Rhabdovirus infection induces public and private T cell responses in teleost fish.

Many viruses induce a strong T cell response that contributes to the elimination of infected cells presenting viral peptides by MHC molecules. The structure and expression of genes encoding molecules homologous to mammalian alphabeta TCRs have been recently characterized in rainbow trout and in several teleost species, but the alphabeta T cell response against pathogens has not been directly demonstrated. To study the modifications of the T cell repertoire during an acute viral infection in rainbow trout, we adapted the immunoscope methodology, which consists of spectratyping the complementarity-determining region 3 length of the TCRbeta chain. We showed that the naive T cell repertoire is polyclonal and highly diverse in the naive rainbow trout. Using viral hemorrhagic septicemia virus (VHSV), which provokes an acute infection in rainbow trout, we identified skewed complementarity-determining region 3 size profiles for several VbetaJbeta combinations, corresponding to T cell clonal expansions during primary and secondary response to VHSV. Both public and private T cell expansions were shown by immunoscope analysis of spleen cells from several infected individuals of a rainbow trout clone sharing the same genetic background. The public response to VHSV consisted of expansion of Vbeta4Jbeta1 T cell, which appeared early during the primary response and was strongly boosted during the secondary response.     

Oligoclonality of rat intestinal intraepithelial T lymphocytes: overlapping TCR beta-chain repertoires in the CD4 single-positive and CD4/CD8 double-positive subsets

Previous studies in humans and mice have shown that gut intraepithelial lymphocytes (IELs) express oligoclonal TCR beta-chain repertoires. These studies have either employed unseparated IEL preparations or focused on the CD8+ subsets. Here, we have analyzed the TCR beta-chain repertoire of small intestinal IELs in PVG rats, in sorted CD4+ as well as CD8+ subpopulations, and important differences were noted. CD8alphaalpha and CD8alphabeta single-positive (SP) IELs used most Vbeta genes, but relative Vbeta usage as determined by quantitative PCR analysis differed markedly between the two subsets and among individual rats. By contrast, CD4+ IELs showed consistent skewing toward Vbeta17 and Vbeta19; these two genes accounted collectively for more than half the Vbeta repertoire in the CD4/CD8 double-positive (DP) subset and were likewise predominant in CD4 SP IELs. Complementarity-determining region 3 length displays and TCR sequencing demonstrated oligoclonal expansions in both the CD4+ and CD8+ IEL subpopulations. These studies also revealed that the CD4 SP and CD4/CD8 DP IEL subsets expressed overlapping beta-chain repertoires. In conclusion, our results show that rat TCR-alphabeta+ IELs of both the CD8+ and CD4+ subpopulations are oligoclonal. The limited Vbeta usage and overlapping TCR repertoire expressed by CD4 SP and CD4/CD8 DP cells suggest that these two IEL populations recognize restricted intestinal ligands and are developmentally and functionally related.     

Intraepithelial lymphocytes express junctional molecules in murine small intestine

Intestinal intraepithelial lymphocytes (IEL) that reside at basolateral site regulate the proliferation and differentiation of epithelial cells (EC) for providing a first line of host defense in intestine. However, it remains unknown how IEL interact and communicate with EC. Here, we show that IEL express junctional molecules like EC. We identified mRNA expression of the junctional molecules in IEL such as zonula occludens (ZO)-1, occludin and junctional adhesion molecule (JAM) (tight junction), beta-catenin and E-cadherin (adherens junction), and connexin26 (gap junction). IEL constitutively expressed occludin and E-cadherin at protein level, while other T cells in the thymus, spleen, liver, mesenteric lymph node, and Peyer's patches did not. Gammadelta IEL showed higher level of these expressions than alphabeta IEL. The expression of occludin was augmented by anti-CD3 Ab stimulation. These results suggest the possibility of a novel role of IEL concerning epithelial barrier and communication between IEL and EC.     

