Structure and Function of the Hematologic System

In adults, plasma accounts for 50% to 55% of blood volume (see Fig. 28.1). Plasma is a complex aqueous liquid containing a variety of organic and inorganic elements (Table 28.1). The concentration of these elements varies, depending on diet, metabolic demand, hormones, and vitamins. Plasma differs from serum in that serum is free of clotting proteins (e.g., fibrinogen). The clotting proteins may interfere with some diagnostic tests.

Plasma contains a large number of proteins (plasma proteins) that make up about 7% of the total plasma weight. These vary in structure and function and can be classified primarily into two major groups, albumin and globulins, plus a small amount of clotting proteins. Most plasma proteins are produced by the liver. The major exception is antibody, which is produced by plasma cells (B cells) in the lymph nodes and other lymphoid tissues (see Chapter 8).

Albumin (about 57% of total plasma protein) serves as a carrier molecule for the normal components of blood and for drugs that have low solubility in water (e.g., free fatty acids, lipid-soluble hormones, thyroid hormones, bile salts). Its most essential role is regulation of the passage of water and solutes through the capillaries. Albumin molecules are large and do not diffuse freely through the vascular endothelium, and thus they maintain the critical colloidal osmotic pressure (or oncotic pressure) that regulates the passage of fluids and electrolytes into the surrounding tissues (see Chapters 1 and 3). Water and solute particles tend to diffuse out of the arterial portions of the capillaries because blood pressure (hydrostatic pressure) is greater in arterial than in venous blood vessels. Water and solutes move from tissues into the venous portions of the capillaries, where the pressures are reversed, oncotic pressure being greater than intravascular pressure or hydrostatic pressure (see Fig. 3.1). In the case of decreased production of albumin (e.g., cirrhosis, other diffuse liver diseases, protein malnutrition) or excessive loss of albumin (e.g., certain kidney diseases, extensive burns), the reduced plasma oncotic pressure leads to excessive movement of water and solutes into the tissue and decreased blood volume.

Most of the remaining plasma proteins are globulins (about 38% of total plasma protein), which are often classified by their properties in an electric field (serum electrophoresis). Under the normal conditions used to perform serum electrophoresis, albumin is the most rapidly moving protein. The globulins are classified into alpha, beta, and gamma groups by their movement relative to albumin. Alpha (α) globulins (those moving most closely to albumin) include high-density lipoproteins [HDLs], prothrombin, and proteins that transport hormones. Beta (β) globulins include low-density lipoproteins (LDLs). Gamma (γ) globulins (those with the least movement) consist primarily of immunoglobulin G (IgG) (see Chapter 8).

Plasma proteins also can be classified by function: clotting, defense, transport, or regulation. The clotting factors or proteins promote coagulation and stop bleeding from damaged blood vessels. Fibrinogen is the most plentiful of the clotting factors and is the precursor of the fibrin clot (see Fig. 28.19). Proteins involved in defense, or protection, against infection include antibodies (immunoglobulins) and complement proteins (see Chapters 8 and 9). Transport proteins specifically bind and carry a variety of inorganic and organic molecules, including iron (transferrin), copper (ceruloplasmin), lipids and steroid hormones (lipoproteins) (see Chapter 1), and vitamins (e.g., retinol-binding protein for vitamin A transport). The plasma lipids, triglycerides, phospholipids, cholesterol, and fatty acids are transported through the blood as complexes with plasma proteins; they are known as lipoproteins (see Chapters 1 and 32). Regulatory proteins include a variety of enzymatic inhibitors (e.g., α₁-antitrypsin) that protect the tissues from damage; precursor molecules (e.g., kininogen) that are converted into active biologic molecules when needed; and protein hormones (e.g., cytokines) that communicate between cells.

Plasma also contains several charged inorganic ions that regulate cell function, osmotic pressure, and blood pH. These include the electrolytes sodium, potassium, calcium, chloride, and phosphate (Electrolytes are described in Chapters 1 and 3).

The cellular elements of the blood are broadly classified as red blood cells (erythrocytes), WBCs (leukocytes), and platelets (thrombocytes) (Fig. 28.2). The components of the blood are listed in Table 28.2.

Erythrocytes

Erythrocytes (red blood cells [RBCs]) are the most abundant cells of the blood, occupying approximately 48% of the blood volume in men and about 42% in women. Erythrocytes are formed in the bone marrow, and there are 4.2 to 6.2 million erythrocytes/mm³ circulating in normal blood. Erythrocytes are primarily responsible for tissue oxygenation. The erythrocyte contains hemoglobin (Hb), which carries the gases, and electrolytes, which regulate gas diffusion through the cell’s plasma membrane. The mature erythrocyte lacks a nucleus and cytoplasmic organelles (e.g., mitochondria), so it cannot synthesize protein or carry out oxidative reactions. Because it cannot undergo mitotic division, the erythrocyte has a limited life span (approximately 100 to 120 days), ages, and is removed from the circulation, primarily in the spleen, to be replaced by new erythrocytes.

The erythrocyte’s size and shape is ideally suited to its function as a gas carrier. It is a small disk with two unique properties: (1) a biconcave shape and (2) the capacity to be reversibly deformed. The flattened, biconcave shape (see Fig. 28.2) provides a surface area/volume ratio that is optimal for gas diffusion into and out of the cell and for deformity. During its life span, the erythrocyte, which is 6 to 8 μm in diameter, repeatedly circulates through splenic sinusoids and capillaries that are only 2 μm in diameter. Reversible deformity enables the erythrocyte to assume a more compact torpedo-like shape, squeeze through the microcirculation (diapedesis), and return to normal (Fig. 28.3).

A photomicrograph shows a ring of endothelium cells, surrounded by red cells. An accompanying illustration shows a similar ring of endothelium cells, surrounded by red cells. A macrophage engulfing R B C is also identified in the illustration.

Erythrocytes are also part of the innate immune system. Erythrocytes catch bacteria in the blood stream by electric charge attraction and kill them by oxycytosis, the release of oxygen from oxyhemoglobin (oxygen-dependent killing). The decomposed bacteria are then cleared in the liver and spleen. Oxycytosis is complimentary to leukocytosis (phagocytosis) as leukocytes cannot recognize and attach bacteria in the blood because of the velocity of blood flow.1

Leukocytes

Leukocytes (WBCs) (see Fig. 28.2) defend the body against organisms that cause infection and also remove debris, including dead or injured host cells of all kinds. The leukocytes act primarily in the tissues but are transported in the circulation. The average adult has approximately 5000 to 10,000 leukocytes/mm³ of blood.

Leukocytes are classified according to structure as either granulocytes or agranulocytes and according to function as either phagocytes or immunocytes (cells that create immunity; see Chapter 8). The granulocytes, which include neutrophils, basophils, eosinophils, and mast cells, are all phagocytes. (Phagocytic action is described in Chapter 7.) Of the agranulocytes, the monocytes and macrophages are phagocytes, whereas the lymphocytes are immunocytes. Fig. 28.4 shows a microscopic image of the various types of leukocytes.

