. Anderson’s Atlas of Hematology, 2003.)  CHAPTER 14 Leukocytes: The Granulocytic and Monocytic Series 241 Nervous tissue Lungs Lymph nodes Bones Spleen FIGURE 14.15 Monoblast. (Reprinted with permission from Kidney Anderson S, Poulsen K. Anderson’s Atlas of Hematology, Philadelphia, PA: Lippincott Williams & Wilkins, 2003.) Connective tissue or histiocytes Liver circulation, cells may be located in a circulating or marginat- ing pool. The ratio of circulating to marginating cells is 1:3.5. FIGURE 14.13 Mononuclear phagocyte system. Phagocytic cells Monocytes are estimated to have a circulatory half-life of are located in many body organs. Promonocyte precursors in the approximately 8.5 hours. The life span of tissue monocytes is bone marrow develop into circulating monocytes in the periph- variable. However, cells in noninflammatory areas have been eral blood. Monocytes ultimately become distributed throughout the body as macrophages. Neutrophils are located in the circulat- demonstrated to live for months to years. ing blood, except when they enter the tissues during acute inflam- mation. (Adapted with permission from Cohen BJ, Wood DL. Morphological Characteristics Memmler’s The Human Body in Health and Disease, 11th ed, Philadelphia, PA: Lippincott Williams & Wilkins, 2009.) Morphological identification of the monocyte (Fig. 14.14) is more difficult than that of the neutrophilic, eosinophilic, or basophilic granulocytes. Monoblasts (Fig. 14.15), promono- have completed their last mitotic division. Monocytes have cytes (young monocytes), and monocytes vary greatly in their no large reserve of cells in a maturation-storage pool as morphological appearance (Table 14.2). The monocytic series do the granulocytes. Once the monocytes have entered the does have a characteristic nuclear chromatin pattern. The chro- matin is clumped, although the clumps are smaller and more elongated than in neutrophils. This pattern can be described as lace-like. The shape of the nucleus of the monocyte may be round or oval, but it is frequently convoluted or twisted. Monoblast Mature monocytes (Fig. 14.16) are the largest mature cells seen in peripheral blood. They may exhibit an irregular cyto- plasmic outline. These cytoplasmic irregularities can include pseudopods. Vacuoles are also commonly observed. Classi- cally, the cytoplasm is blue-gray in color, with fine granula- tion resembling ground glass. Promonocyte REFERENCE RANGES OF GRANULOCYTES AND MONOCYTES Healthy adults and children have a relatively consistent num- ber of each of the granulocytic and monocytic types of leu- kocytes. Normal variations occur along with daily rhythmic fluctuations; racial differences also occur. Monocyte The major function of the granulocytic leukocytes is phagocytosis, a body defense mechanism. Neutrophilic granulocytes are the major phagocytic cell of the circulating FIGURE 14.14 Monocyte maturation series. (Reprinted with blood. Eosinophils and basophils are less effective as phago- permission from Anderson SA, Poulsen KB. Anderson’s Atlas of cytes but have additional specialized functions associated Hematology, 2003.) with body defense. 242 PART 3 Leukocytes TABLE 14.2 Characteristics of Monocytes Monoblast Promonocyte Mature Monocyte Size (mm) 12–20 12–20 12–18 N:C ratio 4:1 3:1–2:1 2:1–1:1 Nucleus Shape Oval, folded Elongated, folded Horseshoe shaped, folded Nucleoli 1–2 or more 0–2 None Chromatin Fine Lace like Lace like Cytoplasm Inclusions Vacuoles variable Vacuoles variable Vacuoles common Granules None None or fine Fine, dispersed Amount Moderate Abundant Abundant Color Blue Blue-gray Blue-gray Shape Monocytes frequently demonstrate irregular cytoplasmic shape with pseudopods Each type of granulocyte and the monocytes have an variation in a person’s eosinophil count is related to the established normal range (Table 14.3) and an average per- fluctuation of the hormone adrenal glucocorticosteroid. centage on a stained blood smear (refer to Chapter 3). Sig- In women, the menstrual cycle affects the eosinophil nificant differences in the total leukocyte count do occur count, with the number of circulating eosinophils dropping between black and white persons. Blacks have a lower total at the time of ovulation and rising during menstruation. The leukocyte count compared with whites, owing to a decreased summer season may produce higher counts in a person with- number of neutrophils. Individual variations in the total out allergies. Exercise produces a brief rise in eosinophils, leukocyte count occur, with a daily fluctuation of as much whereas stress can lower eosinophil counts. as 20%. Peak levels occur in the middle of the night and the early morning. Variations may also occur with specific cells. For example, FUNCTIONAL PROPERTIES OF GRANULOCYTES smoking causes a mild elevation of neutrophils. Monocytes AND MONOCYTES have periodic oscillations of 0.2 × 109/L every 3 to 6 days. Eosinophils have a well-documented daily fluctuation, One of the major functional properties of granulocytes and with a diurnal (time-related) variation in the number of cir- monocytes is phagocytosis. Defense against infectious disease culating cells. The quantity of circulating eosinophils tends is the responsibility of both the phagocytic and the immune to be highest late at night during sleep, decreases during the morning, and begins to rise at midafternoon. This rhythmic Reference Leukocyte Values for a TABLE 14.3 Normal Adult Population WBC Cell Type Reference Range (%) Neutrophils-Bands 0–3 Neutrophils-Segmented 40–74 Eosinophils 1–4 Basophils 0.5–1 Lymphocytes 34 FIGURE 14.16 Blood cell, monocyte. (Reprinted with permission Monocytes 2–6 from Cohen BJ, Wood DL. Memmler’s The Human Body in Health Source : Adapted from Handin, RI, Lux SE, Stossel TP. Blood, 2nd ed, and Disease, 11th ed, Philadelphia, PA: Lippincott Williams & philadelphia, PA: Lippincott williams & Wilkins, 2003, Appendix 6, p. 2194. Wilkins, 2009.)  