DEVELOPEMENT OF NEW IMMUNOSUPPRESSION ON XENOGENEIC TRANSPLANTATION BY COMPLEMENT INHIBITORS

在Dicordant异种移植中，需要克服的最严重的问题是超急性排斥反应，其中补体对异种内皮细胞起主要作用。最近，我们报道了将CD 59导入小鼠成纤维细胞，使其对人补体攻击具有抵抗性。本研究的目的是通过定点突变来研究CD 59抗原的N-糖基化的作用。其次，我们通过在犬肾中引入和表达人CD 59基因来评估异种移植物的修饰。构建了突变型CD 59表达载体，其中Ser-20变为Ala，并将其导入小鼠A31细胞系。对突变型CD 59在小鼠A31细胞系上表达的功能分析表明，突变型pSR α CD 59/A31比pSR α CD 59/A31和pSR/A31更能抵抗人补体攻击。野生型和突变型CD 59的分子量分别为20 kd和14 kd。利用日本血凝病毒（HVJ）脂质体，其中包封的DNA和非组蛋白染色体蛋白高迁移率组1（HMG 1），我们评价了这种混合物通过人CD 59基因转移到狗的肾脏。RT-PCR分析表明CD 59在犬肾脏中有较好的转录。免疫组织化学分析显示，在犬肾小球细胞中检测到CD 59。总之，我们相信，这种技术的出生后的动物将提供一个合适的异种移植给人类。

In Dicordant xenotransplantation, the most serious problem to be overcome is hyperacute rejection, in which complement play a major role on xenogeneic endothelial cells. Recently, we reported the introduction of CD59 to mouse fibroblast showed resistance against human complement attacking. The aim of this study is to investigate the role of N-glycosylation of CD59 antigen by site-directed mutagenesis. Second, we have evaluated the modification of the xenograft by introduction and expression of human CD59 gene in the canine kidney. Mutant CD59 expression vector, in which Ser-20 was changed to Ala, was constructed and introduced into a mouse A31 cell line. The functional analysis of the mutant CD59 expressed on mouse A31 cell line indicated that the mutant pSRalphaCD59/A31 was more registant to human complement attack than the pSRalphaCD59/A31 and pSR/A31. The molecular weight of wild-type and mutant CD59 revealed 20kd and 14kd, respectively. Using the hemagglutinating virus of Japan (HVJ) liposomes, which encapsulated both DNA and nonhistone chromosomal protein high-mobility group1 (HMG1), we evaluated thismixture by human CD59 gene transfer to the canine kidney. RT-PCR analysis indicated CD59 was well transcribed in canine kidney. Immunohistchemical analysis revealed that CD59 was detected in the canine renal glomerular cells. In coclusion, we beleive that such techniques for postnatal animals will provide a suitable xenograft for humans.