Host MHC Class Research Articles

Role of tripeptidyl peptidase II in the processing of Listeria monocytogenes-derived MHC class I-presented antigenic peptides

The effective control of the infection of mice with the facultatively intracellular bacterium Listeria monocytogenes requires CD8 T cells which recognize bacterial antigenic peptides presented in the context of host MHC class I molecules. It is generally accepted that bacterial antigens are processed by the proteasome, a proteolytic cytoplasmic multiprotein complex. We observed that presentation of the L. monocytogenes-derived CD8 T cell epitope LLO 91–99 by infected cells can not be totally suppressed by inhibitors of the proteasome alone. Further analysis revealed that inhibitors of the cytoplasmic tripeptidyl peptidase II suppressed the presentation of the epitopes LLO 91–99 and p60 449–457. While significant suppression of the presentation of LLO 91–99 required the simultaneous inhibition of the proteasome and tripeptidyl peptidase II, presentation of p60 449–457 was suppressed by inhibitors of either the proteasome or TPPII alone. Thus, these data indicate that both, the proteasome and tripeptidyl protease II play a role in the processing of L. monocytogenes-derived antigenic peptides.

MHC haplotypes modulate strength of natural killer (NK) cell licensing (91.10)

Abstract NK cells react to cells that lack self-MHC class I. Yet, since many NK cells cannot recognize self-MHC, mechanisms such as NK cell licensing protect against autoreactivity. To become licensed, i.e. functionally competent to be triggered through its activation receptors, an NK cell must engage host MHC class I via at least one of its MHC class I-specific inhibitory receptors, such as the Ly49 family of receptors in the mouse. However, the general determinants of this process remain largely unknown. Herein, we investigated the licensing impact of the b, d, f, k, q, r, and s H2 haplotypes on Ly49A+ NK cells in MHC-congenic mice. Ex vivo PK136 (anti-NK1.1) stimulation assays indicated that licensing may not be a binary phenomenon as some Ly49A-MHC class I haplotype combinations produced an intermediate licensing phenotype. Ly49A surface accessibility, a measure of cis binding with MHC class I, and Ly49A tetramer binding displayed a similar variability among the MHC haplotypes. Taken together, the data indicate a correlation between strength of licensing and strength of binding of Ly49A receptors with MHC class I, both of which are inversely correlated with surface accessibility of Ly49A receptors. These findings suggest that NK cell licensing is modulated by the strength of the interaction of inhibitory receptors with host MHC class I. Supported by HHMI and grants from the NIH.

A Single CD8+ T Cell Epitope Sets the Long-Term Latent Load of a Murid Herpesvirus

The pathogenesis of persistent viral infections depends critically on long-term viral loads. Yet what determines these loads is largely unknown. Here, we show that a single CD8+ T cell epitope sets the long-term latent load of a lymphotropic gamma-herpesvirus, Murid herpesvirus-4 (MuHV-4). The MuHV-4 M2 latency gene contains an H2-Kd -restricted T cell epitope, and wild-type but not M2− MuHV-4 was limited to very low level persistence in H2d mice. Mutating the epitope anchor residues increased viral loads and re-introducing the epitope reduced them again. Like the Kaposi's sarcoma–associated herpesvirus K1, M2 shows a high frequency of non-synonymous mutations, suggesting that it has been selected for epitope loss. In vivo competition experiments demonstrated directly that epitope presentation has a major impact on viral fitness. Thus, host MHC class I and viral epitope expression interact to set the long-term virus load.

