Introduction: We previously characterized the short-term immune response after pig-to-human kidney xenotransplantation, but little is known about its long-term evolution. Here we aimed to decipher the long-term xenoimmune response in a human decedent model.
Methods: We analyzed a genetically modified pig kidney xenograft implanted for 61 days in a deceased human recipient with a pre-transplant negative xenoreactive crossmatch. We performed morphological evaluation, multi-immunophenotyping, 38-hyperplex complement cascade imaging, bulk transcriptomic profiling, and spatial transcriptomics at cellular and subcellular levels (NanoString CosMx®) at sequential time points (POD10, POD33, POD45, POD49, POD61). Controls included pre-implantation xenograft, wild-type pig kidney autografts, and brain-dead pig kidneys.
Results: From POD10 to POD61, we observed a membranoproliferative glomerulonephritis (MPGN)-like pattern characterized by microvascular inflammation, predominantly in glomeruli, associated with intense linear capillary C4d deposition, glomerular deposition of IgM and IgA, endothelial activation, and early post-xenotransplant de novo positive xenoreactive crossmatch. This pattern was accompanied by glomerular double contours, interstitial edema and endarteritis from POD45. Capillary inflammation primarily consisted of CD68+ and CD15+ innate immune cells throughout all follow-up time points. At POD45, we observed co-localized deposits of IgM, IgA, C1q, C3c, C4d, factor B, and complement factor H regulator along glomerular capillaries. The xenograft exhibited increased expression of genes related to humoral response, including monocyte and macrophage activation, NK cell burden, endothelial activation, interferon-gamma response, and T and B cell interaction, which improved over time after rejection treatment (Figure).

Conclusion: We described for the first time a new entity of xenoantibody-mediated injury characterized by an MPGN-like pattern, mainly driven by complement activation through glomerular xenoimmune complexes, which improved after rejection treatment. These findings support the development of personalized therapies to improve xenotransplantation outcomes.