CD8+ T-Cell Depletion and Rapamycin Synergize with Combined Coreceptor/Stimulation Blockade to Induce Robust Limb Allograft Tolerance in Mice

Abstract

The growing development of composite tissue allografts (CTA) highlights the need for tolerance induction protocols. Herein, we developed a mouse model of heterotopic limb allograft in a stringent strain combination in which potentially tolerogenic strategies were tested taking advantage of donor stem cells in the grafted limb. BALB/c allografts were transplanted into C57BL/6 mice treated with anti-CD154 mAb, nondepleting anti-CD4 combined to either depleting or nondepleting anti-CD8 mAbs. Some groups received additional rapamycin. Both depleting and nondepleting mAb combinations without rapamycin only delayed limb allograft rejection, whereas the addition of rapamycin induced long-term allograft survival in both combinations. Nevertheless, robust donor-specific tolerance, defined by the acceptance of a fresh donortype skin allograft and simultaneous rejection of thirdparty grafts, required initial CD8 + T-cell depletion. Mixed donor-recipient chimerism was observed in lymphoid organs and recipient bone marrow of tolerant but not rejecting animals. Tolerance specificity was confirmed by the inability to produce IL-2, IFN-c and TNF-a in MLC with donor antigen while significant alloreactivity persisted against third-party alloantigens. Collectively, these results show that robust CTA tolerance and mixed donor-recipient chimerism can be achieved in response to the synergizing combination of rapamycin, transient CD8 + T-cell depletion and costimulation/coreceptor blockade.

Pre-treatment of allogeneic bone marrow recipients with the CXCR4 antagonist AMD3100 transiently enhances hematopoietic chimerism without promoting donor-specific skin allograft tolerance

“…All care and handling of animals was carried out in accordance with guidelines provided in the Guide for the Care and Use of Laboratory Animals published by the U.S. Department of Health and Human Services. Full thickness skin from BALB/c donors (for primary grafts) or BALB/c donors and CBA/Ca third party donors (for secondary grafts) was transplanted onto the lateral thoracic wall of recipient mice using standard techniques [5]. After removal of the bandage on day 7 post-transplantation, grafts were monitored daily, and rejection was defined as complete loss of viable donor epithelium.…”

“…However the significant morbidity and mortality associated with myeloablation makes it highly desirable to develop alternative approaches which are mild and efficient. An irradiation-free, transient and mild host immunosuppressive conditioning regimen composed of an anti-T cell antibody cocktail and rapamycin has been reported to induce durable mixed chimerism in mice, but only in the context of “megadose” donor bone marrow infusion or vascularized bone transplantation [4] [5] . Proof of concept for host bone marrow mobilization as a method to enhance mixed chimerism induction after bone marrow transplantation has been reported using the drugs AMD3100 and G-CSF, but only in a congenic mouse bone marrow transplantation system [6]; whether this might also work in the setting of allotransplantation has not yet been reported.…”

Improving the Safety of Tolerance Induction: Chimerism and Cellular Co-Treatment Strategies Applied to Vascularized Composite Allografts

“…T cell depletion prior to VCA transplantation followed by T-cell repopulation after allotransplantation has been associated with allograft acceptance in animal models and in humans [34, 35]. Nonspecific T-cell depletion (lymphodepletion) medications such as ATG [36, 37], Orthoclone mAb OKT3 [38], and humanized Campath-1H CD52 mAb [39] are frequently used clinically for induction therapy before transplantation for prevention and treatment of acute rejection episodes.…”

Interferon gamma licensing of human dendritic cells in T-helper–independent CD8+ alloimmunity

“…Many experimental findings have thus reinforced the view that CD8 ϩ T-cell responses require specific manipulation in addition to CD4 ϩ T-cell responses for effective prevention of organ transplant rejection in both primates and humans. 5,10 However, the mechanisms involved in the activation and memory development of alloreactive CD8 ϩ T cells remain to be elucidated. Greater understanding of this process might therefore help to find new strategies to control CD8 activity and might have important implications for transplantation.…”