Differences in development of lymphocyte subpopulations from gut-associated lymphatic tissue (GALT) of germfree and conventional rats: Effect of aging

The aim of the study was to compare the phenotype of lymphocyte subpopulations of the GALT (gut-associated lymphatic tissue) in germfree (GF) and conventionally (CV) reared rats,i.e. to analyze the effect of microbial colonization on the development of intestinal lymphocyte subsets. Surface marker characteristics were studied in cell suspensions isolated from Peyer’s patches, mesenteric lymph nodes, spleen and the intraepithelial lymphocyte compartment of 2- and 12-month old inbred AVN rats. The pattern of T lymphocyte phenotypes in Peyer’s patches, mesenteric lymph nodes and spleen determined by FACS analysis did not reveal differences between GF and CV rats. In contrast, a 2-month conventionalization of GF rats led to substantial changes in the composition of intestinal intraepithelial lymphocyte subsets (IELs): increase of CD4⁺, CD8α⁺, CD8β⁺, TcR α/β⁺ bearing lymphocytes was observed after colonization of rats with normal microflora. Surprisingly, the relative numbers of lymphocytes bearing TcR γ/δ⁺ did not change during conventionalization. The effect of aging was also studied and differences in IELs composition of aged (GF) and (CV) rats were found to be more pronounced: 6,6% and 30% of lymphocytes bearing TcR α/β were present among IELs in two-month old GF and CV rats, respectively. 30% of IELs in 2-month old GF rats, 80% of IEL from 12-month old CV rats were found to bear TcR α/β. This finding demonstrates that during conventionalization and aging the TcR α/β bearing population of IELs substantially expands. It suggests that mainly this lymphocyte subset responds to microflora stimuli and is probably involved in the protection of the epithelial cell layer of intestinal mucosa.     

Induction by cells of immune responses to intestinal epithelial cell-associated components (ECAC): transfer with cultured murine mesenteric and popliteal/axillary lymph node cells

Although systemic and mucosal immune responses to intestinal epithelial self-antigens occur in several human disorders, there is no model system with which to study the physiology and regulation of the underlying cellular events. Therefore, we undertook to induce an immune response to purified epithelial macromolecules in the Lewis rat; characterize in vitro the reactive cells; and then transfer with immunocytes this antiepithelial reactivity to naive syngeneic rats, identifying the fine specificity and site of humoral and cell-mediated immunity induced in the cell recipient. Donor animals sensitized systemically (via footpad) or locally in gut mucosa (via Peyer's patches) to syngeneic or xenogeneic epithelial antigens generated specific immunoglobulin and were found to have T lymphocytes in the draining nodal areas (including the mesenteric nodes) which were (a) antigen-specific, having a [3H]thymidine uptake in the presence of antigen 30-fold the control; (b) generally of the Thelper/inducer subclass (W3/25+) which, upon further culture, developed phenotype surface markers for activation (IL-2R+); (c) able to induce an antigen-specific humoral and cell-mediated responses upon intravenous injection into naive syngeneic hosts; and (d) demonstrable in gut-associated lymphoid tissue (mesenteric lymph nodes) and, to a lesser extent in spleen, of the cell recipient. Further, lymphocytes cloned from reactive mesenteric lymph node cells demonstrated specificity for a gel-purified subfraction of epithelial antigen, designated P1, containing highly conserved organ-specific macromolecules thought to be autoantigenic for gut.     

Rotavirus-specific cytotoxic T lymphocytes appear at the intestinal mucosal surface after rotavirus infection.

