Role of CD47 in xenograft rejection by macrophages

描述（由申请人提供）：猪异种移植有可能解决人类器官供体日益短缺的问题。由于供者和宿主之间广泛的分子不相容性，先天免疫在异种移植排斥中比在同种异体移植排斥中起更大的作用。CD 47广泛表达并作为SIRP？的配体，巨噬细胞和DC上的抑制性受体。在目前的资助期间，我们证明，缺乏跨物种的相互作用，在CD 47-SIRP？这一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物的排斥。移植CD 47缺陷细胞诱导同基因野生型（WT）小鼠快速先天免疫激活。此外，CD 47-SIRP？需要信号来抑制受体CD 11 hiCD 8？-供体特异性输血（DST）后DC活化和诱导耐受。基于这些和其他数据中提出的申请，我们假设，捐助者CD 47和主机SIRP之间的相互作用？对控制SIRP的激活至关重要？这种相互作用的缺乏激活了宿主巨噬细胞和DC，因此刺激了抗供体T细胞应答。在这里，我们提出了三个具体目标来验证我们的假设。目的：1.阐明CD 47-SIPR？SIRP？+调节中的信号传导异种肝细胞移植后天然免疫细胞的激活我们将CD 47 KO小鼠肝细胞移植到同基因WT小鼠中，以分析先天免疫细胞活化和仅由CD 47差异诱导的移植物排斥反应。我们还将人CD 47转基因与对照猪肝细胞移植到人源化小鼠中，以确定人CD 47表达是否可以抑制人对猪异种移植物的先天免疫应答。目的2是确定“缺失的CD 47”诱导的天然免疫细胞活化在肝细胞移植后T细胞异种免疫反应中的作用。我们将首先在同种异体移植模型中解决这个问题，其中CD 47 KO小鼠将用于模拟CD 47不相容的异种供体。我们将通过体外和过继性T细胞转移试验检查抗供体T细胞应答。然后，我们将使用具有功能性人免疫系统的人源化小鼠研究猪肝细胞上人CD 47表达减弱人T细胞异种免疫应答的潜力。目的3是确定供体细胞上的CD 47促进DST加共刺激阻断治疗的受体中耐受诱导的机制。我们将鉴定在CD 47缺陷DST后快速活化的DC和巨噬细胞群体及其在T细胞活化和耐受诱导中的作用。我们将在CD 47缺陷心脏同种异体移植的小鼠模型和猪-小鼠异种移植模型中解决这些问题，该模型涉及从人CD 47转基因猪到人源化小鼠的DST。这些研究有望为CD 47不相容性激活先天性和适应性异种免疫反应的机制以及使用人CD 47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提供重要见解。 公共卫生相关性：强烈的免疫排斥反应是临床异种移植的主要障碍，这是解决世界范围内人类器官供体严重短缺的一个可能的解决方案。该提案旨在进一步了解异种免疫反应的机制，并制定改善异种移植物存活的策略。

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. PUBLIC HEALTH RELEVANCE: The robust immunological rejection poses a major obstacle to clinical xenotransplantation, a possible solution to the severe worldwide shortage of human organ donors. This proposal aims to further understand the mechanisms of xenoimmune responses and to develop strategies for improving xenograft survival.