Leyden crystals and have lysophospholipase activity. Finally, eosinophils produce and release lipid

mediators such as PAF, prostaglandins, and LTs, which probably contribute to the process of allergic

diseases.4,83,84

Basophils and Mast Cells

Basophils and mast cells are central to the development of allergic inflammation and produce cytokines,

lipid mediators, vasoactive amines, and proteases that can also participate in nonallergic inflammatory

responses (Table 7-1). Both cells share a common progenitor cell, which diverges early during

differentiation. The primary growth factors for basophil development are IL-3 and GM-CSF, whereas

stem cell factor is the primary stimulus for mast cell maturation.85 Although basophils and mast cells are

distinct cell types, they are often grouped together because of the similarity of their granule content,

the stimuli inducing activation, and their effector functions.39

Mature basophils constitute only 0.5% to 1% of the circulating leukocytes. Mast cells are primarily

tissue-fixed residents of connective tissue. Engagement of IgE-bound antigen with the high-affinity IgE

antibody receptor, FceR, provides the primary stimulus for basophil and mast cell activation and

degranulation. In addition, diverse agents such as contrast media, opiates, anaphylatoxins, chemokines,

and neuropeptides may serve as activators. Basophils and mast cells elaborate both preformed, as well

as, newly synthesized inflammatory mediators. The main basophil proteoglycan is chondroitin sulfate A,

whereas mast cells contain heparin, chondroitin sulfate, and chondroitin sulfate E and also store neutral

proteases.86 The lipid mediators LTB4

, LTC4

, and PGD2 are synthesized de novo in response to

stimulation. The LTs are powerful vasoconstrictors, bronchoconstrictors, and chemoattractants for

neutrophils and eosinophils. PGD2

inhibits platelet aggregation and is chemotactic for neutrophils. Both

basophils and mast cells synthesize and secrete histamine. Histamine, or 2-(4-imidazolyl)-ethylamine, is

formed by the carboxylation of histidine. It is stored in preformed granules at acid pH as a complex

with proteins and proteoglycans. Mast cells carry greater quantities of histamine than do basophils. The

classic vasoactive properties of arteriolar dilatation, increased vascular permeability, and

bronchoconstriction are mediated by the H1

receptors. H2

receptors are involved in modulation of the

immune response and stimulating gastric output and mucus secretion. H3

receptors participate in

neuroconduction.39,86,87

Platelets

Though classically considered in the context of hemostasis, studies confirm that platelets are also

integral to normal and pathologic inflammation and link the processes of hemostasis, inflammation, and

tissue repair. They are activated in a variety of inflammatory conditions, such as rheumatoid arthritis

and inflammatory bowel disease, a testimony to their role in inflammation.88 Upon activation, they

release factors that enhance vascular permeability, chemokines, microbicidal proteins, and mitogens for

endothelial cells, smooth muscle cells, and fibroblasts. They assist leukocytes in promoting the

inflammatory reaction and killing microbes by providing an adhesive surface to facilitate emigration, by

stimulating adhered leukocytes, and by further modulating chemokine synthesis (Table 7-5).88,89

Humans possess about 150 to 400 × 109 platelets per liter of blood.88 Thrombopoiesis is regulated by

thrombopoietin as well as a variety of other mediators (IL-3, IL-4, IL-6, IL-7, and IL-11). In fact, human

IL-11 is in clinical use to stimulate thrombopoiesis in patients undergoing chemotherapy.90 After release

from the marrow, platelets circulate with a half-life of approximately 12 days. Interferon α is inhibitory

for megakaryocyte growth, and the elevated levels induced with some viral and inflammatory

conditions may explain the relative thrombocytopenia observed in these conditions.88,91

Recruitment and Activation

Platelets are activated by and aggregate in response to thrombin or activated endothelium. Clot

formation triggers the release of vasoactive agents (PGE2

, PGI2

, PGF2α, and platelet-derived growth

factor [PDGF]) and inflammatory mediators. Serotonin, PGE2

, and PAF increase vascular permeability.

The adhesion molecule PECAM-1 and the β3

integrins mediate platelet plug formation, endothelial cell

adhesion, and leukocyte emigration. The integrin IIb/IIIa and P-selectin facilitate platelet interactions

with other inflammatory cells. The α granules are rich in membrane-bound P-selectin; upon fusion with

the cell surface membrane they enhance the cell surface density of this receptor. On stimulation, both

receptors are expressed on the platelet surface in an activated state. IIb/IIIa binds various adhesive

proteins containing the RGD sequence (i.e., fibrinogen), which serve as a mechanical link to other

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platelets, leukocytes, and the endothelium. Possibly, the immobilization of activated P-selectin–

expressing platelets on the vessel wall may biochemically and functionally promote the adhesion of

neutrophils to endothelial cells.88,91

Table 7-5 Platelet-Derived Mediators

Platelets are a primary source of chemoattractants for neutrophils. Neutral proteinases released from

stimulated platelets cleave complement factor C5, liberating the chemoattractant C5a. PDGF binds

strongly to the extracellular matrix, providing a long-acting source of chemoattractant. Platelet factor 4

