The genetic regulation of natural killer-mediated cell lysis in xenograft

本研究利用HLA-G和HLA-E基因转染仓鼠卵巢肿瘤细胞（CHO）和猪内皮细胞（SEC），研究了HLA-G和HLA-E对NK细胞直接杀伤活性的影响（G1和G3），制备HLA-E和β2-巨球蛋白，亚克隆入pCAGGS HLA-G（G1）和β2-巨球蛋白（β 2-macroglobulin，β2-macroglobulin）的表达得到证实。此外，β2-微球蛋白与HLA-G1共转染使表达水平提高了数倍。HLA-E的表达用流式细胞仪和北方印迹法检测。用HLA-G的信号肽取代HLA-E的信号肽进行表达，并在CHO或SEC上检测转染分子对NK细胞介导的溶解的改善作用，作为体外不协调异种移植物的延迟型排斥模型。HLA-G1和HLA-E可显著抑制NK细胞介导的SEC裂解，HLA-G和/或HLA-E在细胞表面的表达为克服NK细胞介导的异种移植免疫提供了新的途径。

We have investigated the effect of HLA-G and HLA-E to NK-mediated direct cytotoxicity, using hamster ovarian tumor cells (CHO) or swine endothelial cell (SEC) transfectants with these molecules.At first, cDNA of human full-length HLA-G (G1 and G3), HLA-E and β2-macroglobulin were prepared and subcloned into the pCAGGS (β-actin promoter).The surface expression of HLA-G (G1) and β2-macroglobulin were confirmed. In addition, co-transfection of β2-microglobulin with HLA-G1 enhanced the level of expression by several times. On the other hand, HLA-G3 was not expressed on the SEC surface.Concerning HLA-E, the expression was checked by FACS and northern blot. The signal peptide of HLA-E was substituted with that of HLA-G for certain expression.Amelioration of NK cell-mediated lysis by the transfectant molecules on CHO or SEC was then tested as an in vitro delayed-type rejection model of a discordant xenograft. HLA-G1 and HLA-E significantly suppressed the NK cell mediated SEC lysis.The expression of HLA-G and/or HLA-E on the cell surface serves as a new approach in overcoming NK-mediated immunity to xenografts.