Thymus Grafts Research Articles

Intrinsic and Microenvironmental Defects Are Involved in the Age-Related Changes of Lin−c-kit+Hematopoietic Progenitor Cells

The aim of this study was to evaluate through cross-transplantation models the effect of aging on the number of Lin(-)c-kit+ hematopoietic progenitor cells, on their ability to differentiate towards a lymphocyte phenotype, and on the role of the microenvironment in hematopoietic differentiation. The absolute number of purified Lin(-)c-kit+ cells from bone marrow was significantly lower in aged than in young mice. When transplanted in young recipients, Lin(-)c-kit+ hematopoietic progenitor cells from aged mice showed a reduced differentiation capacity in T cells and NK cells, compared to Lin(-)c-kit+ cells from young animals. The role of microenvironment in Lin(-)c-kit+ hematopoietic progenitor cells differentiation was evaluated by injecting young Lin(-)c-kit+ cells in young or aged recipients, the latter transplanted or not with a young thymus. In these conditions, the differentiation of Lin(-)c-kit+ cells from young mice in T and NK cells was less efficient in aged than in young recipients, independently of thymus grafting in aged recipients. In addition to quantitative defects qualitative alterations were also present in Lin(-)c-kit+ cells from aged mice, as evidenced by the fact that the injection of Lin(-)c-kit+ cells from aged donors in young recipients differentiated in CD4+ T cells that retained an interleukin-4 (IL-4) production in-between young and old control values. In conclusion, we have demonstrated that aging is associated with numerical and functional alterations of Lin(-)c-kit+ hematopoietic progenitor cells as well as with an altered microenvironment that is required for Lin(-)c-kit+ cells differentiation toward a lymphocyte phenotype.

Deletion of PKBα/Akt1 Affects Thymic Development

BackgroundThe thymus constitutes the primary lymphoid organ for the majority of T cells. The phosphatidyl-inositol 3 kinase (PI3K) signaling pathway is involved in lymphoid development. Defects in single components of this pathway prevent thymocytes from progressing beyond early T cell developmental stages. Protein kinase B (PKB) is the main effector of the PI3K pathway.Methodology/Principal FindingsTo determine whether PKB mediates PI3K signaling in the thymus, we characterized PKB knockout thymi. Our results reveal a significant thymic hypocellularity in PKBα −/− neonates and an accumulation of early thymocyte subsets in PKBα −/− adult mice. Using thymic grafting and fetal liver cell transfer experiments, the latter finding was specifically attributed to the lack of PKBα within the lymphoid component of the thymus. Microarray analyses show that the absence of PKBα in early thymocyte subsets modifies the expression of genes known to be involved in pre-TCR signaling, in T cell activation, and in the transduction of interferon-mediated signals.Conclusions/SignificanceThis report highlights the specific requirements of PKBα for thymic development and opens up new prospects as to the mechanism downstream of PKBα in early thymocytes.

Redefining epithelial progenitor potential in the developing thymus

Cortical and medullary epithelium represent specialised cell types that play key roles in thymocyte development, including positive and negative selection of the T cell repertoire. While recent evidence shows that these epithelial lineages share a common embryonic origin, the phenotype and possible persistence of such progenitor cells in the thymus at later stages of development remain controversial. Through use of a panel of reagents including the putative progenitor marker Mts24, we set out to redefine the stages in the development of thymic epithelium. In the early embryonic day (E)12 thymus anlagen we find that almost all epithelial cells are uniformly positive for Mts24 expression. In addition, while the thymus at later stages of development was found to contain distinct Mts24(+) and Mts24(-) epithelial subsets, thymus grafting experiments show that both Mts24(+) and Mts24(-) epithelial subsets share the ability to form organised cortical and medullary thymic microenvironments that support T cell development, a function shown previously to be lost in the Mts24(-) cells by E15 when lower cell doses were used. Our data help to clarify stages in thymic epithelial development and provide important information in relation to currently used markers of epithelial progenitors.