(PF4) is a cationic protein that penetrates the vascular wall and is a chemoattractant. Activated platelets

also bind monocytes via P-selectin and PSGL-1 and induce the expression and secretion of monocyte

chemotactic protein-1 (MCP-1) and IL-8. Thrombospondin released from activated platelets mediates

monocyte binding to platelets. Together these substances promote leukocyte margination, activation,

and recruitment to the sites of injury.39

Granules

The vasoactive substances and inflammatory mediators of platelets are either stored in cytoplasmic

granules or synthesized de novo. A platelet contains about 35 α granules and 5 dense bodies. The dense

granules contain adenosine diphosphate (ADP), ATP, serotonin, and calcium that are required during the

earlier stages of inflammation. ADP is the principal platelet agonist during platelet aggregation and

augments the oxidative burst of neutrophils. Serotonin increases vascular permeability and enhances the

superoxide production by macrophages.88

The more abundant α granules contain fibrinogen, RANTES, MIP-1α, thrombospondin, P-selectin,

PF4, PDGF, TGFβ, β-thromboglobulin, high–molecular-weight kininogen (HMWK), and many other

biologically active proteins. PF4 and β-thromboglobulin initiate leukocyte recruitment and activation.

PF4 induces neutrophil adherence to unstimulated endothelium and the release of secondary granules. It

inhibits monocyte apoptosis and promotes macrophage differentiation. PF4 can also stimulate histamine

release from basophils. RANTES is deposited on the endothelium and recruits monocytes from the

circulation. PAF induces platelet aggregation, increases vascular permeability, enhances phagocyte free

radical formation, and the adhesion of platelets to neutrophils. Platelet activation appears to occur in

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allergic asthma and may precede the delayed accumulation of eosinophils in the lung after allergen

exposure. The α granules also carry several important growth factors, including vascular endothelial

growth factor (VEGF), PDGF, and TGFβ. VEGF promotes extravasation and aids recruitment of

leukocytes. PDGF is chemotactic for neutrophils and monocytes. TGFβ is chemotactic for and activates

neutrophils and monocytes early during inflammation, but displays immunosuppressive effects during

later stages of inflammation.88

Platelets are an important source of eicosanoids, including thromboxane, prostaglandins F2α and E2

.

Thromboxane synthetase in platelets is responsible for the production of TXA2

, a potent vasoconstrictor

that also increases vascular permeability and stimulates platelet aggregation. Aspirin and other

nonsteroidal anti-inflammatory drugs (NSAIDS) inhibit platelet function by inhibiting TXA2 production.

As this effect is irreversible, new platelets must be produced (7 to 10 days) to restore normal clot

formation. In emergent circumstances (management of intracranial hemorrhage) platelet administration

is required. Recently, an aspirin response test has been developed that assists in guiding therapy and

targeting transfusion to those with functional evidence of platelet inhibition. It has also assisted in

monitoring the response to transfusion. PGF2α causes vasoconstriction, whereas PGE2 vasodilates and

modulates pain.92,93

Platelets participate in transcellular lipoxygenase (LO) metabolism, which refers to the production of

eicosanoids through interactions with neighboring inflammatory cells. Endothelial cells utilize plateletderived endoperoxides to synthesize PGI2

. Platelets interact with neutrophils in several pathways,

providing a direct link between thrombosis and inflammation. 12-hydroxyeicosatetraenoic acid (12-

HETE) released from activated platelets can be used by unstimulated neutrophils to produce the

chemoattractant 12,20-HETE. 12-HETE and 5-HETE from activated platelets and neutrophils,

respectively, can combine in either cell type to form 5,12 diHETE, an anti-inflammatory compound,

which diverts production away from the proinflammatory LTs. 12-LO from platelets and LTA4

formed

by neutrophils can produce the intermediate 5(6)-epoxytetraene. This intermediate produces lipoxins A4

and B4

that have primarily counterinflammatory functions.39,89,92,94

In addition to modulating inflammation platelets possess some direct microbicidal activity. Platelets

are activated and degranulate when exposed to certain bacteria. Electron microscopic studies have

shown that activated platelets internalize bacteria and viruses. The α granules contain antibacterial

proteins called thrombocidins (TC) in humans that support the killing of adherent bacteria. The two

antibacterial proteins isolated from human platelets (TC-1, TC-2) are bactericidal for Escherichia coli and

S. aureus.88

NONCELLULAR COMPONENTS

Cytokines

Cytokines are soluble protein mediators secreted by the cells of the innate and adaptive immunity in

response to microbes and other antigens, including intra- and extracellular proteins, and mediate many

of the functions of these cells (Table 7-6). They regulate and influence the host response to both PAMPs

on bacterial, viruses, fungi, and parasites and DAMPs released during trauma, burns, allograft rejection,

ischemia/reperfusion injury, and autoimmune disease. They govern lymphocyte differentiation during

adaptive immunity and activate effector cells of both arms of inflammation to eliminate microbes.

