THEME
I. Antigen presentation in the intestine: new rules and regulations*

Lloyd Mayer

Immunobiology Center, Mount Sinai Medical Center, New York, New York 10029

    ABSTRACT
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The recognition that immune responses in the intestine differ from those seen systemically has led to a search for novel pathways involved in mucosal immunoregulation. One cell type that has surfaced as a prime candidate for such a regulatory role is the intestinal epithelial cell. A number of laboratories have documented that intestinal epithelial cells sample luminal antigens and process and present these to primed T cells. However, several unique features have emerged, making their potential role as antigen-presenting cells a critical part of mucosal homeostasis.

intestinal epithelium; antigen presentation; nonclassical major histocompatibility complex

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THE VIEW OF the mucosal immune system as a site for the production of a specialized form of immunoglobulin (secretory IgA) for years overshadowed the finding that the major immune response seen at mucosal sites was a nonresponse. In fact, this state of nonresponsiveness is not a passive process but rather involves the active dampening of immune responses. When viewed in the context of the enormous antigen load in the gastrointestinal tract, it is quite remarkable that continuous overactive inflammation of the gut is not commonplace (although controlled or "physiological" inflammation is the norm). It is only over the past several years that we have come to appreciate the unique features of the mucosal immune system that aid in maintaining what appears to be a generally immunosuppressed tone. Several laboratories have proposed that the intestinal epithelial cell (IEC) is a key regulator of this immunologic state and that novel surface interactions guide this regulation.

IECs have been grouped into a category of nonprofessional antigen-presenting cells (APCs). Such APCs do not constitutively express class II major histocompatibility complex (MHC) molecules and do not activate T cells in a conventional manner (e.g., the lack of expression of specific costimulatory molecules such as B7-1 and B7-2). In fact, IECs are different from other nonprofessional APCs in that they do express class II MHC molecules constitutively in the small bowel and to a lesser extent in the colon (18, 23, 24). These class II molecules are functional and can present antigens to primed T cells or hybridomas (15, 20). However, there are still differences in the class II molecules expressed on the IEC surface, since they have been shown to bear invariant chain (a protein produced in the endoplasmic reticulum that complexes with MHC class II to prevent endogenous peptide binding), raising questions as to whether processing properties in IECs are intact or whether class II MHC on IEC can truly present normally to T cells. Hershberg and colleagues (14) recently addressed this issue using IEC lines T84 and HT29 that had been transfected with DR4beta 0401 cDNA. In the presence of interferon-gamma (IFN-gamma ), these cells processed and presented protein to primed cells, suggesting that appropriate processing could occur. However, others have questioned this in primary IEC, in which both uptake and processing have been quite slow (5). In fact, in the absence of IFN-gamma neither T84 nor HT29 appeared to process antigen normally. This may relate to the presence or absence of appropriate processing enzymes (cathepsins) in the endosomal compartments. If these enzymes are present within normal epithelial cells, they are there in reduced amounts (14). Thus processing of intact protein antigens may be limited, and the ability to generate potent immune responses would be reduced. However, the requirements for complete processing may be rendered moot by events that occur in the gut lumen, resulting in varying degrees of proteolysis or what has been termed luminal preprocessing. In fact, Bland (3) has suggested that the apically expressed class II MHC on enterocytes might serve as peptide transporters for luminally processed peptides. This concept has not been adequately tested but remains intriguing.

If processing does occur, can antigen-specific responses be generated? Primary immune responses require the presence not only of appropriately processed peptide-MHC complexes but also the expression of a number of costimulatory molecules on the surface of the APC to amplify T cell activation. In conventional APCs these have been well characterized and include the important interactions among B7-1 and B7-2 and either CD28 or CTLA-4 as well as CD40 and CD40 ligand (gp39) (16). Binding of these receptors to their appropriate ligands potentiates T cell responses and rescues cells from anergy or apoptosis (16). Thus far there is little evidence to suggest that IECs express any of these costimulatory molecules in vivo and therefore interactions of CD4+ T cells with IEC, even in the presence of an appropriately processed peptide, might result in an anergized cell. Such an outcome would help to protect the host from potentially harmful immune responses against normal luminal organisms.

