Peri‐graft porcine‐specific CD4⁺FoxP3⁺ regulatory T cells by CD40‐CD154 blockade prevented the rejection of porcine islet graft in diabetic mice

Abstract

Background: Anti-CD154 monoclonal antibody (mAb) treatment has been known to have potential to induce immune tolerance in organ transplantation. Several studies have suggested the involvement of CD4 + regulatory T cells (T reg s) in xeno-immune tolerance. However, the characteristics of T reg s and the mechanisms of their regulatory functions in islet xenotransplantation have not been clearly defined. Method:Adult porcine islet cells were isolated and purified, and were transplanted under the kidney capsule of diabetic C57BL/6J mice with the administration of 0.5 mg/mouse of anti-CD154 mAb on 0, 1, 3, 5, and 7 days post-transplantation (DPT). The graft survival was monitored by blood glucose level. The islet graft and recipients' cells were analyzed by immunohistochemistry (IHC), flow cytometry, enzyme-linked immunosorbent spot (ELISPOT) assay, and mixed-lymphocyte reaction.Results: Short-term anti-CD154 mAb monotherapy enabled the porcine islet graft to survive indefinitely in diabetic mice (n = 18). Immunohistochemical staining showed significantly higher ratio of CD4 + Foxp3 + T reg s in the peri-graft site, but not in the spleen and kidney-draining lymph node of the recipient mice. Depletion of T reg s evoked graft rejection, and adoptive transfer of T reg s from anti-CD154 mAb-treated recipients provided protection to the graft from rejection. These T reg s showed more potent suppressive capacity than those from the untreated control and were found to be porcine antigen-specific. Conclusions:In this study, we showed that anti-CD154 mAb monotherapy resulted in indefinite porcine islet graft survival in mice. The porcine-specific CD4 + Foxp3 + T reg s in the peri-graft site played the critical role in the protection of islet graft from rejection, which suggests a prospective immunosuppressive strategy for islet xenotransplantation. K E Y W O R D SCD154, peri-graft site, porcine islet, regulatory T cell, xenotransplantation

Neonatal Pig Sertoli Cells Survive Xenotransplantation by Creating an Immune Modulatory Environment Involving CD4 and CD8 Regulatory T Cells

“…The main highlight of this study was the early and continued presence of CD4 and CD8 Tregs within the NPSC grafts. CD4 Tregs have been implicated in prolongation of allograft survival and recently have been demonstrated to improve survival of xenografts 20 – 24 , 40 . Tregs have been shown to mediate immune tolerance and homeostasis by suppressing T effector cells, inhibiting cell–cell contact with APCs, and secreting regulatory cytokines 41 , 42 .…”

“…Regulatory T cells (Tregs) have potential to suppress cell-mediated responses generated against xenogeneic tissue thus improving graft survival 19 – 21 . Use of CD40-CD154 blockade has led to long-term survival of porcine islet xenografts in diabetic mice 22 – 24 . Using a combination of anti-lymphocyte function associated antigen (LFA)-1 and anti-CD154 mAbs to prolong graft survival, Arefanian et al demonstrated that CD4 + Foxp3 + Tregs (CD4 Tregs) were responsible for this survival as CD4 Treg depletion resulted in graft rejection and reconstitution of B6 rag −/− mice with splenocytes from these tolerant mice resulted in prolonged porcine islet xenograft survival 22 , 23 .…”

“…Using a combination of anti-lymphocyte function associated antigen (LFA)-1 and anti-CD154 mAbs to prolong graft survival, Arefanian et al demonstrated that CD4 + Foxp3 + Tregs (CD4 Tregs) were responsible for this survival as CD4 Treg depletion resulted in graft rejection and reconstitution of B6 rag −/− mice with splenocytes from these tolerant mice resulted in prolonged porcine islet xenograft survival 22 , 23 . Recently, the importance of CD4 Tregs in prolongation of islet xenografts was confirmed using only the anti-CD154 monoclonal antibody (mAb) 24 . They also found Treg depletion led to graft rejection and adoptive transfer of CD4 Tregs from anti-CD154 mAb-treated mice provided prolonged islet xenograft survival in these Treg recipients 24 .…”

CD4+/CD8+T‐cell ratio correlates with the graft fate in pig‐to‐non‐human primate islet xenotransplantation

“…Our group recently observed in mice model that if CD4 + CD25 + regulatory T cells were depleted in the early phase of porcine islet xenotransplantation with CD40-CD154 axis blockade, grafts were not protected by the T cells. 40 It also implies that CD4 + /CD8 + ratios as a biomarker should be interpreted in this context.…”

Xenotransplantation literature update, September/October 2019

“…In the previous issue of Xenotransplantation, Perota et al reported that they had successfully generated a bovine double‐KO model lacking both αGal and Neu5Gc; Chen et al studied the specific role of TLR2 in xenograft rejection of porcine iliac endothelial cells; Yoon et al described the mechanism of how CD154 blockade protects islet xenografts from rejection; Bracey et al assessed the biocompatibility of porcine xenograft‐derived bone scaffold in vitro. Additionally, the abstracts from the International Xenotransplantation Association 2019 Congress are also published here and outline a number of novel projects that were presented at the congress.…”