Introduction: Xenotransplantation is a feasible substitute to resolve shortage of organ for transplantation. To success pig-to-primate xenotransplantation, genetic engineering of donor pig is essential for overcoming pig-to-primate immune rejection. As various genetically modified donor pig such as Knock-out (KO) of xenoantigens, and expression of genes to modulate immunological rejection have been advanced, the survival of pig-to-nonhuman primate xenotransplantation prolonged significantly. However, a deeper understanding of the biological barriers between pig and primate is still needed to accomplish pig-to-primate xenotransplantation. Although swine leukocyte antigen (SLA) has been suggested to be associated with human T cell response and antibody-mediated rejection, the effect of SLA remains unsubstantiated in pig-to-primate xenotransplantation. Therefore, we generated SLA I knock-out pig and SLA null cells to identify response to SLA antigen in human or nonhuman primate T cells and pig-to-nonhuman xenotransplantation.
Methods: SLA I KO pig was generated by disruption of beta-2-microglobulin(β2m) gene based on QKO (quadruple KO, GGTA1/CMAH/A3GalT2/B4GalNT2 KO) using somatic cell nuclear transfer technology. Generation of SLA null fibroblasts were achieved by knock-out of CIITA gene based on PKO (penta KO, GGTA1/CMAH/A3GalT2/B4GalNT2/β2m KO). Expression of SLA I and SLA II was identified by flowcytometry analysis on cells from PKO pig and SLA null fibroblasts. Aorta endothelial cells (AECs) from WT (wild type), QKO and PKO were co-cultured with human or cynomolgus peripheral blood monocyte cells (PBMCs), and proliferation of lymphocytes and CD8 T cell was estimated using cell proliferation kit. The antibody response to SLA antigen was evaluated by incubation of either SLA-positive or SLA-negative fibroblasts and serum from recipient following pig-to-primate kidney transplantation.
Results: Various cells from PKO pig did not express SLA I. SLA-null fibroblasts did not express SLA I and SLA II antigen. The proliferation of human CD8+ T cells were significantly reduced against AECs from PKO pig compared to AECs from WT and QKO. In contrast, cynomolgus CD8+ T cells showed reduced proliferation against AECs from both QKO and PKO pig compared to WT. The donor specific antibody related to SLA antigen especially SLA II was increased in several cases of pig-to-primate kidney transplantation recipient.
Conclusion: Knock-out of β2m and CIITA gene were sufficient to eliminate SLA. Although pig-to-primate xenotransplantation using SLA KO pig have not been performed to assess immune response in this study, our findings imply that SLA may play a potential role in both human T cell response and memory response, leading to delayed immunological rejection