摘要
目的观察高迁移率族蛋白A2(HMGA2)对肾癌细胞ACHN周期及凋亡的影响,为肾肿瘤后续机制探讨提供根据。方法培养肾癌细胞ACHN,选择HMGA2三个不同的基因片段分别为HMGA2-siRNA1、HMGA2-siRNA2、HMGA2-siRNA3与虚拟基因片段(Mock-siRNA),利用RNA干扰技术,瞬时转染肾癌ACHN细胞,采用qRT-PCR和蛋白质印迹法检测HMGA2 mRNA及蛋白表达;PI染色及流式细胞仪检测干扰后ACHN细胞周期的变化。Annexin V-FITC检测干扰后ACHN细胞凋亡效应的影响。结果成功构建使HMGA2基因沉默的肾癌细胞株,并选择HMGA2-siRNA2进行后续试验。细胞周期分析表明,进入Go/G1期的细胞明显增多,而G2/M期的细胞减少,细胞增殖周期阻滞在G1期(P<0.05)。沉默HMGA2基因表达后,肾癌ACHN细胞呈现出明显的早期和晚期凋亡,总体凋亡率从5.69%上升到16.52%(P<0.05)。结论用HMGA2-siRNA干扰ACHN细胞,细胞的周期发生紊乱,凋亡情况明显增强,说明HMGA2在肿瘤的发生、发展、细胞周期和凋亡过程中发挥着重要作用,有望成为治疗肾癌的一个重要靶点,深入研究HMGA2基因对肾癌的靶向治疗具有深远意义。
Objective To investigate the effects of high mobility group A2(HMGA2)on the cell cycle and apoptosis of renal carcinoma ACHN cells.Methods Human renal carcinoma cell lines ACHN was cultured.Three different gene fragments of HMGA2 were selected as HMGA2-siRNA1,HMGA2-siRNA2,HMGA2-siRNA3 and Mock-siRNA,and the ACHN cells were transfected by RNA interference.The mRNA and protein expression of HMGA2 were detected by qRT-PCR and western blot.The changes in cell cycle after transfection in ACHN cells were detected by PI staining and flow cytometry.The effects on apoptosis in ACHN cells were detected by Annexin V-FITC.Results The specific silencing HMGA2 expression of renal carcinoma cell line was successfully constructed,and HMGA2-siRNA2 was selected for subsequent tests.The cell cycle analysis demonstrated that more cells entered the G/Gphase,while less cells entered the G/M phase,showing that the cellular proliferative cycle was blocked in the Gphase(P<0.05).When HMGA2 gene expression was silenced,obvious apoptosis in the early phase and later phase were observed in renal carcinoma ACHN cells.The total apoptotic rate increased from 5.69%to 16.52%(P<0.05).Conclusions When HMGA2-siRNA was used to interfere ACHN cells,the cell cycle was disturbed and apoptosis was significantly enhanced.HMGA2 plays an important role in tumorigenesis,the cell cycle and apoptosis,and is expected to become an important target in the treatment of renal carcinoma.The intensive investigation of the HMGA2 gene may have a far-reaching significance for gene therapy in renal carcinoma.
作者
吕光耀
蒲琳
陈乾
付启忠
刘险峰
董圣芳
杨建勋
刘颖
LV Guangyao;PU Lin;CHEN Qian;FU Qizhong;LIU Xianfeng;DONG Shengfang;YANG Jianxun;LIU Ying(Department of Urology, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China)
出处
《现代泌尿生殖肿瘤杂志》
2021年第2期97-102,共6页
Journal of Contemporary Urologic and Reproductive Oncology
