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Blood and Hemopoiesis
Blood, a specialized connective tissue, is a bright- to dark-red, viscous, slightly alkaline fluid (pH 7.4) that accounts for approximately 7% of the total body weight with a total volume of approximately 5 L in an average adult. It is composed of formed elements— red blood cells (RBCs; erythrocytes) , white blood cells (WBCs; leukocytes) , and platelets —suspended in a fluid component (the extracellular matrix), known as plasma ( Figs. 10.1 and 10.2 ).
Photomicrograph of circulating blood. Note the abundance of erythrocytes, as well as the three leukocytes. Also, observe the presence of numerous platelets that appear as small dots interspersed among the erythrocytes. (×270)
Diagram of cells and platelets of circulating blood.
Because blood circulates throughout the body within the confines of the circulatory system, it is an ideal vehicle for the transport of materials. The primary functions of blood include conveying nutrients from the gastrointestinal system to all of the cells of the body and, subsequently, delivering the waste products of these cells to specific organs for elimination. Numerous other metabolites, cellular products (e.g., hormones and other signaling molecules), and electrolytes are also ferried by the bloodstream to their final destinations. Oxygen (O 2 ) is carried by hemoglobin within erythrocytes from the lungs for distribution to the cells of the organism and carbon dioxide (CO 2 ) is conveyed both by hemoglobin and by the fluid component of plasma (as bicarbonate ion, HCO 3 − , as well as in its free form) for elimination by the lungs.
Blood also helps regulate body temperature and aids in maintaining the acid–base and osmotic balance of the body fluids. Finally, blood acts as a pathway for migration of WBCs between various connective tissue compartments of the body.
The fluid state of blood necessitates the presence of a protective mechanism, coagulation ( clotting ), to stop its flow in case of damage to the vascular tree. The process of coagulation is mediated by platelets and blood-borne factors that transform blood from a liquid to a gel state.
When blood is removed from the body and placed in a test tube, clotting occurs unless the tube is coated by an anticoagulant such as heparin. Upon centrifugation of liquid blood, the formed elements settle to the bottom of the tube as a red precipitate (44%) covered by a thin translucent layer, the buffy coat (1%), and the fluid plasma remains on top as the supernatant (55%). The red precipitate is composed of RBCs, and the total RBC volume is known as the hematocrit , whereas the buffy coat consists of WBCs and platelets.
The finite life span of blood cells requires their constant renewal to maintain a steady circulating population. This process of blood cell formation from established blood cell precursors is called hemopoiesis (also referred to as hematopoiesis ).
Blood
Blood is composed of a fluid component (plasma) and formed elements, consisting of the various types of blood cells, as well as platelets.
Light microscopy identifying circulating blood cells relies on either the Wright or Giemsa modifications of the original Romanovsky technique of using a mixture of methylene blue and eosin. Methylene blue stains acidic cellular components blue, and eosin stains alkaline components pink. Still other components are colored a reddish blue by binding to azures , substances formed when methylene blue is oxidized.
Plasma
Plasma is a yellowish fluid in which cells, platelets, organic compounds, and electrolytes are suspended and/or dissolved.
During coagulation, some of the organic and inorganic components of plasma become integrated into the clot. The remaining straw-colored fluid, which no longer has the clot-forming components dissolved or suspended in it, is known as serum .
Plasma is composed of approximately 90% water, 9% protein, and 1% inorganic salts, ions, nitrogenous compounds, nutrients, and gases. The types, origins, and functions of the blood proteins are listed in Table 10.1 . Capillaries and small venules are leaky and allow plasma to enter the connective tissue spaces, where it is known as extracellular fluid , whose composition of electrolytes and small molecules is similar to that in plasma. However, the concentration of proteins in extracellular fluid is much lower than that in plasma because not even small proteins, such as albumin, can escape the endothelial lining of a capillary or that of a venule.
Clinical Correlations
Albumin , one of the proteins in plasma, is chiefly responsible for the establishment of blood’s colloid osmotic pressure , the force that maintains normal blood volume by opposing the movement of fluid from the capillaries and venules into the interstitial spaces.