The gastrointestinal tract is constantly exposed to a variety of potentially invasive bacteria and viruses. The first line of defense of the host against these pathogens is the intestinal mucosal surface, which consists of epithelial cells, intraepithelial lymphocytes (IELs), mucus, and secretory immunoglobulins. Little is known about the function, memory, or trafficking of IELs after intestinal infection. We found that IELs obtained 6 days after oral inoculation of mice with the intestinal pathogen rotavirus (simian strain RRV) lysed rotavirus-infected target cells; cytotoxic T lymphocytes (CTLs) were responsible for rotavirus-specific cytotoxic activity. Rotavirus-specific cytotoxic activity by IELs was (i) eliminated by treatment with Thy 1.2-specific immunoglobulin M plus complement, (ii) restricted by proteins encoded at the major histocompatibility complex, and (iii) absent in mock-infected animals. Oral inoculation of mice with RRV also induced rotavirus-specific CTLs in splenic and intestinal lymphocytes (mesenteric lymph nodes, Peyer's patch). Parenteral inoculation induced rotavirus-specific CTLs in splenic, intestinal (IELs, mesenteric lymph nodes, Peyer's patch), and nonintestinal lymphocytes (inguinal nodes). Therefore, presentation of rotavirus to the intestinal mucosal surface was not necessary to induce IELs with virus-specific cytotoxic activity. At 4 weeks after oral or parenteral inoculation of mice with RRV, rotavirus-specific CTL precursors appeared among splenic, Peyer's patch, inguinal, and mesenteric node lymphocytes, but not among IELs. IELs with rotavirus-specific cytotoxic activity may be generated from precursors at a site other than the intestinal mucosal surface. Part of the response of the host to enteric infection may include surveillance and lysis of virus-infected villus epithelial cells by IELs.     

Differential contribution of Fas- and perforin-mediated mechanisms to the cell-mediated cytotoxic activity of naive and in vivo-primed intestinal intraepithelial lymphocytes

Intestinal intraepithelial lymphocytes (IELs) are known to exert strong constitutive cytotoxic activity. In the present study we compared the Ag-specific cytotoxic activity and the effector mechanisms involved in non-Ag-primed, naive and in in vivo-primed IELs and splenic CD8 T cells. Ex vivo isolated naive CD8alphaalpha TCRalphabeta IELs, CD8alphabeta IELs, and splenocytes from lymphocytic choriomeningitis virus (LCMV)-specific TCR transgenic mice exert Ag-specific cytotoxic activity in a long-term, but not in a short-term, cytotoxicity assay. This cytotoxic activity is mainly Fas-Fas ligand mediated and is significantly reduced in the presence of 20 microg/ml Fas-Fcgamma1 fusion protein. Both CD8alphabeta IELs and CD8alphabeta splenocytes isolated from LCMV-infected C57BL/6 mice exert potent perforin-dependent cell-mediated cytotoxicity. CD8alphaalpha TCRalphabeta IELs from LCMV-infected animals, however, show only minimal Ag-specific cytotoxicity. The potent cytotoxic activity of in vivo activated CD8alphabeta IELs is not affected by the addition of Fas-Fcgamma1. Nevertheless CD8alphabeta IELs from LCMV-infected perforin-deficient mice exert Ag-specific cytotoxicity in a short-term cytotoxicity assay, and this cytotoxicity is almost completely blocked by the addition of Fas-Fcgamma1. These results demonstrate that naive CD8alphabeta IELs exert Ag-specific, Fas-Fas ligand-mediated, constitutive cytotoxic activity in a long-term cytotoxicity assay, whereas primed CD8alphabeta IELs primarily use the perforin-dependent exocytosis pathway to exert their potent cytotoxic activity. Furthermore, these results clearly illustrate the requirement for Ag-specific determination of IEL-mediated cytotoxicity, because the elevated, but variable, frequencies of memory-type T cells in this compartment may lead to ambiguous results when polyclonal activation or redirected assays are used.     