CHAPTER 14 Leukocytes: The Granulocytic and Monocytic Series 243 systems in humans. The activities of these two systems are somewhat coordinated and interdependent. Phagocytosis is the process that enables particular cells to engulf and disable particles, such as bacteria. Defects in phagocytosis (e.g., chronic granulomatous disease, in which there is defective killing of phagocytosed microorganisms) can be fatal. Body defense systems include phagocytic cells (neutro- phils, monocyte, and macrophages), cells that release inflam- matory mediators (basophils, mast cells, and eosinophils), and natural killer cells. The molecular components of body defenses include complement, acute-phase proteins, and cytokines (e.g., interferons). Upon initiation of inflammation, local inflammatory sig- nals, for example, chemokines, cytokines, and adhesion mol- ecules, initiate an orchestrated process of actively recruiting neutrophils into the tissue. Neutrophils act quickly in body defense by recognizing pathogen-associated molecular pat- FIGURE 14.17 Bone marrow aspirate from a patient with AIDS terns (PAMPs) through encoded toll-like receptors (TLRs). and progressive disseminated histoplasmosis. Highly characteristic Human neutrophils express the majority of TLRs. Fur- image shows macrophage filled with 1- to 2-mm yeast cells (Wright thermore, they act in concert with other components of stain, original magnification ×400). (Reprinted with permission the immune system, by expressing receptors for antibody from Crapo JD, et al. Baum’s Textbook of Pulmonary Diseases, (fraction crystallizable receptors [FcRs]) and complement 7th ed, Philadelphia, PA: Lippincott Williams & Wilkins, 2004.) (complement receptors [CRs]). Recently, interleukin-17A (IL-17A) and IL-17F, two cytokines produced by a subset in phagocytosis, they possess less phagocytic activity. Their of T helper cells have been found to indirectly induce the ineffectiveness is owing to both the small number of these recruitment of neutrophils in inflammation. cells in the circulating blood and the lack of powerful Human peripheral blood monocytes are a heterogeneous lytic enzymes. Eosinophils and basophils are functionally population of leukocytes, distinguishable by the expression important in body defense in other ways. of CD14 and CD16 membrane markers. Monocyte subset phenotypes include inflammatory and resident monocytes. The Role of Macrophages These subsets differentially express cell adhesion molecules and chemokine receptors that implies alternate recruitment Macrophages (Fig. 14.17) participate in the phagocytic mechanisms to a site of inflammation. Monocytes are not process and are particularly important in the processing just phenotypically different but subsets use alternate sets of of antigens as part of the immune response. Macrophages cell adhesion molecules and chemokine receptors that con- exist as either fixed or wandering cells. Fixed macrophages tribute to different functions and are recruited at different line the endothelium of capillaries and the sinuses, the time points after initiation of inflammation. Classical mono- bone marrow and organs, spleen, and lymph nodes. Spe- cytes with CD14+CD16− membrane markers produce lower cialized macrophages, such as the pulmonary alveolar mac- amounts of proinflammatory cytokines but contribute more rophages, are the dust phagocytes of the lung and function effectively to bacterial clearance by phagocytosis compared as the first line of defense against inhaled foreign particles with nonclassical monocytes. Nonclassical CD14loCD16+ and bacteria. Macrophages and their known precursor, monocytes are more potent in presenting antigens in the monocytes, migrate freely into the tissues from the blood immune response. to replenish and reinforce the macrophage population. When there is tissue damage and inflammation, cellular General Characteristics activity increases with the release of substances that attract macrophages. Although the major cells associated with phagocytosis are the neutrophilic leukocytes (neutrophils) and the monocytes- Acute Inflammatory Response macrophages, the neutrophils are the principal leukocytes involved in a localized inflammatory response. The inflam- Acute inflammation is of short duration and is characterized matory exudate (pus), which develops rapidly in an inflam- by vascular and cellular changes. In an acute inflammatory matory response, is primarily composed of neutrophils and response, cells and molecules of the immune system move monocytes. into the affected site (Figs. 14.18 and 14.19). The process Neutrophils are steadily lost to the respiratory system, the of movement of fluids, proteins, and leukocytes (primarily gastrointestinal system, and the urinary system, where they neutrophils) into the interstitial tissue is called exudation. participate in generalized phagocytic activities. Although Activation of complement generates C3b, which coats the the eosinophils and basophils are capable of participating surface of the pathogen. The neutrophil chemoattractant and 244 PART 3 Leukocytes FIGURE 14.18 Acute inflammatory disease. (Reprinted with permission from Delves PJ, Roitt IM. The Immune Sys- tem, Part I, N Engl J Med, 343(1):37–49, 2000. Copyright © 2000 Massachusetts Medical Society. All rights reserved.) activator C5a is also produced and, together with C3a and Substances released from a pathogen and from damaged C4a, triggers the release of histamine by degranulating mast tissues upregulate the expression of adhesion molecules cells. This in turn causes the contraction of smooth muscles on the vascular endothelium, alerting passing cells to the and a rapid increase in vascular permeability. presence of infection. The cell surface molecule L selec- tin on neutrophils recognizes carbohydrate structures, for example, sialyl-Lewis, on the vascular adhesion molecules. Vasodilation The neutrophil rolling along the vessel wall is arrested in Edema its course by these interactions. As the neutrophil becomes Neutrophils activated, it rapidly sheds L selectin from its surface and replaces it with other cell surface adhesion molecules (e.g., Intensity Monocytes and integrins). These integrins bind the molecule E selectin, macrophages which appears on the blood vessel wall under the influence of inflammatory mediators (e g., bacterial lipopolysaccha- ride and the cytokines, interleukin-1 and tumor necrosis factor-a). Complement components and