The development of xenograft

我们建立了几个猪内皮细胞（SEC）系，通过转染cDNA表达人MCP、DAF和MCP/DAF杂交，并评估了这些转染分子在c介导的细胞裂解中的功能，作为猪对人异种移植物的体外超急性排斥反应模型。1) DAF可有效保护SEC免受c介导的细胞裂解。特别是在高表达克隆中，抑制率达到80%以上。另一方面，MCP也能有效抑制C活性，但作为c3步调节剂的效率不如DAF。杂交显示约60%的c介导的细胞裂解抑制。其效果明显优于单独使用MCP，但由于各克隆中转染分子的表达率不同，与单独使用DAF效果不明显。2) CD59在充分表达时对SEC有保护作用。然而，DC59在保护SEC膜免受人C攻击方面不如DAF有效，因为SEC在人补体级联的C5b-8步被裂解。为了获得高水平表达人mcp的转基因动物，人们对其启动子和增强子进行了多项研究。通过PCR和Southern blotting检测，我们获得了11个只缺失polyA信号的独立MCP转基因小鼠系和5个不缺失整个3'UT的MCP转基因小鼠系。MCP在携带3′ut长度为120bp的MCP cDNA的小鼠中表达量很少，而携带3′ut长度为120bp的MCP cDNA的小鼠在许多器官中表达量都很明显；尤其是在肌肉、心脏和胰腺中。

We established several swine endothelial cell (SEC) lines, expressing human MCP,DAF,and an MCP/DAF hybrid by transfection of cDNA,and assessed the function of these transfectant molecules on C-mediated cell lysis as an in vitro hyperacute rejection model of swine to human discordant xenograft.1) DAF is quite effective to protect SEC from C-mediated cell lysis. Especially, over 80% suppression was observed in high expression clone. On the other hand, MCP is also effective in suppressing C activity, but the less efficient as the C3-step regulator than DAF.Hybrid showed approximate 60% of suppression of C-mediated cell lysis. It is more effective than MCP alone clearly, but indistinct from DAF alone because of the difference in the expression rate of transfectant molecules in each clone.2) CD59 is effective in protection of SEC when it is sufficiently expressed. However, DC59 is not so efficient in protecting SEC membrane from human C attack as DAF,because SEC is lysed at the C5b-8 step of the human complement cascade.TRANSGENICTo obtain the transgenic animals expressing human-MCP at high level, several studies concerning the promoter and enhancer have done. We obtained 11 independent lines of MCP transgenic mice lacking just the polyA signal and 5 lines without the entire 3'UT,as assessed by PCR and Southern blotting. While the expression of MCP in mice carrying MCP cDNA with 120bp of 3'UT was minimal, that in mice carrying MCP cDNA without total 3'UT was evident in many organs ; especially in muscle, heart, and pancreas.