TABLE 10.1
Proteins of Plasma
| Protein | Size | Source | Function |
|---|---|---|---|
| Albumin | 60,000–69,000 Da | Liver | Maintains colloid osmotic pressure and transports certain insoluble metabolites |
| Globulins α - and β -Globulins |
80,000–1 × 10 6 Da | Liver | Transport metal ions, protein-bound lipids, and lipid-soluble vitamins |
| γ -Globulin | Plasma cells | Antibodies of immune defense | |
| Clotting Proteins | |||
| (e.g., prothrombin, fibrinogen, accelerator globulin) | Varied | Liver | Formation of fibrin threads |
| Complement proteins C1 through C9 |
Varied | Liver | Destruction of microorganisms and initiation of inflammation |
| Plasma Lipoproteins | |||
| Chylomicrons | 100–500 μm | Intestinal epithelial cells | Transport of triglycerides to liver |
| Very low-density lipoprotein (VLDL) | 25–70 nm | Liver | Transport of triglycerides from liver to body cells |
| Low-density lipoprotein (LDL) | 3 × 10 6 Da | Liver | Transport of cholesterol from liver to body cells |
Formed Elements
RBCs, WBCs, and platelets constitute the formed elements of blood.
Erythrocytes
Erythrocytes (RBCs) are the smallest and most numerous cells of blood; they have no nuclei and are responsible for the transport of oxygen and carbon dioxide to and from the tissues of the body.
Each erythrocyte ( RBC ) resembles a biconcave-shaped disk that is 7.5 μm in diameter, 2.0 μm thick at its widest region, and less than 1 μm thick at its center. When stained with Giemsa or Wright stains, erythrocytes display a salmon-pink color ( Figs. 10.3 and 10.4 ). This shape provides the cell with a large surface area relative to its volume, enhancing its capability for gaseous exchange. Although precursor cells of the erythrocyte possess nuclei and organelles during their development and maturation, they expel them prior to entering the circulation. Thus, mature, circulating erythrocytes have neither nuclei nor organelles, but they do have soluble enzymes in their cytosol. Within the erythrocyte, the enzyme carbonic anhydrase facilitates the formation of carbonic acid from CO 2 and water. This acid dissociates to form bicarbonate (HCO 3 − ) and hydrogen (H + ). It is as bicarbonate that most of the CO 2 is ferried to the lungs for exhalation. The ability of bicarbonate to cross the erythrocyte cell membrane is mediated by the integral membrane protein band 3 , a coupled anion transporter that exchanges intracellular bicarbonate for extracellular Cl − ; this exchange is known as the chloride shift . Additional enzymes include those of the glycolytic pathway ( Embden-Meyerhoff pathway ) as well as enzymes that are responsible for the pentose monophosphate shunt ( hexose monophosphate shunt ) for the production of the high-energy molecule reduced nicotinamide adenine dinucleotide phosphate (NADPH) , a reducing agent. The pentose monophosphate shunt does not require the presence of oxygen and is the chief method whereby the erythrocyte produces adenosine triphosphate (ATP), necessary for its energy requirement.
Clinical Correlations
- 1.
During its 120-day life, each erythrocyte negotiates the entire circulatory system at least 100,000 times and, therefore, must pass through innumerable capillaries whose lumen is smaller than the cell’s diameter. To navigate through such small-bore vessels, the erythrocyte undergoes deformations of its shape and becomes subject to tremendous shear forces. It is the erythrocyte cell membrane and the underlying cytoskeleton that contribute to the ability of the RBC to maintain its structural and functional integrity. As erythrocytes reach their 120-day life span, they become fragile and display on their surface a group of oligosaccharides that act as signals for macrophages of the liver, bone marrow, and spleen to destroy these old RBCs.
- 2.
Males have more erythrocytes per unit volume of blood than do females (5 × 10 6 vs. 4.5 × 10 6 per mm 3 ), and members of both sexes living at higher altitudes have correspondingly more RBCs than residents living at lower altitudes.
Hemoglobin
Hemoglobin is a large protein composed of four polypeptide chains, each of which is covalently bound to a heme group.
