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ＣＤ５９分子是广泛分布于各组织细胞表面的１８ｋＤａ糖蛋白，通过肌醇磷脂酰聚糖（ＧＰＩ）锚固于细胞膜，具有抑制同源补体攻膜复合物（ＭＡＣ）形成和参与介导Ｔ细胞活化等多种功能。本文应用ＲＴ－ＰＣＲ方法，从Ｊｕ－ｒｋａｔ细胞的总ＲＮＡ中扩增得到３９６ｂｐ的ｃＤＮＡ片段，经测序证实该片段包括２５ａａ信号肽在内的全部的ＣＤ５９编码序列。进一步将此ＣＤ５９的ｃＤＮＡ重组于逆转录病毒载体ｐＬＸＳＮ，电穿孔转染ＰＡ３１７细胞，并用病毒上清感染小鼠成纤维母细胞ＮＩＨ３Ｔ３及小鼠胸腺瘤细胞ＥＬ－４。经Ｇ４１８加压筛选，ＦＡＣＳ检测获得表达ＣＤ５９的阳性细胞克隆。补体杀伤实验结果表明：表达ＧＰＩ－型ＣＤ５９分子的ＮＩＨ３Ｔ３和ＥＬ－４细胞对人血清补体溶破的抵抗作用较空载体转染的非表达细胞明显增强。证实了用逆转录病毒载体可成功地将人ＣＤ５９基因导入异源细胞，使表达ＣＤ５９的异源细胞获得抑制人补体溶破的功能。本研究为探讨ＣＤ５９分子与细胞活化的关系及其信号转导机制建立了良好的细胞模型，并为进一步应用于异种器官移植，或对由于ＣＤ５９遗传缺损所致ＰＮＨ进行基因治疗奠定了基础。
The CD59 molecule is an 18 kDa glycoprotein widely distributed on the surface of various tissue cells. It is anchored to the cell membrane through inositol phosphatidylglycerols (GPIs) and inhibits the formation of homologous complement membrane formation (MAC) and is involved in the activation of T cells A variety of functions. In this paper, a 396-bp cDNA fragment was amplified by RT-PCR from total RNA of Ju-rkat cells. The entire CD59 coding sequence, including the 25aa signal peptide, was confirmed by sequencing. Further, the cDNA of CD59 was recombined into the retroviral vector pLXSN, electroporated into PA317 cells, and the mouse fibroblasts NIH3T3 and mouse thymoma cells EL-4 were infected with virus supernatant. The positive cells clones expressing CD59 were obtained by pressurized screening with G418 and detected by FACS. The results of complement-killing assay showed that the resistance of NIH3T3 and EL-4 cells expressing GPI-type CD59 molecules to complement-depleted human serum was significantly increased compared with the non-expressing cells transfected with empty vector. It is confirmed that the retrovirus vector can successfully transfer the human CD59 gene into the heterologous cells so that the heterologous cells expressing CD59 have the function of inhibiting human complement dissolution. This study established a good cell model for exploring the relationship between CD59 molecule and cell activation and its signal transduction mechanism and laid the foundation for further application in xenotransplantation or gene therapy of PNH caused by CD59 genetic defect.