Host MHC class II+ antigen-presenting cells and CD4 cells are required for CD8-mediated graft-versus-leukemia responses following delayed donor leukocyte infusions

AbstractFollowing bone marrow transplantation, delayed donor leukocyte infusions (DLIs) can induce graft-versus-leukemia (GVL) effects without graft-versus-host disease (GVHD). These antitumor responses are maximized by the presence of host hematopoietic antigen-presenting cells (APCs) at the time of DLI. Using a tumor-protection model, we demonstrate here that GVL activity following administration of DLIs to established mixed chimeras is dependent primarily on reactivity to allogeneic MHC antigens rather than minor histocompatibility or tumor-associated antigens. CD8+ T-cell–dependent GVL responses against an MHC class II–negative tumor following delayed DLI require CD4+ T-cell help and are reduced significantly when host APCs lack MHC class II expression. CD4+ T cells primed by host APCs were required for maximal expansion of graft-versus-host reactive CD8+ T cells but not their synthesis of IFN-γ. In contrast, the GVL requirement for CD4+ T-cell help was bypassed almost completely when DLI was administered to freshly irradiated recipients, indicating that the host environment is a major factor influencing the cellular mechanisms of GVL.

Streptococcus pyogenes: Insight into the function of the streptococcal superantigens

The group A streptococcus produces a number of highly potent exoproteins that act as superantigens. The cascade of pro-inflammatory events that follow invasive streptococcal infection is greatly enhanced by production of such toxins, leading to profound hypotension and multi-organ failure in some cases. Superantigens such as streptococcal mitogenic exotoxin Z (SMEZ) interact with host MHC class II and the T cell receptor, leading to activation events in both cells. In vitro, these interactions lead to expansion and cytokine production by specified T cell subsets. Studies using humanised HLA class II transgenic mice and isogenic streptococcal strains have characterised the in vivo responses to superantigens produced in the context of live infection. Notwithstanding the obvious deleterious role of superantigens in toxic shock, the evolutionary advantage conferred by these toxins remains a subject of speculation.

88: ALFA-1 induced cardiac allograft survival: Tolerance induction requires host MHC class I expression

The reliable induction of tolerance to transplanted organs is a major goal in transplantation research to obviate chronic anti-rejection therapy. Lymphocyte function-associated antigen-1 (LFA-1) has been identified as one likely component of the T-cell activation process that may be interrupted to lead to graft tolerance without the negative side effects of non-specific immunosuppression. NKT-cells restricted by non-classical class I CD1 molecule have been implicated in peripheral tolerance induction.

CD4+CD25+ T Cells Can Inhibit CD8 T Cell Mediated GVHD: Requirement for In Vivo Recognition of Allogeneic Host MHC Class II Antigens.

CD4 regulatory T (Treg) cells have shown promise in the transplantation mileu including the ability to inhibit the development of graft vs host disease (GVHD) following allogeneic hematopoietic stem cell transplants (HCT). The antigen specificity of the Treg population(s) involved is not yet clear nor is the role of their activation following transplant. We are interested in determining the requirement for recognition of host MHC antigens following infusion of CD4+CD25+ T cells in an experimental model of GVHD. To clearly distinguish the requirements of regulatory vs GVH reactive cells, a model of CD8 T cell mediated GVHD was developed using highly purified BALB/c (H2d) donor CD8+ T cells (Miltenyi column, 95-98%). CD8 T cells were transplanted together with T cell depleted (TCD) BALB/c BMC into 12.0 GY (6.0 Gy split dose) TBI conditioned C57BL/6 (B6, H2b) recipients. To support development of GVHD by these cells, resistance was inhibited by treatment of recipients with anti-NK1.1mab (PK136) at Days -1, 0 and +7. BALB/c CD8+ T cells at doses of 5.0x106 but not 2.5x106 induced weight loss and some lethality in B6 recipients. 5x106 CD8+ T cells were then transplanted into B6-MHC class II−/ − recipients. GVHD symptoms including weight loss and lethality were readily apparent in these mice post-transplant. Interestingly, GVHD was consistently more severe with respect to the induction of weight loss and lethality in MHC Class II−/ − vs B6-wt recipients. Highly enriched BALB/c CD4+CD25+ T cells (> 95%) were produced from spleen and lymph node cells following negative (B-cells, CD8 and NK) and positive (CD25) selection using Miltenyi magnetic bead columns. Co-transplant of 1x106 CD4+CD25+ T cells together with BALB/c CD8+ T cells into B6 recipients inhibited GVHD as assessed by the absence of weight loss and lethality compared to B6 recipients of CD8+ T cells alone. In contrast, BALB/c CD4+CD25+ T cells failed to protect B6-MHC class II−/ − recipients from severe CD8+ T cell mediated GVHD. These findings demonstrate that donor CD4+ T regulatory cells can suppress GVHD inducing CD8+ T cells after the former recognize host class II alloantigen following transplant. We hypothesize that activated CD4+CD25+ T regulatory cells inhibit GVH reactive T cells at the host APC interface. Future studies in this model can be designed to examine ex-vivo activated and expanded CD4+CD25+ T regulatory populations. Transplant of such cells will enable us to address questions regarding the importance of in vivo recognition of host class II in the regulation of GVHD by these cells.