Granulocytes

The granulocytes are formed in the red bone marrow and have a nucleus with several lobes (polymorphonuclear) and many membrane-bound granules in their cytoplasm. These granules contain enzymes capable of killing microorganisms and catabolizing debris ingested during phagocytosis. The granules also contain powerful biochemical mediators with inflammatory and immune functions. These mediators, along with the digestive enzymes, are released from granulocytes in response to specific stimuli and affect other cells in the circulation. The biochemical mediators have vascular and intercellular effects, and the enzymes participate in the breakdown of debris from sites of infection or injury. Granulocytes are capable of amoeboid movement, by which they migrate through vessel walls (diapedesis) and then to sites where their action is needed. They have a life span of only a few hours or days.

The neutrophil (polymorphonuclear neutrophil [PMN]) is the most numerous and best understood of the granulocytes. Neutrophils constitute about 65% to 75% of the total leukocyte count in adults. The results vary by laboratory and populations studied.

The cytoplasm of neutrophils contains small lysosomal granules and a central nucleus with two to five distinct lobes. Immature neutrophils are called bands or stabs. Mature neutrophils are called segmented neutrophils because of the characteristic appearance of their nucleus (see Fig. 28.4). Neutrophils reach a fully mature state in the bone marrow, where these mature neutrophils are called the marrow neutrophil reserve. Normally it takes about 14 days for neutrophils to develop from early precursors, but this process is accelerated by infection and treatment with colony-stimulating factors.

Neutrophils are the chief phagocytes of early inflammation. Soon after bacterial invasion or tissue injury, neutrophils migrate out of the capillaries and into the damaged tissue, where they phagocytize and destroy contaminating microorganisms and debris. Neutrophils also release nuclear DNA onto extracellular pathogens to control their growth and proliferation and aid the killing of bacteria. These extracellular DNA deposits are termed neutrophil extracellular traps (NETs), and are a phagocytosis-independent anti-microbial pathway. In addition to their crucial role in fighting infection, neutrophils play a complex role in tumors. (see Emerging Science Box: Tumor Associated Neutrophils). Neutrophils are sensitive to the environment in damaged tissue (e.g., low pH, enzymes released from damaged cells) and die in 1 or 2 days. The breakdown of dead neutrophils releases digestive enzymes from their cytoplasmic granules. These enzymes dissolve cellular debris and prepare the site for healing. (This final function, called débridement, is described in Chapter 7.)

Eosinophils, which have large, coarse granules, constitute only about 2% to 5% of the normal leukocyte count in adults. Using a spectrum of pattern-recognition receptors, eosinophils are capable of amoeboid movement and phagocytosis. Unlike neutrophils, eosinophils ingest antigen-antibody complexes and are induced by immunoglobulin E (IgE)–mediated hypersensitivity reactions to attack parasites by release of toxic molecules. Eosinophils also mediate immunity against bacterial, viral, and fungal infections by presenting bacterial, viral, and parasitic antigen to cytotoxic T cells. IL-5 promotes the maturation and migration of eosinophils during the inflammatory response. Eosinophilic inflammation mediated by IL-5, particularly from T helper cells, and other cytokines, contributes to the type 2 hypersensitivity response associated with asthma and other chronic eosinophilic inflammatory diseases (see Chapter 9). During inflammation, eosinophil death and cytolysis is known as extracellular trap cell death (ETosis), which differs from the processes of accidental necrosis or apoptosis. Eosinophil extracellular trap products consist of damage-associated molecular patterns (DAMPs) that are able to activate both the innate and adaptive immune systems (see Chapters 7 and 8).² Eosinophils also release IL-4 important for tissue repair and regeneration in muscle and liver.³

Eosinophil secondary granules contain toxic chemicals (e.g., major basic protein, eosinophil cationic protein, eosinophil peroxidase, eosinophil-derived neurotoxin) that are highly destructive to parasites and viruses. Eosinophil granules also contain a variety of enzymes (e.g., histaminase) that help control inflammatory processes. Eosinophils also release proinflammatory molecules, including leukotrienes, prostaglandins, platelet-activating factor (PAF), and a variety of cytokines (e.g., IL-1, IL-4, IL-6, tumor necrosis factor-alpha [TNF-α], and granulocyte-macrophage colony-stimulating factor [GM-CSF]). Type I and type II hypersensitivity allergic reactions and asthma are characterized by high circulating eosinophil counts (see Chapters 9 and 36). The eosinophils may be involved in a dual role: regulation of inflammation and contribution to the destructive inflammatory processes observed in allergic reactions (e.g., the lungs of persons with asthma and eosinophilic esophagitis).

Basophils, which make up less than 1% of leukocytes, are structurally similar to the mast cells (see Fig. 28.4). Basophils contain cytoplasmic granules that have an abundant mixture of biochemical mediators, including histamine, chemotactic factors, proteolytic enzymes (e.g., elastase, lysophospholipase), and an anticoagulant (heparin). Stimulation of basophils also induces synthesis of vasoactive lipid molecules (e.g., leukotrienes) and cytokines, including IL-6, which affects differentiation of type 1 T-helper cells (Th1 cells) (focus on intracellular parasites) and type 2 T-helper cell (Th2 cells) (focus on extracellular parasites). Basophils are a particularly rich source of the cytokine IL-4, which preferentially guides B-cell differentiation toward plasma cells that secrete IgE (see Chapter 8).

The number of basophils is often increased at sites of allergic inflammatory reactions and parasitic infection, particularly ectoparasites (e.g., ticks). IgE receptors on the basophil induce degranulation at sites of IgE-mediated hypersensitivity reactions and contribute to the local inflammatory response.

Mast cells are highly similar to basophils but are generated from a different set of precursor cells in the bone marrow, from which they migrate in an immature form into tissues. Mast cells reside in vascularized connective tissues just beneath body epithelial surfaces, including the submucosal tissues of the respiratory and gastrointestinal tracts, as well as in the dermal layer that lies just below the surface of the skin. Mast cells play a central role in inflammation. Activation and degranulation of the mast cells affect a great number of other cells, including those involved in inflammation (e.g., vascular endothelial cells, smooth muscle cells, circulating platelets and leukocytes, nerves) and healing (e.g., fibroblasts), as well as glandular cells and cells of the immune system. Their activation contributes greatly to increased permeability of blood vessels and synthesis of mediators producing smooth muscle contraction (see Fig 7.4).

Agranulocytes

The agranulocytes—monocytes, macrophages, and lymphocytes—contain relatively fewer granules than granulocytes. Monocytes and macrophages make up the mononuclear phagocyte system (MPS) (see Chapter 7). Both monocytes and macrophages participate in the immune and inflammatory response, being powerful phagocytes. They also ingest dead or defective host cells, particularly blood cells.