Cover Picture: Redefining epithelial progenitor potential in the developing thymus – Eur. J. Immunol. 9/2007

AbstractThe immunohistochemical analysis of a frozen section of a thymus graft is taken from the article by Rossi et al. (pp. 2411–2418). The image shows cytokeratin expression in green and Aire expression (medullary epithelial cells) in red. The graft shown is generated from the EpCAM1+Mts24+ thymic epithelial subset obtained at embryonic day 16 (BALB/c mice). In their study, Rossi et al. show that, in contrast to previous reports, Mts24 is not a progenitor marker for embryonic epithelial cells capable of creating the required thymic microenvironment for T cell development.

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Porcine Thymic Grafts Protect Human Thymocytes from HIV‐1–Induced Destruction

Human immunodeficiency virus type 1 (HIV-1) infection depletes thymocytes and destroys thymic structure. Functional, tolerant human T cells develop in vivo in immunodeficient mice receiving porcine thymus and human fetal liver fragments under the kidney capsule. In this model, we evaluated the potential of porcine thymus to protect human thymocytes from the effects of HIV-1. Compared with that observed in control mice with human thymic grafts, porcine thymus attenuated human thymocyte depletion by the CCR5-tropic isolate JR-CSF without preventing thymocyte infection. Porcine thymus protected human thymocytes from infection and depletion by a CXCR4-tropic HIV-1 isolate without reducing peripheral blood viral loads or T cell infection. Human thymocytes from human but not porcine grafts showed decreased Bcl-2 expression and increased apoptosis after NL4.3 infection. Thus, porcine thymus protects human thymocytes from the cytopathic effect of HIV-1, suggesting a possible approach to achieving immune restoration in patients with acquired immunodeficiency syndrome who have incomplete responses to antiretroviral therapy. The model allows analysis of the mechanisms of HIV-mediated thymic dysfunction.

Index of Suspicion in the Nursery

A 15-day-old girl has abnormally shaped low-set ears, left facial microsomia, and micrognathia suggestive of Goldenhar syndrome. Magnetic resonance imaging (MRI) of the brain reveals colpocephaly, membranous cleft palate, and fluid in the left middle ear cavity. Echocardiography documents normal heart anatomy and function. The infant is feeding well and has appropriate weight gain, but she has occasional decreases in oxygen saturation during feedings. Upper gastrointestinal radiographic study shows massive reflux, prompting the administration of ranitidine and positioning with her head elevated. Despite the reflux precautions, the baby continues to have episodes of low oxygen saturation while feeding, which now occur even at rest. The episodes last for less than 1 minute and are relieved by bag-and-mask ventilation. Twenty days after birth, she has a generalized clonic seizure. At this time, her glucose concentration is 67 mg/dL (3.7 mmol/L) and ionized calcium concentration is 2.48 mg/dL (0.62 mmol/L) on a capillary specimen, which is confirmed by a similarly low venous blood determination of total calcium of 4.3 mg/dL (1.08 mmol/L). She also develops a staphylococcal abscess in the left leg and left femoral vein thrombosis. Calcium gluconate infusion resulted in resolution of the seizure. However, calcium concentrations …

424: Anti-CD20 prevents alloantibody production and attenuates cardiac allograft vasculopathy in monkeys treated with anti-CD154