Erythrocytes are packed with hemoglobin , a large tetrameric protein (68,000 Da) composed of four polypeptide chains, each of which is covalently bound to an iron-containing heme . The iron is protected from oxidation by the globin chain even though oxygen can bind to it. The globin moiety of hemoglobin releases CO 2 in regions of high oxygen concentration, such as in the lungs, and O 2 binds to the iron of each heme. However, in oxygen-poor regions, as in body tissues, hemoglobin releases O 2 and binds CO 2 . This property of hemoglobin makes it ideal for the conveyance of respiratory gases. Hemoglobin carrying O 2 is known as oxyhemoglobin , and hemoglobin carrying CO 2 is called carbaminohemoglobin (or carbamylhemoglobin ).
Clinical Correlations
- 1.
Hypoxic tissues release 2,3-diphosphoglyceride, a carbohydrate that facilitates the release of oxygen from the erythrocyte. Hemoglobin also binds NO, a neurotransmitter substance that causes dilation of blood vessels, permitting RBCs to release more oxygen and pick up more CO 2 within the tissues of the body.
- 2.
Carbon monoxide ( CO ) has a much greater affinity, about 250-fold, than O 2 for the heme portion of hemoglobin, and when CO binds to the iron of the heme, the hemoglobin molecule is transformed to its (R-) Hb form and increases its affinity to oxygen so that it cannot be released to the tissues, even in hypoxic regions. People who are trapped in areas of poor ventilation with a running gasoline-powered engine or in a building on fire frequently succumb to CO poisoning. Many such victims, especially those who are fair-skinned, instead of being cyanotic (with a bluish pallor) present with healthy-looking, cherry-red skin because of the color of the CO-hemoglobin complex (carbon monoxyhemoglobin) even though they are dead.
On the basis of the amino acid sequences, there are four normal human polypeptide chains of hemoglobin, designated α , β , γ , and δ . The principal hemoglobin of the fetus, fetal hemoglobin ( HbF ), composed of two alpha chains and two gamma chains, is replaced shortly after birth by adult hemoglobin ( HbA ). There are two types of normal HbAs, HbA1 ( α 2 β 2 ) and the much rarer form, HbA2 ( α 2 δ 2 ). In an adult, approximately 96% of the hemoglobin is HbA 1 , 2% is HbA 2 , and the remaining 2% is HbF.
Clinical Correlations
- 1.
Several hereditary diseases result from defects in the genes encoding the hemoglobin polypeptide chains. Diseases referred to as thalassemia are marked by decreased synthesis of one or more hemoglobin chains. In β -thalassemia, synthesis of the β -chains is impaired. In the homozygous form of this disease, which is most prevalent among persons of Mediterranean descent, HbA is missing and high levels of HbF persist after birth. In the past couple of years, gene therapy has demonstrated great success in treating this condition. Inactivated lentivirus was used to infect the patients’ own immature stem cells retrieved from their bone marrow with the normal globin gene in vitro. Then, the patients underwent chemotherapy to destroy their mutated stem cells, and the genetically modified stem cells were introduced. The modified stem cells migrated to the bone marrow and started producing normal hemoglobin. The clinical trial consisted of 22 patients, nine of whom had a severe case of thalassemia. Of these patients, three no longer required transfusions and six required 74% fewer transfusions than prior to treatment. The remaining 13 patients who had milder cases of thalassemia no longer required transfusions. Clinical trials are currently progressing to verify and extend the results.
- 2.
Sickle cell anemia is the result of a point mutation at a single locus of the β -chain (valine is incorporated into the sequence instead of glutamate), forming the abnormal hemoglobin HbS. When the oxygen tension is reduced (e.g., during strenuous exercise), HbS changes shape, producing abnormal-shaped (crescent-shaped) erythrocytes that are less pliant, more fragile, and more prone to hemolysis than normal cells. Sickle cell anemia is prevalent in the black population, especially in those whose ancestors lived in regions of Africa where malaria is endemic. In the United States, about 1 of 600 newborn African-American babies is stricken with this condition. Individuals with sickle cell anemia are more resistant to malaria , a disease caused by a parasite, than people whose hemoglobin is not mutated. It was believed that sickle-shaped erythrocytes were more resistant to the entry of the parasite Plasmodium falciparum , the most fatal of the five Plasmodium species that cause malaria. However, it has been reported recently that the erythrocytes of individuals with sickle cell anemia manufacture an increased amount of the enzyme heme oxygenase-1. This enzyme produces CO, a gas that not only protects hemoglobin from degradation, but also limits the toxic ability of P. falciparum , thus providing more time for the individual’s immune system to fight the parasitic invasion.