MHC-Mismatched Islet Allografts Are Vulnerable to Autoimmune Recognition In Vivo

When transplanted into type 1a diabetic recipients, islet allografts are subject both to conventional allograft immunity and, presumably, to recurrent autoimmune (islet-specific) pathogenesis. Importantly, CD4 T cells play a central role both in islet allograft rejection and in autoimmune disease recurrence leading to the destruction of syngeneic islet transplants in diabetic NOD mice. However, it is unclear how NOD host MHC class II (I-A(g7))-restricted, autoreactive CD4 T cells may also contribute to the recognition of allogeneic islet grafts that express disparate MHC class II molecules. We hypothesized that islet-specific CD4 T cells can target MHC-mismatched islet allografts for destruction via the "indirect" (host APC-dependent) pathway of Ag recognition. To test this hypothesis, we determined whether NOD-derived, islet-specific CD4 T cells (BDC-2.5 TCR transgenic cells) could damage MHC-mismatched islets in vivo independent of conventional allograft immunity. Results demonstrate that BDC-2.5 CD4 T cells can vigorously destroy MHC class II-disparate islet allografts established in NOD.scid recipients. Tissue injury is tissue-specific in that BDC-2.5 T cells destroy donor-type islet, but not thyroid allografts established in the same NOD.scid recipient. Furthermore, BDC-2.5 CD4 T cells acutely destroy MHC class II-deficient islet allografts in vivo, indicating that autoimmune pathogenesis can be completely independent of donor MHC class II expression. Taken together, these findings indicate that MHC-mismatched islet allografts can be vulnerable to autoimmune pathogenesis triggered by autoreactive CD4 T cells, presumably through indirect autoantigen recognition in vivo.

MHC CLASS I ALLOCHIMERIC MOLECULE MODULATES ALLOANTIBODY RESPONSE AND UPREGULATES EXPRESSION OF PROTECTIVE GENES: INFLUENCE ON DEVELOPMENT OF GRAFT ACCOMMODATION