Purpose: Blockade of CD40-154 prolongs cardiac allograft survival in cynomolgus macaques, but cardiac allograft vasculopathy (CAV) causes graft failure, and is closely associated with appearance of anti-donor antibody (Ab). We hypothesized that depletion of CD20 B-cells might interfere with formation of anti-donor plasma cells and alloantibodies, and thus prevent CAV. Methods and Materials: Thirty MLR-mismatched heterotopic cardiac cynomolgus allograft recipients were treated with anti-CD154 high intensity monotherapy ( CD154; n 17, 6 with ATG) or CD154 with additional CD20 (20mg/kg q wk for 4 weeks: CD154 CD20; n 13, 11 with ATG). Some animals received an additional donor thymus graft or bone marrow infusion. Acute rejection was usually treated with steroids; graft survival was censored at 90 days. Results: 10 animals died with beating grafts, mainly with ATG-associated lung pathology, and are excluded from the survival analysis. Graft survival with CD154 CD20 (median 90d) and proportion of grafts surviving to 90 days (5/5) was significantly increased relative to CD154 (median 43d, IQR(25-75) 35-82d, p 0.007; 3/15 90d). With CD154, 15/15 (100%) developed anti-donor IgM and 14/15 (93%) IgG, usually weeks before graft failure, whereas no IgG was detected in samples analyzed to date from the CD154 CD20 group, and weak IgM was transiently detected in only one animal. All CD154 grafts had severe CAV (score 3 on 0-4 scale), in many instances with fibrosis and unscorable vessels due to infarction. In stark contrast, CD154 CD20 was associated with CAV 1 in 4 of 5 grafts, and CAV 2 in the 5, and preserved myocardial architecture. Conclusions: Using CD20 with CD154 is associated with decreased elaboration of alloAb, prolonged graft survival to 90 days, and significant attenuation of CAV. Concomitant thymus transplant or ATG induction did not appear to be necessary to this effect; ATG is associated with significant morbidity. These findings suggest that CD20 may reduce the incidence of CAV, perhaps by inhibiting alloAb.

Thymic rejuvenation and the induction of tolerance by adult thymic grafts

The thymus, the site of origin of T cell immunity, shapes the repertoire of T cell reactivity through positive selection of developing T cells and prevents autoimmunity through negative selection of autoreactive T cells. Previous studies have demonstrated an important role for the thymus not only in central deletional tolerance, but also in the induction of peripheral tolerance by vascularized renal allografts in juvenile miniature swine recipients. The same protocol did not induce tolerance in thymectomized recipients nor in recipients beyond the age of thymic involution. We subsequently reported that vascularized thymic lobe grafts from juvenile donors were capable of inducing tolerance in thymectomized juvenile hosts. However, the important question remained whether aged, involuted thymus could also induce tolerance if transplanted into thymectomized hosts, which, if true, would imply that thymic involution is not an intrinsic property of thymic tissue but is rather determined by host factors extrinsic to the thymus. We report here that aged, involuted thymus transplanted as a vascularized graft into juvenile recipients leads to rejuvenation of both thymic structure and function, suggesting that factors extrinsic to the thymus are capable of restoring juvenile thymic function to aged recipients. We show furthermore that rejuvenated aged thymus has the ability to induce transplant tolerance across class I MHC barriers. These findings indicate that it may be possible to manipulate thymic function in adults to induce transplantation tolerance after the age of thymic involution.

GalT knockout pig to baboon xenotransplantation

The success and widespread acceptance of clinical allotransplantation has led to a shortage of cadaveric organs and a resurgence of interest in xenotransplantation. Although swine are generally regarded as the preferred source of xenogeneic organs, the severe immunologic response to swine organs has so far precluded their clinical use. However, we have recently obtained pigs not expressing the alpha 1,3-galactosyltransferase gene (GalT-KO swine). We have reported initial results using GalT-KO swine as donors to baboons which demonstrated survival of a life-supporting renal graft for up to 83 days without rejection when combined with a regimen designed to induce tolerance by vascularized donor thymic grafts. Although further improvements will be needed to warrant a clinical trial of xenotransplantation, the initial results are encouraging. In addition to genetic modification of donor pigs, it is hoped that our tolerance-inducing regimen will allow successful clinical xenotransplantation by overcoming additional xenogeneic immunological barriers.

