The generation of CD8+ cytotoxic T cells is enhanced by both TH1 and TH2 cytokines. IL-2, IFN-y, IL-4, and IL-5 all enhance the generation of CTLs although IL-2 is most effective.142-145

TH1 cytokines are important mediators of delayed type hypersensitivity (DTH)146 (see below). TH1 and TH2 cytokines cross-regulate. TH1 and TH2 cytokines tend to be mutually inhibitory136,147 regardless of what cell is producing them. IFN-y inhibits the proliferation of TH2 clones and IL-10 suppresses both cytokine production and proliferation of TH1 clones. IL-10 inhibits IFN-y production by TH1 clones by 90% and inhibits production of TH1 cytokines by CTL clones and LGL. IL-10 acts at the level of antigen presenting cells and their relatives such as skin Langerhans cells.148

T cells from IL-2 knockout mice have disturbed cytokine production121 in vitro, overproducing IL-4, IL-6, and IL-10. In vivo, such mice have increased serum levels of IgG1 and IgE due to increased IL-4 production. For IL-10 knockouts the results are less clearcut.

Revising the Concept of "Suppressor T cells"

Mixing nonresponding and responding populations of T cells can shut off the responders. This "contagious" unresponsiveness used to be attributed to a special class of "suppressor T cells", typically carrying CD8 markers, but it is now clear that antigen-specific T cells with unique suppressor function cannot be isolated. Suppression is a cell population phenomenon, not attributable to unique specialized cell class. All cytokines have both positive and negative effects, and the cells which produce them cannot be assigned a uniquely positive or negative function except in relationship to one specific set of circumstances and one specific target system. For example, TH2 cytokines such as IL-4 can suppress DTH responses but help IgE responses. Thus T cells producing IL-4 can have many simultaneous functions, positive and negative, depending on where the IL-4 is received. So suppression by IL-4 is really a characteristic of the cell that receives IL-4, not the cell that produces it. Similarly, cells producing TGF-p are negative regulators in some types of inflammation and positive in others.149-155

Thus negative regulation or suppression can often be explained without postulating the existence of specialized suppressor cells. In clinical transplantation, where negative regulation is vital to success, the agenda has shifted from suppressor T cells to a detailed analysis of the role of particular molecules in negative regulation of graft injury and inflammation, such as TH2 cytokines and TGF-p.

The Alloantibody Response Against MHC Antigens B-cell activation normally takes place in germinal centers (GC) of draining lymph nodes or spleen, but may occur in the graft infiltrate of a transplant. The surface Ig of the B cell, sIg, engages the polymorphic regions of the donor MHC, particularly the a-helices, in the native, nondenatured, unprocessed form. The MHC antigen is probably shed from donor cells. This leads to B cell triggering and internalization of the antigen. The mechanisms of signalling through sIg involves a receptor complex on the B cell similar to the CD3 complex on T cells.156

The result is that a signal for B cell triggering is delivered, activating intracellular pathways which include the calcium-dependent pathway.

To recruit antigen-specific T cell help, B cells must present peptides of allogeneic MHC antigen in the groove of its class II antigens. To accomplish this, the allogeneic MHC antigen bound by sIg is endocytosed, through proteins around the receptor termed a and p,156 and presented as peptides in the class II groove of the B cell. Host B cells thus present peptides of donor MHC to host CD4 T cells. Antigen presentation by a B cell is crucial for the T-cell response.157 T cell-B cell interactions are weak unless the T cell recognizes its cognate antigen on the B cell. The CD4 T cells may initially be sensitized by antigen on host or donor dendritic cells158 because antigen specific B cells are uncommon in the early stages of the response before they are triggered and undergo clonal expansion. CD4 T cells, B cells, and DCs presumably interact in multi-cell complexes.

T cell-B cell engagement involves a variety of adhesion and signalling interactions, including CD4 with class II, CD40 ligand with CD40, LFA1 with ICAM-1, and CD2 with LFA3, CD5 with CD72, etc. Several cytokines are also transmitted from the CD4 cell to the B cell and B-cell signalling molecules such as CD40 help to trigger the T cell. The expression of adhesion molecules, cytokines, and cytokine receptors increases. The signals to the B cell from the T-helper determine whether the B cell will progress towards antibody production and memory, or toward anergy/apoptosis (programmed suicide).159 Apoptosis is regulated by the gene bcl-2 in lymphocytes:160 mice with knockouts of bcl-2 gene have spontaneous suicide of their lymphoid tissues and lymphocytes.161

If the signals are correct, the B cell undergoes massive clonal expansion and differentiation. Ig, initially expressed on the B cell membrane, can now be released in large quantities as circulating antibody against MHC and other alloantigens.

What Sites on the MHC Does Alloantibody Recognize?

Alloantibody recognizes the "nonself" sites in the a-helix and the ends of the p-pleated sheets that are due to the effect of polymorphic amino acids. The most abundant and important Ig class produced is IgG, which has two antigen binding sites. Each IgG can engage only one site in the MHC molecule. The other binding site of the alloantibody can engage the same region of another MHC molecule. The alloantibody usually binds to the side of an MHC sheet and helix domain or to the top of one a-helix, not across the groove like the TCR. However, one IgG molecule will not fix complement efficiently: an adjacent IgG molecule is needed. The best way of assuring that such IgG complexes will be assembled is to have multiple clones responding to different sites in the mismatched molecule. This is usually the case with clinically important anti-MHC responses: they are polyclonal and react with several sites on the MHC molecule.

Does the peptide in the MHC groove influence antibody binding to the MHC? Perhaps, because the peptide may alter the shape of the domain, as well as possibly directly contacting the antibody in a few cases. Alloantibodies specific for the MHC allele plus a specific peptide are known162 and would escape detection in our usual antibody screening programs. Alloantibody which required a specific peptide would usually react with too few MHC molecules to be quantitatively important. It is conceivable that alloantibody recognizing abundant tissue specific peptides in MHC alleles could act as tissue specific alloantibodies in rejection, e.g., anti-endothelial antibodies. This could help to resolve the old problem of tissue specific alloantibodies such as anti-endothelial antibodies.163

Cytotoxic CD8 Response

Whether the naive CD8 cell requires an APC for its primary stimulation is less well established than for the CD4 cell. The presence of the CD4 cytokines and possibly direct contact from CD4 have been suggested to be necessary for the CD8 cell to be triggered. However, CD8 cells can also be directly triggered without CD4 cells at times, as shown in CD4 deficient or class II deficient mice.

With time and clonal expansion, the CD8 cell acquires the ability to be cytotoxic for target cells. Cytotoxicity is direct lysis of target cells in suspension with the targets undergoing programmed cell death (apoptosis). Functional cytolytic ability correlates with the expression of serine esterases (granzymes) and perforins.164 Although both are sequestered in cytoplasmic granules, perforins and granzymes are regulated differently. Another mechanism of target cell lysis is the interaction of a TNF-like molecule on the T cell (Fas ligand) with a TNF-receptor-like molecule on the target (Fas). Cytolytic ability also requires adhesion molecule interaction between the cytotoxic T cell and the target cell.

The Possible Role of Natural Killer (NK) Cells in Allorecognition

NK cells can lyse cells with little or no class I, apparently being inhibited by expression of class I. This may reflect recognition of the class I groove by an NK receptor.165-167 Little is known about such receptors, and the role of NK cells in transplantation is uncertain.

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