other inflamma- tory mediators all contribute to the recruitment of inflam- 1 2 3 4 5 matory cells as does an important group of chemoattractant Acute, short- Days cytokines called chemokines. term injury Activated neutrophils pass through the vessel walls, mov- FIGURE 14.19 Timing of acute inflammatory events. The graph ing up the chemotactic gradient to accumulate at the site of shows both intensity and duration for vasodilation, edema, neu- infection, where they are positioned to phagocytize any C3b- trophils, and monocytes and macrophages. (Reprinted with per- coated microbes. Mutations in the genes for a number of dif- mission from McConnell TH. The Nature Of Disease Pathology for ferent adhesion molecules have been described in patients the Health Professions, Philadelphia, PA: Lippincott Williams & with leukocyte-adhesion deficiencies, some of which are Wilkins, 2007.) associated with life-threatening infections.  CHAPTER 14 Leukocytes: The Granulocytic and Monocytic Series 245 FIGURE 14.20 Early biomechanical events in sepsis. (Reprinted with permission from Wheeler AP, Bernard GR. Treating patients with severe sepsis, N Engl J Med, 340(3):208, 1999. Copyright © 2005 Massachusetts Medical Society. All rights reserved.) Sepsis are tumor necrosis factor, interleukin-1 and interleukin-8, which are generally proinflammatory, and interleukin-6 Neutrophils have been regarded as a help and a hindrance. and interleukin-10, which tend to be anti-inflammatory. Although neutrophils were believed to be essential for the A trigger (e.g., microbial toxin) stimulates the production killing of pathogens, excessive release of oxidants and pro- of tumor necrosis factor and interleukin-1, which in turn teases by neutrophils is also believed to be responsible for promote endothelial cell–leukocyte adhesion, release of injury to organs. proteases and arachidonate metabolites, and activation of Sepsis is an infection-induced syndrome defined as the clotting. Interleukin-8, a neutrophil chemotaxis, may have presence of two or more of the following features of systemic an especially important role in promoting tissue inflamma- inflammation: tion (Fig. 14.21). Fever or hypothermia Leukocytosis or leukopenia Steps in Phagocytosis Tachycardia and tachypnea Phagocytosis can be divided into three stages (Fig. 14.22). Supranormal minute ventilation These stages are movement of cells, engulfment, and digestion. Early biochemical events in sepsis (Fig. 14.20) include the key If microorganisms are not effectively immobilized, stage 4, element, cytokines. The most widely investigated cytokines subsequent phagocytic activity may take place. Neutrophils Monocytes Phagocytosis (e.g., microbes) Phagocytosis Chemokine and (e.g., microbes, cytokine synthesis apoptotic cells) Endothelial permeability Chemokine and Mechanisms of cytokine synthesis PMN-mediated monocyte recruitment granule protein- chemokine synthesis ‘find me’ SIGNALS Onset of mediated adhesion IL-6 transsignaling of apoptotic neutrophils inflammation and emigration 1h 12h 24h....days.... FIGURE 14.21 Summary of leukocyte recruitment pattern. 246 PART 3 Leukocytes without impeding regional alveolar blood flow, thus accommodating the marginated pool. The marginating pool of neutrophils, adhering to the endothelial lining of capillar- ies, migrates through the vessel wall to the intestinal tissues. This amoeboid movement is called diapedesis. Stage 2: Engulfment After the phagocytic cells have arrived at the site of injury, the invading microorganisms or particles can be engulfed through active membrane invagination. Leukocyte-specific CD18 integrins mediate neutrophil adhesion during migra- tion and function as phagocytic receptors for bacterial uptake and killing. It is important to realize that phagocytosis is an active process that requires a large expenditure of energy by the cells. The required energy is primarily provided by anaerobic glycolysis. The principal factor in determining whether phagocyto- sis can occur is the physical nature of the surfaces of both FIGURE 14.22 Phagocytosis. Step 1 depicts the movement of a seg- the foreign particle and the phagocytic cell. Bacteria must mented neutrophil toward the site of bacterial invasion, chemotaxis. Step 2 depicts the initiating event in engulfment, the adherence of be more hydrophobic than the phagocyte. Some bacteria, the phagocytic membrane to the bacterial cell wall. This process can such as Streptococcus pneumoniae, possess a hydrophilic cap- be enhanced by opsonization. If the surface tensions of the mem- sule and are not normally phagocytized. Most nonpatho- branes are conducive to engulfment, the phagocytic cell membrane genic bacteria are easily phagocytized because they are very invaginates and engulfs the bacterium (step 3), and a phagosome is hydrophobic. formed (step 4). The phagosome fuses with one or more lysosomal Certain soluble factors, including complement (a plasma granules (step 5). Digestion of the bacterium occurs (step 6) and protein), coupled with antibodies, and substances such as normally results in autolysis of the phagocyte. acetylcholine, enhance the phagocytic process. Enhancement of phagocytosis through the process of opsonization, the coat- Stage 1: Movement of Cells ing of a particle with immunoglobulin and/or complement, Various phagocytic cells continually circulate throughout the speeds up the ingestion of particles. If the surface tensions blood, lymph, gastrointestinal system, and respiratory tract. are conducive to engulfment, the phagocytic cell membrane The physical occurrence of damage to tissues and inflamma- invaginates, a process that leads to the formation of an iso- tion due to trauma or microbial multiplication releases sub- lated vacuole, a phagosome, within the cell. stances that attract phagocytic cells. Cells are guided to the site of injury by the concentration Stage 3: Digestion gradient of chemotactic substances. This event is termed Digestion follows ingestion of particles, the required energy chemotaxis. Chemotactic factors polarize and orient attached being provided primarily by anaerobic glycolysis. The neutrophils for locomotion. Cells acquire a characteristic vacuole