- 3.
Hemoglobin A1c ( HbA1c or A1c ) is a glycated hemoglobin that forms when plasma glucose levels are elevated and glucose molecules attach to the N-terminus of hemoglobin’s beta chain. This is a nonreversible reaction, so, once attached, the hemoglobin molecules do not lose their glucose. If the plasma glucose levels remain elevated, glycated hemoglobin levels also increase; therefore, a measure of A1c levels is indicative of the blood sugar concentrations for the previous 2 to 3 months. The normal glycated hemoglobin level in adults is approximately 4% to 5.6%. Individuals with diabetes have higher A1c levels because their blood sugar concentrations are usually greater than those of individuals without diabetes. For a patient with diabetes, an A1c level below 7% is considered to be very good, indicating a good control of blood glucose levels. A1c levels higher than 7% is worrisome because the higher the level, the greater the probability of acquiring diabetes-related conditions.
Erythrocyte Cell Membrane
The cell membrane of the erythrocyte and the underlying cytoskeleton are highly pliable and can withstand great shear forces.
The RBC plasma membrane, a typical lipid bilayer, is composed of about 50% protein, 40% lipids, and 10% carbohydrates. Most of the proteins are transmembrane proteins, principally glycophorin A (as well as lesser quantities of glycophorins B, C, and D), ion channels (calcium-dependent potassium channels and Na + -K + ATP), and the anion transporter band 3 protein , which transports Cl − and HCO − 3 . It also acts as an anchoring site for ankyrin , band 4.1 protein, hemoglobin, and glycolytic enzymes ( Fig. 10.5 ). The RBC membrane also possesses the peripheral proteins: spectrin, ankyrin, band 4.1 protein, and actin. Band 4 . 1 protein acts as an anchoring site for spectrin, band 3 protein, and glycophorins. Thus, ankyrin, band 3 protein, and band 4.1 protein anchor the cytoskeleton—a hexagonal lattice composed chiefly of spectrin tetramers , actin , and adducin —to the cytoplasmic aspect of the RBC plasmalemma (see Chapter 2 ). This subplasmalemmal cytoskeleton helps maintain the biconcave disk shape of the erythrocyte.
Clinical Correlations
- 1.
Defects in the cytoskeletal components of erythrocytes result in various conditions marked by abnormally shaped cells. Hereditary spherocytosis , for instance, is caused by synthesis of an abnormal spectrin, band 3 protein, and protein 4.2. RBCs of patients with this condition are more fragile and transport less oxygen compared with normal erythrocytes. Moreover, these spherocytes are preferentially destroyed in the spleen, leading to anemia.
- 2.
Deficiency of glycophorin C is responsible for elliptocytic RBCs without the resultant hemolytic anemia. These cells are unstable and fragile and are less capable of deformation than normal erythrocytes.
The extracellular surface of the RBC plasmalemma has specific inherited carbohydrate chains that act as antigens and determine the blood group of an individual for the purposes of blood transfusion. The most notable of these are the A , B , and H antigens , which determine the four primary blood groups, A , B , AB , and O , in which H antigen determines the O blood type ( Table 10.2 and Fig. 10.6 ). People who lack either the A or B antigen, or both, have antibodies against the missing antigen in their blood; if they undergo transfusion with blood containing the missing antigen, the donor erythrocytes are attacked by the recipien’s serum antibodies and are eventually lysed. Because type O blood has neither A nor B antigen, everyone can receive type O blood, making type O people “universal donors,” and because type AB individuals have no antibodies against A, B, or H antigens, they can accept blood from anyone, therefore, they are “universal acceptors.”
Clinical Correlations
- 1.