Cytokines play important roles in tumor biology and angiogenesis. Though essential for a normal

immune response, excessive cytokine release underlies a variety of pathophysiological inflammatory

states such as ARDS and MODS.95–98

Table 7-6 Cytokines

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Cytokine secretion is brief, and as cytokines typically are not stored preformed, they must be

transcribed and synthesize de novo synthesis, which is also transient as the messenger RNA is unstable.

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Their functions are pleiotropic and redundant, which has been interpreted to represent a teleological

safety mechanism. However, such protective characteristics greatly limit the therapeutic utility of either

cytokine administration or blockade. In fact, over 30 randomized controlled clinical trials have been

conducted to assess the efficacy of modulating inflammation, on reducing mortality in a variety of

contexts: trauma, sepsis. The majority have involved manipulating systemic cytokine concentrations.

Only one, activated protein C, was briefly FDA approved for use; it was recently removed from the

market due to lack of benefit.99

Cytokines often influence the synthesis and actions of other cytokines, such as IL-1–induced T-cell IL-2

production; this interaction may be synergistic (additive or multiplicative) or antagonistic. They may

function in an autocrine, paracrine, or even endocrine function, although the inability to correlate

plasma cytokine concentrations with the extent of tissue damage suggests that they are designed for

local rather than systemic inflammation.4,39,95

Cytokine actions are mediated by specific membrane receptors on target cells to which they bind with

high affinity (kD α 10−10 to 10−12). Hence, only minute quantities are necessary to elicit a response.

Such efficiency is accompanied by a narrow therapeutic index and potential for unwanton global effects

(i.e., MODS) with even small systemic concentrations. The receptors and the cells expressing them are

regulated by external stimuli, which provide some degree of specificity to the response, even though the

cytokines themselves are not antigen specific. Receptor binding alters cellular gene transcription

(induction or suppression) that may result in proliferation, differentiation, and the acquisition of new or

suppression/enhancement of pre-existing functions.4

All cytokine receptors consist of at least one transmembrane protein with an extracellular portion for

ligand binding and an intracellular domain mediating signal transduction. Current receptor classification

is based on structural homologies among the extracellular cytokine binding domains. Type I cytokine

receptors contain a domain with two conserved pairs of cysteine residues and a membrane proximal

sequence of tryptophan-serine-X-tryptophan-serine, where X is any amino acid (WSXWS). Type II

resemble type I receptors, however the WSXWS motif is absent. The Ig superfamily consists of receptors

with extracellular Ig domains. TNF receptors belong to a family of receptors with conserved cysteinerich extracellular domains. Finally seven-transmembrane α-helical receptors mediate the functions of

cytokines called chemokines through GTP-binding (G) proteins.4

Translating ligand engagement to the signaling events that alter cellular phenotype involves a variety

of signaling cascades dependent upon the structure of the cytoplasmic tail of the particular receptor

(Table 7-6). This is another method by which to classify cytokine receptors. These individual signaling

cascades will be discussed in the context of the various cytokines and respective receptors employing

them.

Early Cytokines/Innate Immunity

TNF-α

TNFα is the principal initiating mediator of the inflammatory response to gram-negative bacteria and

other infectious pathogens. It is an important early mediator of inflammation and is essential for the

normal initiation, maintenance, and repair of tissue injury. However, aberrant production may underlie

the pathologic sequelae of many inflammatory and infectious states (i.e., MODS, rheumatoid arthritis,

Crohn’s disease). Mononuclear phagocytes are the principle source of TNFα, usually in response to the

potent prototypical stimulus lipopolysaccharide of gram-negative organisms.100,101 TNFα production

may be augmented by concomitant stimulation with IFNγ, IL-4 and IL-10, whereas, corticosteroids are

suppressive.102 Secondary sources include mast cells, NK cells, and T and B lymphocytes. Other stimuli

inducing production and release include complement C5a, GM-CSF, hypoxia, IL-1, NO, ROS, and TNFα

itself.39

TNFα is expressed on the cell membrane as a bioactive protein and is cleaved into its soluble form by

a specific TNFα converting enzyme. There are 2 distinct TNF receptors present on virtually all cell types

(Fig. 7-6).103 Ligand binding to TNF-RII leads to recruitment of TNF receptor-associated factors (TRAFs)

that ultimately activate the transcription factors NFκB and AP-1, and thereby influence gene expression

and cellular phenotype.4 Alternatively, binding to TNF-RI activates caspases and triggers apoptosis, in

addition to signaling transcription factors. The manner by which either a proinflammatory or apoptotic

path is chosen is unknown, although the net effect of TNFα binding is probably the culmination of

differential adaptor protein expression and downstream signal transduction cascades. TNF-RI appears to

be more important in host defense as knockout TNF-RI mice show a greater degree of impairment in

host defense than mice lacking TNF-RII.4

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