formed during the engulfment process fuses with asymmetric shape with the formation of pseudopodia. The one or more lysosomal granules that contain various lytic movement of neutrophils is a process called extravasation. enzymes. The granules of neutrophils contain various anti- Extravasation of polymorphonuclear leukocytes (PMNs) to bacterial substances such as lysosomal hydrolases, lysozyme, the site of inflammation precedes a second wave of emigrat- and myeloperoxidase. The action of the oxygen-dependent ing monocytes. Monocyte subsets are affected differently by myeloperoxidase-mediated system, hydrogen peroxide, and signals generated from PMNs. PMN granule proteins induce an oxidizable cofactor serve as major factors in the actual adhesion as well as emigration of inflammatory monocytes to killing of bacteria within the vacuole. Other oxygen-inde- the site of inflammation involving b2-integrins and formyl- pendent systems, such as alterations in pH, lysozymes, lacto- peptide receptors. PMNs create an environment, including ferrin, and the granular cationic proteins, also participate in rapidly undergoing apoptosis, that is favorable for extrava- the bactericidal process. sation and accumulation of inflammatory monocytes at the An energy-dependent respiratory burst accompanies site of inflammation. phagocytosis. The respiratory burst generates oxidizing com- Actively motile segmented neutrophils are able to gather pounds through the hexose-monophosphate shunt. Oxidiz- at the site of inflammation quickly, but monocytes are slower ing compounds produced from partial oxygen reduction are to arrive. Segmented neutrophils can be found in the begin- important in bacteriocidal activity. Utilization of reduced ning exudate in less than 1 hour. nicotinamide adenine dinucleotide phosphate (NADPH) or Large numbers of intracapillary neutrophils are retained reduced nicotinamide adenine dinucleotide (NADH) as an elec- in the narrow capillaries of the pulmonary circulation tron donor subsequent to the activation of a membrane-bound  CHAPTER 14 Leukocytes: The Granulocytic and Monocytic Series 247 - oxidase produces superoxide (O 2) from oxygen. Hydrogen peroxide (H2O2) is either generated from superoxide sponta- neously or catalyzed by superoxide dismutase. - 2 O2 + NADPH ® 2 O 2 + NADP+ + H+ - 2 O 2 + 2 H+ ® H2O2 + O2 The killing effect of H2O2 is potentiated by the forma- tion of peroxide-halide. This reaction requires the enzyme myeloperoxidase, found in the primary granules of neutro- philic granulocytes. H2O2 + C1- myeloperoxidase ® HOC1 + H2O Monocytes are particularly effective as phagocytic cells because of the large amounts of lipase in their cytoplasm. Although monocytes accumulate at a site of acute inflam- FIGURE 14.23 A spleen imprint shows many Leishman-Donovan mation more slowly, they persist longer. Their metabolic (L-D) bodies, which are composed of a round nucleus and a rod-shaped kinetoplast. Giemsa, ×3300. burst is less extreme than neutrophils, but their capacity to kill many microbes is more diverse compared with that of neutrophils. Lipase digests the lipid-rich cell wall of bacteria multiply in the phagocytic cells. Repeated multiplication such as Mycobacterium tuberculosis. Monocytes are further produces extensive numbers of parasitized macrophages and able to bind and destroy cells coated with complement-fixing monocytes (Fig. 14.23). Infection with the pathogenic fungus antibodies because of the presence of membrane receptors Histoplasma capsulatum, which also infiltrates the mononu- for specific components or types of immunoglobulin. clear phagocytic system and appears within macrophages of As a result of the release of lytic enzymes, the cytoplasmic the bone marrow, can mimic kala-azar. membrane of the phagocytic cell is usually ruptured, and the engulfing cell itself is phagocytized by macrophages. Mac- rophage digestion proceeds without risk to the cell unless the Specialized Functions of Granulocytes ingested material is toxic. However, if the ingested material Eosinophils damages the lysosomal membrane, the macrophage will also be destroyed because of the release of lysosomal enzymes. The eosinophil is considered to be a homeostatic regula- tor of inflammation that leaves the circulating blood when adrenal cortical hormone increases. Functionally, this means Stage 4: Subsequent Phagocytic Activity that the eosinophil attempts to suppress inflammatory tissue If invading microorganisms are not phagocytized at entry reactions to prevent the excessive spread of inflammation. into the body, they may establish themselves in secondary Eosinophils proliferate in response to antigenic stimula- sites, such as the lymph nodes or various body organs. Undi- tion and contain substances that inactivate factors released by gested bacteria produce a secondary inflammation, where mast cells and basophils. The primary function of eosinophils neutrophils and macrophages again congregate. If bacteria appears to be their reactions with products from mast cells, escape from secondary tissue sites, bacteremia will develop. lymphocytes, and other soluble substances in the blood, such In patients whose conditions are unresponsive to antibiotic as the coagulation factors, complement, and hormones. intervention, this situation can be fatal. Although eosinophils are ineffective in protecting the body In cases of acquired immunodeficiency syndrome (AIDS), against invading foreign particles, they do play a role in the researchers at the National Cancer Institute have found body defense mechanism. Eosinophils regulated allergic dis- evidence of the virus in mononuclear phagocytes. Infected eases and granulomatous and fibrotic disorders, and eosino- phagocytes were found in brain and lung tissue specimens phil recruitment in response to helminthic parasite infection from AIDS patients, indicating that the brain infection might has been well documented. Eosinophils have the ability to have been caused by the phagocytes. interact with the larval stages of some helminthic parasites Phagocytes harboring AIDS virus were found to be more and to damage them by means of oxidative mechanisms. powerfully infective than T lymphocytes (discussed