P1028 Aims: Specific transplantation tolerance can be induced in rat recipients treated during peri-transplant period with MHC class I antigens in which 4 donor a.a. are substituted onto host MHC class I sequences. The tolerance characterized with upregulation of cyto-protective and anti-apoptotic genes and accumulation of IgG type alloantibodies in the vessel endothelium. This study was designated to investigate possible influence of this therapy on development of graft accommodation through passive transfer of alloantibodies to a new cohort of recipients. Methods: 1 ml of serum from tolerant ACI rats after allochimeric protein therapy was injected through the portal vein to lightly irradiated (130 Rad) ACI recipients of WF hearts. Grafts were evaluated at 100 days. Results: Passive transfer of sera from rats conditioned with allochimeric therapy, but not from naive rats, to lightly irradiated syngeneic ACI recipients of WF hearts resulted in indefinite survival of cardiac allografts (MST 135+21.9, n=13). Histopathology examination revealed intact myocardial architecture, absent or minimal intramiocardial inflammation, and no evidence of neointimal hyperplasia. The degree of neointimal proliferation and TVS within each vessel was quantitated by the neointimal index (NI). Serum conditioned hosts demonstrated NI of 6.5 with 32% of vessels effected. In contrast, control group receiving low dose radiation in conjunction with naive ACI serum had NI of 39 with 70% of vessels affected. Upregulation of genes for Th2-type cytokines (IL-10 and IL-4) was found in tolerated grafts in comparison to control. FACScan analysis of serum from passive transferred tolerated ACI rats revealed low level of IgM alloantibodies. IgG production was markedly abrogated but noticeably higher than in naive control, IgG isotype analysis showed upregulation for IgG1 and IgG2a alloantibodies. Immunohistology staining of tolerant grafts with anti-IgM and IgG antibodies showed deposits of IgG granules around of internal intima when in control hearts after high doses of CsA such patterns were not revealed. To detect level of apoptotic cells within myocardium of long term survived tolerant grafts in compare to chronic rejected tissue in situ hybridization was performed (TUNEL). Analysis showed decreased number of apoptotic cells in tolerant hosts as compared to controls (7.5+1.2; 13.4+ 2.5, per 5-6 fields, respectively). Conclusions: Allochimeric MHC class I therapy orchestrates the development of donor-specific alloantibodies than can be transferred to new recipient and might cause allograft vascular accommodation, characterized by inhibition of apoptotic cell death, upregulation of cyto-protective genes and IgG antibody binding to vascular endothelium.

A major role for host MHC class I antigen presentation for promoting islet allograft survival

The purpose of this study was to determine the role for CD8 T cells versus generalized MHC class I–restricted antigen presentation in islet allograft rejection and tolerance. Diabetic C57BI/6 (B6, H-2 b) controls, C57BI/6 CD8-deficient (CD8 KO), or MHC class I–deficient C57BI/6 (β2m KO) recipients were grafted with allogeneic BALB/c (H-2 d) islets. Islet allografts were acutely rejected in untreated B6, CD8 KO, and in β2m KO mice, indicating that neither CD8 T cells nor host MHC class I is required for allograft rejection. We then determined the efficacy of costimulation blockade in these same strains. Costimulation blockade with anti-CD154 therapy facilitated long-term islet allograft survival in both B6 and in CD8 KO recipients. However, anti-CD154 treated β2m KO recipients were completely refractory to anti-CD154 therapy; all treated animals acutely rejected islet allografts with or without therapy. Also, anti-NK1.1 treatment of wild-type B6 mice abrogated graft prolongation following anti-CD154 therapy. Taken together, results show a dramatic distinction between two forms of MHC class I–restricted pathways in allograft prolongation. Although anti-CD154-induced allograft survival was CD8 T-cell independent, an intact host MHC class I-restricted (β2m-dependent) pathway is nevertheless necessary for allograft survival. This pathway required NK1.1+ cells, implicating NK and/or NKT cells in promoting allograft prolongation in vivo.

Death of peripheral CD8+ T cells in the absence of MHC class I is Fas-dependent and not blocked by Bcl-xL.

Productive immune responses require an appropriate environment to support peripheral CD8(+) T cell survival. Although host MHC class I molecules appear to be required for this process, the cellular and molecular requirements have not been comprehensively studied. Using adoptive transfer of 2C/recombinase-activating gene-2 (RAG-2)(-/-) TCR-transgenic T cells, we found that the survival of both naive and effector CD8(+) T cells was dependent upon host expression of the same MHC class I alleles that supported thymic selection. Expression of appropriate MHC class Iby either bone marrow- or non-bone-marrow-derived cells was sufficient, suggesting that professional antigen-presenting cells were not mandatory. In contrast to MHC class I, neither T cell expression of CD28 nor host expression of ICAM-1 was required for peripheral T cell survival. Finally, T cell death in the absence of appropriate host MHC class I was overcome by elimination of Fas signaling but not by overexpression of Bcl-x(L) by CD8(+) T cells. These results suggest that, in the absence of a survival signal provided by engagement of host MHC/self peptide complexes, CD8(+) T cells die via a Fas-dependent, mitochondria-independent pathway.

