Intrathymic Inoculation Research Articles

Intrathymic inoculation of donor antigen: an ineffective strategy for prolonging xenograft survival.

The aim of this study was to determine whether intrathymic inoculation of the recipient with donor antigen and short-term depletion of peripheral lymphocytes would lead to donor-specific unresponsiveness to rat pancreatic islet xenografts. The results were compared directly with an allograft model to determine whether there were substantial differences in the mechanisms of graft prolongation between allografts and xenografts using an identical conditioning regimen. Streptozotocin-induced diabetic C57B6 mice were injected with up to 0.3 ml of rat anti-mouse lymphocyte serum (ALS) 1 day before intrathymic injection of donor splenocytes. DA rat islet xenografts or Balb/c mouse islet allografts were transplanted 3 days later. ALS depleted CD3+ and CD4+ peripheral blood T lymphocytes to less than 5% of values in control mice by 24 h. Median islet xenograft survival (MGS) was 9 days in untreated mice, 28 days in mice receiving 0.2 ml of ALS and 32 days in mice receiving 0.3 ml of ALS alone. Intrathymic injection of 5 x 10(6) DA splenocytes plus 0.2 ml of ALS did not improve islet xenograft survival beyond 28 days. Increasing the intrathymic inoculum to 10(7) DA splenocytes with or without a higher dose of ALS (0.3 ml) did not increase MGS beyond 26 days, although 2 out of 18 animals survived beyond 100 days. These long-term surviving mice rejected a second DA rat islet graft in less than 22 days, indicating that tolerance was not achieved. To confirm the efficacy of this treatment regimen in allotransplantation, diabetic C57B6 mice received 10(7) Balb/c splenocytes intrathymically and 0.3 ml of ALS. A Balb/c islet allograft was performed 3 days later. Allograft survival was similar to that of rat islet xenografts with 40%, (4 out of 10) of grafts surviving beyond 100 days. In contrast to the xenograft recipients, a second Balb/c islet allograft survived indefinitely, indicating that tolerance was achieved. Histology of the long-surviving allografts showed intact islets with a sparse cellular infiltrate, whereas the long-surviving xenografts (> 100 days) showed a large cellular infiltrate and significant islet destruction. To investigate further the role of the thymus, adult thymectomized C57B6 mice were treated with 0.3 ml of ALS and received a DA rat islet xenograft. The median graft survival was 52 days and no graft survived beyond 80 days, suggesting that peripheral xenoreactive T cells remained after ALS treatment and greater T-cell depletion was necessary to obtain permanent engraftment. These results show that peripheral xenoreactive T cells which remain after profound T-cell depletion are capable of rejecting an islet xenograft despite intrathymic inoculation of donor antigen. The T-cell-mediated xenograft response appears to be stronger and more difficult to suppress than the allograft response using this strategy.

A clinical trial of intrathymic inoculation of donor bone marrow cells with concomitant pediatric heart transplantation: preliminary observations.

A clinical trial of intrathymic inoculation of donor bone marrow cells with concomitant pediatric heart transplantation: preliminary observations.

Intrathymic inoculation of donor B cells prolongs cardiac allograft survival by non-deletional mechanisms

Resting B (rB) cells and splenic dendritic cells (DCs) were purified from splenocytes as described. A typical preparation of rB cells as analysed by flow cytometry was found to be 95.7% major histocompatibility class (MHC) II, 1.2% CD80, 2.5% CD86, and 92.3% sIg. A typical preparation of splenic DCs was 92.2% MHC II, 68.8% CD80, 88.2% CD86, 68% N418, and 9.4% sIg. In functional assays, DCs but not rB cells were able to stimulate proliferation of allogeneic T cell and generate cytotoxic T cells. T cells were made from spleen using a nylon wool column as described and were sorted negatively by FACS Vantage with turbo sort option (Becton Dickinson, Mountain View, Calif). Typical preparations as analysed by FACS was found to be .90% CD3, ,0.5% IA, and ,1% Ig.