Parathyroid Gland Transplantation without Posttransplantation Immunosuppression

RATIONALE: Determine if combined thymus and parathyroid gland transplantation in a patient with DiGeorge syndrome will result in function of both organs. METHODS: The patient had both athymia with a total T cell count of 29 cells/μL and profound hypoparathyroidism with a parathyroid hormone level below the limit of detection. Allogeneic postnatal cultured thymus and a maternal parathyroid gland were transplanted after conditioning with 3 doses of rabbit antithymocyte globulin. The mother, thymus donor, and patient had partial HLA matching. RESULTS: The parathyroid hormone level was normal when first tested on day +17 and has since remained normal. The patient weaned off all calcium supplementation. On day +139, she developed a bacterial infection of her central line, during which she maintained ionized calcium levels above 1.10 mmol/L without the requirement for calcium supplementation. At day +213, her ionized calcium levels remain within normal limits (1.24 mmol/L). Thymic function has been excellent. A thymus graft biopsy performed on day +80 showed evidence of thymopoiesis. Naïve T cells were detectable in the blood on day +86, with progressively increasing numbers over time (naïve CD4+ 114 cells/μL and naïve CD8+ 42 cells/μL on day +202). The total T cell count increased over time (258 cells/μL on day +202), as well. Normal PHA responses were seen on days +157 and +202. The patient did not require any posttransplantation immunosuppression. CONCLUSIONS: Parathyroid transplantation, when combined with thymus transplantation in this patient with complete DiGeorge syndrome, led to parathyroid function without posttransplantation immunosuppression.

Adult Human Hematopoietic Cells Differentiate into Mature T Cells Via a CD3-4+8- Intermediate within the Mouse Thymic Microenvironment; a New Model System for the Study of Human Thymocyte Development.

Normal human T lymphocyte differentiation occurs within the thymus. We previously determined that human thymocyte precursors develop via a novel CD3-4+8- intermediate population, utilizing SCID-hu thymic grafts (Kraft, Weissman &Waller, JEM). We have recently worked to develop a more convenient and robust model of human hematopoiesis utilizing RAG2/Common gamma chain double knock out mice (RAG2 DKO) transplanted with hematopoietic stem and progenitor cells from adult human donors.Methods: Human peripheral blood derived CD34+ cells were obtained to >90% purity by magnetic bead positive selection following apheresis from healthy GCSF mobilized donors. 200,000-800,000 human CD34+ cells were injected intrahepatically into 0–2 day old RAG2/CG DKO pups following 2Gy x 2 of irradiation. At serial time points following intrahepatic transplantation, human CD45+ chimerism and donor derived phenotype was measured within the bone marrow, peripheral blood, spleen, liver, lymph node and thymus by flow cytometry. Thymuses were further analyzed utilizing antibodies to human CD3, CD4 and CD8. Results: Robust human thymopoeisis was observed in CD34+ transplanted mice. The degree of human engraftment increased with the number of CD34+ cells transplanted. Overall >70% of recipient mice successfully engrafted with >10% human CD45+ expression in analyzed tissues, with a mean of 63% of cells within the thymus being human derived. At earlier time points (4–6 weeks post transplant) the thymus of recipient mice were found to contain high fractions of immature CD45+ CD3-4-8- cells (making up 31–43% of human cells within the thymus) and the CD3-4+8- intermediate (22–30%) and CD4+8+ double positive (34–71%) populations with very rare mature CD3+4+8- or CD3+4-8+ T-cells (figure A). At later time points the fraction of immature CD3-4-8- and CD3-4+8- populations declined and increasing fractions of mature CD3+4+8- and CD3+4-8+ populations were identified (Figure B), in distributions similar to those found in normal human thymus.Conclusions: Human T-cells can differentiate within the mouse thymus derived from adult human CD34+ cells, and development appears to progress normally within a murine thymic microenvironment. The early developement of a CD3-4+8- intermediate suggests that T-cell development occurs via this population, unlike the CD3-4-8+ intermediate found in murine thymopoesis, suggesting that the pathway of human T-cell differentiation is intrinisic to the human thymocytes, and is independant of whether the thymic stroma is human (as found in SCID-hu) or murine (in Rag2 DKO). This robust model system enabling study of human thymopoesis utilizing hematopoietic stem cells from normal and diseased adults human donors may provide significant advantages for the study of human intrathymic T-cell differentiation and function in-vivo. [Display omitted]

