Abstract
Vascularized composite allotransplantation (VCA) allows tissue replacement after devastating loss but is currently limited in application and may be more widely performed if maintenance immunosuppression was not essential for graft acceptance. We tested whether peri-transplant costimulation blockade could prolong VCA survival and required donor bone-marrow cells, given that bone-marrow might promote graft immunogenicity or graft-versus-host disease. Peritransplant CD154 mAb/rapamycin (RPM) induced long-term orthotopic hindlimb VCA survival (BALB/c->C57BL/6), as did CTLA4Ig/RPM. Surprisingly, success of either protocol required a bone-marrow-associated, radiation-sensitive cell population, since long-bone removal or pre-transplant donor irradiation prevented long-term engraftment. Rejection also occurred if Rag1−/− donors were used, or if donors were treated with a CXCR4 inhibitor to mobilize donor BM cells pre-transplant. Donor bone-marrow contained a large population of Foxp3+ T-regulatory (Treg) cells, and donor Foxp3+ Treg depletion, by diphtheria toxin administration to DEREG donor mice whose Foxp3+ Treg cells expressed diphtheria toxin receptor, restored rejection with either protocol. Rejection also occurred if CXCR4 was deleted from donor Tregs pre-transplant. Hence, long-term VCA survival is possible across a full MHC disparity using peritransplant costimulation blockade-based approaches, but unexpectedly, the efficacy of costimulation blockade requires the presence of a radiation-sensitive, CXCR4+ Foxp3+ Treg population resident within donor BM.
Highlights
Vascularized composite allotransplantation (VCA) allows tissue replacement after devastating loss but is currently limited in application and may be more widely performed if maintenance immunosuppression was not essential for graft acceptance
CD154 monoclonal antibody (mAb) (MR-1) plus donor splenocyte transfusion (DST, 5 × 106 cells, i.v.) is very effective at inducing tolerance of various kinds of allografts, including that of hearts, kidneys or islets transplanted across fully MHCdisparate combinations such as BALB/c->C57BL/6 mice (H-2d->H-2b)[6,7]
This protocol, while able to prolong VCA survival to >30 days, was unable to induce long-term (>100 days) VCA survival (Fig. 1A), and use of either agent alone led to only 2–3 days of prolongation of allograft survival
Summary
Vascularized composite allotransplantation (VCA) allows tissue replacement after devastating loss but is currently limited in application and may be more widely performed if maintenance immunosuppression was not essential for graft acceptance. We hypothesized that a clinically relevant protocol such as peri-transplant delivery of some form of COB plus RPM might promote long-term VCA survival by inhibiting the allo-activation of conventional T cells and allowing the development of Treg-dependent mechanisms that maintain allograft survival. Corollaries of this over-arching hypothesis are that potentially both donor and recipient Tregs might contribute to graft survival; that Tregs must prevent potent host responses to allogeneic skin that is typically considered highly immunogenic; and that therapeutic agents that negatively or positively affect Treg function might impair or further promote outcomes, respectively. We sought to test these concepts in murine limb transplant models that are ideally suited to mechanistic investigations
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