It is interesting to note that the carbohydrate chains of the A and B blood groups are identical, with the exception of their terminal sugar molecules and that people possessing the O blood group also have the same carbohydrate chains but do not have the terminal sugar molecule present in the A or B blood types (i.e., they have one fewer member in their carbohydrate group than A or B antigens). The addition of the terminal sugar group of A and B antigens each requires a specific enzyme, and neither of these two enzymes is present in individuals with the O blood group. For the type A blood group, the enzyme A-transferase (N-acetylgalactosamine transferase) adds a terminal N-acetylgalactosamine to the type H antigen. In the type B blood group, the enzyme terminal B-transferase (galactose transferase) adds a terminal galactose to the type H antigen. In blood type AB, both enzymes are present and form both type A and type B. In blood type O, neither enzyme is present; therefore, neither galactose nor N-acetylgalactosamine is added to the H antigen.
- 2.
Recent reports have demonstrated that individuals with the AB blood group have a greater prevalence of impaired cognitive function related to aging than individuals possessing other blood groups. The reason for this occurrence is not as yet understood.
- 3.
In an experiment, a group of volunteers drank water containing E. coli isolated from a patient with diarrhea. Of the volunteers whose blood type was A or AB, 81% had diarrhea whereas only 50% of type O and Type B volunteers had diarrhea. These blood type antigens are present also on the cells lining the intestines. Apparently, the E. coli are more likely to attach to the type A antigen than to type H or type B antigens. Once attached, they release their toxin, which causes diarrhea.
TABLE 10.2
ABO Blood Group System
| Blood Group | Antigens Present | Miscellaneous |
|---|---|---|
| A | Antigen A | |
| B | Antigen B | |
| AB | Antigens A and B | Universal acceptor |
| O | H antigen only but neither antigen A nor B | Universal donor |
Another important blood group, the Rh group, is so named because it was first identified in rhesus monkeys. This complex group comprises almost 50 antigens, although many are relatively rare. One of the Rh antigens, D antigen ( Rh factor ), is so common in the human population that the erythrocytes of 85% of Americans have D antigen on their surface; these individuals are thus said to be Rh positive ( Rh+ ). The remaining 15% of the population does not have the antigen and is said to be Rh negative ( Rh− ).
Clinical Correlations
When an Rh − pregnant woman delivers her first Rh + baby, enough of the baby’s blood is likely to enter her circulation to induce the formation of anti-Rh antibodies in the mother. Because the first antibody to be produced is immunoglobulin M (IgM), it is too large to be able to cross the placental barrier (see Chapter 12 on the lymphoid [immune] system). During a subsequent pregnancy with an Rh + fetus, the new fetus’s RBCs enter the mother’s bloodstream. This second exposure elicits the formation of IgG antibodies, which are smaller and are able to cross the placental barrier. These antibodies attack the erythrocytes of the fetus, causing erythroblastosis fetalis, a condition that may be fatal to the newborn. Prenatal and postnatal transfusions of the fetus are necessary to prevent brain damage and death of the newborn unless the mother has been treated with anti-D globulin (RhoGAM) before or shortly after the birth of the first Rh + baby. The anti-D globulin complexes with D antigen, preventing the mother’s immune system from recognizing it as an antigenic molecule. Therefore, the mother’s immune system does not produce antibodies that would otherwise attack the fetus’s erythrocytes, preventing the occurrence of erythroblastosis fetalis.
Leukocytes
Leukocytes are white blood cells that are classified into two major categories: granulocytes and agranulocytes.
The number of leukocytes ( WBCs ) is much smaller than that of RBCs. In fact, in a healthy adult, there are only 6500 to 10,000 WBCs per mm 3 of blood. Unlike erythrocytes, leukocytes do not function within the bloodstream but rather use it as a means of traveling from one region of the body to another. When leukocytes reach their destination, they leave the bloodstream by migrating between the endothelial cells of the blood vessels ( diapedesis ), enter the connective tissue spaces, and perform their function. Within the bloodstream as well as in smears, leukocytes are round; in connective tissue, they are pleomorphic. They generally defend the body against foreign substances.
WBCs are classified into two groups:
-
Granulocytes, which have specific granules in their cytoplasm
-
Agranulocytes, which lack specific granules in their cytoplasm
Both granulocytes and agranulocytes possess nonspecific ( azurophilic ) granules, now known to be lysosomes .
There are three types of granulocytes, differentiated according to the color of their specific granules after application of Romanovsky-type stains: neutrophils, eosinophils, and basophils. There are two types of agranulocytes: lymphocytes and monocytes. The differential leukocyte count and various properties of the leukocytes are detailed in Table 10.3 .