Life-supporting pig-to-baboon renal transplantation using GalT knockout donors: Benefit of cotransplanting a vascularized donor thymic graft

The use of animal organs for human transplantation could potentially alleviate the worldwide critical shortage of donor organs, and thus has stimulated investigation of novel strategies directed at making xenotransplantation applicable to the clinic. The latter will likely require overcoming barriers of rejection through strategies utilizing either immune suppression or tolerance induction, or a combination of the two. We have previously reported that vascularized donor thymic tissue grafts, transplanted either as composite thymokidneys or vascularized thymic lobe grafts, induced tolerance across fully allogeneic barriers in miniature swine. Recently, we have applied the strategy of vascularized thymic lobe transplantation in a model of hDAF pig-to-baboon xenotransplantation and demonstrated that these grafts induced donor-specific unresponsiveness for up to 2 to 3 months following transplantation, at which time the grafts were lost from apparent humoral rejection. We have now extended these studies using cloned α-1,3-galactosyltransferase (GalT) knockout pigs as donors. We report here our initial experience in a preclinical model of pig-to-baboon xenotransplantation. This abstract presents clinical data on recipients of GalT knockout kidneys, especially focusing on four long-term recipients of GalT knockout kidney plus thymus transplantation in detail. Briefly, these four baboons receiving either xeno-thymokidneys or vascularized thymic lobe plus kidney maintained longer than 2 months. Three of four long-term survivors maintained normal plasma creatinine levels for 56, 68, and 83 days, respectively, before expiring from unexpected causes (anesthetic complication during surgery to replace an infected intravenous catheter (day 68), and myocardial infarction, possibly drug-induced on days 56 and an arterial catheter trouble on day 83. Cytotoxic T-lymphocyte assays on day 78 in one long-term acceptor showed that the baboon maintained anti-allo cytotoxic T-lymphocyte responses but lost antipig cyctotoxic T-lymphocyte responses, indicating donor-specific unresponsiveness. The fourth baboon experienced an apparent rejection crisis between days 53 and 65, which reversed with anti-T-cell rejection therapy; subsequently the animal expired on day 81 from pneumonia. Neither hyperacute nor accelerated acute rejection was observed in any animal. Use of GalT knockout donors markedly extends the survival of vascularized thymus plus renal xenografts in baboons. Although the induction regimen still needs to be modified to reduce complications, these initial results are encouraging with regard to the potential of cotransplanting vascularized thymic tissue with organ xenografts as a means of achieving long-term tolerance across pig-to-primate barriers.

Life-supporting pig-to-baboon renal transplantation using GalT knockout donors: Benefit of cotransplanting a vascularized donor thymic graft

Life-supporting pig-to-baboon renal transplantation using GalT knockout donors: Benefit of cotransplanting a vascularized donor thymic graft

Heart and en-bloc thymus transplantation in miniature swine

Donor-specific tolerance to organ allografts might be induced by cotransplantation of a sufficient amount of vascularized donor thymus. To facilitate donor thymus-induced cardiac allograft tolerance, we have developed a novel technique for heart and en-bloc thymus transplantation in swine. Donor heart and en-bloc thymus grafts were prepared by a technique that preserves the entire arterial supply and venous drainage of the right thymic lobe. En-bloc grafts (n = 4) were transplanted heterotopically into the abdomens of major histocompatibility complex-matched miniature swine. Recipients received 12 days of cyclosporine intravenously. Grafts were monitored by palpation, electrocardiographic monitoring, and periodic open biopsy. Engraftment of the donor thymus was demonstrated by measuring the proportion of recipient-type thymocytes in the donor thymus with flow cytometry. All of the heart and en-bloc thymus grafts had normal cardiac contractility and immediate perfusion of the thymus. All en-bloc grafts were accepted for more than 200 days without significant acute cellular rejection or cardiac allograft vasculopathy. Thymic tissue of en-bloc grafts displayed normal architecture and supported thymopoiesis of recipient-type cells. We have validated a new technique of donor thymus transplantation that could have utility in human heart transplantation.

