Donor Lymphocyte Infusion-Mediated Graft-versus-Host Responses in a Preclinical Swine Model of Haploidentical Hematopoietic Cell Transplantation.

We previously described successful hematopoietic stem cell engraftment across MHC barriers in miniature swine without graft-versus-host disease (GVHD) using novel reduced-intensity conditioning regimens consisting of partial transient recipient T cell-depletion, thymic or low-dose total body irradiation, and a short course of cyclosporine A. Here we report that stable chimeric animals generated with these protocols are strongly resistant to donor leukocyte infusion (DLI)-mediated GVH effects. Of 33 total DLIs in tolerant chimeras at clinical doses, 21 failed to induce conversion to full donor hematopoietic chimerism or cause GVHD. We attempted to overcome this resistance to conversion through several mechanisms, including using sensitized donor lymphocytes, increasing the DLI dose, removing chimeric host peripheral blood cells through extensive recipient leukapheresis before DLI, and using fully mismatched lymphocytes. Despite our attempts, the resistance to conversion in our model was robust, and when conversion was achieved, it was associated with GVHD in most animals. Our studies suggest that delivery of unmodified hematopoietic stem cell doses under reduced-intensity conditioning can induce a potent, GVHD-free, immune tolerant state that is strongly resistant to DLI.

In vivo observations of cell trafficking in allotransplanted vascularized skin flaps and conventional skin grafts.

The problem of allogeneic skin rejection is a major limitation to more widespread application of clinical composite tissue allotransplantation (CTA). Previous research examining skin rejection has mainly studied rejection of conventional skin grafts (CSG) using standard histological techniques. The aim of this study was to objectively assess if there were differences in the immune response to CSG and primarily vascularized skin in composite tissue allotransplants (SCTT) using in vivo techniques in order to gain new insights in to the immune response to skin allotransplants. CSG and SCTT were transplanted from standard Lewis (LEW) ad Wistar Furth (WF) to recipient transgenic green fluorescent Lewis rats (LEW-GFP). In vivo confocal microscopy was used to observe cell trafficking within skin of the transplants. In addition, immunohistochemical staining was performed on skin biopsies to reveal possible expression of class II major histocompatibility antigens. A difference was observed in the immune response to SCTT compared to CSG. SCTT had a greater density cellular infiltrate than CSG (p<0.03) that was focused more at the center of the transplant (p<0.05) than at the edges, likely due to the immediate vascularization of the skin. Recipient dendritic cells were only observed in rejecting SCTT, not CSG. Furthermore, dermal endothelial class II MHC expression was only observed in allogeneic SCTT. The immune response in both SCTT and CSG was focused on targets in the dermis, with infiltrating cells clustering around hair follicles (CSG and SCTT; p<0.01) and blood vessels (SCTT; p<0.01) in allogeneic transplants. This study suggests that there are significant differences between rejection of SCTT and CSG that may limit the relevance of much of the historical data on skin graft rejection when applied to composite tissue allotransplantation. Furthermore, the use of novel in vivo techniques identified characteristics of the immune response to allograft skin not previously described, which may be useful in directing future approaches to overcoming allograft skin rejection.

Skin tolerance: in search of the Holy Grail.

In 1943, Gibson and Medawar opened the modern era of transplantation research with a paper on the problem of skin allograft rejection. Ten years later Billingham, Brent and Medawar demonstrated that it was possible to induce selective immune acceptance of skin grafts in mice, a state of tolerance. After over six decades, however, the precise mechanism of skin allograft rejection remains still ill-defined. Furthermore, it has not been possible to achieve reliably clinical tolerance allowing the widespread application of skin allotransplantation techniques. The first successful applications of skin allotransplantation have included the hand and face. However, complications from the chronic immunosuppression regimens limit the application of these techniques. Induction of tolerance to skin (and the other tissues in the allograft) would be the most effective way to overcome all these difficulties, but this is yet to be achieved reliably, stimulating some to look for other ways to surmount the current limitations. This paper summarizes alternatives to enlarge the scope of skin allotransplantation techniques, current understanding of mechanisms of skin rejection, and the utility and limitations of animal models used to study skin rejection and tolerance induction. Finally, manipulation strategies to achieve skin tolerance are outlined.

Recipient damage after musculocutaneous transplant rejection.

BACKGROUND: In the event of a composite allograft failure, damage to the recipient tissues may make retransplantation impossible. This study aimed to quantify the damage after composite allograft failure to assess whether retransplantation is feasible. METHODS: Rats (n=6) in the group I received composite musculocutaneous flap allotransplants (WF-->Lew) with immunosuppression allowing healing-in of the allograft before being tapered allowing rejection. At full rejection, the recipient vascular pedicle was examined, biopsies of the recipient tissue bed were taken and graded for damage, and in vitro assays were performed. Groups II (allograft without immunosuppression, n=7), III (isograft with immunosuppression, n=5), and IV (isograft without immunosuppression, n=6) were included to attempt to identify the contributions of the rejection process, immunosuppression, and healing to recipient tissue damage. RESULTS: The vascular pedicle was patent to within 1 mm of the anastomosis in all rejected allografts. Furthermore, it was possible to retransplant after full rejection. There was minimal damage to the recipient tissues at the time of full rejection in group I. In contrast, group II had significantly more damage (P<0.05) to recipient muscle and skin. This correlated with more severe immune reaction with more than 100 times antibody production in group II compared with group I. Groups III and IV had little recipient tissue damage. CONCLUSIONS: These results suggest that it is possible to retransplant after rejection of a musculocutaneous transplant while on immunosuppression. Furthermore, the second transplant will not be limited in form or function by recipient tissue bed damage.