TABLE 10.3
Leukocytes
| Granulocytes | Agranulocytes | ||||
|---|---|---|---|---|---|
| Features | Neutrophils | Eosinophils | Basophils | Lymphocytes | Monocytes |
| Number/mm 3 | 3500–7000 | 150–400 | 50–100 | 1500–2500 | 200–800 |
| % of WBCs | 60–70% | 2–4% | <1% | 20–25% | 3–8% |
| Diameter (μm) | |||||
| Section | 8–9 | 9–11 | 7–8 | 7–8 | 10–12 |
| Smear | 9–12 | 10–14 | 8–10 | 8–10 | 12–15 |
| Nucleus | 3–4 lobes | 2 lobes (sausage shaped) | S shaped | Round | Kidney shaped |
| Specific granules | 0.1 μm, light pink a | 1–1.5 μm, dark pink a | 0.5 μm, blue/black a | None | None |
| Contents of specific granules | Type IV collagenase, phospholipase A 2 , lactoferrin, lysozyme, phagocytin, alkaline phosphatase, vitamin B 12 –binding protein | Aryl sulfatase, histaminase, β -glucuronidase, acid phosphatase, phospholipase, major basic protein, eosinophil cationic protein, neurotoxin, ribonuclease, cathepsin, peroxidase | Histamine, heparin, eosinophil chemotactic factor, neutrophil chemotactic factor, peroxidase, neutral proteases, chondroitin sulfate | None | None |
| Surface markers | Fc receptors, platelet-activating factor receptor, leukotriene B 4 receptor, leukocyte cell adhesion molecule-1 | IgE receptors, eosinophil chemotactic factor receptor | IgE receptors | T cells: T-cell receptors, CD molecules, IL receptors B cells: Surface immunoglobulins |
Class II HLA, Fc receptors |
| Life span | < 1 wk | < 2 wk | 1–2 y (in murines) | A few months to several years | A few days in blood, several months in connective tissue |
| Function | Phagocytosis and destruction of bacteria | Phagocytosis of antigen–antibody complex; destruction of parasites | Similar to mast cells to mediate inflammatory responses | T cells: Cell-mediated immune response B cells: Humorally mediated immune response |
Differentiate into macrophage: phagocytosis, presentation of antigens |
CD, Cluster of differentiation; HLA, human leukocyte antigen; IgE, immunoglobulin E; IL, interleukin; WBC, white blood cell.
Neutrophils
Neutrophils compose most of the white blood cell population; they are avid phagocytes, destroying bacteria that invade connective tissue spaces.
Neutrophils ( polymorphonuclear leukocytes, polys ) are granulocytes and are the most numerous of the WBCs, constituting 60% to 70% of the total leukocyte population. In blood smears, neutrophils are 9 to 12 μm in diameter and have a multilobed nucleus (see Figs. 10.2 and 10.3 ). The lobes, joined by slender connections, increase in number as the cell ages. In females, the nucleus presents a characteristic small appendage, the “ drumstick ” ( Barr body or sex chromosome ), which contains the condensed, inactive second X chromosome; it is not evident in every cell. The neutrophil plasmalemma possesses complement receptors, as well as Fc receptors for IgG. They are among the first defensive cells to appear in acute bacterial infections.
Neutrophil Granules
Neutrophils possess specific, azurophilic, and tertiary granules.
Three types of granules are present in the cytoplasm of neutrophils: small specific granules (0.1 μm in diameter), larger azurophilic granules (0.5 μm in diameter), and tertiary granules.
Specific granules contain enzymes and pharmacological agents that aid the neutrophil in performing its antimicrobial functions (see Table 10.3 ). In electron micrographs, these granules appear to be somewhat oblong ( Fig. 10.7 ).
Azurophilic granules are lysosomes that house acid hydrolases, myeloperoxidase (MPO), the antibacterial agent lysozyme, bactericidal permeability–increasing protein, cathepsin G (an enzyme that may contribute to the destruction and degradation of pathogens phagocytosed by these cells), elastase, and nonspecific collagenase.
Tertiary granules contain gelatinase and cathepsins, as well as glycoproteins that are inserted into the plasmalemma.