A Major Role for Host MHC Class I Antigen Presentation for Promoting Islet Allograft Survival.

Barbara Davis Center, Departments of Medicine and Immunology, University of Colorado Health Sciences Center, Denver, Colorado, USA.

Acquisition of MHC-specific receptors on murine natural killer cells.

NK cells in adult mice express two families of MHC class I-specific receptors, namely, Ly49 and CD94/NKG2. Co-expression of these receptors in various combinations generates diverse receptor repertoires. The expression of individual receptors is mostly stochastic and independent of each other. NK cells acquire the receptors as they develop from progenitors in the bone marrow in adult mice. In vivo as well as in vitro studies have shown that the acquisition of the receptors is ordered and regulated by the host MHC class I. Developing NK cells first acquire CD94/NKG2 and subsequently various Ly49 receptors in an ordered manner. Unlike adult NK cells, most fetal and neonatal NK cells express CD94/NKG2 but not Ly49. During the first several weeks after birth, NK cells expressing various Ly49 receptors slowly accumulate, while CD94/NKG2+ NK cells decrease to approximately 50% of the population. The acquisition of NK receptors following hematopoietic stem cell transplantation is also a slow and apparently preprogrammed process, mimicking the ontogeny, regardless of whether stem cells from fetal liver or adult bone marrow are used as donors. The regulation of the transcription of individual receptor genes is rather complex, since two promoters have been identified for the genes encoding Ly49 and CD94.

Inhibition Of Natural, Interleukin-2 Stimulated and Bacillus Calmette-Guerin Enhanced Cytotoxicity With Anti-CD16 Antibodies

We studied the cytolytic mechanism of nonstimulated, bacillus Calmette-Guerin (BCG) stimulated and interleukin (IL)-2 (Chiron Corp., Amsterdam, The Netherlands) stimulated peripheral blood mononuclear cells. We inhibited the cytotoxicity of nonstimulated, BCG stimulated and IL-2 stimulated peripheral blood mononuclear cells against various target cells using 3 monoclonal antibodies directed against the CD16 receptor of natural killer cells or alternatively monoclonal antibodies against the alpha and beta subunits of the IL-2 receptor complex (IL-2R). The main target cell was the poorly differentiated transitional cell line T24. Of the 3 anti-CD16 antibodies tested only CLB FcR-gran/1 effectively inhibited natural, IL-2 stimulated and BCG enhanced cytotoxicity. Cytotoxicity was also markedly diminished after depletion of CD16+CD56+/- cells with CLB FcR-gran/1. An hour of pretreatment with CLB FcR-gran/1 was enough to reduce significantly the level of cytotoxicity evoked by overnight stimulation with BCG or IL-2. Simultaneous administration of anti-IL-2Ralpha and anti-IL-2Rbeta significantly decreased the killing of target cells by BCG stimulated and IL-2 stimulated peripheral blood mononuclear cells. Within the stimulation times chosen the same killing mechanisms seemed to explain the nonstimulated, BCG stimulated and IL-2 stimulated cytotoxicity with CD16 positive cells as central effectors. Anti-CD16 antibodies may deliver a target cell independent down-regulatory signal to natural killer cells or alternatively mimic a nonIg ligand and block the detection of the target cell.

A two-phase pathogenesis of graft-versus-host disease in mice.

Activation of donor T cells is required for the development of graft-versus-host disease (GVHD), a major complication of bone marrow transplantation. We investigated a murine model of GVHD across major and minor histocompatibility barriers. BALB/c recipients were lethally irradiated and transplanted with 10(7) bone marrow and 5 x 10(6) spleen cells from C57BL/6 donors. There were two separate phases of clinical disease. The first phase was most severe on day 7 after transplant. Weight and condition improved until day 12 and then a second phase of clinical GVHD commenced, which persisted until euthanasia. IL-2 mRNA expression, as a measure of T cell activation, was determined by quantitative PCR. The two phases of clinical GVHD were preceded by two separate peaks of IL-2 mRNA in the spleen. Host MHC class II(+) cells became undetectable by flow cytometry 7 days after transplantation, whereas donor MHC class II(+) cells increased during the first 9 days after transplantation. Removal of donor MHC class II(+) cells from the graft had no effect on the first phase. Possible roles for host and donor antigen-presenting cells (APC) in the two phases of the disease are discussed.