in Certain proteins released from eosinophilic granules are Chapter 16). The infected phagocytes may be responsible known to damage antibody-coated Schistosoma parasites. for passing the virus back to the rest of the immune system, with other infected cells passing the virus back to healthy Basophils phagocytes. Basophils and mast cells (in tissues) share significant phe- Protozoan organisms, such as Leishmania donovani (pop- notypic and functional properties. Both cell lines possess ularly referred to as kala-azar), are cleared by the mononu- metachromatic granules containing histamine and proteogly- clear phagocyte cells of the liver, spleen, and bone marrow. cans and express the high-affinity immunoglobulin E (IgE) The Leishman-Donovan bodies, which are oblong or round, receptor through which they can be activated to degranulate 248 PART 3 Leukocytes and synthesize inflammatory mediators. Histamine plays an neutrophils is considered to be 3% in adults; newborn infants important role in acute, systemic allergic reactions. Degran- have a somewhat higher normal average. A neutrophilic band ulation (loss of granules) occurs when an antigen, such as count greater than 11% is considered to be consistent with an pollen, binds to two adjacent immunoglobulin E (IgE)-type inflammatory condition. When the percentage of band forms antibody molecules located on the surface of basophils. The and other immature neutrophils such as metamyelocytes and release of the contents of these basophilic granules results in myelocytes increases, the condition is sometimes referred to as increased vascular permeability, smooth muscle spasm, and a shift to the left. The normal average for segmented neutro- vasodilation. If this reaction is severe, it can result in anaphy- phils is 56% and approximately 4% for monocytes. lactic shock. A class of compounds, the leukotrienes, mediates the Absolute Cell Counts inflammatory functions of leukocytes. The observed sys- temic reactions that are related to these compounds were The absolute number of segmented neutrophils and bands is previously attributed to the slow-reacting substance of ana- considered to be a less specific index of inflammation than phylaxis (SRS-A). other tests because the total leukocyte count drops in many patients with overwhelming infection. This condition results Monocytes from the movement of circulating granulocytes into the tissue In addition to phagocytosis, monocytes are able to synthesize sites of infection. However, the absolute count may be valuable various biologically important compounds, including trans- in other cases of inflammation. An example of the method of ferrin, complement, interferon, and certain growth factors. calculating an absolute cell count is presented in Box 14.1. In cellular immunity, monocytes assume a killer role. In this role, they are activated by sensitized lymphocytes to phago- cytize offending cells or antigen particles. This is important Erythrocyte Sedimentation Rate in fields such as tumor immunology. The ESR, or sed rate, is a nonspecific indicator of disease with increased sedimentation of erythrocytes in acute and chronic inflammation and malignancies. Although this procedure ASSESSMENT METHODS is nonspecific, it is one of the most commonly performed laboratory tests. Inflammation almost always follows acute tissue damage. Very few tests have as long a history as the ESR. A Swedish Diagnostic categories of acute inflammation can include physician, Fahraeus, is credited with the discovery of this test bacterial causes and nonbacterial causes such as trauma, in 1915. However, the sedimentation of blood was one of the chronic inflammation, and viral disease. Among the many principles on which ancient Greek medicine was based. The laboratory tests that have been advocated for the diagnosis Greek philosophy of the four humors (fluids) in the human of inflammation, the total leukocyte count, the percentage of band and segmented neutrophils determined by a differential leukocyte count, the absolute neutrophil cell count, and the erythrocyte sedimentation rate (ESR) are the most common. BOX 14.1 Other tests include direct cell counts of eosinophils or baso- phils, the leukocyte alkaline phosphatase (LAP) cytochemi- cal stain, and neutrophilic function tests. Absolute Cell Counts Total Leukocyte Count Absolute counta = absolute cell value = total leukocyte count × percentage of cell type The total leukocyte count can be elevated above 10 × 109/L in PATIENT DATA conditions such as pregnancy or strenuous exercise. The total Total leukocyte count: 15.0 × 109/L count may be depressed because of overwhelming bacterial Differential blood smear results: bands 12%, segmented infection (sepsis) or immunosuppressive agents. A diagnosis of neutrophils 80%, lymphocytes 8% acute inflammation is generally based on a total leukocyte count greater than 10.5 × 109/L in combination with other factors. SAMPLE CALCULATION Absolute segmented neutrophil value = total leukocyte count × % of segmented neutrophils Differential Blood Smear Evaluation Absolute value = 15.0 × 109/L × 0.80 = 12.0 × 109/L Some authorities advocate doing away with the identification of segmented neutrophils band forms on the leukocyte differential procedure (see Chap- a This formula can be used to determine the absolute value of any cell ter 26) because of individual variability in cell identification and appearing on a leukocyte differential blood smear. limited usefulness. Patients with stress conditions can demon- Normal absolute values include segmented neutrophils 1.4–6.5 × 109/L, strate an increase in the number of band forms in the presence bands 0–0.7 × 109/L, lymphocytes 1.2–3.4 × 109/L. of a normal total leukocyte count. The normal average for band  CHAPTER 14 Leukocytes: The Granulocytic and Monocytic Series 249 body was established in the 5th century B.C. and further Functional abnormalities expressed by patients with con- developed by Aristotle. This belief proposed that these flu- genital neutropenia include defective migration, bacterial ids formed the body. On the basis of this philosophy, each killing, or increased apoptosis. person had a predisposition for a particular disease