Neutrophil Functions
Neutrophils phagocytose and destroy bacteria by using the contents of their various granules.
Neutrophils interact with chemotactic agents to migrate to sites invaded by microorganisms. They accomplish this by entering postcapillary venules in the region of inflammation and adhering to the various selectin molecules located on the luminal cell membranes of the endothelial cells of these vessels by use of their selectin receptors . The interaction between the neutrophil’s selectin receptors and the selectins of the endothelial cells causes the neutrophils to roll slowly along the vessel’s endothelial lining. As the neutrophils are slowing their migrations, interleukin-1 ( IL-1 ) and tumor necrosis factor ( TNF ) induce the endothelial cells to express intercellular adhesion molecule type 1 (ICAM-1) , to which the integrin molecules of neutrophils avidly bind.
When binding occurs, the neutrophils stop migrating in preparation for their passage through the endothelium of the postcapillary venule to enter the connective tissue compartment ( Fig. 10.8 ) . Once there, they destroy the microorganisms by phagocytosis and by the release of hydrolytic enzymes (and respiratory burst ). In addition, by manufacturing and releasing leukotrienes , neutrophils assist in the initiation of the inflammatory process. The sequence of events is as follows:
- 1.
The binding of neutrophil chemotactic factor (NCF), released by mast cells and basophils to NCF receptors of the neutrophil’s plasmalemma, facilitates the release of the contents of tertiary granules into the extracellular matrix.
- 2.
Gelatinase and cathepsins degrade the basal lamina, facilitating neutrophil migration. Glycoproteins present in the tertiary granules become inserted in the cell membrane and aid the process of phagocytosis.
- 3.
The contents of the specific granules are also released into the extracellular matrix, where they attack the invading microorganisms and aid neutrophil migration.
- 4.
Microorganisms, phagocytosed by neutrophils, become enclosed in phagosomes ( Fig. 10.9A,B ). Enzymes and pharmacological agents of the azurophilic granules are usually released into the lumina of these intracellular vesicles, where they destroy the ingested microorganisms. Because of their phagocytic functions, neutrophils are also known as microphages to distinguish them from the larger phagocytic cells, the macrophages.
- 5.
Bacteria are killed not only by the action of enzymes but also by the formation of reactive oxygen compounds within the phagosomes of neutrophils. These are superoxide (O 2 − ), formed by the action of NADPH oxidase on O 2 in a respiratory burst; hydrogen peroxide, formed by the action of superoxide dismutase on superoxide; and hypochlorous acid, formed by the interaction of MPO and chloride ions with hydrogen peroxide (see Figs. 10.9C,D ).
- 6.
Frequently, the contents of the azurophilic granules are released into the extracellular matrix, causing tissue damage, but usually catalase and glutathione peroxidase limit the tissue injury by degrading hydrogen peroxide.
- 7.
Once neutrophils perform their function of killing microorganisms, they also die, resulting in the formation of pus (the accumulation of dead leukocytes, bacteria, and extracellular fluid).
- 8.
Not only do neutrophils destroy bacteria, but they also synthesize leukotrienes from arachidonic acids in their cell membranes. These newly formed leukotrienes aid in the initiation of the inflammatory process.
Clinical Correlations
- 1.
Children with hereditary deficiency of NADPH oxidase are subject to persistent bacterial infections because their neutrophils cannot form a respiratory burst response to the bacterial challenge. Their neutrophils cannot generate superoxide, hydrogen peroxide, or hypochlorous acid during phagocytosis of bacteria.
- 2.
Individuals suffering from neutropenia, low levels of neutrophils in circulating blood, have problems fighting bacterial infections. This condition may be acute, lasting less than 3 months, or chronic, lasting more than 3 months. Neutropenia may be mild (1000–1500 neutrophils per mm 3 of blood), moderate (500–1000 neutrophils per mm 3 of blood), or severe (less than 500 neutrophils per mm 3 of blood). The causes of neutropenia may be decreased neutrophil production by the bone marrow or excess neutrophil destruction outside the bone marrow.
Eosinophils
Eosinophils phagocytose antigen–antibody complexes and kill parasitic invaders.