Induction of tolerance to an “untolerizable” strain by double-strain intrathymic inoculation in experimental rat cardiac transplantation

Induction of tolerance to an “untolerizable” strain by double-strain intrathymic inoculation in experimental rat cardiac transplantation

Immune nonresponsiveness to cardiac allografts by intrathymic inoculation of donor class I allopeptides is associated with high levels of transcripts for Th2 cytokines in the graft

Immune nonresponsiveness to cardiac allografts by intrathymic inoculation of donor class I allopeptides is associated with high levels of transcripts for Th2 cytokines in the graft

Simultaneous inoculation of donor lymphocyte subsets with transplantation in a rat cardiac allograft model

Intrathymic (IT) inoculation of soluble alloantigens (Ag) obtained from 3 M KCI extracts of resting T-cells induces donor-specific tolerance to cardiac allografts and islet allografts. This study examined the cellular basis of induction of transplantation tolerance by IT injection of soluble Ag. Our results show that while IT inoculation of 2 mg soluble donor Ag on Day -7 relative to Lewis islet transplantation induced specific unresponsiveness to islet allografts (>200 days) in naive diabetic recipients, IT inoculation of 2 mg soluble Ag on the same day as islet transplantation did not prolong islet allograft survival in the same Lewis-to-WF rat combination. To define the role of donor APCs in intrathymic tolerance, we showed that IT injection of an admixture of 1 × 104 donor DC and 2 mg soluble Ag caused acute islet graft rejection. In contrast, addition of 1 × 104 recipient-type DC to the IT inoculum did not prevent long-term graft survival. This finding suggests that while the presence of donor APCs in the inoculum does not appear neccessary for IT-alloantigen to induce peripheral tolerance, presentation of the soluble Ag in the thymus is dependent on host APCs. This conclusion is supported by our in vitro MLR experiments which showed that in vivo WF-Ag-primed Lewis T-cells proliferated specifically to WF-soluble Ag and that the response was enhanced 14-fold by the addition of responder-type DC. Addition of anti-Lewis MHC class II mAb specifically blocked the alloresponse, thus suggesting that in vivo Ag-primed T-cells are capable of recognizing and proliferating in response to allopeptides presented by responder APCs. We also showed that adoptive transfer of syngeneic naive T-cells into unresponsive recipients failed to break tolerance to long-term surviving islet allografts. This finding suggests that tolerance in this model is not due to a lack of T help. On the other hand, the adoptive transfer of spleen cells, but not sera, from the unresponsive WF recipients bearing long-term (> 120 days) functioning Lewis islets resulted in prolonged survival of donor-type but not third-party islet allografts in secondary syngeneic hosts. Our data suggest that the tolerogenic effect of IT inoculation of soluble Ag is dependent on the indirect pathway of Ag presentation and clonal deletion of alloreactive T-cells in the thymus, while suppresser/regulatory mechanism may be involved in the maintenance of peripheral tolerance.

Experimental Goodpasture's syndrome in Wistar-Kyoto rats immunized with α3 chain of type IV collagen

Glomerulonephritis and lung hemorrhage of autoimmune Goodpasture syndrome develop due to immune reactions against epitope(s) of the non-collagenous (NC1) domain of alpha3-chain of type IV collagen [alpha3(IV) NC1]. Whether thymic mechanisms have a role in the loss of tolerance to the Goodpasture epitope has not been established. We studied the renal and pulmonary effects of immunization with different forms (monomer, dimer, or hexamer) of alpha3(IV) NC1 collagen in Wistar-Kyoto (WKY) rats, and assessed whether the intrathymic inoculation of the antigen may protect against anti-GBM disease. WKY rats were immunized with bovine alpha3(IV) monomer, dimer, or hexamer, or with alpha3(IV) NC1 synthetic peptide. Renal function, kidney and lung immunohistology, and circulating and tissue bound antibodies to type IV collagen chains were analyzed. Effects of intrathymic inoculation of antigen on subsequent disease induction were analyzed in WKY rats given alpha3(IV) NC1 dimer or GBM preparation intrathymically 48 hours before immunization. Proteinuria, linear IgG deposition in GBM, and crescentic glomerulonephritis developed in WKY rats immunized with alpha3(IV) NC1 dimer or hexamer. Lesions were dose-dependent upon injections of 10 to 100 microgram dimer. The alpha3(IV) NC1 monomer induced less severe proteinuria and no crescents. Pulmonary hemorrhage was detectable in 35% of rats immunized with 25 to 100 microgram alpha3(IV) NC1 dimer; alpha3(IV) synthetic peptide (36 carboxyl terminal) did not induce disease. Rats injected intrathymically with up to 100 microgram alpha3(IV) NC1 dimer or with GBM 48 hours before immunization were not protected against subsequent development of proteinuria and glomerulonephritis. These findings document that glomerulonephritis and lung hemorrhage can be elicited in WKY rats by immunization with alpha3(IV) NC1. Failure of the intrathymic inoculation of antigen to prevent disease suggests that immunological tolerance cannot be achieved by this intervention, in contrast to other autoimmune conditions, and may imply independent roles for cellular and humoral nephritogenic pathways in anti-GBM nephritis.

