Approaches to avoid immune responses induced by repeated subcutaneous injections of allogeneic umbilical cord tissue-derived cells.

BACKGROUND: Cellular treatments for repairing diseased tissues represent a promising clinical strategy. Umbilical cord tissue-derived cells (UTC) are a unique source of cells with a low immunogenic profile and potential for tissue repair. By using UTC from miniature swine, we previously demonstrated that despite their low immunogenic phenotype, UTC could induce an immune response under certain inflammatory conditions and after multiple subcutaneous (SC) injections. Given that repeat dosing of cells may be necessary to achieve a lasting therapeutic benefit, in this study, we examined approaches to avoid an immune response to multiple SC injections of UTC. METHODS: By using in vitro and in vivo measures of sensitization to SC cellular injections, we assessed the effects of varying the location of administration site, prolongation of timing between injections, and use of immunosuppressive treatments on repeated cellular injections in Massachusetts General Hospital major histocompatibility complex-defined miniature swine. RESULTS: Although under normal conditions, a single SC injection of major histocompatibility complex-mismatched UTC did not induce a detectable immune response, multiple SC injections of UTC demonstrated rapid humoral and cell-mediated immune responses. Avoidance of an immune response to repeat SC injection was achieved by concurrent immunosuppression with each dose of UTC. CONCLUSIONS: UTC and other similar cell types believed to be nonimmunogenic have the potential to induce immune responses under certain conditions. These studies provide important considerations and guidelines for preclinical studies investigating allogeneic cellular therapies.

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.

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.

Immunogenicity of umbilical cord tissue derived cells.

Umbilical cord tissue provides a unique source of cells with potential for tissue repair. Umbilical cord tissue-derived cells (UTCs) are MHC class I (MHCI) dull and negative for MHC class II (MHCII), but can be activated to increase MHCI and to express MHCII with IFN-gamma stimulation. Mesenchymal stem cells with similar characteristics have been inferred to be nonimmunogenic; however, in most cases, immunogenicity was not directly assessed. Using UTC from Massachusetts General Hospital MHC-defined miniature swine, we assessed immunogenicity across a full MHC barrier. Immunogenicity was assessed by in vitro assays including mixed lymphocyte reaction (MLR) and flow cytometry to detect serum alloantibody. A single injection of MHC-mismatched unactivated UTCs did not induce a detectable immune response. When injected in an inflamed region, injected repeatedly in the same region or stimulated with IFN-gamma prior to injection, UTCs were immunogenic. As clinical cellular repair strategies may involve injection of allogeneic cells into inflamed regions of damaged tissue or repeated doses of cells to achieve the desired benefit, our results on the immunogenicity of these cells in these circumstances may have important implications for optimal success and functional improvement for this cellular treatment strategy for diseased tissues.