Previous studies support a role for intestinal epithelial cells (IEC) as antigen-presenting cells in mucosal immune responses. T cells activated by IEC are CD8 ϩ , suppressor in function, and dependent upon CD8-associated p56 lck activation. A 180-kD glycoprotein (gp180) recognized by mAbs B9 and L12 has been identified and shown to be important in CD8 ϩ T cell activation by IEC. Since IEC derived from patients with inflammatory bowel disease (IBD) are incapable of activating CD8 ϩ T cells, we asked whether this correlated with gp180 expression. While frozen sections of normal bowel revealed bright gp180 staining on all IEC, both inflamed and uninflamed ulcerative colitis (UC) specimens showed patchy staining. In Crohn's disease (CD), staining was faint to absent. Flow cytometry confirmed immunohistochemical data. The staining patterns correlated with the ability of IEC to activate CD8-associated p56 lck. Normal IEC induced phosphorylation of p56 lck in CD8 ␣ but not CD4 ϩ transfectants. In contrast, both UC and CD IEC activated CD4 and, to a much lesser extent, CD8-associated p56 lck. Thus, gp180 expression by IBD IEC appears to be altered, and correlates with a functional alteration of lck activation. This defect may reflect a more proximal event in the pathogenesis of IBD. (
Previous studies have shown that normal human intestinal epithelial cells stimulate CD8؉ suppressor T cell proliferation in an allogeneic mixed epithelial/T cell coculture system, which is neither restricted by class I or class II major histocompatibility complex antigens nor by any soluble factors from epithelial cells. Two epithelial specific monoclonal antibodies (mAb), mAb B9 and mAb L12, are potent inhibitors of this mixed epithelial/T cell reaction but not of conventional mixed lymphocyte reactions. While phenotypically distinct by tissue staining, both mAbs recognize a 180-kDa epithelial membrane glycoprotein (gp180). Further characterization of gp180 revealed the following. 1) The protein migrated between 150 and 180 kDa in SDS-polyacrylamide gel electrophoresis and could be resolved by Western blot using mAb B9 or mAb L12. 2) The molecule has two forms, an apically sorted glycosylphosphatidylinositolanchored form and a basolateral transmembrane form. 3) gp180 is heavily N-glycosylated, since N-glycanase treatment results in a >50% reduction in size. 4) Purified gp180 can bind to peripheral blood T cells and activate p56 lck . 5) gp180 can activate p56 lck in 3G8 (a murine T cell hybridoma transfected with human CD8␣ cDNA) but not in 3G4 (CD4 transfectant), suggesting that gp180 binds to CD8. Thus, gp180 appears to be a novel regulator of mucosal immune responses.The mucosal immune system of the gastrointestinal tract is distinct from the systemic immune system by the general immunologically suppressed tone in the gut and the induction of specific immune unresponsiveness after antigen challenge, termed oral tolerance. It is believed that CD8 ϩ suppressor T cells primed in the gut may play an important role in oral tolerance (1-7). However, the mechanism underlying the generation of the CD8 ϩ suppressor T cells has not been completely delineated. Previous studies in our lab as well as others have shown that intestinal epithelial cells are able to process and present antigens to T cells (8, 9). More interestingly, intestinal epithelial cells are able to selectively induce CD8 ϩ suppressor T cell proliferation in an allogeneic mixed epithelial cell/T cell reaction (METR) 1 system (8), and conventional restriction elements do not appear to be involved, since neither antibodies against class I nor class II MHC block proliferation (10). Furthermore, the proliferation is not driven by soluble factors from the epithelial cells (8).One molecule that appears to be clearly involved in this interaction is CD8, since there is activation of CD8-associated p56 lck and antibody to CD8 blocks CD8 ϩ suppressor T cell proliferation in this system (10).We have hypothesized that the generation of CD8 ϩ suppressor T cells is attributed to the expression of a unique CD8 binding ligand on the surface of intestinal epithelial cells (10). Therefore, a series of epithelial cell-specific monoclonal antibodies were generated and screened for their ability to inhibit CD8 ϩ T cell proliferation in our culture system. Two of these mAbs...
SummaryThe activation of CD8 + suppressor T cells by normal intestinal epithelial cells in antigen-specific or allogeneic mixed cell culture systems has significant implications for the regulation of mucosal immune responses. In this study, we found that the capacity of epithehal cells to induce CD8 + suppressor T cell activation appeared to be linked to the binding of CD8 molecules on the T cell surface. This appears to be mediated by a non-class I molecule expressed on the epithelial cell surface, which binds to CD8 and results in the activation of the CD8-associated src-like tyrosine kinase, p56 lck. Epithehal cell-stimulated p56 lck activation is an early event (in contrast to monocytes) and is essential for T cell activation, since proliferation could be completely abrogated by pretreatment of T cells with genestein or herbamycin, both of which are protein tyrosine kinase inhibitors. Pretreatment ofT cells with anti-CD8 or of intestinal epithelial cells with an anti-epithelial cell mAb B9 inhibited p56 lck activation and further confirmed that CD8 on the T cell and a CD8 ligand on the epithelial cell were involved in this T cell activation event. The specificity of this reaction was confirmed in experiments in which murine transfectants 3G4 and 3G8, expressing CD4 or CD8, respectively, were used. Coculture of 3G8 with epithelial cells but not with monocytes activated p56 lck in this cell line, whereas p56 lck was preferentially activated in 3G4 cells when monocytes were used as the stimulator cells. Although stimulation through CD8-and CD8-associated p561ok was important for epithelial cell-induced T cell activation, T cell proliferation could not be induced by crosslinking CD8 alone with monoclonal antibody anti-CD8. These data suggest that a second signal, possibly through the T cell antigen receptor since activation of the T cell receptor-associated kinase fyn was also seen, is required for epithelial cell-driven T cell proliferation. O ver the past several years, it has become increasingly apparent that the rules governing mucosal immune responses differ from those of the peripheral immune system. First, antigen priming via the oral route most frequently results in the development of immunologic suppression or oral tolerance rather than in an active immune response. Second, the cell types in the gastrointestinal tract are not typical of what one would find systemically. Intraepithelial lymphocytes are predominantly CD8 + T cells that are, in general, anergic despite exposure to inordinate numbers of disparate foreign antigens in the gut lumen (1-3). In contrast, lamina propria T cells are activated memory cells, a rich source of cytokines, but with limited prohferative capacity to specific antigen (4, 5). An understanding of the underlying mechanisms responsible for these phenomena has been slow to evolve, partly because of our limited This manuscript is in partial fulfillment of a Ph.D. thesis (3(in Li and Xian Yang Yio). knowledge of how the primary trigger of an immune response, antigen, is handled b...
The immunologic tone of the intestinal tract is one of suppressed or highly regulated responses. While there are several components (intrinsic and extrinsic to the gut-associated lymphoid tissue) responsible for this immunologically suppressed tone, the intestinal epithelial call (IEC) has been proposed as a key player in this process. IECs can take up and process antigen but distinct surface molecules and restriction elements allow them to present these antigens to unique regulatory T cells. These include the expression of the class Ib molecule CD1d as well as a novel CD8 ligand, gp180. These molecules come together to activate a subpopulation of CD8+ regulatory cells whose function is to suppress immune responses in an antigen non-specific fashion most likely through cognate interactions. This form of regulation may be unique to the gut-associated lymphoid tissue which is consistent with the unusual demands upon this part of the immune system.
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