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Mere antigen exposure is insufficient for activation of naïve T cells. Proliferation and differentiation

require costimulatory signals provided by molecules on APCs. The best characterized costimulator

pathway involves the T-cell surface molecule CD28 and its counterligands B7-1 and B7-2 expressed on

activated APCs.4,39 CD28 delivers signals that enhance T-cell survival, by increasing expression of the

antiapoptotic protein Bcl-X, the production of cytokines such as IL-2 and the IL-2 receptor, and the

differentiation of immature T cells. In vitro, purified populations of CD4+ cells challenged with antigen

by APCs that express B7, proliferate and secrete cytokines. This does not occur if B7 is absent. The

costimulatory signal must come from the same APC that provides the initial signal. DCs are the most

potent APC because they express both classes of MHC molecules and the B7 molecules, whereas

macrophages and B cells must be activated to express the costimulatory molecules. This expression of

costimulators is regulated so as to ensure that T-cell activation is temporally and spatially appropriate.

For instance, during T-cell activation, engagement of CD40 ligand with CD40 induces upregulation of

B7 costimulators on the APCs. In addition, it increases the secretion of cytokines such as IL-12 that

promote T-cell differentiation, and cytokines are secreted that promote T-cell differentiation and

activation. A protein CTLA-4, expressed on activated T cells, is homologous to CD28 and binds B7-1 and

B7-2. Unlike CD28, CTLA-4 functions to terminate T-cell responses and plays a role in self-tolerance. On

the basis of many experimental studies of costimulators, antagonists against B7 molecules and CD40L

are in clinical trials to prevent the rejection of organ allografts.4,39

The differentiation of naïve CD8+ T cells to CTL requires a stronger costimulatory signal. This can be

provided by either DC, as they have the greatest intrinsic costimulatory activity, or by a CD4+ helper T

lymphocyte. Naïve helper T cells attached to the same APC as the CD8+ T cell can be activated to

elaborate IL-2. Attached effector T helper cells can stimulate the APC to express more costimulatory

molecules. In the case of virulent viruses, cytotoxicity can substitute for CD28 costimulation, and so the

typical costimulatory signal is not required for activation. For less virulent viruses, costimulation is

necessary for CTL induction.39,76 The absence of costimulation results in an unresponsive, or anergic T

cell. Recently this has been shown to be mediated by the serine/threonine kinase calcium/calmodulindependent protein kinase (CaMK) II (see below).77 Anergic T cells do not produce IL-2 and therefore

cannot proliferate and differentiate into effector cells even when presented with antigen at a later

time.39

The affinity of most TCRs for peptide–MHC complexes is low, with dissociation constants of the order

of 10−5 to 10−7 and an estimated TCR–antigen interaction of less than 10 seconds. Furthermore, on any

APC fewer than 1000 of the 105 available MHC molecules are likely to be displaying any one peptide at

any particular time. Therefore, one APC can engage a small fraction of the 104 to 105 antigen receptors

on a single T cell.4 Activation of an individual T cell may require multiple sequential engagements of

that cell’s antigen receptors by peptide–MHC complexes on APCs. With engagement, there is clustering

of membrane receptors, tyrosine phosphorylation of several proteins, and recruitment and activation of

adaptor proteins.

TCRs are devoid of enzymatic activity and must utilize other signal-transducing proteins.4 After TCR–

MHC engagement, several membrane surface proteins and intracellular signaling proteins are rapidly

recruited: TCR complex, CD4 or CD8, receptors for costimulators such as CD28, and enzymes and

adaptor proteins.4 After TCR clustering, activated tyrosine kinases associated with and phosphorylate

tyrosine residues on CD3 and TCR (Fig. 7-5). These phosphorylation sites provide docking sites for

other tyrosine and protein kinases, such as Lck, an Src family tyrosine kinase, and ZAP-70, a tyrosine

kinase. These kinases become activated with phosphorylation. Activated ZAP-70 phosphorylates several

