The Role Of Donor CD47 On Innate Immune Cell Activation And T Cell Alloresponses And Its Mechanistic Study

The balance between the stimulatory and inhibitory signaling regulate innate immune cell activation. Interaction between Signal regulatory protein-alpha(SIRP?) and its ligand CD47 provide a “don't eat me” signal to the macrophage to prevent phagocytosis, CD47 was incompatible between different species, interspecies incompatibility of CD47 plays an important role in triggering rejection of xenogeneic hematopoietic cells by macrophages. However, whether CD47 incompatibility also induces rejection of non-hematopoietic cellular xenografts remains unknown. Herein, we have addressed this question in a mouse model of hepatocyte transplantation in which CD47(-/-) hepatocytes were used to resemble xenografts for CD47 incompatibility. in order to track the implanted hepatocyte GFP transgenic mice were used for both the CD47 KO and WT mice. GFP+ hepatocyte from WT or CD47(-/-) B6 mice were isolated and intrasplenically transplanted into na?ve B6 mice, isolated hepatocyte survival over 90% was confirmed under microscope with trypan blue staining. Before the transplantation. We show that intrasplenic transplantation of CD47(-/-), but not wild-type(WT) hepatocytes, into partially hepatectomized syngeneic WT mice resulted in a rapid increase in Mac-1(+) cells with an activation phenotype(i.e., Mac-1(+)CD14(+) and Mac-1(+)CD16/32(high)), compared to non-transplant controls at early time(measured at 3 and 5 days after transplantation). In addition, CD47(-/-) hepatocytes were more severely damaged than WT hepatocytes as indicated by the greater AST and ALT serum levels in these mice. Furthermore, long-term donor hepatocyte survival and liver repopulation were observed in mice receiving WT hepatocytes, whereas CD47(-/-) hepatocytes were completely rejected within 2 weeks. So lack of CD47 on donor hepatocytes promote recipient myeloid innate immune cell activation and subsequent associated graft loss in syngeneic recipients. In order to further investigate the role of donor CD47 in regulation of T-cell alloresponses. We addressed this question by assessing OVA-specific immune responses in mice following hepatocyte transplantation from CD47-competent or-deficient OVA-transgenic donors. Compared to sham-operated controls, intrasplenic transplantation of CD47-deficient OVA+ hepatocytes significantly accelerated rejection of OVA+ skin grafted 7 days after hepatocyte transplantation. In contrast, mice receiving CD47-competent OVA+ hepatocytes showed prolonged and even indefinite survival of OVA+ skin allografts. With histology examination, the grafted skin on recipient of WT hepatocyte transplantation did not show sign of significant inflammatory cell infiltration while robust inflammatory cell infiltration was observed in the grafted skin on the recipient of CD47(-/-) hepatocytes transplantation. IHC have shown the grafted skin on the recipient of CD47(-/-) hepatocytes transplantation was infiltrated by large amount of both CD4 and CD8 T cell. T cells from mice receiving CD47-deficient, but not CD47-competent, OVA+ hepatocytes showed significantly enhanced responses to OVA+ stimulators compared to sham-operated controls. In contrast to the production of tolerogenic cytokines(IL-4 and IL-10) in the recipients of CD47-competent hepatocytes, mice receiving CD47-deficient hepatocytes showed elevated production of IFN-? and IL-1?. Moreover, significant expansion of myeloid-derived suppressor cells(MDSCs)was detected in the recipients of CD47-competent hepatocytes, Gemcitabine can effectively deplete MDSCS in mice, WT hepatocyte transplantation failed to induced tolerance in Gemcitabine treatment mice. So MDSCs is required for tolerance induction in WT hepatocyte transplantation recipients mice. Thus, donor CD47 plays an important role in the control of T-cell alloresponses and tolerance induction following hepatocyte transplantation. Further study of the CD47 may provide us a new direction and filed about the hepatocyte xenotransplantation or other cell and organ xenotransplantation. Manipulation of CD47 may significantly improve xenotransplantation survival,CD47 may become a new therapeutic target that may draw xenotransplantation one step closer toward the clinical use.to Our data also suggest that intrasplenic hepatocyte transplantation may provide a means to induce allograft tolerance.