Upregulation of CD80 on glomerular podocytes plays an important role in development of proteinuria following pig-to-baboon xeno-renal transplantation - an experimental study

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Background Our initial studies utilizing a galactosyl‐α1‐3‐galactosyltransferase gene knockout (GalTKO) pig‐to‐baboon renal transplant model demonstrated that the early development of nephrotic syndrome has been a significant obstacle to the long‐term survival of baboon recipients. We have recently documented that sphingomyelin phosphodiesterase‐3 (SMPDL3b) and CD80 expressed on podocytes in porcine kidney grafts contribute to this complication. We have hypothesized that one regulator of immune function is CD47 and that incompatibilities in CD47 between pig and baboon could potentially affect macrophage function, increasing the susceptibility of the kidney grafts to immunologically induced injury. Methods In order to address this hypothesis in vitro, we isolated and cultured porcine podocytes and ECs from GalTKO alone, human CD47 (hCD47)/hCD55 expressing transgenic (Tg) GalTKO swine, and GalTKO hCD46/hCD55 Tg swine along with baboon or human macrophages. Results We found that baboon macrophages phagocytosed porcine ECs in a similar manner to human macrophages, and this response was significantly reduced when porcine ECs and podocytes expressed hCD47/hCD55 but not hCD46/hCD55 without hCD47. Furthermore, masking hCD47 by anti‐hCD47 antibody on hCD47/hCD55Tg ECs restored phagocytosis. These results are consistent with the hypothesis that CD47 incompatibility plays an important role in promoting macrophage phagocytosis of endogenous cells from the transplanted kidney. Conclusions The similar levels of phagocytosis of porcine cells by baboon and human macrophages suggest that the expression of hCD47Tg on glomerular cells in donor porcine kidneys may prove to be a key strategy for preventing proteinuria following kidney xenotransplantation in a pig‐to‐human as well as a pig‐to‐baboon model.

Section: Introductionmentioning

confidence: 83%

Section: Introductionmentioning

confidence: 99%

Transgenic expression of human CD47 reduces phagocytosis of porcine endothelial cells and podocytes by baboon and human macrophages

Nomura

Ariyoshi

Watanabe

et al. 2019

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“…Using these two techniques, we have demonstrated that vascularized thymic tissue can successfully induce tolerance and support thymopoiesis across fully allogeneic barriers in MGH miniature swine . By extending this strategy to pig‐to‐baboon models, we have recently achieved survival of a life‐supporting TK for over 6 months . Notably, multiple recipients bearing a functional TK for over 3 months developed donor‐specific unresponsiveness at the T‐ and B‐cell levels .…”

Section: Introductionmentioning

confidence: 99%

“…[23][24][25] By extending this strategy to pig-to-baboon models, we have recently achieved survival of a life-supporting TK for over 6 months. 26 Notably, multiple recipients bearing a functional TK for over 3 months developed donor-specific unresponsiveness at the Tand B-cell levels. 26,27 Although new thymic emigrants (CD4 + /CD31 + / CD45 + ) were developed (Yamada et al manuscript in preparation) beyond 2 months following transplantation, we have not yet been able to fully stop immunosuppression, although we have been successful with tapering it markedly.…”

Section: Introductionmentioning

confidence: 99%

“…26 Notably, multiple recipients bearing a functional TK for over 3 months developed donor-specific unresponsiveness at the Tand B-cell levels. 26,27 Although new thymic emigrants (CD4 + /CD31 + / CD45 + ) were developed (Yamada et al manuscript in preparation) beyond 2 months following transplantation, we have not yet been able to fully stop immunosuppression, although we have been successful with tapering it markedly. 26 Therefore, additional strategies are required for the rapid induction of tolerance in the pig-to-NHP model.…”

Section: Introductionmentioning

confidence: 99%

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Preparation of hybrid porcine thymus containing non‐human primate thymic epithelial cells in miniature swine

Sekijima

Sahara

Shimizu

et al. 2019

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Background We have achieved greater than a 6‐month survival of a life‐supporting kidney co‐transplanted with a vascularized thymic graft into non‐human primates (NHPs). Although we have achieved pig‐specific unresponsiveness in vitro, immunosuppression was not able to be fully weaned. Studies in mice and humanized mice suggest that a hybrid pig thymus (Hyb‐thy)‐containing host thymic epithelial cells (TECs) can optimize intra‐thymic selection, achieving xenograft tolerance with improved reconstitution of T‐cell function. Methods We have tested the feasibility of the preparation of a Hyb‐thy that contains NHP TECs in the donor thymic grafts. We first prepared the Hyb‐thy in the donor pigs 2‐3 weeks before xeno‐Tx. We performed six cases of Hyb‐thy preparation in six juvenile miniature swine. Two pigs received non‐manipulated cynomolgus monkey thymic cells that were isolated from an excised atrophic thymus via injection into their thymic lobes (Group 1). The remaining four received thymic cells that were isolated from non‐atrophic thymic glands (Groups 2 and 3). Pigs in Group 2 received unmanipulated thymic cells in one thymic lobe, as well as CD2‐positive cell‐depleted TEC‐enriched cells in the contralateral lobe. Pigs in Group 3 received TEC‐enriched cells alone. Results All thymus‐injected pigs received tacrolimus and rapamycin until endpoint (POD16). We detected cynomolgus monkey TEC networks in pig thymus from Groups 1 and 3, while pigs in Group 2 rejected the thymic cells. We demonstrated the preparation of Hyb‐thy in pigs using tacrolimus plus rapamycin therapy. Conclusions Our results suggest that the enrichment of TEC from the excised NHP thymus facilitated NHP TEC engraftment in pig thymus.

Co-transplantation of Vascularized Thymic Graft with Kidney in Pig-to-Nonhuman Primates for the Induction of Tolerance Across Xenogeneic Barriers

Yamada

Ariyoshi

Pomposelli

et al. 2020

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Using advanced gene editing technologies, xenotransplantation from multi-transgenic alpha-1,3galacto-syltransferase knockout pigs has demonstrated marked prolongation of renal xenograft survival, ranging from days to greater than several months for life-supporting kidneys and >2 years in a heterotopic non-life-supporting cardiac xenograft model. However, continuous administration of multiple immunosuppressive drugs continues to be required, and attempts to taper immunosuppression have been unsuccessful. These data are consistent with previous reports indicating that the human-anti-porcine T cell response is similar or stronger than that across allogeneic barriers. Due to the strength of both the innate and adaptive immune responses in xenotransplantation, the level of continuous immunosuppression needed to control these responses and prolong xenograft survival has been associated with prohibitive morbidity and mortality. These facts provide compelling rationale to pursue a clinically applicable strategy for the induction of tolerance. Mixed chimerism and thymic tissue transplantation have both achieved xenogeneic tolerance in pig-to-mouse models, and both have recently been extended to pig-to-baboon models. Although these strategies are promising in small animal models, neither direct intravenous injection of porcine bone marrow cells nor direct fetal thymic tissue transplantation into recipients was able to achieve >2 days chimerism following BM Tx or the engraftment of thymic tissues across xenogeneic barriers in pig-to-nonhuman primate models. Several innovative procedures have been largely developed by Kazuhiko Yamada to overcome these failures. These include vascularized thymic transplantation, combined with either thymokidney (TK) or vascularized thymic lobe (VTL) transplantation. Utilizing the strategy of transplanting vascularized thymic grafts with kidney from the same GalT-KO donor without further gene modification, we have achieved longer than 6 months survival of life-supporting kidneys in a baboon. Notably, the recipient became