The mononuclear phagocyte system (MPS), also known as the reticuloendothelial system or macrophage system, consists of specialized endothelial cells: promonocytes and their precursors in the bone marrow; monocytes in the peripheral blood; and a portion of macrophages that reside in the tissues, remaining there for months or perhaps years. Cells of the MPS ingest and destroy (by phagocytosis) unwanted materials, such as foreign protein particles, circulating immune complexes, microorganisms, debris from dead or injured cells, defective or injured erythrocytes, and dead neutrophils.

Monocytes are immature macrophages. They are the largest normal blood cell and have a horseshoe-shaped nucleus (see Fig. 28.4). Monocytes are formed and released by the bone marrow into the bloodstream. As they mature, monocytes migrate into a variety of tissues (e.g., liver [Kupffer cells], spleen, lymph nodes, peritoneum, gastrointestinal tract, brain [microglia], and bone) and differentiate into tissue macrophages with tissue-specific functions (Table 28.3). Monocytes also differentiate into myeloid linage dendritic cells and disseminate to lymphoid organs and body surfaces such as the skin and mucous membranes. Both macrophages and dendritic cells are part of the MPS. Other monocytes may migrate out of blood vessels and differentiate into macrophages in response to infection or inflammation. Macrophages are generally larger and are more active as phagocytes than monocytes. Dendritic cells frequently extend projections (dendrites) into the tissue and assume a “neuron-like” appearance. The origin and turnover of many of the tissue macrophages are not precisely known. It seems clear that once monocytes leave the circulation, they do not return. They can survive many months or even years.

Table 28.3

Mononuclear Phagocyte System

Name of Cell	Location
Committed Stem Cells	Bone marrow
Monoblasts	Bone marrow
Promonoblasts	Bone marrow
Monocytes	Bone marrow and peripheral blood
Macrophages	Tissue
Kupffer cells	Liver macrophages
Alveolar macrophages	Lung
Histiocytes	Connective tissue
Macrophages	Bone marrow
Fixed and free macrophages	Spleen and lymph nodes
Pleural and peritoneal macrophages Adipose macrophages	Serous cavities Adipose (fat) tissue
Microglial cells	Nervous system
Mesangial cells	Kidney
Osteoclasts	Bone
Langerhans cells	Skin
Dendritic cells	Lymphoid tissue, lining of respiratory and gastrointestinal tracts

Modified from Kumar V, et al. Robbins & Cotran pathologic basis of disease, 9th edition. Philadelphia: Saunders; 2015.

The normal role of macrophages is to remove old and damaged cells and large molecular substances from the blood. Cellular targets of macrophage phagocytosis include circulating senescent or damaged erythrocytes and platelets (removed primarily in spleen), dead neutrophils (in the circulation and at sites of inflammation), and cells undergoing apoptosis. Noncellular targets include antigen-antibody complexes, cellular debris, products of coagulation, and macromolecules (such as lipids and carbohydrates synthesized by the body as the result of faulty metabolism, as in storage diseases). Macrophages remove and kill contaminating microorganisms in the blood (mostly in the liver and spleen) and at sites of infection. Macrophages and, particularly, dendritic cells are the major “antigen-processing” and “antigen-presenting” cells that initiate immune responses (see Chapter 8). Macrophages initiate wound healing and tissue remodeling and, if activated by cytokines from T cells, secrete a large array of biologically active chemicals that if uncontrolled result in chronic inflammation and tissue injury (see Chapter 7).

Lymphocytes constitute about 20% to 25% of the total leukocyte count and are the primary cells of the immune response (see Fig. 28.4 and Chapter 8). Lymphocytes arise from lymphoid stem cells in the bone marrow and subsequently develop and reproduce in the lymphatic tissues as mature T cells, B cells, or plasma cells. The lymphocytes do not contain any enzyme-filled digestive vacuoles. The life span of the lymphocyte can be days, months, or years, depending on its type and subtype.

Natural killer (NK) cells, which resemble large granular lymphocytes, recognize cells that do not express “self” proteins (self-antigens) on their plasma membrane or that contain foreign or abnormal antigens such as cancer cells or some virus-infected cells without prior exposure to these antigens (see Chapters 7 and 8). Hence, they are named natural killer cells as they do not require exposure to antigen for activation to differentiate them from T-cytotoxic cells which are induced by antigen. NK cells also have the capacity to activate T cells and phagocytes and produce a variety of cytokines that can regulate immune responses. The predominant form of NK cells develops in the bone marrow and circulates in the blood, and is found mainly in the peripheral blood and spleen where it accounts for 5% to 10% of the circulating lymphoid pool. NK cells develop independent of the thymus, although some NK precursors are found in the thymus and may develop into NK T cells, which will then have markers of both NK cells and T cells.⁴

Platelets

Platelets (thrombocytes) are not true cells, but rather irregularly discoid-shaped cytoplasmic fragments that lack a nucleus and deoxyribonucleic acid (DNA) and are incapable of mitotic division. Platelets are essential for blood coagulation and control of bleeding. They are formed in the bone marrow by fragmentation of very large cells (40 to 100 μm in diameter) known as megakaryocytes. They contain cytoplasmic granules (e.g., dense granules, alpha granules, and lysosomes) and can release adhesive proteins and coagulation and growth factors when stimulated by injury to a blood vessel. Platelets can assume different shapes and form adhesive pseudopodia (long surface extensions) that increase their surface area and promote interconnectedness and adherence to collagen fibers in damaged vascular walls, plugging vascular openings to control bleeding (Fig. 28.5).

The normal platelet concentration is about 150,000 to 400,000 platelets/mm³ of circulating blood, although the normal ranges may vary slightly from laboratory to laboratory. An additional one-third of the body’s available platelets are in a reserve pool in the spleen. A platelet circulates for approximately 8 to 11 days, and is then removed by macrophages, mostly in the spleen.

The spleen is a concave, encapsulated organ that weighs about 150 g and is about the size of a fist. It is located in the left upper abdominal cavity, curved around a portion of the stomach. Strands of connective tissue (trabeculae) extend throughout the spleen from the splenic capsule, dividing it into compartments that contain masses of lymphoid tissue called splenic pulp (white and red) (Fig. 28.6). The spleen is interlaced with many blood vessels, some of which can distend to store blood. Blood that circulates through the spleen comes from the splenic artery, which branches from the descending aorta and reenters the circulatory system through the splenic vein and into the portal vein.