Induction of long-term heart graft survival in HLA class I transgenic mice by intrathymic injection of HLA class I peptides

CD4+CD25+ regulatory T cells are selected in the thymus to control autoreactive thymic escapees preventing autoimmunity that cannot be achieved by negative selection or deletion alone, thus playing an important role in the maintenance of immunological homeostasis. Not only significant in preventing autoimmunity, CD4+CD25+ regulatory T cells have also been shown to be involved in allograft tolerance in organ transplantation. We have formerly introduced two lines of HLA class I transgenic mice to elucidate the role of HLA class I molecules in transplantation biology. Using a heterotopic cardiac transplantation model, we show herein that intrathymic inoculation of donor HLA class I–derived synthetic peptide results in the generation of CD4+CD25+ regulatory T cells, which induce graft specific tolerance without any preconditioning of the recipient or use of immunosuppressive drugs. This study provides evidence of the novel therapeutic potential of CD4+CD25+ regulatory T cells for clinical transplantation.

Intrathymic administration of B cells induces prolonged survival of fully allogeneic cardiac grafts without prolonged deletion of donor-specific thymocytes

Intrathymic (IT) injection of alloantigen has been shown to induce unresponsiveness to allografts although the exact mechanisms of tolerance induction remains unclear, C57BL/10 (H2 b) cardiac allografts were accepted in C3H/He (H2 k) mice pretreated with IT inoculation of donor splenocytes (1 × 10 6) in combination with a depleting anti-CD4 monoclonal antibody 27 days before cardiac transplantation. To investigate which cell types were responsible for tolerance induction by IT injection of alloantigen, resting B (rB) cells or dendritic cells were used as the thymic inoculum instead of whole splenocytes. IT injection of rB cells induced indefinite graft prolongation in all recipients while only 20%of mice that had received IT injection of dendritic cells accepted grafts for over 100 days. In contrast, IT injection of dendritic cells resulted in significant deletion of donor-specific thymocytes whereas rB cells were relatively ineffective. IT deletion is not essential for the induction of tolerance by IT injection of rB cells; nondeletional mechanisms can be involved.

Intrathymic Inoculation of Donor Bone Marrow at the Time of Transplantation Plus a Short Course of Tacrolimus Induce Long-Term Acceptance to Rat Lung Allografts

Intrathymic Inoculation of Donor Bone Marrow at the Time of Transplantation Plus a Short Course of Tacrolimus Induce Long-Term Acceptance to Rat Lung Allografts

NONRESPONSIVENESS TO CARDIAC ALLOGRAFTS INDUCED BY INTRATHYMIC INJECTION OF CLASS I ALLOPEPTIDES IS ASSOCIATED WITH LOCAL AND SYSTEMIC EXPRESSION OF TH2 CYTOKINES

487 Intrathymic inoculation of donor antigens results in the induction of tolerance to allografts. We have recently demonstrated that cardiac allograft rejection in the PVG.R8-to-PVG.1U rat strains combination that is disparate for an isolated class I (RT1.Aa) MHC molecule, involves the recognition of this antigen as peptides presented by the recipient MHC molecules. Three synthetic peptides corresponding to variable portions ofα-helices of the α-1 and α-2 domains of the RT1.Aa molecule serve as CD4+ T cell epitopes for graft rejection. Intrathymic inoculation of the recipient with a mixture of these peptides results in immune nonresponsiveness to allografts in the majority of graft recipients (>75%). To study the underlying mechanisms of immune nonresponsiveness in this model, we studied intragraft as well as systemic expression of cytokines and intragraft deposition of alloantibodies in long-term grafts (>185 days), grafts with delayed (35-82 days) and normal rate of rejection (6-7 days). All the rejecting grafts had deposition of IgM, IgG1, IgG2a, IgG2b, and IgG2c. In contrast, long-term grafts had moderate deposition of only Th2 associated IgG1 and IgG2a and totally lacked other antibody subclasses. Rejected grafts had high levels of intragraft TNF-α and IFN-γ and moderate levels of IL-4. In marked contrast, long-term grafts had high levels of IL-4 and moderate to undetectable levels of IFN-γ and TNF-α. We also detected high levels (<10 nM) of IL-10 in the sera of long-term graft recipients. There was a gradual increase in the serum level of IL-10 starting 7-12 days after transplantation, which reached a plateau after 120 days and then declined to background levels after 200 days post-transplantation in the majority of graft recipients. Animals with acute rejection did not have detectable levels of IL-10. Regardless of the status of the graft, no systemic expression of IL-2, IL-4, and IFN-γ was found in any of the graft recipients. Taken together, our data provide clear evidence for the predominance of the Th2 pathway in long-term graft recipients. This pathway may plan an active role in maintaining graft survival by blocking the effector mechanisms of graft destruction induced by the Th1 pathway. This study was supported in part by grants from National Institutes of Health (AI 33587) and American Heart Association (9650229N).

