中文摘要：

目的探讨在T系急性白血病的Jurkat细胞中,GPI类锚固蛋白CD59通过LAT介导的细胞增殖、活化、凋亡等相关的生物学效应,研究GPI锚固类膜蛋白分子对细胞调控的作用方式。方法分别构建LAT-EGFP、neg-EGFP融合蛋白慢病毒载体,转染Jurkat细胞,建立稳定表达株（LAT-EGFP作为实验组,neg-EGFP转染空病毒组作为对照组）。利用CD59单克隆抗体刺激实验组及对照组细胞后,通过CCK-8检测细胞的增殖活性,利用流式细胞术检测细胞凋亡状况,并通过激光共聚焦显微镜观察LAT与CD59分子在细胞中的定位表达。最后利用免疫印迹技术检测细胞内相关信号转导蛋白的表达情况。结果 CD59单抗交联刺激后,细胞增殖活性增加。实验组细胞出现明显晚期凋亡甚至坏死现象。而免疫荧光显示LAT-EGFP组细胞中LAT分子高亮聚集于脂筏区,相较于对照组细胞呈现明显的戒指环状。Western blot显示,CD59抗体交联后,ZAP70、Fyn、LCK的表达均下降且LATEGFP组蛋白表达量低于对照组。结论 GPI锚固蛋白CD59通过T细胞活化连接蛋白LAT对T细胞信号转导发挥正向调控作用,转染LAT-EGFP的Jurkat细胞处于活化状态,LAT呈戒指环状定位于脂筏。

英文摘要：

This study desigend to observe the biological effects of GPI anchor protein CD59 in cell proliferation, activation and apoptosis via LAT on Jurkat cell and to further study the regulation way of GPI anchor proteins. LAT-EGFP and neg-EGFP fusion protein lentivirus vectors were constructed and transfected into Jurkat cells to establish stably expressing cell line. Then CCK8 and flow cytometry were used to detect the cell proliferation activity and cell apoptosis; laser scanning confocal microscope was used to observe the location of LAT; Western blot was used to detect the levels of ZAP70, Fyn and Lck. The data revealed that after CD59 antibody cross-linked stimulation, the cell proliferation activity increased; the LAT-EGFP group cells appeared obvious late apoptosis, even necrosis. And immunofluorescence showed that LAT molecules highlighted in the lipid rafts of LAT-EGFP group which present obvious ring shape; Western blot showed that the levels of ZAP70,Fyn, and Lck in LAT-EGFP group were lower than those of the neg-EGFP group after CD59 antibody cross-linking. Therefore, we concluded that GPI anchor protein CD59 plays a positive role on T cell signal transduction via LAT. Jurkat cells that transfected with LAT-EGFP are in active state, and the LAT present clusters form and locate in lipid rafts.