Three illustrations, A, B, C, and D, shows the splenic architecture. Top-left panel, A. The illustration shows a cross-section of the spleen and highlights the following structures: trabecula showing trabecular vessels, trabeculae, red pulp, white pulp, and capsule. Top-right panel, B. The illustration shows and labels the following subcapsular splenic structures, clockwise from the top: lymphoid nodule, capsule, red pulp (pulp cords, venous sinusoids), and white pulp (germinal center, corona of B cells, periarterial lymphatic sheath, trabecula, and trabecular vein. Middle panel, C. The illustration shows magnified view of the red pulp and the white pulp. The red pulp comprises venous sinusoid, terminal arterial capillary, sheathed arteriole, venous sinusoid, penciller artery, terminal artery capillary, sheathed arteriole, and pulp arteriole. The white pulp comprises lymphocytes, lymphoid nodule corona of B cells, germinal center, corona, central artery, marginal sinusoid, marginal zone (B cells), periarterial lymphatic sheath (T cells), and marginal zone (B cells). Bottom panel, D. The illustration shows the splenic arteries and the trabecular arteries. The following structures on the illustration are labeled, marginal zone (M Z B cells), primary follicle (T h 2, mature B cells), secondary follicle (mantle zone and germinal center), and periarterial lymphatic sheath (T cells).

Arterial blood that enters the spleen first encounters the white splenic pulp, which consists of masses of lymphoid tissue containing macrophages and lymphocytes, primarily T lymphocytes in proximity to the arterioles (the periarterial lymphoid sheath) (see Fig. 28.6). Cellular clumps (lymphoid follicles) are formed in the white pulp around the splenic arterioles. The lymphoid follicles consist primarily of B lymphocytes and are the chief sites of immune function within the spleen. Here blood-borne antigens encounter lymphocytes, initiating the immune response and the conversion of lymphoid follicles into germinal centers in the middle of the follicle where B cells proliferate and differentiate during the humoral immune response (see Chapter 8).

Some of the blood that enters the terminal capillaries continues through the microcirculation and enters highly distensible storage areas, called venous sinuses, in the red pulp of the spleen. The venous sinuses (and the red pulp) can store more than 300 mL of blood. Passive dilation of the venous sinuses enables the spleen to increase its storage capacity as needed by the body. Sudden reductions in blood pressure cause the sympathetic nervous system to stimulate constriction of the sinuses and expel as much as 200 mL of blood into the venous circulation, helping restore blood volume or pressure in the circulation and increasing the hematocrit by as much as 4%.

The endothelial lining of the venous sinuses is discontinuous (having gaps between endothelial cells) and therefore extremely permeable, allowing blood cells to exit the circulation. The red pulp contains a system of loosely interconnected resident macrophages that provide the principal site of splenic filtration. Because of the slow circulation in the sinuses, the macrophages easily phagocytose old, damaged, or dead blood cells of all kinds (but chiefly erythrocytes), microorganisms, macromolecules, and particles of debris. Hemoglobin from phagocytosed erythrocytes is catabolized, and heme (iron) is stored in the cytoplasm of the macrophages or released back into the blood (see Fig. 28.15). The macrophages also can remove particulate inclusions containing denatured hemoglobin (Heinz bodies) from erythrocytes without harming the cells themselves. Blood that filters through the red pulp then moves through the venous sinuses and into the portal circulation (blood flowing to the liver).

The spleen is not absolutely necessary for life or for adequate hematologic function. However, splenic absence as a result of any cause (atrophy, traumatic injury, or removal because of disease) has several secondary effects on the body. For example, leukocytosis (high levels of circulating leukocytes) often occurs after splenectomy, suggesting that the spleen exerts some control over the rate of proliferation of leukocyte stem cells in the bone marrow or their release into the bloodstream. Circulating levels of iron also may decrease, reflecting the spleen’s role in the iron cycle (see the Iron Cycle section). The immune response to encapsulated bacteria (e.g., Streptococcus pneumoniae [pneumococcus], Neisseria meningitidis [meningococcus], and Haemophilus influenzae), which is primarily an IgM response, may be severely diminished, resulting in increased susceptibility to disseminated infections and sepsis. An increase in morphologically defective blood cells within the circulation after the loss of the spleen helps confirm the spleen’s role in removing old or damaged cells. Thrombocytosis and thrombosis are complications of splenectomy and requires anticoagulation therapy.⁵

Functionally, lymph nodes are part of the hematologic and immune systems and are the primary site for the first encounter between antigen and lymphocytes. Lymphocytes enter the lymph node from the blood through specialized postcapillary venules called high endothelial venules (HEVs) by means of diapedesis across the endothelial lining (Fig. 28.7). Within the secondary lymph nodes, naïve T and B lymphocytes are segregated into different regions. B lymphocytes tend to migrate preferentially to nodes in the cortex and medulla, whereas T lymphocytes predominantly migrate to the paracortex. Macrophages reside in the lymph node; help filter the lymph of debris, foreign substances, and microorganisms; and provide antigen-processing functions. The dendritic cells encounter and process antigens (see Chapter 8) and microorganisms in other tissues, then enter the lymph node through the afferent lymph vessels, and migrate throughout the nodes. The reticular network provides adhesive surfaces for trapping large numbers of phagocytes and lymphocytes and facilitates their organization into follicles or primary nodules. The presence of antigen, either removed from the lymph by macrophages or presented on the surface of dendritic cells, results in the production of secondary nodules containing germinal centers. In the germinal centers lymphocytes, particularly B cells, respond to antigenic stimulation by undergoing proliferation and further differentiation into memory cells and plasma cells (see Chapter 8). Plasma cells migrate to the medullary cords. The B-lymphocyte proliferation in response to a great deal of antigen (e.g., during infection) may result in lymph node enlargement and tenderness (reactive lymph node), which can be palpable during a physical examination.

Three illustrations, A, B, and C, show the architecture of the lymph node. Left panel, A. The illustration shows a cross-section of the lymph vessel. The following structures on the illustration are labeled, clockwise from the top: afferent lymphatic vessels, interdigitating dendritic cells in the paracortical zone, trabeculae, capsule, intermediary sinus, capillary web, medullary sinus, efferent lymphatic vessel, artery, vein, high endothelial venules (H E Vs), paracortical zone (T-cell zone), secondary follicle (B-cell zone), follicular dendritic cells in a secondary follicle, and subcapsular cortical sinus. Top-right panel, B. The illustration shows the cellular composition of the lymph node. The following structures on the illustration, clockwise from the top: sinus lining cell, capsule, subcapsular sinus and lymph flow, fibroblastic reticular cell (F R C) conduit, reticular fiber, fibroblastic reticular cell, subcapsular macrophage or dendritic cell, trabecular sinus and lymph flow, dendritic cell, H E V, and perivenular channel. Bottom-right panel, C. The illustration shows the subcapsular sinus and labels the following parts: subcapsular or cortical sinus cell, lymphocyte, capsule, subcapsular or cortical sinus cell, macrophage, and subcapsular or cortical sinus.

Erythropoiesis is the development of RBCs. In the confines of the bone marrow, progenitor cells proliferate and differentiate into large, nucleated proerythroblasts, which are committed to producing erythroid cells (Fig. 28.11). The proerythroblast, which has ribosomes and can produce protein, differentiates through several intermediate forms of erythroblast (sometimes called normoblast) while progressively eliminating most intracellular structures (including the nucleus), synthesizing hemoglobin, and becoming more compact, eventually assuming the shape and characteristics of an erythrocyte.

