Role of CD47 in xenograft rejection by macrophages

描述（由申请人提供）：猪的异种移植有可能解决人类器官供体日益短缺的问题。由于供体和宿主之间存在广泛的分子不相容，先天免疫在异种移植排斥反应中起着比同种异体移植排斥反应更大的作用。CD47普遍表达，并作为SIRP？巨噬细胞和树突状细胞的抑制受体。在目前的资助期内，我们证明了CD47-SIRP？该途径在很大程度上解释了巨噬细胞介导的造血和非造血细胞异种移植的排斥反应。同源野生型（WT）小鼠移植cd47缺陷细胞可诱导快速先天免疫激活。此外,CD47-SIRP吗?信号抑制受体CD11hiCD8？- DC激活和诱导供者特异性输血（DST）后耐受。基于这些和应用中提供的其他数据，我们假设供体CD47和宿主SIRP？是控制SIRP激活的关键？并且这种相互作用的缺失激活宿主巨噬细胞和dc，从而刺激抗供体T细胞反应。在这里，我们提出3个具体目标来检验我们的假设。目的1是阐明CD47-SIPR的作用。信号在SIRP调控中的作用？+肝细胞异种移植后的先天免疫细胞激活。我们将CD47 KO小鼠肝细胞移植到同基因WT小鼠体内，分析CD47差异引起的先天免疫细胞活化和移植物排斥反应。我们还将把人CD47转基因与对照的猪肝细胞移植到人源化小鼠体内，以确定人CD47表达是否会抑制人对猪异种移植物的先天免疫反应。目的2是确定“缺失CD47”诱导的先天免疫细胞激活在肝细胞移植后T细胞异种反应中的作用。我们将首先在同种异体移植模型中解决这个问题，在该模型中，CD47 KO小鼠将被用来模拟CD47不相容的异种供体。我们将通过体外和过继T细胞转移试验来检查抗供体T细胞反应。然后，我们将利用具有功能性人类免疫系统的人源化小鼠，研究人CD47在猪肝细胞上表达以减弱人T细胞异种免疫反应的潜力。目的3是确定供体细胞上的CD47促进DST +共刺激阻断治疗受体耐受诱导的机制。我们将确定cd47缺陷DST后快速激活的DC和巨噬细胞群体及其在T细胞激活和耐受性诱导中的作用。我们将在cd47缺陷心脏移植小鼠模型和猪-小鼠异种移植模型中解决这些问题，该模型涉及从人类cd47转基因猪到人源化小鼠的DST。这些研究有望对CD47不相容激活先天和适应性异种免疫反应的机制提供重要的见解，以及使用人类CD47转基因猪作为供体促进异种耐受诱导和异种移植物存活的潜力。

DESCRIPTION (provided by applicant): Xenotransplantation from pigs has the potential to resolve the growing shortage of human organ donors. Because of the extensive molecular incompatibilities between the donor and host, innate immunity plays a much greater role in xenograft rejection than in allograft rejection. CD47 is ubiquitously expressed and serves as a ligand of SIRP?, an inhibitory receptor on macrophages and DCs. During the current funding period, we demonstrated that the lack of cross-species interaction in CD47-SIRP? pathway largely accounts for macrophage-mediated rejection of hematopoietic and non-hematopoietic cellular xenografts. Transplantation of CD47-deficient cells induces rapid innate immune activation in syngeneic wild-type (WT) mice. Furthermore, CD47-SIRP? signal is required to repress recipient CD11hiCD8?- DC activation and induce tolerance after donor-specific transfusion (DST). Based on these and other data presented in the application, we hypothesize that the interaction between donor CD47 and host SIRP? is essential for controlling activation of SIRP?+ macrophages and DCs, and that the absence of this interaction activates host macrophages and DCs, hence stimulating anti-donor T cell responses. Here, we propose 3 specific aims to test our hypothesis. Aim 1 is to elucidate the role of CD47-SIPR? signaling in the regulation of SIRP?+ innate immune cell activation after hepatocyte xenotransplantation. We will transplant CD47 KO mouse hepatocytes into syngeneic WT mice to analyze innate immune cell activation and graft rejection induced solely by CD47 disparity. We will also transplant human CD47 transgenic vs. control pig hepatocytes into humanized mice to determine whether human CD47 expression may inhibit human innate immune responses to pig xenografts. Aim 2 is to determine the role of "missing CD47"-induced innate immune cell activation in T cell xenoresponses after hepatocyte transplantation. We will first address this question in allotransplant models, in which CD47 KO mice will be used to mimic CD47-incompatible xenogeneic donors. We will examine anti-donor T cell responses by in vitro and adoptive T cell transfer assays. We will then investigate the potential of human CD47 expression on pig hepatocytes to attenuate human T cell xenoimmune responses using humanized mice with a functional human immune system. Aim 3 is to determine the mechanisms by which CD47 on donor cells facilitates tolerance induction in DST plus costimulatory blockade-treated recipients. We will identify the DC and macrophage populations that are rapidly activated after CD47-deficient DST and their roles in T cell activation and tolerance induction. We will address these questions in a mouse model of CD47-deficient cardiac allotransplantation, and a pig-to-mouse xenotransplantation model that involves DST from human CD47-transgenic pigs into humanized mice. These studies are expected to provide significant insights into the mechanisms by which CD47 incompatibility activates innate and adaptive xenoimmune responses, and the potential of using human CD47 transgenic pigs as donors to facilitate xenotolerance induction and xenograft survival.