*Purpose: Xenotransplantation, the use of genetically modified pigs as organ donors for human transplant, is a potential strategy to address the organ shortage. Antibody-mediated rejection remains a barrier to consistent, long-term success in non-human primate models of xenotransplantation. The antigen target (glycan, MHC, or other) of the xenoantibodies during xenograft rejection is currently unknown. Given the first clinical xenotransplant trials may be in highly sensitized recipients, the development of assays to determine xenoantibody targets will be essential to xenograft success.
*Methods: Five maximally MHC-mismatched NHPs underwent swapping skin transplants to induce MHC sensitization, followed by bilateral nephrectomy and kidney transplantation from an αGal knockout, human CD55 transgenic (GGTA1/CD55) pig (National Swine Resource and Research Center, Columbia, MO). A flow cytometry crossmatch (FCXM) was performed in which donor splenocytes were incubated with recipient sera and anti-IgM or anti-IgG secondary antibodies were utilized to measure antibody binding. To evaluate the potential contribution of anti-glycan antibodies, sera was first absorbed on wild-type (WT) pig red blood cells (pRBC) prior to the FCXM.
*Results: All NHPs demonstrated a positive pre-transplant IgM and IgG FCXM to the donor pig. All developed graft failure and were euthanized at days 12, 15, 27, and 55. Histology at the time of sacrifice demonstrated active antibody-mediated rejection with thrombotic microangiopathy in all cases. WT pRBC absorption significantly decreased IgM antibody binding at the naïve and pre-transplant time points, however this difference was not present at the time of graft rejection (Figure 1A). IgG antibody binding increased significantly at the time of graft rejection, with no significant change following WT pRBC absorption (Figure 1B).
*Conclusions: In a highly-sensitized NHP model of xenotransplantation, it appears anti-glycan IgM xenoantibodies decline while anti-MHC IgG antibodies increase at the time of antibody-mediated rejection. Further studies are needed to determine if these are de novo or represent an amnestic response from prior allosensitization.

Joseph Ladowski

Duke University

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