Eosinophils are granulocytes that constitute less than 4% of the total WBC population. They are round cells in suspension and in blood smear (10–14 μm in diameter) but they may be pleomorphic during their migration through connective tissue. They have a sausage-shaped, bilobed nucleus in which the two lobes are linked by a thin connecting strand (see Figs. 10.2 and 10.3 ) and their cell membrane has receptors for IgG, IgE, and complement. Electron micrographs display a small, centrally located Golgi apparatus, a limited amount of rough endoplasmic reticulum (RER), and only a few mitochondria that are usually located in the vicinity of the centrioles near the cytocenter. Eosinophils are produced in the bone marrow, and interleukin-5 ( IL-5 ) causes proliferation of their precursors and their differentiation into mature cells. In the absence of IL-5, basophils develop instead of eosinophils.
Eosinophil Granules
The specific granules of eosinophils possess an externum and an internum.
Eosinophils possess specific granules and azurophilic granules. Specific granules are oblong (1.0–1.5 μm in length, < 1.0 μm in width) and stain deep pink with Giemsa and Wright stains. Electron micrographs demonstrate that specific granules have a crystal-like, electron-dense center, the internum , surrounded by a less electron-dense externum ( Fig. 10.10 ). The internum houses major basic protein , eosinophilic cationic protein , and eosinophil-derived neurotoxin , the first two of which are highly efficacious agents in combating parasites. The externum houses the enzymes listed in Table 10.3 .
The nonspecific azurophilic granules are lysosomes (0.5 μm in diameter) containing hydrolytic enzymes similar to those found in neutrophils. These enzymes function both in the destruction of parasitic worms and in the hydrolysis of antigen–antibody complexes internalized by eosinophils.
Eosinophil Functions
Eosinophils assist in the elimination of antibody–antigen complexes and in the destruction of parasitic worms.
The binding of histamine, leukotrienes, and eosinophil chemotactic factor (released by mast cells, basophils, and neutrophils) to eosinophil cell membrane receptors induces the migration of these cells to the site of allergic reaction, inflammatory reaction, or parasitic worm invasion. Eosinophils degranulate their major basic protein or eosinophil cationic protein on the surface of the parasitic worms, killing them by forming pores in their pellicles, thus facilitating access of agents such as superoxides and hydrogen peroxide to the parasite cell membrane and cytoplasm.
Eosinophils also release substances that inactivate the pharmacological initiators of the inflammatory response, such as histamine and leukotriene C. Additionally, they engulf antigen–antibody complexes that pass into the eosinophil’s endosomal compartment for eventual degradation. The ribonucleases in the azurophilic granules of eosinophils combat viral pathogens. Moreover, eosinophils participate in the degradation of fibrin.
Clinical Correlations
Connective tissue cells in the vicinity of antigen–antibody complexes release the pharmacological agents histamine and IL-5, causing increased formation and release of eosinophils from the bone marrow. In contrast, elevation of blood corticosteroid levels depresses the number of eosinophils in circulation.
Basophils
Basophils are similar to mast cells in function even though they originate from different precursors in the bone marrow.
Basophils are granulocytes that constitute less than 1% of the total leukocyte population. They are round cells in suspension but may be pleomorphic during migration through connective tissue. They are 8 to 10 μm in diameter (in blood smears) and have an S -shaped nucleus, which is commonly masked by the large specific granules present in the cytoplasm (see Figs. 10.2 and 10.3 ). Electron micrographs demonstrate the presence of the small Golgi apparatus, few mitochondria, extensive RER, and occasional glycogen deposits. Basophils have a number of surface receptors on their plasmalemma, including immunoglobulin E ( IgE ) receptors (FcεRI) . Basophils originate in the bone marrow but can be formed only in the absence of IL-5.
Basophil Granules
Basophils possess specific and azurophilic granules.
The specific granules of basophils stain dark blue to black with Giemsa and Wright stains. They are approximately 0.5 μm in diameter and frequently press against the periphery of the cell, creating the basophil’s characteristic “roughened” perimeter, as seen by light microscopy. These granules contain heparin, histamine, eosinophil chemotactic factor, neutrophil chemotactic factor, neutral proteases, chondroitin sulfate, and peroxidase (see Table 10.3 ). The nonspecific azurophilic granules are lysosomes, which contain enzymes similar to those of neutrophils.
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