Tolerance induction by intrathymic inoculation prevents chronic renal allograft rejection.

These experiments investigated the ability of the donor-specific unresponsiveness created by the intrathymic inoculation of donor alloantigen to effectively prevent chronic rejection in an established rat model of chronic renal allograft rejection. Three study groups were examined: (1) Allograft controls--F-344 rats received a Lewis renal allograft plus 10 days of low-dose cyclosporine (CsA); (2) isograft controls--F-344 rats received an F-344 renal isograft and low-dose CsA; (3) experimental group--F-344 rats received a T-cell depleted syngeneic bone marrow transplant and intrathymic injection of Lewis bone marrow. Twenty-one days after bone marrow transplant, these animals received a Lewis renal allograft. Allograft controls demonstrated severe parenchymal fibrosis; isograft controls and intrathymic (IT) animals failed to develop this lesion. Immunohistochemical analysis revealed increased CD4+ T cells infiltrating the cortex of the allograft controls. Cytokine interferon-gamma and interleukin-2 transcripts were strongly positive in allograft controls and were absent from isograft controls and IT allografts as determined by reverse transcriptase-polymerase chain reaction. Analysis of tolerant grafts by flow microfluorimetry and genomic DNA amplification could not detect chimerism to a level of < 0.1%. IT inoculation of donor alloantigen can confer long-term unresponsiveness and prevent the development of the characteristic lesions of chronic rejection.

Induction of allograft nonresponsiveness after intrathymic inoculation with donor class I allopeptides. I. Correlation of graft survival with antidonor IgG antibody subclasses.

We have recently demonstrated that cardiac allograft rejection in the PVG.R8-to-PVG.1U rat strain combination involves the recognition of a isolated class I (RT1.Aa) molecules as peptides in the context of the recipient MHC molecules. Three synthetic peptides (P1, P2, and P3) corresponding to the alpha-helices of the RT1.Aa molecule served as T-cell epitopes for graft rejection. In this study, we demonstrate that two of these peptides (P2 and P3) are sufficient to induce immune nonresponsiveness (median survival time >237 days) to cardiac allografts when presented to the recipient immune system in the thymus 7 days before transplantation. This effect was time dependent, as intrathymic inoculation 60 days before transplantation did not prolong graft survival (median survival time=12 days). Previous studies have demonstrated a critical role for alloantibody responses in mediating graft rejection in this rat strain combination. We, therefore, studied the role alloantibody responses may play in the observed immune nonresponsiveness. The titers of alloantibody in serum samples harvested from graft recipients at different times after transplantation were measured. We used recipient primary aortic endothelial cells genetically manipulated to express the donor RT1.Aa molecule as targets in an enzyme-linked immunosorbent assay. High titers of anti-RT1.Aa IgM antibody were detected in unmanipulated controls at the time of graft rejection. The IgM antibody switched to high IgG titers in intrathymically inoculated rats with accelerated or delayed rejection. Graft rejection in intrathymically manipulated recipients that had achieved a transient state of immunological nonresponsiveness correlated with higher titers of the IgG2b alloantibody. In marked contrast, the long-term graft survivors expressed undetectable or low levels of the IgG2b antibody and moderate to high levels of the IgG1 and IgG2a subclasses. These data suggest that the IgG2b alloantibody may contribute to the rejection reaction, whereas IgG1 and IgG2a may be involved in active enhancement of graft survival.