A series of illustration shows the differentiation of erythrocyte. The sequence of differentiation is as follows. • Erythroid progenitor. • Erythroid progenitor in the presence of erythropoietin develops into committed proerythroblast. • Committed proerythroblast (hemoglobin synthesis begins). • Normoblast (nucleus shrinks and is reabsorbed). • Reticulocyte (cell leaves marrow and enters blood-stream). • Erythrocyte (cell achieves final size and shape; hemoglobin synthesis ceases).

The last immature form is the reticulocyte, which is anucleate and contains a mesh-like (reticular) network of ribosomal ribonucleic acid (rRNA). The reticulocyte contains polyribosomes (for globin synthesis) and mitochondria (for oxidative metabolism and heme synthesis). The reticulocyte matures into an erythrocyte within 24 to 48 hours. During this period, mitochondria and ribosomes disappear and the cell becomes smaller and more disc-like. With these final changes, the erythrocyte loses its capacity for hemoglobin synthesis and oxidative metabolism. Reticulocytes remain in the marrow approximately 1 day and are released into the venous sinuses. They continue to mature in the bloodstream and may travel to the spleen for several days of additional maturation. The normal reticulocyte count is 1% of the total RBC count. Approximately 1% of the body’s circulating erythrocyte mass normally is generated every 24 hours. Therefore, the reticulocyte count is a useful clinical index of erythropoietic activity and indicates whether new red cells are being produced.

Regulation of erythropoiesis

In healthy humans, the total volume of circulating erythrocytes remains surprisingly constant. Most steps of erythropoiesis are primarily under the control of a feedback loop involving erythropoietin (EPO) and other cytokines (see Table 28.4). In conditions of tissue hypoxia, EPO is secreted primarily by the peritubular cells of the kidney (Fig. 28.12). Rising circulating levels of EPO cause a compensatory increase in erythrocyte production. The normal steady-state rate of production of 2.5 million erythrocytes per second can increase to 17 million per second under anemic or low-oxygen states, such as high-altitude environments or pulmonary disease. Thus the body responds to reduced oxygenation of blood in two ways: (1) by increasing the intake of oxygen through increased respiration and (2) by increasing the oxygen-carrying capacity of the blood through increased erythropoiesis.

An illustration shows the role of erythropoietin in regulation of erythropoiesis. 1. Decreased R B Cs, decreased hemoglobin synthesis, decreased blood flow, hemorrhage, and increased oxygen consumption by tissues stimulate the kidney. 2. Oxygen level decreases. 3. E P O increases. 4. Triggers the bone marrow. 5. Erythrocytes increase. 6. Oxygen level increases. 7. E P O decreases.

Recombinant human erythropoietin (r-HuEPO) is used in individuals with anemia secondary to decreased EPO from chronic renal failure. An immediate effect of EPO administration is an increase in the blood reticulocyte count, followed by increasing levels of erythrocytes. The most significant side effect is increased blood pressure.

Hemoglobin synthesis

Erythrocytes form hemoglobin in the bone marrow. Hemoglobin (Hb), the oxygen-carrying protein of the erythrocyte, constitutes approximately 90% of the cell’s dry weight. Hb-packed blood cells take up oxygen in the lungs and exchange it for carbon dioxide in the tissues. A single erythrocyte can contain as many as 300 hemoglobin molecules. Hb increases the oxygen-carrying capacity of blood by 100-fold. Each Hb molecule is composed of two pairs of polypeptide chains (the globins) and four colorful complexes of iron plus protoporphyrin (the hemes), which are responsible for the blood’s ruby-red color and oxygen-carrying capacity (Fig. 28.13).

Each polypeptide chain contains approximately 150 amino acids and is arranged in a knotted-sausage configuration. The chains assemble to form a tetrahedron containing two pairs of identical chains. Several variants of Hb exist (Table 28.5), and they differ only slightly in primary structure based on the use of different polypeptide chains: alpha, beta, gamma, delta, epsilon, or zeta (α, β, γ, δ, ε, or ζ, respectively). Hb A, the most common type in adults, is composed of two α- and two β-polypeptide chains (α₂β₂) (see Fig. 28.13). A normal variant, fetal Hb (Hb F), is a complex of two α- and two γ-polypeptide chains (α₂γ₂) that binds oxygen with a much greater affinity than adult Hb. Other variants are genetic mutations associated with various diseases (see Chapter 29).¹⁰

Table 28.5

Structure of Normal Hemoglobin Molecules

Type of Hemoglobin (Hb)	Identity of Polypeptide Chain	Significance
HbA	α2β2	92% of adult Hb
HbA1c	α2 (β-NH-glucose)	5% of adult Hb; increased in diabetes (see Chapter 22)
HbA2	α2δ2	2% of adult Hb; increased in β-thalassemia (see Chapter 29)
HbF	α2γ2	Major fetal Hb from the third through ninth month of gestation; promotes oxygen transfer across platelets; increased in β-thalassemia
Hb Gower I	ε4 or ζ2β2	Present in early embryo; function unknown
Hb Gower II	α2ε2	Present in early embryo; function unknown
Hb Portland	ζ2γ2	Present in early embryo; function unknown

NH, Amine.

Heme is a large, flat, iron-protoporphyrin disc that is synthesized in the mitochondria and can carry one molecule of oxygen (O₂). Thus, an individual Hb molecule with its four hemes can carry four oxygen molecules. If all four oxygen-binding sites are occupied by oxygen, the molecule is said to be saturated. Through a series of biochemical reactions, protoporphyrin, a complex four-ringed molecule, is produced and bound with ferrous iron. It is crucial that the iron be correctly charged; reduced ferrous iron (Fe²⁺) can bind oxygen in the lungs and release it in the tissues, whereas ferric iron (Fe³⁺) cannot. Binding of O₂ to ferrous iron temporarily oxidizes Fe²⁺ to Fe³⁺ (oxyhemoglobin), but after the release of O₂ the body reduces the iron to Fe²⁺ and reactivates the Hb (deoxyhemoglobin [reduced hemoglobin]). Without reactivation, the Fe³⁺-containing Hb (methemoglobin) cannot bind O₂. An excess of ferric iron occurs with certain drugs and chemicals, such as nitrates and sulfonamides, and reduces oxygen-carrying capacity.

Hb undergoes a conformational change when binding O₂. When one of the iron molecules binds O₂, the porphyrin ring changes shape, increasing the exposure of the three remaining iron atoms to O₂. This greatly increases the affinity for the oxygen-carrying capacity of Hb, as occurs in the lungs. When oxygen is unloaded from Hb, the oxygen-carrying capacity of hemoglobin is low, facilitating the transport of carbon dioxide back to the lungs.