Intestinal CD8 alpha alpha and CD8 alpha beta intraepithelial lymphocytes are thymus derived and exhibit subtle differences in TCR beta repertoires

Intraepithelial lymphocytes (IEL) of the small intestine are anatomically positioned to be in the first line of cellular defense against enteric pathogens. Therefore, determining the origin of these cells has important implications for the mechanisms of T cell maturation and repertoire selection. Recent evidence suggests that murine CD8 alpha alpha intestinal IELs (iIELs) can mature and undergo selection in the absence of a thymus. We analyzed IEL origin by cell transfer, using two congenic chicken strains. Embryonic day 14 and adult thymocytes did not contain any detectable CD8 alpha alpha T cells. However, when TCR(+) thymocytes were injected into congenic animals, they migrated to the gut and developed into CD8alphaalpha iIELs, while TCR(-) T cell progenitors did not. The TCR V beta 1 repertoire of CD8 alpha alpha(+) TCR V beta 1(+) iIELs contained only part of the TCR V beta 1 repertoire of total iIELs, and it exhibited no new members compared with CD8(+) T cells in the thymus. This indicated that these T cells emigrated from the thymus at an early stage in their developmental process. In conclusion, we show that while CD8 alpha alpha iIELs originate in the thymus, T cells acquire the expression of CD8 alpha alpha homodimers in the gut microenvironment.     

Differential effects of vasoactive intestinal peptide, substance P, and somatostatin on immunoglobulin synthesis and proliferations by lymphocytes from Peyer's patches, mesenteric lymph nodes, and spleen

We examined the effect of vasoactive intestinal peptide, substance P, and somatostatin on concanavalin A (1 microgram/ml)-induced lymphocyte proliferation and immunoglobulin (IgA, IgM, and IgG) synthesis by cells from spleens, Peyer's patches, and mesenteric lymph nodes. These neuropeptides (10(-7) to 10(-12) M) modulated immune responses in a dose-dependent manner. For a comparative study, neuropeptides were used at 10(-8) M concentration. Both vasoactive intestinal peptide and somatostatin significantly decreased DNA synthesis (30 to 50%), whereas substance P increased synthesis (40%) in lymphocytes from all organs tested. IgA synthesis was significantly altered by all of the neuropeptides tested, whereas IgM synthesis was less affected and IgG synthesis was virtually unchanged. Somatostatin inhibited IgA (20 to 50%) and IgM (10 to 30%) synthesis in lymphocytes from all three organs. Substance P increased IgA synthesis in mesenteric lymph nodes (50%), spleens (70%), and Peyer's patches (300%). It also increased IgM synthesis in Peyer's patches (20%) and spleens (30%), but was without effect on IgM synthesis in mesenteric lymph nodes. Vasoactive intestinal peptide increased the IgA response in mesenteric lymph nodes (20%) and spleens (30%), but inhibited IgA synthesis in lymphocytes from Peyer's patches (60%). Interestingly, in Peyer's patches, IgM synthesis was increased by vasoactive intestinal peptide (80%), whereas it was unchanged in mesenteric lymph nodes and spleen. Thus, not only did these neuropeptides have different effects on the production of different immunoglobulin isotypes, but their effect was also organ-specific. Because neuropeptides which are abundant in the intestine can modulate IgA and other immunoglobulin synthesis in vitro, they may play a significant regulatory role in mucosal immune responses in vivo.     

Selective emigration of suppressor T cells from Peyer's patches

The emigration of Peyer's patch lymphocytes to mesenteric lymph nodes was studied by injecting fluorescein isothiocyanate (FITC) directly into Peyer's patches. Using double immunofluorescence it was demonstrated that at 2 and 4 hr after FITC injection 70% of the labeled cells that migrated to mesenteric lymph nodes were T lymphocytes, although rat Peyer's patches contain only 15-20% T lymphocytes. At later time points after FITC injection this percentage of T cells derived from Peyer's patches gradually declined, most likely caused by selective interaction and/or retention inside the mesenteric lymph node. Determination of helper and suppressor T-cell subsets within this emigrating population showed an increased number of T suppressor cells migrating into mesenteric lymph nodes. The putative role of suppressor T cells in inducing systemic tolerance after oral antigen administration was discussed.     