Aging reversibility: from thymus graft to vegetable extract treatment-- application to cure an age-associated pathology.

Neonatal thymus graft and thymus calf extract (TME) in vivo treatment exert similar corrective actions on different mouse age-related alterations. The aim of the present paper is to investigate whether a vegetal extract, wheat sprout extract (WESPRE), could mimic the thymus action on recovering age-related alterations and if this extract can cure an age-associated pathology, the cataract in dogs. Present experiments were carried out by using WESPRE and TME in vivo in old mice to check their ability to recover the altered DNA synthesis in hepatocyte primary cultures. Old mice treated with WESPRE and TME showed a recovery of hepatocyte DNA synthesis levels when compared with the old untreated ones. The increase of DNA and protein contents observed in aged animals is reduced by WESPRE treatments to levels observed in young mice hepatocytes. We measured also WESPRE phosphorylation activity by endogenous kinase: it was from 10 to 40 times higher with respect to wheat seeds. Old dogs were orally treated for a month and the lens opacity analysed before and after the treatment. Results showed a reduction from 25 to 40% of lens opacity. The efficacy of wheat sprouts in the recovery of age-related alterations and in treating age-associated pathologies could be due to the contemporary presence of small regulatory acid peptides, a remarkable level of highly energetic phosphoric radicals and antioxidant molecules, peculiarities that may be, to some extent, related to the aging process regulation.

Role of the Thymus and Kidney Graft in the Maintenance of Tolerance to Heart Grafts in Miniature Swine

The authors have examined the mechanism whereby co-transplantation of a kidney and heart from the same donor induces and maintains tolerance to both organs in miniature swine. Transplants were performed across a major histocompatibility complex class I mismatch, and recipients received cyclosporine for 12 days. Group 1 animals received heart transplants alone (n=5), and all other groups received both heart and kidney allografts. Group 2 animals received no further intervention (n=2). Group 3 animals underwent transplant nephrectomy 8 days after heart and kidney co-transplantation (n=2). Group 4 animals underwent transplant nephrectomy 100 days after co-transplantation (n=2). Skin grafts were placed on group 4 animals, on one group 3 animal, and on two animals from group 2. Group 5 animals underwent thymectomy 100 days after co-transplantation (n=4). Group 1 animals developed cardiac allograft vasculopathy (CAV) and rejection. Group 2 animals never developed CAV and demonstrated in vitro donor-specific unresponsiveness. Group 3 animals suffered CAV and rejection. Group 4 animals developed CAV without concomitant donor-specific cell-mediated lympholysis reactivity, interstitial rejection, or cessation of graft function. Skin grafts on group 3 and group 4 animals led to fulminant rejection of heart and skin grafts, in contrast to grafts on group 2 animals that had no in vivo effect. Group 5 animals developed CAV but no significant increase in interstitial infiltrates. Both the kidney and thymus were necessary for maintenance of tolerance to heart allografts.

Pigs as xenogeneic donors

The success and widespread acceptance of clinical allotransplantation has led to a shortage of cadaveric organs and a resurgence of interest in xenotransplantation. Although swine are generally regarded as the preferred source of xenogeneic organs, the severe immunologic response to swine organs has so far precluded their clinical use. However, the past 10 years have seen many advances in the field, especially the use of genetically modified swine. These include animals transgenic for human complement regulatory proteins, and most recently, animals not expressing the alpha 1,3-galactosyltransferase gene (GalT-KO swine), produced by nuclear transfer. The initial results using GalT-KO swine as donors to nonhuman primates demonstrated survival of a life-supporting renal graft for up to 83 days without rejection when combined with a regimen designed to induce tolerance by vascularized donor thymic grafts. Although the initial results are encouraging, further improvements will be needed to warrant a clinical trial of xenotransplantation. In this review, we discuss immunologic barriers to xenotransplantation and recent attempts to overcome them, strategies to induce tolerance across a xenogenic barrier, and issues that will need to be considered before a clinical trial is attempted.