Several other molecules can competitively bind to deoxyhemoglobin. Carbon monoxide (CO) directly competes with O₂ for binding to Fe²⁺ with an affinity that is about 200-fold greater than that of O₂. Thus even a small amount of CO can dramatically decrease the ability of Hb to bind and transport O₂. Hb also binds carbon dioxide (CO₂), but at a binding site separate from where O₂ binds. In the lungs, CO₂ is released, allowing hemoglobin to bind O₂.

Erythrocytes may also play a role in the maintenance of vascular relaxation and blood flow. Nitric oxide (NO) produced by blood vessels is a major mediator of relaxation and dilation of the vessel walls. In the lungs, Hb can concurrently bind O₂ to the Fe²⁺ and NO to cysteine residues in the globins. As Hb transfers its O₂ to tissue, it may also shed small amounts of NO, contributing to dilation of the blood vessels and helping transfer of the O₂ into tissues.

Normal destruction of senescent erythrocytes

After about 120 days in the circulation, old erythrocytes are removed by tissue macrophages, primarily in the spleen.11 Although mature erythrocytes lack nuclei, mitochondria, and the endoplasmic reticulum, they do have cytoplasmic enzymes capable of glycolysis (anaerobic glucose metabolism) and production of small quantities of adenosine triphosphate (ATP). ATP provides the energy needed to maintain cell function and keep its plasma membrane pliable. Metabolic processes diminish as the erythrocyte ages, so less ATP is available to maintain plasma membrane function. The aged or senescent red cell becomes increasingly fragile and loses its reversible deformability due to disruption of the anchorage between the cytoskeleton and the plasma membrane, thus becoming susceptible to rupture while passing through narrowed regions of the microcirculation.

Additionally, the plasma membrane of senescent red cells undergoes phospholipid rearrangement, signaling macrophages (primarily in the spleen) to selectively remove and sequester the senescent red cells. If the spleen is dysfunctional or absent, macrophages in the liver (Kupffer cells) assume control of this process. The erythrocytes are digested by proteolytic and lipolytic enzymes in the phagolysosomes (digestive vacuoles) of the macrophage. The heme and globin dissociate easily, and the globin is broken down into its component amino acids. The iron in hemoglobin is oxidized, forming Fe³⁺ (methemoglobin), and recycled.

During digestion of Hb in the macrophage, porphyrin reduces to unconjugated bilirubin, which is conjugated (made water soluble) and excreted from the liver into the intestine or into bile (Fig. 28.14). Bacteria in the intestinal lumen transform conjugated bilirubin into urobilinogen. Although a small portion is reabsorbed and excreted by the kidneys, most urobilinogen is excreted in feces. Conditions causing accelerated erythrocyte destruction increase the load of serum unconjugated bilirubin, increased liver conjugation of bilirubin, and increased urinary excretion of urobilinogen. Gallstones (cholelithiasis) can result from a chronically elevated rate of bilirubin excretion.

An illustration shows the metabolism of bilirubin released by heme. Erythrocytes from the bone marrow, at 120 days, become aged, abnormal, or damaged. 1. Erythrocytes are attracted to the macrophages (M P S), where they are destructed as heme or globin. 2. Heme undergoes catabolism releasing free bilirubin by 250 milligrams per day. 3. Uptake of complex. 4. Conjunction (becomes water soluble). 5. Excretion. 6. Enterohepatic circulation of urobilinogen. 7. Urobilinogen is excreted as stool. Conjunction also triggers the kidney, supporting the renal excretion of urobilinogen of 4 milligrams per day.

Iron cycle

Approximately 67% of total body iron is bound to heme in erythrocytes (Hb), and about 5% to 10% is bound to heme-containing myoglobin in muscle cells. Approximately 30% is stored in mononuclear phagocytes (i.e., macrophages) and hepatic parenchymal cells as either ferritin (iron bound to protein) or hemosiderin (intracellular bound iron). The remaining 3% (less than 1 mg) is lost daily in urine, sweat, bile, sloughing of epithelial cells from the skin and intestinal mucosa, and minor bleeding, as occurs with menstruation in women. There is no excretory mechanism for iron. Unbound, or free, iron is toxic to human cells. Approximately 25 mg of iron is required daily for erythropoiesis; only 1 to 2 mg of iron is dietary, and the remainder is obtained from continual recycling of iron from erythrocytes through a process known as the iron cycle.12 Pregnant women require more iron to meet the needs of the placenta and growing fetus.

The methemoglobin released from the breakdown of senescent or damaged erythrocytes is dissociated by the enzyme heme oxygenase, and the iron is released into the bloodstream, where it binds again to transferrin or is stored in the macrophage’s cytoplasm as ferritin or hemosiderin (Fig. 28.15). A minute amount of iron is stored in muscle cells by the heme-containing protein myoglobin. Unavailable stores of iron are present in cytochromes, catalases, and peroxidase enzymes.

An illustration shows the iron cycle. Erythrocytes from the bone marrow enter the blood stream and tend to become aged, abnormal, or damaged (iron released from hemoglobin). These erythrocytes are consumed by macrophages (M P S), where the hemoglobin is destructed as heme or globin. The heme destructs as bilirubin, which is stored in the liver or secreted with bile, or as iron, which is released directly. Dietary iron absorbed from the intestine is: • Stored in the liver or released into the bone marrow. • Stored in the spleen or released into the bone marrow. • Iron plus transferrin (transports iron in plasma). • Iron reused in the synthesis of new hemoglobin in the bone marrow.

Ferritin is the major intracellular iron storage protein. Apoferritin is ferritin without attached iron and can store thousands of atoms of iron. Apoferritin binds to free Fe²⁺ and stores it as Fe³⁺, becoming ferritin. Excess accumulations of iron produce large intracellular iron storage complexes, known as hemosiderin. Hemosiderin is visible as an iron-based pigment under a light microscope as cell inclusions. The iron within deposits of hemosiderin is poorly available because it is not water-soluble, but it is available when iron requirements increase. The most common cause of hemosiderin deposition is simple bruising. Hemosiderin in small amounts within iron-rich tissues (i.e., spleen, liver, bone marrow) is considered normal. Large aggregates or its presence in tissue, such as the lungs or subcutaneous tissue, suggests a pathologic condition known as hemosiderosis.

Iron from either dietary sources, release of iron stores, or erythrocyte catabolism is transported in the blood bound to apotransferrin, thus becoming transferrin. Under normal conditions, only one-third of the iron-binding sites on transferrin molecules are occupied. Apotransferrin is a glycoprotein synthesized primarily by hepatocytes in the liver but also produced in small quantities by tissue macrophages, submaxillary and mammary glands, and ovaries or testes. Iron for hemoglobin production is carried by transferrin to the bone marrow, where it binds to transferrin receptors on erythroblasts. Transferrin receptors are on the plasma membrane of all nucleated cells, but at particularly high levels on erythroid precursors and rapidly proliferating cells (e.g., lymphocytes), and are thought to be the only route of cellular entry for transferrin-attached iron. Transferrin is recycled (transferrin cycle) by intracellular dissociation of iron from transferrin and secretion of the resultant apotransferrin to the bloodstream. The iron is transported to the erythroblast’s mitochondria (the site of Hb production), where the enzyme heme synthetase inserts Fe²⁺ into protoporphyrin to form heme. Heme then is bound to globin to form Hb. Iron not used in erythropoiesis is stored temporarily as ferritin or hemosiderin and later excreted or recycled.