IFN-gamma-producing dendritic cells are important for priming of gut intraepithelial lymphocyte response against intracellular parasitic infection

The importance of intraepithelial lymphocytes (IEL) in immunoprotection against orally acquired pathogens is being increasingly recognized. Recent studies have demonstrated that Ag-specific IEL can be generated and can provide an important first line of defense against pathogens acquired via oral route. However, the mechanism involved in priming of IEL remains elusive. Our current study, using a microsporidial model of infection, demonstrates that priming of IEL is dependent on IFN-gamma-producing dendritic cells (DC) from mucosal sites. DC from mice lacking the IFN-gamma gene are unable to prime IEL, resulting in failure of these cells to proliferate and lyse pathogen-infected targets. Also, treatment of wild-type DC from Peyer's patches with Ab to IFN-gamma abrogates their ability to prime an IEL response against Encephalitozoon cuniculi in vitro. Moreover, when incubated with activated DC from IFN-gamma knockout mice, splenic CD8(+) T cells are not primed efficiently and exhibit reduced ability to home to the gut compartment. These data strongly suggest that IFN-gamma-producing DC from mucosal sites play an important role in the generation of an Ag-specific IEL response in the small intestine. To our knowledge, this report is the first demonstrating a role for IFN-gamma-producing DC from Peyer's patches in the development of Ag-specific IEL population and their trafficking to the gut epithelium.     

Inhibitory effect of dietary n-3 polyunsaturated fatty acids to intestinal IL-15 expression is associated with reduction of TCRalphabeta+CD8alpha+CD8beta-intestinal intraepithelial lymphocytes

Intestinal intraepithelial lymphocytes (IELs) and their cytokines play an important role in the regulation of gut immune response and take part in gut immune barrier function. n-3 polyunsaturated fatty acid (PUFA) is an immunoregulator that has been shown to influence the process of gut inflammation. Interleukin (IL)-15 is a T-cell growth factor that has been shown to influence the differentiation of IEL. The aim of this study was to analyze the effects of dietary n-3 PUFA on IEL. IEL phenotype and cytokine (TNF-alpha, IFN-gamma, IL-4, IL-10 and TGF-beta1) profile were measured by FACS and real-time RT-PCR in healthy adult rats fed with fish oil diet for 90 days. Rats fed with corn oil diet served as controls. Intestinal IL-15 expression was measured by immunohistochemistry and real-time RT-PCR. The results demonstrated a decrease of intestinal IL-15 expression in the fish oil group. Associated with this deduction, n-3 PUFA significantly decreased the proportion of TCRalphabeta+CD8alpha+CD8beta- cells and IEL-derived TNF-alpha, IFN-gamma, IL-4 and IL-10. In conclusion, n-3 PUFA could inhibit intestinal mucosal expression of IL-15 and may influence phenotype and function of IEL through this mechanism.     

Reovirus serotype 1/strain Lang-stimulated activation of antigen-specific T lymphocytes in Peyer's patches and distal gut-mucosal sites: activation status and cytotoxic mechanisms

Intraduodenal priming of mice with reovirus serotype 1/strain Lang (reovirus 1/L) stimulates gut lymphocytes and generates precursor and effector CTLs. Our earlier studies demonstrated that germinal center and T cell Ag (GCT) is a marker which identifies reovirus 1/L-specific precursor CTL and effector CTL in Peyer's patches (PP) of reovirus 1/L-inoculated mice. In this study, we characterized the expression of the activation markers, GCT and CD11c, on reovirus 1/L-stimulated gut lymphocytes and the effector mechanisms involved in reovirus 1/L-specific cytotoxicity. We found that intraduodenal reovirus 1/L inoculation of mice induced the expression of both GCT and CD11c on PP lymphocytes (PPL), intraepithelial lymphocytes (IEL), and lamina propria lymphocytes (LPL), and these activated cells expressed Fas ligand (FasL). The majority of the GCT+ CD11c+ IEL and LPL expressed a phenotype, TCRalphabeta+ Thy-1+ CD8+ similar to that expressed on reovirus 1/L-stimulated PPL. However, splenic lymphocytes expressed GCT but not CD11c after stimulation with reovirus 1/L. Perforin, Fas-FasL, and TRAIL pathways were found to be involved in PPL, IEL, and LPL cytotoxic activity against reovirus 1/L-infected targets. In PPL, perforin and Fas-FasL pathways were more effective than TRAIL. In IEL, all three cytotoxic mechanisms were equally as effective. However, LPL prefer Fas-FasL and TRAIL over perforin. Further, we demonstrated the preferential migration of GCT+ PPL to the intraepithelial compartment and the lamina propria. These results suggest that GCT and CD11c can be used as activation markers for gut lymphocytes and CD11c can also be used to differentiate between activated gut and systemic lymphocytes.     