depend- ing on the predominance of one of the four fluids: blood, Neutrophilic Hypersegmentation Index phlegm, yellow bile, or black bile. In 1836, Nasse recognized that a property of plasma, later Mature segmented neutrophils have from two to five nuclear identified as increased proteins, produced an increased sink- lobes (segments). Counting the number of lobes can be per- ing speed of erythrocytes in whole blood. The work of Nasse formed to determine the neutrophilic hypersegmentation went unnoticed for nearly a century because medicine was index (NHI). A right shift or increase in the number of lobes undergoing a radical reform, moving away from the humoral to five or more occurs in various conditions, for example, philosophy of the Greeks toward the cellular pathology theo- sepsis, chronic nephritis. The NHI is clinically useful in ries of Virchow (see Chapter 18). With the reestablishment vitamin B12 deficiency (pernicious anemia) and folic acid by Fahraeus of the significance of the empirical basis of diagnosis. Greek medicine, Alf Westergren began working concurrently Three methods exist for calculating the NHI: on refining the technique. 1. Lobe average. This is determined by counting the number Except for some refinements, the ESR procedure contin- of lobes in a number of neutrophils, for example, 200, and ues to be an established parameter of inflammation in the dividing by the total number of neutrophils for the aver- modern clinical laboratory. The Westergren method (see age number of lobes. The reference value is 2.5 to 3.3. Chapter 24) has been selected by the CLSI as the standard 2. Percentage of neutrophils with five or more lobes. Count method of choice. the number of lobes in randomly selected segmented neutrophils, for example, 200. Add up the total number Assessment of Eosinophils and Basophils of lobes for each segmented neutrophil counted and di- vide by the total number of cells counted. The reference Examination of a peripheral blood smear normally demon- range is greater than 3%. strates an average of approximately 4% eosinophils. Because 3. Hypersegmentation index. To calculate this index, use a this method of estimation is only semiquantitative, an abso- minimum of 200 segmented cells. lute eosinophil count, either by manual chamber counting (see Chapter 24) or by the use of automated equipment, Number of neutrophils with 5 or more lobes × 100 is preferred. This procedure is required only if an extreme Number of neutrophils with 4 lobes increase in eosinophils is demonstrated on a peripheral blood smear or if clinical symptoms suggest an increase. Values greater than 16.9 are considered to indicate The basophil is the least numerous of the granulocytes. hypersegmentation. This method is considered to be the Normally, differential smears of normal blood have only one most sensitive method. basophil, if any. An increase in basophils is very significant and is seen in conditions such as chronic myelogenous leu- kemia and polycythemia vera. CHAPTER HIGHLIGHTS Leukocyte Alkaline Phosphatase Test The following types of leukocytes are found in peripheral blood, in order of frequency: neutrophils, lymphocytes, This procedure is discussed in detail in Chapters 21 and monocytes, eosinophils, and basophils. The function of the 26. The value of this cytochemical stain is in differentiating entire leukocytic system is to defend the body against disease, malignant disorders from leukemoid reactions. with each type of leukocyte having a unique function. Neutrophilic Function The Granulocytic Series A number of diseases are associated with leukocyte dysfunc- The cellular elements of the blood are derived from a single, tions related to locomotion, chemotaxis, adhesion, or the multipotential stem cell. This stem cell undergoes differentia- ability of cells to destroy infectious organisms. In vitro assays tion, multiplication, and maturation within the bone marrow. of the rate of cell movement and the directional orientation of After a progenitor cell becomes committed to a specific cell line, the movement as well as the ability of granulocytes to destroy mitosis and early development take place in the proliferative organisms have been in existence for more than 20 years. compartment of the bone marrow. A neutrophilic granulocyte A defect in cell adhesiveness, for example, leukocyte adhesion matures in the following sequence: stem cell, myeloblast, pro- defect (LAD), can lead to decreased cell locomotion. myelocyte, myelocyte, and metamyelocyte. Once the metamy- A test that assesses the killing ability of granulocytes is elocyte stage has been reached, the proliferative phase comes the nitroblue tetrazolium (NBT) test. In the routine clinical to an end. In the next phase of bone marrow development, laboratory, this procedure is infrequently performed. the maturation-storage compartment, metamyelocytes, and 250 PART 3 Leukocytes most band-type neutrophils mature into segmented granulo- system. The neutrophils defend the body against infectious cytes or polymorphonuclear neutrophils. agents and local noninfectious challenges. Macrophages Release of granulocytes from the bone marrow is influ- participate in the phagocytic process and are important enced by a variety of factors. These factors are chemical and in the processing of antigens as part of the immune sys- physical. Chemical factors include neutrophil-releasing fac- tem. Macrophages may be of either fixed or wandering tor and leukocytosis-inducing factor. Physical factors include type. Fixed types are located in the mononuclear phago- greater flexibility and mobility of cells, allowing the cells to cyte system. pass through the sinusoid barrier of the bone marrow. Infection with a pathogen triggers an acute inflamma- The earliest identifiable neutrophil-eosinophil-basophil tory response involving cells and molecules of the immune precursor in the bone marrow is the myeloblast. This cell has a system. Sepsis is an infection-induced syndrome with classic high N:C ratio and frequently contains Auer rods in the cyto- features. Tumor necrosis factor and certain interleukins act plasm. The second stage of development is the promyelocyte, as proinflammatory or