Splenic red pulp macrophages are specialized for iron recycling with increased expression of proteins for the uptake of hemoglobin, the breakdown of heme, and the export of iron.13 The body’s iron homeostasis is primarily controlled by the hormone hepcidin. Hepcidin is the regulator of the systemic transportation of iron.¹⁴ It is a protein synthesized in the liver and released into the plasma, where it is bound with high affinity to α₂-macroglobulin and with less affinity to albumin. Hepatocellular hepcidin production is regulated physiologically by the levels of iron in the body, rate of erythropoiesis, and percentage of oxygen saturation. Hepatocytes (liver cells) sense levels of circulating iron by means of receptors for transferrin. Excess iron is stored in hepatocytes and macrophages, and hepatocytes sense these levels by means of receptors for bone morphogenetic protein (BMP), most likely BMP-6, which is a growth factor produced to a large extent by bone marrow sinusoid endothelial cells. Hepcidin production also can be induced by inflammation through IL-6 and other inflammatory cytokines, produced in response to infection or chronic conditions such as cancer.

Hepcidin regulates iron levels through its binding capacity to ferroportin, which is a transmembrane iron exporter found in the plasma membrane of cells that transport or store iron, including macrophages, hepatocytes, enterocytes (intestinal cells), and erythrocytes. The body’s total iron balance is maintained through controlled absorption rather than excretion. Dietary iron (primarily as Fe²⁺) is transported directly across the membranes of enterocytes in the duodenum and proximal jejunum and is carried in the plasma by transferrin. (Transport mechanisms are described in Chapter 1.) Hepcidin induces internalization and degradation of ferroportin, thus leading to increased intracellular iron stores, decreased dietary iron absorption, and decreased levels of circulating iron. Decreased production of hepcidin leads to release of stored iron and increased dietary absorption. Thus, if the body’s iron stores are low or the demand for erythropoiesis increases, dietary iron is transported rapidly through the epithelial cell and into the plasma. If body stores are high and erythropoiesis is not increased, iron transport is stopped. Hepcidin production also can be induced by inflammation. During inflammation, increased levels of hepcidin lead to sequestering of iron as a result of down-regulation of ferroportin expression on macrophages and enterocytes and thus can cause anemia.

Leukocytes consist of lymphocytes, granulocytes, and monocytes (their pathways of differentiation are shown in Fig. 28.10). Most of the leukocytes arise from stem cells in the bone marrow. HSCs differentiate into two populations of progenitor cells: common lymphoid progenitors and common myeloid progenitors. Lymphopoiesis (lymphocytopoiesis) is the generation of lymphocytes from lymphoid progenitor cells released into the bloodstream to undergo further maturation in the primary and secondary lymphoid organs (see Chapter 8).

Myelopoiesis is the development of granulocytes (neutrophils, eosinophils, and basophils) and monocytes from the differentiation of myeloid progenitor cells in the bone marrow. The common myeloid progenitors differentiate into progenitors for basophils, mast cells, eosinophils, and megakaryocytes, and into granulocyte/monocyte progenitors. The granulocyte/monocyte progenitors further differentiate into monocyte progenitors and granulocyte progenitors, which develop into monocytes/macrophages and neutrophils, respectively. Development from HSCs to common granulocyte-monocyte progenitors primarily is under the control of SCF, IL-3, and GM-CSF, whereas further differentiation into granulocytic and monocytic progenitors is controlled by granulocyte colony stimulating factor (G-CSF) and macrophage stimulating factor (M-CSF), respectively (see Table 28.4 and Fig. 28.10).

Monocytic progenitors undergo development into monocytes within 24 hours and are released into the circulation. Monocytes mature into various forms of macrophages, a process that is usually completed within 1 or 2 days after their release.

Progenitor cells for granulocytes normally fully mature in the bone marrow into neutrophils, eosinophils, and basophils. The ultimate phenotype is determined by relative local bone marrow concentrations of early- and late-acting cytokines, including GM-CSF, G-CSF, IL-3, IL-5, SCF, and others (see Table 28.4). Granulocytes are released into the blood within 10 to 14 days of development. The bone marrow selectively retains immature granulocytes as a reserve pool that can be rapidly mobilized in response to the body’s needs.

Most leukocytes exist in the body from days to years, depending on type (e.g., mature neutrophils live for hours and some macrophages live for years). Maintenance of optimal levels of granulocytes and monocytes in the blood depends on the availability of pluripotent stem cells in the marrow, induction of these into committed stem cells, timely release of new cells from the marrow, and mobilization of the granulocyte reserve pool. Leukocyte production increases in response to infection, to the presence of steroids, and to reduction or depletion of reserves in the marrow. It is also associated with strenuous exercise, convulsive seizures, heat, intense radiation, paroxysmal tachycardias, pain, nausea and vomiting, and anxiety.

Thrombopoiesis is the development of platelets. Platelets (thrombocytes) are derived from stem cells and progenitor cells that differentiate into megakaryocytes (see Fig. 28.10). During thrombopoiesis, the megakaryocyte nucleus enlarges and becomes extremely polyploidy (up to 100-fold or more of the normal amount of DNA) without cellular division. Concurrently, the numbers of cytoplasmic organelles (e.g., internal membranes, granules) increase and the cell develops cellular surface elongations and branches that progressively fragment into platelets. A single large (up to 100 μm) megakaryocyte may produce thousands of smaller platelets (2 to 3 μm). Like erythrocytes, platelets released from the bone marrow lack nuclei but contain granules that promote their stickiness. About two-thirds of platelets enter the circulation, and the remainder resides in the splenic pool.

Thrombopoietin (TPO), a hormone growth factor, stimulates the production and differentiation of megakaryocytes and is the main regulator of the circulating platelet numbers.¹⁵ TPO is primarily produced by the liver and induces platelet production in the bone marrow. Platelets express receptors for TPO and, when circulating platelet levels are normal, TPO is adsorbed onto the platelet surface and prevented from accessing the bone marrow and initiating further platelet production. When platelet levels are low, however, the amount of TPO exceeds the number of available platelet TPO receptors, and free TPO can enter the bone marrow. During inflammation IL-6 induces increased production of TPO, which increases production of newly formed platelets, which are more thrombogenic.

Platelets circulate in the bloodstream for about 8 to 10 days before beginning to lose their ability to perform thrombogenic activity and biochemical reactions. Senescent platelets are sequestered and destroyed in the spleen by macrophage phagocytosis.