adaptor proteins that subsequently induce a variety of signal transduction cascades. Adaptor proteins

contain structural domains that bind other proteins and thereby facilitate the correct spatial orientation

that is required for signal transduction. The ras pathway is also activated, which is an early step in the

activation of the mitogen-activated protein kinases (MAPK) that can activate a variety of transcription

factors. Ras is a member of a family of guanine nucleotide-binding proteins (GDP/GTP) that are

involved in diverse activation responses in different cell types. This pathway is an amplification process

by which few upstream kinases lead to the activation of several downstream kinases. Ultimately

activation of the terminal extracellular regulated kinase (ERK 1/2) leads to the phosphorylation of the

protein ELK, which stimulates the transcription of fos, a component of the activation protein 1 (AP-1)

transcription factor (Table 7-4). Concomitantly, c-Jun N-terminal kinase (JNK) is activated, which

phosphorylates c-Jun, the second component of AP-1. The third member of the MAPK family, p38, is

also activated. The activities of the MAPKs are terminated by specific protein tyrosine/threonine

phosphatases that are regulated by the MAPKs themselves. Hence, the entire system is self-regulated by

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a negative feedback system.4

Figure 7-5. T-cell signaling and activation. (Redrawn from Abbas AK, Lichtman AH. Cellular and Molecular Immunology. 5th ed.

Philadelphia, PA: Saunders; 2003.)

Table 7-4 Transcription Factors

Activation of TCR also leads to the induction of PLC, in particular PLCγ1 (Fig. 7-5). Phosphorylated

PLCγ1 catalyzes the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2

) into inositol 1,4,5-

triphosphate (IP3

) and diacylglycerol (DAG), and activates enzymes that generate additional active

transcription factors. IP3

increases cytosolic calcium that leads to a large influx of both intracellular and

extracellular calcium with subsequent activation of calcium- and calmodulin-dependent proteins.

Calcineurin, a calcium/calmodulin-dependent phosphatase is integral to T-cell activation via modulation

of the activation of the transcription factors nuclear factor of activated T cells (NFAT) and NFκB (Table

7-4).4,75 These transcription factors are essential for the induction of cytokine transcription, in particular

IL-2 production. However, in the absence of costimulation, activation of the CaMK II opposes the

actions of calcineurin as described above and produces an anergic cell.77 DAG activates protein kinase C,

which activates additional transcription factors. The role of PKC and calcium in T-cell function is made

evident by studies in which pharmacologic activation of PKC and/or elevation of intracellular calcium

concentration stimulates T-cell cytokine secretion and proliferation.4 Regulation of those kinases

operant in T-cell signaling involves protein tyrosine phosphatases. Through dephosphorylation they

modulate TCR signaling. Two phosphatases induced with TCR clustering are SHP-1 and SHP-2.

The ultimate goal of all these signaling transduction pathways is to activate transcription factors that

bind to promoter regions and enhance transcription. Three transcription factors that are activated in T

cells and appear critical for most T-cell responses are NFAT, AP-1, and NFκB (Table 7-4).

A third mechanism of T-cell activation involves lipid antigens, such as cell wall protein from

intracellular bacteria. These antigens bind CD1, a MHC-related cell surface molecule that presents these

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antigens to certain subtypes of T cells. A superantigen is an unprocessed bacterial or retroviral product

that binds the MHC molecule and the TCR outside the usual antigen-binding sites. This engagement

leads to a polyclonal and nonspecific stimulation of a large proportion of the T-cell population. An

overwhelming activation of all arms of the immune system ensues and underlies much of the

pathophysiology of toxic shock syndrome (TSS). Intravenous immunoglobulin (IVIG), by binding this

antigen, is thought to be of therapeutic benefit.