CD4 T cell–mediated cardiac allograft rejection requires donor but not host MHC class II

Numerous studies indicate that CD4 T cells are required for acute cardiac allograft rejection. However, the precise role for CD4 T cells in this response has remained ambiguous owing to the multipotential properties of this T-cell subpopulation. In the current study, we demonstrate the capacity of CD4 T cells to serve as direct effector cells of cardiac allograft rejection. We show that CD4 T cells are both necessary and sufficient for acute graft rejection, as indicated by adoptive transfer experiments in immune-deficient SCID and rag1(-/-) recipients. We have analyzed the contribution of direct (donor MHC class II restricted) and indirect (host MHC class II restricted) antigen recognition in CD4-mediated rejection. Acute CD4 T cell-mediated rejection required MHC class II expression by the allograft, indicating the importance of direct graft recognition. In contrast, reciprocal experiments indicate that CD4 T cells can acutely reject allogeneic cardiac allografts established in rag1(-/-) hosts that were also MHC class II deficient. This latter result indicates that indirect presentation of donor antigens by host MHC class II is not required for acute CD4-mediated rejection. Taken together, these results indicate that CD4 T cells can serve as effector cells for primary acute cardiac allograft rejection, predominantly via direct donor antigen recognition and independent of indirect reactivity.

Role of Qa-1(b)-binding receptors in the specificity of developing NK cells.

NK cells acquire the ability to recognize MHC class I molecules during development. Studies with Qa-1(b) tetramers (Qa-1 tetramers) showed that nearly all NK1.1(+) cells from newborn C57BL/6 mice express Qa-1-binding receptors. Cytotoxic activity of these cells is fully inhibited by Qa-1 ligands on target cells. In contrast, neither receptors for H-2K(b) nor H-2D(b) were observed on NK1.1(+) cells from newborn mice. After birth, frequencies of Qa-1 tetramer(+)/ NK1.1(+) cells gradually decrease as the number of Ly49(+) /NK1.1(+) cells increases. Cell transfer studies showed that Qa-1 tetramer(+) cells from newborn mice do not lose expression of Qa-1 receptors, but that they further acquire expression of Ly49 molecules. Acquisition of Qa-1-binding receptors appears largely independent of host MHC class I molecules, as observed in studies using beta2-microglobulin-deficient (beta2m(-/-)) mice as well as K(b)/ D(b-/-) and K(b)/D(b)/beta2m(-/-) mice. The present results suggest that Qa-1-binding receptors play an important role in the specificity of developing NK cells, and suggest that these cells rely mainly on inhibitory receptors specific for non-classical MHC class I molecules to maintain self tolerance during the first weeks of life.

The fate of donor T-cell receptor transgenic T cells with known host antigen specificity in a graft-versus-host disease model.