anti-inflammatory factors in the acti- a heavily granulated cell, which may retain visible nucleoli vation of clotting, or tissue inflammation. in the nucleus. Following the promyelocyte, the myelocyte is Physical trauma initiates the events of phagocytosis. The the third stage. In this stage, granules that distinguish neu- attraction of phagocytic cells to the site of injury is termed trophils, eosinophils, and basophils begin to become appar- chemotaxis. The neutrophils are the most abundant of the ent. Myelocytes mature into metamyelocytes. The nucleus of cells participating in phagocytosis and arrive at the site rap- the myelocyte progresses from an indented structure to an idly. Monocytes are slower in arriving. Engulfment of foreign elongated structure as it matures. The last two maturational particles, such as bacteria, is the next step. Digestion of bac- stages are the band and segmented forms. Band forms have teria follows engulfment. Antibacterial enzymes contained a condensed chromatin pattern in the nucleus and a thin, in granules, and alterations within the cell such as a change elongated, curved nuclear shape. The segmented neutrophil in pH, destroy the engulfed bacteria. Lytic enzymes disrupt is typified by the multiple segments of the nucleus that are the cellular membrane of the phagocytic cells, which are in attached to one another by fine filaments. turn phagocytized by macrophages. Macrophage digestion proceeds without risk to the cell unless the ingested mate- The Monocytic-Macrophage Series rial is toxic. If bacteria are not phagocytized or destroyed at entry, they may establish themselves at secondary sites, and Cells of the mononuclear phagocyte system include the bacteremia develops. monocytes and their bone marrow precursors, the mac- Both the eosinophils and basophils can participate in the rophages. Macrophages have a variety of names, depending phagocytic process, but they are relatively ineffective, for a on their tissue location. Collectively, they are referred to as variety of reasons. However, eosinophils and basophils have the mononuclear phagocyte system. separate important and specialized functions. Eosinophils Morphological identification of monocytes is more dif- prevent the excessive spread of inflammation. Histamine ficult than granulocyte identification. A convoluted nuclear found in basophils is important in acute allergic reactions, shape is typical of monocytes. The promonocyte, or young the most important being anaphylactic shock. monocyte, has a greater N:C ratio than the mature monocyte. The cytoplasm is gray-blue, with fine granules that resemble Assessment Methods ground glass. Vacuoles can frequently be observed. Several laboratory tests can be used to assess the inflamma- Normal Values and Functional Properties of tory response. Two important indicators are the total leu- Granulocytes and Monocytes kocyte count and the differential count of leukocytes on a peripheral blood smear. Other assessments include the abso- Defense against infectious disease is the responsibility of lute cell count, ESR, absolute counts of eosinophils and baso- both the phagocytic and the immune (antigen-antibody) phils, the LAP test, and neutrophilic function tests. CASE STUDY CASE 14.1 c. Total RBC 4.0 × 1012/L An 18-year-old woman came to the emergency department d. Total WBC 28.5 × 109/L because of severe abdominal pain. She had no fever or nau- A peripheral blood smear examination revealed the sea. Her periods had been regular. following: Laboratory Data e. Segmented PMNs 26% a. Hemoglobin 13.5g/dL f. Band neutrophils 42% b. Hematocrit 40% g. Lymphocytes 32% (continued)  CHAPTER 14 Leukocytes: The Granulocytic and Monocytic Series 251 CASE STUDY (continued) A few nucleated red blood cells were observed per 100 stress from a variety of nonmalignant disorders (e.g., WBCs on the peripheral blood smear. severe infection or inflammation, ovarian cysts, or inflammatory bowel disease). Questions 3. To distinguish between a leukemic condition and a 1. What abnormalities appeared in the patient’s laboratory leukemoid reaction, the LAP test could be used. An values? increased score is associated with a leukemoid reaction 2. What could cause this type of abnormality? or a variety of malignancies (e.g., lymphoma, multiple 3. How could leukemia be differentiated from a leukemia- myeloma, polycythemia vera). Decreased scores are like, leukemoid, reaction or lymphoma? associated with chronic myelogenous leukemia and Discussion some other conditions. 1. A leukoerythroblastic reaction is evident because of the presence of both immature granulocytes and nucleated DIAGNOSIS: Leukemoid Reaction Subsequent to erythrocytes. Ovarian Cysts 2. This type of reaction could be associated with malignant diseases (e.g., leukemias or lymphomas) or exogenous REVIEW QUESTIONS 1. The granulocyte cells that are believed to descend from a 5. The stages of neutrophilic granulocyte development are common multipotential stem cell in the bone marrow are A. promyelocyte, myeloblast, myelocyte, metamyelocyte, A. neutrophils and eosinophils and band and segmented neutrophils B. basophils and lymphocytes B. myeloblast, promyelocyte, myelocyte, metamyelocyte, C. lymphocytes and monocytes and band and segmented neutrophils D. Both A and B C. myelocyte, myeloblast, promyelocyte, metamyelocyte, 2. The types of granulocytic leukocytes found in the prolif- and band and segmented neutrophils erative compartment of the bone marrow are D. myeloblast, promyelocyte, metamyelocyte, A. myeloblasts, myelocytes, and metamyelocytes myelocyte, and band and segmented neutrophils B. myeloblasts, promyelocytes, and myelocytes 6. Marginating granulocytes in the peripheral blood can be C. myeloblasts, promyelocytes, myelocytes, and found metamyelocytes A. in the circulating pool D. myeloblasts, promyelocytes, myelocytes, metamy- B. in the tissues elocytes, and band neutrophils C. adhering to the vascular endothelium 3. The types of granulocytic leukocytes found in the D. all of the above maturation-storage compartment of the bone marrow 7. The major function of neutrophilic granulocytes is are A. antibody production A. metamyelocytes, band form neutrophils, segmented