Eosinophils

Eosinophils are marrow-derived granulocytes that share some properties of neutrophils, act in

conjunction with basophils and mast cells as primary effectors in allergen inflammation and are

involved in the eradication of helminthic infections (Table 7-1). Upon exiting the marrow, their

intravascular half-life is but a few hours, whereafter they enter the mucosa of the lung, gastrointestinal,

and genitourinary tracts.39,78 IL-3, GM-CSF, and IL-5 promote eosinophil differentiation, the induction of

effector functions, and survival, the last by inhibiting apoptosis.78 Emigrating through inflamed

endothelium they release inflammatory mediators and toxic agents from cytoplasmic granules. They

generate superoxide anion and hydrogen peroxide, though less efficiently than neutrophils. They express

IgE receptors and stimulate histamine release from basophils and mast cells through major basic protein

(MBP). They can also regulate basophil and mast cell function by releasing enzymes that inactivate

histamine and slow-reacting substance of anaphylaxis (SRS-A).

Recruitment and Activation

Eosinophils are recruited primarily to sites of parasitic infection and allergen challenge. Mast cells and

macrophages responding to either secrete mediators (IL-5, PAF, LTB-4) that upregulate expression of

endothelial adhesion molecules. Eosinophils themselves are more responsive to CC chemokines (MIP-1

α, RANTES, and MCP-3) and the cytokines produced with TH2 activation. Engagement of eosinophil β1

integrin VLA4 with VCAM-1 and fibronectin on the endothelium initiates the process of emigration. IL-4

can activate both the binding and the upregulation of endothelial cell VCAM-1. Diapedesis and tissue

infiltration also involves members of the β2

integrin family (Mac-1 and LFA) similar to that which

mediates neutrophil recruitment.39,79

Eosinophils are activated by IL-3, GM-CSF, IL-5, PAF, CC chemokines, and C3a and C5a of

complement. IL-5 is a potent activating agent and enhances the ability of eosinophils to release granule

contents on FcR cross-linking. A positive feedback cycle ultimately is established, in which recruited

eosinophils produce cytokines and chemokines that recruit more eosinophils and other leukocytes.

Granules

Eosinophils possess a compartmentalized armamentarium of toxic substances that assist in the

elimination of organisms, in particular helminths. Their specific granules contain GM-CSF and MBP, the

latter of which is cytotoxic to parasites and normal cells. Further, it functions as a stimulus for

histamine release from mast cells and basophils.80 The granule matrix contains eosinophil peroxidase

(EPO), eosinophil-derived neurotoxin, lysosomal enzymes, catalase, TNFα, TGFβ, and eosinophilic

cationic proteins that stimulate formation of transmembrane pores to increase target cellular

permeability.81 EPO is released extracellularly on target cell surfaces where it generates hydrogen

peroxide and hydrogen halides. Approximately 30% of oxygen consumed by stimulated eosinophils is

utilized in the formation of halogenating species. Thiocyanate may be the major halide for the EPO–

H2O2 system.82 If ingested by a neighboring phagocyte, EPO can combine with H2O2 and halides to

form hypohalous acids. EPO also stimulates neutrophil aggregation and adhesion to endothelial cells.

Although they express Fc receptors for IgG, IgA, and IgE, they are relatively insensitive to activation by

antigen-mediated cross-linking of these receptors. However, they can kill microorganisms by antibodydependent cell-mediated cytotoxicity.4,39

The primary targets of eosinophils are extracellular parasites. The size of these pathogens prohibits

phagocytosis, and their integument is relatively resistant to the microbicidal products of neutrophils and

macrophages; however they can be killed by MBP, which is released after cross-linking of Fc-bound IgE

coating the parasite. The TH2 response to parasitic invasion produces IL-4, IL-5, and IL-13. IL-4

stimulates the production of specific IgE antibodies, which opsonize helminths. IL-5 activates

eosinophils, which bind to the IgE-coated helminthes via Fc receptors. Activated eosinophils then release

their granule contents and generate ROS and hypohalous acids. EPO also stimulates neutrophil

aggregation and adhesion to endothelial cells. The sparse uncompartmental granules contain Charcot–

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