Host thymectomy and cyclosporine lead to unstable skin graft tolerance after class I mismatched allogeneic neonatal thymic transplantation in mice

Background Our laboratory has demonstrated that xenogeneic porcine thymus tissue grafted in thymectomized (ATX) and T cell-depleted mice induces donor-specific tolerance. Recipient thymectomy is essential for the success of tolerance induction. In contrast, studies in pigs grafted with non-vascularized allogeneic class I mismatched thymus tissue under the cover of CyA have shown that removal of host thymus is detrimental to thymic graft survival. To determine the requirements for nonvascularized allogeneic class I-mismatched thymic engraftment in mice, we performed thymic allotransplantation under the cover of CyA. Materials and methods Euthymic and ATX B10.MBR mice received class I mismatched B10.AKM neonatal mouse thymus (NMTHY) tissue under the kidney capsule with or without a short course of CyA. The grafts were allowed to engraft for two and a half months before exploratory laparotomy was performed to evaluate them. Three months after the thymic transplant, mice were challenged with donor-specific skin grafts to assess tolerance. One month after donor-specific skin grafting, they received third party B10.BR skin grafts. Cellular anti-donor immune responses were studied at the time of euthanasia. Results CyA-treated ATX and euthymic control mice showed good engraftment of the allogeneic thymic tissue at the time of exploratory laparotomy, whereas non-CyA-treated ATX and euthymic controls had rejected the grafts. The CyA-treated ATX B10.MBR mice accepted donor-specific skin grafts, but rejected them following a challenge with third party B10.BR skin grafts. Untreated ATX and euthymic mice and 6 of 7 CyA-treated euthymic mice rejected donor skin within 15 days. Mixed lymphocyte reactions did not show an increased anti-donor response, but CML clearly showed sensitization and increased killing activity against donor-type targets in these mice. Conclusion Allogeneic thymic transplantation across a class I MHC barrier under the cover of CyA induces a metastable state of tolerance in mice. To achieve this state, ATX of the recipient is required.

Induction and Maintenance of T-dependent or T-independent Xenotolerance by Nonprimarily-Vascularized Skin or Thymus Grafts

The success of clinical xenotransplantation will depend on induction of xenotolerance. We have previously shown that combined xenothymus and vascularized xenoheart transplantation under the coverage of a tolerizing regimen (TR) can induce and maintain full xenotolerance. Here, induction/maintenance of xenotolerance using nonprimarily-vascularized thymus and/or skin grafts was investigated. Hamster skin or thymus or combined skin and thymus transplantation was performed in nude rat recipients with or without administering a TR (NK cell depletion, day -14; xenoantigen infusion, day -14; Leflunomide, day -14 through +14). Xenotolerance was confirmed by subsequent transplantation of a vascularized hamster heart, measurement of xenoantibody formation, or mixed lymphocyte reaction (MLR). Skin grafts were as effective as vascularized heart grafts to induce/maintain T-independent xenotolerance. Even without TR and despite being rejected themselves, xenoskin grafts lead to progressively developing xenononreactivity. Xenothymus transplantation induced xenotolerance in the T-dependent but not in the T-independent immune compartment, leading to rejection of subsequently transplanted hamster hearts by T-independent mechanisms (production of IgM but not IgG xenoantibodies (Xabs), presence of antihamster MLR nonresponsiveness). Combined skin and thymus xenotransplantation sensitized the T-cell compartment, leading to hyperacute rejection of subsequently transplanted hamster hearts. This was not the case when the skin grafts were transplanted late (2 months) after the thymus grafts. Xenogeneic skin and xenogeneic thymus grafts have opposite xenotolerance inducing capacities in the T-independent as compared to the T-dependent immune compartment. Thymus grafts induce and maintain T-dependent but not T-independent xenotolerance. Skin grafts alone induce T-independent xenotolerance but sensitize the T-cell compartment when transplanted concomitantly with thymus grafts.