B. destruction of parasites neutrophils, mature eosinophils, and mature baso- C. phagocytosis phils D. suppression of inflammation B. only band form neutrophils, segmented neutrophils, 8. The half-life of circulating granulocytes in normal blood mature eosinophils, and mature basophils is estimated to be C. metamyelocytes, band form neutrophils, segmented A. 2.5 to 5 hours neutrophils, mature eosinophils, and mature baso- B. 7 to 10 hours phils C. 24 hours D. segmented neutrophils, immature and mature D. 2 days monocytes, and mature lymphocytes 9. Identify the cell with these characteristics: prominent 4. Release of neutrophils from the bone marrow is believed primary granules that are rich in myeloperoxidase and to be influenced by chloroacetate esterase and has a diameter of 14 to 20 mm. A. CSF A. myeloblast B. interleukins B. promyelocyte C. interferon C. myelocyte D. all of the above D. promonocyte (continued) 252 PART 3 Leukocytes REVIEW QUESTIONS (continued) 10. The earliest granulocytic maturational stage in which 19. The immediate precursor of the macrophage is the secondary or specific granules appear is A. myeloblast A. myeloblast B. monoblast B. monoblast C. promonocyte C. promyelocyte D. monocyte D. myelocyte 20. The correct sequence(s) of events in successful 11. The mature granulocytes seen in the peripheral blood of phagocytosis is (are) healthy persons include A. chemotaxis, opsonization, phagosome formation, A. band form and segmented neutrophils and the action of antibacterial substances B. eosinophils and basophils B. opsonization, chemotaxis, phagosome formation, C. lymphocytes and monocytes and the action of antibacterial substances D. Both A and B C. engulfment, opsonization, digestion, and destruction 12. The granules of segmented neutrophils contain of bacteria or particulate matter A. lysosomal hydrolases D. Both A and C B. lysozymes 21. The major function of eosinophils is C. myeloperoxidase A. suppression of inflammatory reactions D. all of the above B. destruction of protozoa 13. Which of the following are contents of basophilic C. participation in anaphylaxis granules? D. phagocytosis A. Heparin 22. Monocytes are capable of B. Histamine A. phagocytosis C. Myeloperoxidase B. synthesis of biologically important compounds D. Both A and B C. assuming a killer role 14. The tissue basophil can be referred to as a/an D. All of the above A. mast cell 23. The hematology tests that are useful in the early diagnosis B. macrophage of acute inflammation are the C. mononuclear cell A. total leukocyte count and total erythrocyte count D. antibody-producing cell B. total leukocyte count and white blood cell differential 15. A leukocyte with the morphological characteristics of being count the largest normal mature leukocyte in the peripheral C. ESR and absolute neutrophil cell count blood and having a convoluted or twisted nucleus is the D. Both B and C A. myelocyte 24. The total leukocyte count can be increased in certain B. metamyelocyte states. Select the conditions when this is not true. C. promonocyte A. Strenuous exercise D. monocyte B. Overwhelming bacterial infection 16. The reference range of PMN neutrophil count in adults C. Sepsis is D. Use of immunosuppressive agents A. 20 – 40% 25. Acute inflammation is based on B. 40 – 60% A. total leukocyte count >10.5 × 109/L C. 60 – 80% B. neutrophilic band count 20% 60 years Secondary AML Favorable Prognostic Factors in Existing disabilities TABLE 19.12 Features of multidrug resistance Acute Leukemia Peripheral blood leukocyte count >20,000 × 109/L Acute myeloid leukemia FACTORS USED TO PREDICT A RELAPSE Cytogenetic factors (patients aged 16–59 y) Unfavorable karyotype Age >60 years t(15;17) Delayed response to induction chemotherapy t(8;21) Features of multidrug resistance inv(16)/t(16:16) Peripheral blood leukocyte count >20,000 ×109/L Acute lymphoblastic leukemia Female gender AML, acute myeloid leukemia. Children aged 1–10 y 328 PART 3 Leukocytes be treated with more aggressive therapy, including very high includes administration of daily oral doses of 6-mercap- doses of cytarabine (HIDAC) alone or in combination with topurine with weekly infusions of methotrexate. Mainte- amsacrine, mitoxantrone, or etoposide. nance treatment usually lasts for approximately 2 years. Once remission is achieved, intensive treatment of patients The long-term survival rate after maintenance therapy is with AML is essential to prevent a relapse. approximately 30% to 40%. Allogenic stem cell transplant from an HLA-matched related or unrelated donor is the only chance for a cure. Postremission therapy for AML yields a median leuke- Stem Cell Transplant mia-free duration of 12 to 18 months, with approxi- AMLs and ALLs are treatable by stem cell transplantation. mately 20% to 25% of these patients being cured of their Progenitor blood cells are considered to be pluripotent disease. because they have the ability to evolve into different types of cells, for example, granulocytes, lymphocytes, etc. Some Relapse progenitor cells circulate in the blood stream and are called peripheral blood stem cells (PBSCs). PBSCs are found in When treatment fails in patients with AML, the available much smaller quantities in the circulating blood than in the options are determined by age, duration of the first remis- bone marrow but are becoming a popular mode of allogenic sion, and cytogenetic findings, among other factors. Patients transplantation. The CD34 antigen identifies a population of with favorable cytogenetic characteristics—t(15;17), stem cells that can reconstitute hematopoiesis after myeloab- t(8;21), or inv(16) mutation—who were in remission for lative chemotherapy. more than 1 year before relapse have an approximately 20% chance of survival after subsequent therapy. For children and younger adults who have a first relapse or who do not Future Trends Vaccines have a complete response to first-line induction therapy, the recommended option is marrow-ablative (high-dose) Vaccines for cancer, including AML, are under investigation. cytotoxic treatment followed by hematopoietic stem cell An ongoing clinical trial of peptide vaccines, derived from transplantation, including autog