HBx基因瞬时转染对 SUDHL-4细胞增殖的影响

摘要

Aim:To investigate the effects of HBx transient transfection on proliferation , cell cycle and apoptosis in lymphoma cell SUDHL-4.Methods:The HBx eukaryon expression vector pcDNA 3.1-X was established through recombi-nation DNA technique.The reconstituted plasmid pcDNA3.1-X and empty vector pcDNA3.1 were transfected into SUDHL-4 cells which were named SUDHL-4-HBx and SUDHL-4-con.Untransfected SUDHL-4 and SUDHL-4-con were used as control.The CCK8 experiment was performed to detect the reproductive ability of cells in 24, 48, 72 and 96 h,and growth curves were obtained .Cell apoptosis and cell cycle were detected by flow cytometry .Results: The reconstituted plasmid pcDNA3.1-X was identified by restriction enzyme digestion and nucleotide sequencing , the expressions of HBx mRNA and protein were confirmed in the SUDHL-4 cells transfected with HBx .Compared with SUDHL-4 group and SUDHL-4-con group, the proliferative capacity of cells in SUDHL-4-HBx group was obviously decreased .Flow cytometry analysis showed that HBx transfection significantly increased G 0/G1 phase proportion and cell apoptosis rate when comparing with SUDHL-4 and SUDHL-4-con groups(P<0.05).Conclusion:HBx transient transfection could inhibit SUDHL-4 cell proliferation by blocking cell cycle in G 0/G1 phase and increasing cell apoptosis .%目的：研究HBx基因瞬时转染对淋巴瘤细胞SUDHL-4增殖、细胞周期及凋亡的影响。方法：用分子克隆方法构建HBx基因的真核载体pcDNA3．1-X，利用脂质体转染法将pcDNA3．1-X转入淋巴瘤细胞SUDHL-4中，构建整合有HBx基因的淋巴瘤细胞株SUDHL-4-HBx，以SUDHL-4及转染空载体的细胞SUDHL-4-con作为对照，利用CCK8法检测细胞转染24、48、72和96 h后的增殖活性，绘制生长曲线；应用流式细胞仪检测细胞周期和凋亡情况。结果：重组质粒双酶切结果及测序结果表明成功构建pcDNA3．1-X，转染后SUDHL-4细胞有HBx mRNA及蛋白的表达。与SUDHL-4组及SUDHL-4-con组相比，SUDHL-4-HBx组细胞的增殖能力下降，G0／G1期细胞比例增加，S期细胞比例减少，凋亡率升高（P＜0．05）。结论：瞬时转染HBx基因后，SUDHL-4细胞增殖减缓，细胞周期阻滞于G0／G1期，凋亡率增加。