No abstract

Xenoantibody production directed at a wide variety of T lymphocyte-dependent and T lymphocyte-independent xenoantigens remains the major immunologic obstacle for successful xenotransplantation. The B lymphocyte subpopulations and their helper factors, involved in T-cellindependent xenoantibody production are only partially understood, and their identification will contribute to the clinical applicability of xenotransplantation. Here we show, using models involving T-celldeficient athymic recipient mice, that rapidly induced, T-cell-independent xenoantibody production is mediated by marginal zone B lymphocytes and requires help from natural killer ( IntroductionSuccessful xenotransplantation may alleviate the ever-increasing need for donor organs, but severe immunologic barriers, of which xenoreactive antibodies represent the most important 1, still hamper (pre)clinical application. Pigs are now generally considered the most suitable organ donors for clinical xenotransplantation, 1 but transplantation of pig organs into nonhuman primates, currently constituting the best validated preclinical model, leads to xenograft rejection within a few hours. This hyperacute rejection is mediated by pre-existing, so-called "natural antibodies," which are directed at a specific Gal␣1,3Gal␤1,4GlcNAc (Gal) oligosaccharide, that is present on (especially endothelial) proteins of most lower animal species, but not of nonhuman primates or man. [2][3][4] Anti-Gal natural antibodies can rapidly activate the complement system of the recipient, leading to hyperacute rejection. 5 Several procedures have been explored in recent years to solve the problem of hyperacute rejection. 6,7 The most appealing ones were the development of genetically modified donor pig strains that either expressed transgenes of human complement regulatory proteins, able to interfere with complement activation, 8 or that lacked the ␣1,3-galactosyltransferase gene to synthesise ␣Gal. 9,10 The latter pig strain seemed particularly attractive, as Gal antigens had been found to be also strongly involved in acute vascular xenograft rejection, a second type of xenograft rejection that develops within a few days in situations where hyperacute rejection is prevented. 7,11 It was hoped, therefore, that elimination of Gal epitopes would prevent both hyperacute rejection and acute vascular rejection.A number of recent reports on the transplantation of ␣1,3-galactosyltransferase-ko pig kidney or heart grafts in nonhuman primates have shown that loss of Gal expression in donor pig organs could indeed prevent hyperacute rejection, but that T-celldependent IgG xenoantibodies were induced against non-Gal xenoantigens, unless a treatment regimen was used that resulted in specific T-cell xenotolerance. [12][13][14] In the latter case, long-term functional renal xenograft survival was obtained, but after a few months, mild focal thrombotic microangiopathy was observed, suggesting that additional modifications to the treatment regimen are needed to permit the development of the kin...

After transplantation of primarily vascularized xenografts (Xgs), T-independent mechanisms may lead to Xg rejection before T-cell activation even takes place. The possibility of achieving T-independent xenotolerance was evaluated in nude rats that normally reject hamster cardiac Xgs within 4 days by non-T cell-mediated mechanisms. After donor antigen infusion, temporary NK-cell depletion and a 4-week administration of Leflunomide, hamster heart grafts survived even after withdrawal of immunosuppression. Tolerant rats accepted second hamster hearts, but promptly rejected mouse heart Xgs. In vivo immunization and in vitro cytotoxicity assays indicated that this species-specific tolerance was based on B-lymphocyte and NK-cell tolerance respectively.

We studied the effect of CTLA-4 blockade on graft-versusleukemia and graft-versus-host responses in a mouse model of minor histocompatibility-mismatched bone marrow transplantation. Early CTLA-4 blockade induced acute graft-versus-host disease. Delayed CTLA-4 blockade resulted in a lethal condition with lymphosplenomegaly, but with stable mixed T-cell chimerism, unchanged alloreactive T-cell frequencies and absent anti-host reactivity in vitro. In contrast, multiorgan lymphoproliferative disease with autoimmune hepatitis and circulating anti-DNA auto-antibodies were documented. Splenic lymphocytes exhibited ex vivo spontaneous proliferation and a marked proliferative response against host-type dendritic cells pulsed with syngeneic (host-type) tissue-peptides. Both phenomena were exclusively mediated by host and not donor T cells, supporting an autoimmune pathogenesis. Selectively hostderived T-cell immune reactivity was equally documented against leukemia-peptide-pulsed dendritic cells, and this was paralleled by a strong in vivo antileukemic effect in anti-CTLA-4-treated and subsequently leukemia-challenged chimeras. In conclusion, delayed CTLA-4 blockade induced a host-derived antileukemic effect, occurring in the context of an autoimmune syndrome and strictly separated from graft-versus-host disease. Both antileukemic and autoimmune responses depended on the allogeneic component, as neither effect was seen after syngeneic bone marrow transplantation. Our findings reveal the potential of using CTLA-4 blockade to establish antileukemic effects after allogeneic hematopoietic stem cell transplantation, provided autoimmunity can be controlled.

Successful grafting of vascularized xenografts (Xgs) depends on the ability to reliably induce both T cell-independent and -dependent immune tolerance. After temporary NK cell depletion, B cell suppression, and pretransplant infusion of donor Ags, athymic rats simultaneously transplanted with hamster heart and thymus Xgs developed immunocompetent rat-derived T cells that tolerated the hamster Xgs but provoked multiple-organ autoimmunity. The autoimmune syndrome was probably due to an insufficient development of tolerance for some rat organs; for example, it led to thyroiditis in the recipient rat thyroid, but not in simultaneously transplanted donor hamster thyroid. Moreover, grafting a mixed hamster/rat thymic epithelial cell graft could prevent the autoimmune syndrome. These experiments indicate that host-type thymic epithelial cells may be essential for the establishment of complete self-tolerance and that mixed host/donor thymus grafts may induce T cell xenotolerance while maintaining self-tolerance in the recipient.