Changes in antigen distribution and lymphocyte migration pattern after peroral immunization

The migration pattern of lymphoid cells in long-term p.o. immunized and control mice using the transfer of 51Cr-labelled cells from spleen, Peyer's patches and mesenteric or peripheral lymph nodes was studied. There are no differences between the homing activity of spleen of PLN cells to different organs of recipient animals. Peyer's patch cells from SRBC-fed mice home significantly more to the gut of antigen-fed mice; also the MLN cells from these mice exhibit a higher localization in the gut of SRBC-fed mice. There were no differences between the localization of antigen (SRBC) in different organs of SRBC-fed and control mice. The clearance of this antigen was higher in SRBC-fed animals.     

Multipotential acceptance of Peyer's patches in the intestine for both thymus-derived T cells and extrathymic T cells in mice

Peyer's patches (PP) are important inductive sites for the mucosal immune response. It is well known that lymphocytes that migrate into PP are mainly of T-cell lineage from thymus-derived cells (i.e. alphabetaTCR(high) cells). In this study, we further characterized the properties of PP lymphocytes in mice using a mouse model of colitis induced by dextran sulphate sodium (DSS). Although the major site of the inflammation induced by DSS is known to be the large intestine, the small intestine was also damaged. When mice developed DSS-induced colitis, CD3+CD8+B220+ gammadelta T cells increased in PP in the small intestine. These gammadelta T cells, which are not seen in the PP of normal mice, resembled intraepithelial lymphocytes (IEL) in the small intestine in terms of their expression of CD5, CD103 and Thy1.2. In addition, the Vgamma/delta repertoire of these gammadelta T cells was similar to that of gammadelta IEL. When DSS-treated mice were injected with IEL isolated from normal mice, IEL including gammadelta T cells preferentially migrated to PP, raising the possibility that B220+ T cells seen in PP of diseased mice may derive from IEL in the small intestine. Our present study suggests that PP might be able to accept T-cell lineages from intestinal IEL as well as from thymus-derived T cells.     

MHC class I allele dosage alters CD8 expression by intestinal intraepithelial lymphocytes

The development of TCR alphabeta(+), CD8alphabeta(+) intestinal intraepithelial lymphocytes (IEL) is dependent on MHC class I molecules expressed in the thymus, while some CD8alphaalpha(+) IEL may arise independently of MHC class I. We examined the influence of MHC I allele dosage on the development CD8(+) T cells in RAG 2(-/-) mice expressing the H-2D(b)-restricted transgenic TCR specific for the male, Smcy-derived H-Y Ag (H-Y TCR). IEL in male mice heterozygous for the restricting (H-2D(b)) and nonrestricting (H-2D(d)) MHC class I alleles (MHC F(1)) were composed of a mixture of CD8alphabeta(+) and CD8alphaalpha(+) T cells, while T cells in the spleen were mostly CD8alphabeta(+). This was unlike IEL in male mice homozygous for H-2D(b), which had predominantly CD8alphaalpha(+) IEL and few mostly CD8(-) T cells in the spleen. Our results demonstrate that deletion of CD8alphabeta(+) cells in H-Y TCR male mice is dependent on two copies of H-2D(b), whereas the generation of CD8alphaalpha(+) IEL requires only one copy. The existence of CD8alphabeta(+) and CD8alphaalpha(+) IEL in MHC F(1) mice suggests that their generation is not mutually exclusive in cells with identical TCR. Furthermore, our data imply that the level of the restricting MHC class I allele determines a threshold for conventional CD8alphabeta(+) T cell selection in the thymus of H-Y TCR-transgenic mice, whereas the development of CD8alphaalpha(+) IEL is dependent on, but less sensitive to, this MHC class I allele.     