Induction of allograft nonresponsiveness after intrathymic inoculation with donor class I allopeptides. II. Evidence for persistent chronic rejection despite high levels of donor microchimerism.

We have recently demonstrated that three synthetic peptides corresponding to the donor class I RT1.Aa molecule induce long-term survival of cardiac allografts in the PVG.R8-to-PVG.1U rat strain combination (disparate for one isolated class I, RT1.A, molecule) when presented to the recipient immune system in the thymus. Long-term graft survivors had measurable levels of donor-reactive alloantibodies in their serum. In this study, we examined long-term allografts for the presence of chronic rejection and donor microchimerism to assess whether this regimen of immune modulation establishes true tolerance and whether this tolerance is dependent upon the presence of donor-recipient microchimerism. Histological examination of long-term heart grafts (>100 days) demonstrated chronic rejection, including a mild degree of myocardial infiltration by mononuclear cells, mild to moderate myocardial fibrosis, and various vascular changes ranging from focal intimal thickening to total vascular lumen blockade due to smooth muscle cell proliferation. In contrast, long-term syngeneic hearts transplanted under similar experimental conditions lacked these pathological manifestations. Donor microchimerism was analyzed using the polymerase chain reaction with a pair of oligonucleotides specific for the donor class I RT1.Aa gene and genomic DNA harvested from various tissues from graft recipients. We detected high levels of donor microchimerism in the heart, kidney, liver, skin, bone marrow, thymus, and lymph nodes of long-term graft recipients. Donor microchimerism was also detected in unmanipulated control graft recipients at rejection (7 days) and in intrathymically manipulated recipients that rejected allografts in a delayed fashion (12-82 days). These data clearly demonstrate that intrathymic inoculation of donor class I allopeptides induces long-term graft survival but does not prevent chronic rejection. Allograft rejection occurred despite high levels of donor microchimerism, providing direct evidence that donor-recipient microchimerism is not sufficient for the prevention of acute or chronic rejection in this model.

Mechanisms of intrathymic tolerance induction to isolated rat hepatocyte allografts.

Intrathymic injection of alloantigen in young adult rats is capable of mediating long-lived transplantation tolerance. In this study, we use a well-defined model of isolated hepatocyte transplantation to define the mechanisms of intrathymic induced tolerance. The recipient rats are Nagase analbuminemic rats (NAR) that are deficient in albumin, to allow for following transplant acceptance using metabolic and genetic markers. Tolerance to allogeneic hepatocyte transplants could be mediated by intrathymic injection of live allogeneic splenocytes, lethally irradiated splenocytes, or isolated hepatocytes. Intrathymic injection of live allogeneic splenocytes, but not of hepatocytes or irradiated splenocytes, resulted in donor microchimerism in peripheral lymphoid organs, with preferential expansion of CD4-positive T cells in the recipient spleens. Tolerance could be adoptively transferred from tolerant animals to naive recipients, but only from those animals that had been inoculated with intrathymic donor splenocytes. We conclude that donor microchimerism is found after intrathymic inoculation of live splenocytes, but is not required for tolerance induction and that microchimerism is not an absolute requirement for the generation of regulatory cells.

Mechanisms of intrathymic tolerance induction to isolated rat hepatocyte allografts

Intrathymic injection of alloantigen in young adult rats is capable of mediating long-lived transplantation tolerance. In this study, we use a well-defined model of isolated hepatocyte transplantation to define the mechanisms of intrathymic induced tolerance. The recipient rats are Nagase analbuminemic rats (NAR) that are deficient in albumin, to allow for following transplant acceptance using metabolic and genetic markers. Tolerance to allogeneic hepatocyte transplants could be mediated by intrathymic injection of live allogeneic splenocytes, lethally irradiated splenocytes, or isolated hepatocytes. Intrathymic injection of live allogeneic splenocytes, but not of hepatocytes or irradiated splenocytes, resulted in donor microchimerism in peripheral lymphoid organs, with preferential expansion of CD4-positive T cells in the recipient spleens. Tolerance could be adoptively transferred from tolerant animals to naive recipients, but only from those animals that had been inoculated with intrathymic donor splenocytes. We conclude that donor microchimerism is found after intrathymic inoculation of live splenocytes, but is not required for tolerance induction and that microchimerism is not an absolute requirement for the generation of regulatory cells. (Hepatology 1997 Nov;26(5):1287-95)

Prophylactic intervention in RadLV-induced lymphomagenesis with an anti-IL-4 monoclonal antibody.