Normally platelets are generally observed “rolling” along the margins of vessels. At sites of vessel injury, however, platelets become adherent to the site of endothelial damage, where the subendothelial matrix is exposed and endothelial cells have released vWF and decreased their antithrombotic activities (Fig. 28.17). Platelet adhesion is mostly mediated by the binding of platelet surface receptor glycoprotein Ib (GPIb) to vWF. The vWF protein is found in the subendothelial matrix and is released by endothelial cells and platelets. Platelet adhesion narrows the diameter of the blood vessel, resulting in increasing shear forces that could strip platelets off the vessel surface. However, those same forces induce conformational changes in the vWF molecule that result in increased affinity with the adhesive GPIb, thus stabilizing the adherent platelet.

A series of illustrations show blood vessel damage, platelet activation, blood clot, and clot dissolution. Subendothelial exposure. The illustration shows collagen, P G I sub 2, platelets, and endothelial sloughing. • Occurs after endothelial sloughing. • Platelets begin to fill endothelial gaps. • Promoted by thromboxane A sub 2 (T X A sub 2). • Inhibited by prostacyclin I sub 2 (P G I sub 2). • Platelet function depends on many factors, especially calcium. Adhesion. The illustration shows collagen and sticky platelets. An accompanying magnified view identifies a G P 1 b with G P 1 a or 2 a and G P 2 b or 3 a on the endothelium and collagen with v W F attached to fibrinogen. Adhesion is initiated by loss of endothelial cells (or rupture or erosion of atherosclerotic plaque), which exposes adhesive glycoproteins such as collagen and von Willebrand factor (v W F) in the subendothelium. v W F and, perhaps, other adhesive glycoproteins in the plasma deposit on the damaged area. Platelets adhere to the subendothelium through receptors that bind to the adhesive glycoproteins (G P 1 b, G P 1 a or 2 a, G P 2 b or 3 a). Activation. The illustration shows platelets concentrated at the collagen. • After platelets adhere they undergo an activation process that leads to a conformational change in G P 2 b or 3 a receptors, resulting in their ability to bind adhesive proteins, including fibrinogen and v W F. • Changes in platelet shape. • Formation of pseudopods. • Activation of arachidonic pathway. Aggregation. The illustration shows platelets attached to the R B C. Thrombin converted to fibrin, which forms a mesh on the platelets. • Induced by release of T X A sub 2. • Adhesive glycoproteins bind simultaneously to G P 2 b or 3 a on two different platelets. • Stabilization of the platelet plug (blood clot) occurs by activation of coagulation factors, thrombin, and fibrin. • Heparin neutralizing factor enhances clot formation. Platelet plug formation. The illustration shows a fibrin mesh covering a cluster of platelets attached to R B Cs. RBCs and platelets enmeshed in fibrin. Clot retraction and clot dissolution. The illustration shows a thick fibrin mesh. Thrombin activates protein and triggers the mesh generating plasminogen activators that produce plasmin for fibrin degradation. • Clot retraction, using large number of platelets, joins the edges of the injured vessel. • Clot dissolution is regulated by thrombin and plasminogen activators.

Platelet adhesion is also facilitated by other interactions between platelet receptors and exposed molecules of the subendothelial matrix. For instance, adhesion is increased through binding of the platelet collagen receptors GPVI and integrin α₂β₁ to exposed collagen in the matrix.

As a result of interactions with the endothelium or the subendothelial matrix, as well as exposure to inflammatory mediators produced by the endothelium and other cells, the platelets are activated. Activation results in reorganization of the platelet cytoskeleton, leading to dynamic changes in platelet shape from smooth spheres to those with spiny projections (increases surface area) (see Fig. 28.5) and degranulation (also called the platelet-release reaction) and resulting in the release of various potent biochemicals.

Platelets contain three types of granules: dense bodies, alpha granules, and lysosomes. The contents of the dense bodies and alpha granules are particularly important in hemostasis. The dense bodies contain ADP, serotonin, and calcium. ADP recruits and activates other platelets through specific receptors. During activation the platelet plasma membrane experiences several important changes, including becoming ruffled and sticky; undergoing cellular spreading to make tight contacts between neighboring platelets, causing the platelet plug to seal the injured endothelium; and externalizing the phospholipid phosphatidylserine, which provides a matrix for activation of clotting factors. Serotonin is a vasoactive amine that functions like histamine and increases vasodilation and vascular permeability. Calcium is necessary for many of the adhesive interactions as well as for intracellular signaling mechanisms that control platelet activation.

Alpha granules contain a mixture of clotting factors (e.g., fibrinogen), growth and angiogenic factors (e.g., platelet-derived growth factor [PDGF], vascular endothelial growth factor [VEGF], basic fibroblast growth factor), and angiogenesis inhibitors (e.g., platelet factor 4, thrombospondin, inhibitors of metalloproteinases). Platelet factor 4 also is a heparin-binding protein and enhances clot formation at the site of injury. Depending on the particular stimulus, platelets may selectively release promoters or inhibitors of angiogenesis. Many of these mediators either promote or inhibit platelet activity and the eventual process of clot formation. PDGF stimulates smooth muscle cells and promotes tissue repair.

Platelet lysosomes have a role in the digestion of phagocytic and cytosolic components, similar to that in nucleated cells. Lysosomal contents also may have important extracellular functions, such as supporting receptor cleavage, fibrinolysis and degradation of extracellular matrix components, and remodeling of the vasculature.¹⁸

Platelets also initiate production of the prostaglandin derivative thromboxane A2 (TXA2), which counters the effects of PGI₂ produced by endothelial cells. TXA₂ causes vasoconstriction and promotes the degranulation of platelets, increases expression of platelet fibrinogen receptors, and stimulates platelet aggregation, whereas PGI₂ promotes vasodilation and inhibits platelet degranulation. Arachidonic acid metabolism by cyclooxygenase (COX) is a major pathway for blood platelet activation. An isoform of cyclooxygenase (COX-1) converts arachidonic acid to TXA₂ in platelets. Aspirin at low doses specifically and irreversibly inactivates COX-1, decreasing production of TXA₂ and decreasing platelet activation and aggregation.¹⁹ Daily intake of low doses of aspirin leads to more than 95% inhibition of TXA₂ in just a few days.

Other stimuli of platelet activation include epinephrine, thrombin, and collagen. Thrombin and collagen are particularly strong stimuli. Thrombin cleaves the extracellular domain of G-protein–coupled protease-activated receptors (PARs), thereby initiating transmembrane signaling.

If blood vessel injury is minor, hemostasis is achieved temporarily by formation of the platelet plug within 3 to 5 minutes of injury. Platelet plugs adhere to damaged vessel walls and seal the many minute ruptures that occur daily in the microcirculation, particularly in capillaries. With too few platelets, numerous small hemorrhagic areas called purpuras develop under the skin and throughout the tissues (see Chapter 29). If primary hemostasis is inadequate to prevent further bleeding, the process proceeds through secondary hemostasis to create a larger complex of more tightly interactive platelets within a matrix created by activation of the clotting system.