Yet it has been evident in several studies that IECs are capable of taking up soluble protein antigens. This has been documented in both in vitro and in vivo systems (8, 11, 12, 21). However, the kinetics of uptake are slow compared with conventional APCs, although the intracellular pathways of trafficking are similar (endolysosomal pathway) (Fig. 1). The results do vary, though, depending on the cell system used, and only two studies have examined this capacity in vivo (11, 12). More recently, Berin et al. (2) have described an ex vivo system in which antigen trafficking through the IEC could be accelerated, resulting in an effector response. Berin et al. (2) reported that systemic priming with a given antigen increased the transcellular transport of that antigen. This was associated with the induction of mast cell degranulation in the lamina propria. Such a finding raises the possibility that antigen trafficking is specific, potentially aided by immune-mediated mechanisms and promoted by a distinct cytokine milieu. From all of these studies, it is clear that transcellular transport of antigen does occur in intracellular processing compartments and this sets the stage for antigen presentation.


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Fig. 1.   Colocalization of mannose-6-phosphate receptor (red phycoerythrin) and tetanus toxoid (green fluorescein) 60 min after pulsing a monolayer of HT-29 human colon carcinoma cells with fluorescein isothiocyanate-tetanus toxoid. Yellow fluorescent dots in panel at right represent colocalization of antigen (TT) in the late endosomal compartment (M6PR+). This is a classical class II pathway but is also the site where CD1d is located.

This is where the similarities to classic APCs end. Although the class II molecules expressed on IEC can be functional, several laboratories have reported (4, 19) that the cells proliferating in normal IEC-T cell cocultures are predominantly CD8 and are not the CD4+ T cells one would expect in class II restricted events. CD8+ T cell activation in this system involves the CD8 molecule itself and is associated with the activation of its associated tyrosine kinase p56lck (17). Because MHC class I molecules, the conventional ligands for CD8, are not found in the endosomal compartments in which soluble proteins travel through the epithelium and because blockade of MHC class I molecules does not inhibit IEC-induced T cell proliferation, other molecules must be involved that govern these interactions. Several years ago, Terhorst and colleagues (6, 7) published the finding that nonclassical class I molecules (CD1) were expressed on murine and human IEC. Hershberg and colleagues (13) described one additional class Ib molecule, TL, expressed by normal IEC in the mouse. These class Ib molecules were shown to be functional in that cytolytic activity by intraepithelial lymphocytes could be inhibited by antibodies to these restriction elements (1). Furthermore, antibodies to CD1d inhibited the proliferation of CD8+ T cells in an allogeneic IEC-T cell coculture system (22). A model started to evolve when it was reported that CD1d was endosomally localized and capable of binding peptides larger than those bound by conventional class I and class II molecules (10). A deep hydrophobic pocket noted after the crystal structure of CD1d was resolved supported the concept that limited antigen processing within the endosome in an IEC could favor peptide binding to CD1d rather than class II. This could have been the complete story except for the finding that CD1d fails to bind to CD8 (unlike classical class I MHC) and would not necessarily result in CD8+ T cell activation. When a new molecule was identified that appeared to associate with CD1d it became possible to resolve this issue. Campbell et al. (9) and Yio and Mayer (25) described a 180-kDa glycoprotein (gp180) recognized by two monoclonal antibodies, which is capable of binding to CD8 and activating its associated kinase, as well as binding to CD1d. The complex of gp180 and CD1d becomes class I-like, with CD1d able to bind both processed peptide and the T cell receptor, while the associated gp180 binds to CD8, helping to form a coreceptor complex (Fig. 2).


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Fig. 2.   Expression of CD1d coupled to gp180 on the surface of the intestinal epithelial cell, forming a complex that is class I-like, capable of binding to both the antigen receptor and CD8. This coreceptor complex allows for the appropriate signaling molecules to come into proximity and maximizes phosphorylation of intracellular substrates and kinases. Major histocompatibility complex (MHC) class I can do this by itself [i.e., bind to the T cell receptor (TcR) and CD8], but a distinct pathway of activation is induced (see text).

It is unclear where the association of gp180 and CD1d occurs (endosome vs. surface). Which cells are interacting with IECs is not well defined (intraepithelial lymphocytes vs. lamina propria lymphocytes vs. recently migrated peripheral lymphocytes). However, these findings do underscore the unique nature of immunoregulation at mucosal sites: the use of unconventional APCs, restriction elements, and costimulatory molecules. It is not surprising that the rules are different for mucosal immune responses. Clearly, how to manipulate this system to generate effective mucosal vaccines is a focus for future research.

    ACKNOWLEDGEMENTS

I thank Debbie Matz for help in the preparation of this manuscript.

    FOOTNOTES

* First in a series of invited articles on Current Concepts in Mucosal Immunity.

This work was supported by National Institute of Allergy and Infectious Diseases Grants AI-23504 and AI-24671, as well as by the Kennedy-Leigh Charitable Trust.

Address reprint requests to Dept. of Immunobiology, Medicine, and Microbiology, Mt. Sinai Medical Center, 1 Gustave Levy Place, New York, NY 10029.

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