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

Abstract

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.

Antibody reactivity with new antigens revealed in multi‐transgenic triple knockout pigs may cause early loss of pig kidneys in baboons

“…Our recent data indicated that hCD47 expression on porcine endothelial cells or glomerular podocytes inhibited phagocytosis by baboon or human phagocytes 6 . None of the baboons that received mTg GalT‐KO kidneys including hCD47 Tg developed proteinuria, including these two rejectors.…”

“…Our initial study demonstrated an 83‐day survival of a baboon bearing a life‐supporting GalT‐KO pig kidney graft without rejection, when a vascularized donor pig thymus was co‐transplanted from the same GalT‐KO pig 4 . In fact, survival of up to 193 days has been achieved with a modified immunosuppressive regimen that minimized the development of proteinuria, which is associated with activation of podocytes in kidney grafts during the induction period 5,6 . Recently, after overcoming the early development of proteinuria in baboons, 5,7 we have had multiple baboon recipients bearing GalT‐KO kidney plus vascularized thymic grafts that maintained renal graft function longer than 4 months, even without complement inhibitors (Yamada et al manuscript in preparation).…”

Intra‐bone bone marrow transplantation from hCD47 transgenic pigs to baboons prolongs chimerism to >60 days and promotes increased porcine lung transplant survival

“…We have successfully produced hCD47 Tg GalTKO miniature swine that express hCD47 in all blood cell lineages . Endothelial cells isolated from these hCD47 pigs markedly reduced phagocytosis by both human and baboon macrophages …”

Immunogenetics of xenotransplantation

“…Proteinuria has been described to be triggered by the loss of sphingomyelin phosphodiesterase acid‐like 3b expression on porcine podocytes. Treatments as inoculation of anti‐CD20 antibodies (Rituximab) or transgene expression of human CD47 are reported to prevent proteinuria (Nomura et al., 2019; Shah et al., 2018). Indeed, transgenic pig cells expressing human CD47 avoid phagocytosis by baboon macrophages, preventing xenograft injury and proteinuria (Nomura et al., 2019).…”