Inhibition of in vivo lymphocyte migration to inflammation and homing to lymphoid tissues by the TA-2 monoclonal antibody. A likely role for VLA-4 in vivo.

The adhesion receptors, LFA-1 and VLA-4, on lymphocytes mediate lymphocyte adherence to cytokine-activated endothelial cells (EC) in vitro. Based on our previous data, which suggested that the mAb TA-2 reacted with rat VLA-4, the effect of TA-2 on lymphocyte migration out of the blood was examined. Small peritoneal exudate lymphocytes (sPEL) preferentially migrate to cutaneous inflammatory reactions, whereas lymphocytes from peripheral lymph nodes (PLN) migrate poorly to inflammatory sites but home avidly to PLN. Treatment of sPEL with TA-2 inhibited sPEL migration to DTH, LPS, poly I:C, IFN-gamma, IFN-alpha/beta, and TNF-alpha by 35 to 65% and their accumulation in PLN by 50%. The homing of PLN lymphocytes to PLN was not inhibited by TA-2. Spleen T cell migration to cutaneous inflammatory sites was inhibited but homing to PLN was not affected. Systemic treatment with TA-2 inhibited sPEL migration to inflamed or cytokine-injected skin by up to 70%. Similarly, TA-2 strongly inhibited the migration of Ag-stimulated PLN lymphoblasts to skin and to PLN. The migration of lymphocytes from all sources, including the peritoneum, spleen, PLN, mesenteric nodes, and Peyer's patches, to mesenteric lymph nodes and Peyer's patches was inhibited by 80% and 95%, respectively. In conclusion, our results suggest that VLA-4 and possibly other alpha 4 integrins mediate the migration of the inflammation-seeking sPEL and Ag-activated lymphoblasts to cutaneous inflammatory sites and lymph nodes but do not affect the homing of PLN lymphocytes to PLN. These integrins also appear to be necessary for the migration of all types of lymphocytes to Peyer's patches and mesenteric lymph nodes.     

Resistance to viral infection by intraepithelial lymphocytes in HIV-1 P18-I10-specific T-cell receptor transgenic mice

For the analysis of mucosal immunity to HIV-1, we have recently established a line of transgenic (Tg) mice expressing the TCRalpha and TCRbeta genes of the murine CTL clone RT1 specific for P18-I10 (RGPGRAFVTI), an immunodominant gp160 envelope-derived epitope of IIIB isolate, restricted by the H-2D(d) MHC-I molecule. Here we examine those cells bearing specific TCR among the intraepithelial lymphocytes (IELs), with flow cytometric analysis using H-2D(d)/P18-I10 tetramers. We observed three distinct CD3(+), tetramer positive populations among the IELs: extra-thymic CD8alphabeta(+), alphabetaTCR T-cells; CD8 alphaalpha+, gammadeltaTCR T-cells; and thymus-derived CD8alphabeta+, alphabetaTCR T-cells. Challenge of these Tg mice with P18-I10 encoded by a vaccinia virus vector, either intrarectally (i.r.) or intraperitoneally (i.p.), revealed that the intraepithelial compartment seems to be a major site for prevention of the spread of viral infection. Such immunity appears due to the thymus-derived, CD8alphabeta+ antigen-specific CTLs together with CD8alphaalpha+ gammadelta cells, which regulate virus spread. This model system for studying CTL based immunity at mucosal sites should prove helpful in developing rational approaches for HIV control.