The expansion of transgenic donor CD8+ T-cells with known allospecificity against a host MHC class I alloantigen was examined in a murine graft-versus-host disease (GVHD) model. Lethally irradiated, Ld+ BALB/c mice received bone marrow cells and spleen cells from anti-Ld 2C T-cell receptor (TCR)-transgenic B6 mice, alone or with normal B6 spleen cells. Transgenic TCR-bearing T-cell expansion, apoptosis, and function were monitored at various time points and were correlated with clinical outcome. Fifteen-fold clonal expansion of 2C CD8 cells occurred by day 4 after bone marrow transplantation. Between days 4 and 7, increasing proportions of 2C CD8 cells underwent apoptotic cell death, coincident with a 7-15-fold decline in their numbers. CD8 and TCR expression were down-regulated on 2C CD8 cells by day +4 after bone marrow transplantation, and they were anergic to TCR-mediated stimulation. Clinically, the BALB/c recipients of 2C spleen cells exhibited only minimal chronic GVHD. In contrast, lethally irradiated BALB/c mice receiving similar numbers of non-transgenic B6 bone marrow cells and spleen cells exhibited severe GVHD (median survival time: 28 days). The addition of a small number of 2C spleen cells to the inoculum accelerated GVHD mortality, and 2C CD8 cells showed a similar time course of expansion and decline to that observed in recipients of larger numbers of 2C cells alone. Initial clonal expansion, down-regulation of CD8 and TCR, anergy, and later deletion of graft-versus-host-reactive CD8 cells via apoptosis occurs in lethally irradiated recipients. Expansion of a single CD8 clone produces much less severe GVHD than that induced by a polyclonal, mixed CD4 plus CD8 response. These results have implications for GVHD pathogenesis and its sometimes self-limited nature.

The MHC Class I Molecule H-2Dp Inhibits Murine NK Cells via the Inhibitory Receptor Ly49A

MHC class I molecules strongly influence the phenotype and function of mouse NK cells. NK cell-mediated lysis is prevented through the interaction of Ly49 receptors on the effector cell with appropriate MHC class I ligands on the target cell. In addition, host MHC class I molecules have been shown to modulate the in vivo expression of Ly49 receptors. We have previously reported that H-2Dd and H-2Dp MHC class I molecules are able to protect (at the target cell level) from NK cell-mediated lysis and alter the NK cell specificity (at the host level) in a similar manner, although the mechanism behind this was not clear. In this study, we demonstrate that the expression of both H-2Dd and H-2Dp class I molecules in target cells leads to inhibition of B6 (H-2b)-derived Ly49A+ NK cells. This inhibition could in both cases be reversed by anti-Ly49A Abs. Cellular conjugate assays showed that Ly49A-expressing cells indeed bind to cells expressing H-2Dp. The expression of Ly49A and Ly49G2 receptors on NK cells was down-regulated in H-2Dp-transgenic (B6DP) mice compared with nontransgenic B6 mice. However, B6DP mice expressed significantly higher levels of Ly49A compared with H-2Dd-transgenic (D8) mice. We propose that both H-2Dd and H-2Dp MHC class I molecules can act as ligands for Ly49A.

Tolerance to Antigen-Presenting Cell-Depleted Islet Allografts Is CD4 T Cell Dependent

Pretreatment of pancreatic islets in 95% oxygen culture depletes graft-associated APCs and leads to indefinite allograft acceptance in immunocompetent recipients. As such, the APC-depleted allograft represents a model of peripheral alloantigen presentation in the absence of donor-derived costimulation. Over time, a state of donor-specific tolerance develops in which recipients are resistant to donor APC-induced graft rejection. Thus, persistence of the graft is sufficient to induce tolerance independent of other immune interventions. Donor-specific tolerance could be adoptively transferred to immune-deficient SCID recipient mice transplanted with fresh immunogenic islet allografts, indicating that the original recipient was not simply "ignorant" of donor antigens. Interestingly, despite the fact that the original islet allograft presented only MHC class I alloantigens, CD8+ T cells obtained from tolerant animals readily collaborated with naive CD4+ T cells to reject donor-type islet grafts. Conversely, tolerant CD4+ T cells failed to collaborate effectively with naive CD8+ T cells for the rejection of donor-type grafts. In conclusion, the MHC class I+, II- islet allograft paradoxically leads to a change in the donor-reactive CD4 T cell subset and not in the CD8 subset. We hypothesize that the tolerant state is not due to direct class I alloantigen presentation to CD8 T cells but, rather, occurs via the indirect pathway of donor Ag presentation to CD4 T cells in the context of host MHC class II molecules.