CD200调控巨噬细胞活化参与异种肝移植急性血管性排斥的机制研究

以基因编辑猪为供体的异种肝移植，尽管克服了超急性免疫排斥，但仍表现为移植物内大量受体来源巨噬细胞浸润和剧烈炎症反应。研究表明，跨膜糖蛋白CD200可抑制单核-吞噬细胞激活，在多种疾病中发挥抗吞噬、抗炎等作用。申请人前期实施了GalT-KO猪-猴肝移植，并行RNA-seq发现：移植物内CD200表达下调，且受体来源巨噬细胞浸润明显，组织中凋亡细胞增加；相关性分析揭示巨噬细胞浸润程度和移植物损伤呈正相关；血清学证实炎性细胞因子亦分泌增加。体外猪内皮（EC）-人PBMCs共培养时，CD200过表达可减少EC凋亡以及炎性因子分泌。本项目拟建立猪源或人源CD200分子过表达的GalT-KO猪EC细胞系，利用异种EC-巨噬细胞混合培养体系、人源化小鼠异种EC移植+人单核、淋巴细胞输注、以及猪-猴肝移植等，探索和验证CD200负性调控异种急性血管性排斥的细胞分子机制，为供体猪基因改造提供有价值靶点。

Liver xenotransplantation from gene-edited pigs, despite overcoming hyperacute immune rejection, still presents a large number of receptor-derived macrophage infiltration and a severe inflammatory response within liver graft. Studies have shown that transmembrane glycoprotein CD200 can inhibit the activation of mononuclear-phagocytic cells, and play an anti-phagocytic and anti-inflammatory role in a variety of diseases. In the early studies of GalT-KO pig-to-monkey liver transplantation performed by applicant, the RNA-seq results showed that the expression of CD200 within the graft was down-regulated, the infiltration of receptor-derived macrophages was obvious, and the apoptotic cells in the tissue were increased. Correlation analysis revealed that the degree of macrophage infiltration was positively correlated with graft injury. Serology confirmed that the secretion of inflammatory cytokines also increased. In vitro pig endothelium (EC) - human PBMCs co-culture, CD200 over-expression can reduce EC apoptosis and inflammatory cytokine secretion. This project intends to establish GalT-KO pig EC cell lines over-expressing pig or human CD200, using pig EC-human macrophage mixed culture system, pig EC transplant within humanized mice + monocyte and lymphocyte infusion, and pig-to-monkey liver transplantation, to explore and validate cell-molecular mechanism of CD200 in negative regulation of xenogeneic acute vascular rejection, providing valuable target for donor pig genetic modification.