Intrathymic inoculation of the radiation leukemia virus (RadLV) into C57BL/6 mice induces thymic lymphomas after 4-6 months. During premalignant latency, a population of RadLV-infected prelymphoma (PL) cells (whose survival is dependent on autostimulation with IL-4) persists in the thymus. PL cells explanted from RadLV-inoculated mice can be propagated in cultures containing IL-4, and in vitro growth of PL cells is effectively inhibited by anti-IL-4 antibodies. We subjected RadLV-inoculated mice to prophylactic treatment with anti-IL-4 antibodies and a virus-specific immunotoxin (IT). Administration of IT delayed the onset of lymphoma but was not curative. Anti-IL-4 antibodies had a similar effect when administered at low doses. High doses of anti-IL-4, given 3-5 weeks after virus inoculation, provided complete protection against lymphoma. These results demonstrate the effectiveness of prophylactic intervention during premalignancy by using antagonists that restrain the growth of PL cells.

Localization of tolerogenic epitopes in the α1 helical region of the rat class I major histocompatibility complex molecule

Induction of tolerance to organ allografts remains an elusive goal in transplant biology. Tolerance toward heart allografts was induced by preoperative intrathymic (IT) inoculation of extracted soluble histocompatibility antigens (HAg) in combination with a single dose of anti-T-cell antibodies. 1 In contrast, peritransplant intravenous (iv) or intraportal administration of HAg combined with a short course of cyclosporine (CyA) prolonged allograft survival but did not induce transplantation tolerance. 2 IV injection of peptides corresponding to amino acid (AA) residues 75–84 of the human HLA-B7 protein combined with a short course of CyA before and after grafting induced donor-specific tolerance to heart allograft in rats. 3 The present experiments examined the localization of tolerogenic epitopes in rat class I major histocompatibility complex RTI.A u and RT1.A 1 molecules.

Long-term maintenance of donor-specific unresponsiveness in NIH miniature swine by intrathymic inoculation with allogeneic islets

Long-term maintenance of donor-specific unresponsiveness in NIH miniature swine by intrathymic inoculation with allogeneic islets

Induction of peripheral chimerism in an allogenic model of small bowel transplantation with intrathymic inoculation of donor-derived cells

A SELECTED population of patients with intestinal failure and unable to support the iatrogenic consequences of long-term parenteral nutrition would greatly benefit from intestinal transplantation. Despite preliminary trials to establish this technique, early graft loss and subsequent infections are very common, and the clinical result has been very deceptive.’ Better than aggressive, general immunosuppression therapies, the implementation of more selective, toleranceinducing, therapies also represents a current challenge in the field of clinical transplantation. Recently and in various models related to solid organ transplants and autoimmunity, the intrathymic (IT) injection of donor-derived cells/ Ags (or autologous targets in the case of autoimmunity) has been shown efficient in the induction of Ag-specific tolerance.2-4 The rational of this manipulation is based in the attempt to negatively select the newly emerging alloreactive T-cell specificities when confronted with their targets, as it happens normally inside the thymus.” We present some preliminary findings in a new experimental system of heterotopic intestinal grafts and IT injections of donor-derived cells (splenocytes + bone marrow cells) in inbred mouse strains. The experimental groups have been also submitted to several combinations of lethal X-radiation and syngenic bone marrow (BM) transplantation and to short courses of parenteral cyclosporine A (CyA). The evolution of grafted tissues, quantification of the degree of peripheral chimerism, and modification of allospecific T-cytotoxic lymphocytes have been evaluated during 3 to 4 month periods of follow-up. Very high percentages of donor MHC class I-positive cells are present in the periphery, but not inside the thymus, of the grafted mice for at least 2 to 3 months. Surprisingly, the presence of injected cells inside the thymus is undetectable after 1 week of the IT manipulation. Also, clonal deletion of recipient thymocytes reacting against donor-derived superantigens (Mls-la) is not observed. The vast majority of chimeric cells represent (Ag-presenting) dendritic cells, likely coming from both the grafted intestine and the IT inoculum. Allospecific T-cell cytotoxic reactivities are significantly reduced in the mice receiving IT injections. This suggests