猫肉瘤相关蛋白对前列腺癌细胞周期及凋亡的影响及其机制  

Influence of feline sarcoma-related protein on cell cycle and apoptisis of human prostate cancer cells and the mechanism

作　　者：胡旭东[1] 于德新[1] 张志强[1] 谢栋栋[1] 王毅[1] 张涛[1] 

机构地区：[1]安徽医科大学第二附属医院泌尿外科,合肥230601

出　　处：《中华实验外科杂志》2015年第6期1288-1290,共3页Chinese Journal of Experimental Surgery

摘　　要：目的 探讨小RNA干扰技术（siRNA）沉默猫肉瘤相关蛋白（Fer）的表达对前列腺癌细胞周期及凋亡的影响及其机制.方法 设计合成3组针对Fer特异的siRNA转染人前列腺癌PC-3细胞,应用实时定量反转录聚合酶链反应（RT-qPCR）和Western blot法检测转染前后PC-3细胞中Fer mRNA及蛋白表达水平,并筛选出干扰效率最高的小干扰RNA进行后续实验,用流式细胞仪检测各组细胞的细胞周期变化并分析细胞凋亡,观察Fer基因表达抑制后细胞周期及凋亡相关基因表达水平的变化.结果 利用RT-qPCR检测siRNA-1、siRNA-2、siRNA-3组的Fer mRNA相对表达量分别为0.288、0.369、0.623,与阴性对照（NC）组比较差异均具有统计学意义（以NC组FermRNA表达量为l,P＜0.05）,Western blot方法同样证实3组小RNA干扰组显著抑制PC-3细胞的Fer蛋白的表达,其中siRNA-1干扰效率最高（P＜0.01）.流式细胞仪结果显示RNA干扰（Si）组细胞G0/G1期细胞比例为（52.340±2.049）％,显著高于NC组（40.230±2.065）％,而在S期细胞比例为（35.040 ±3.810）％,显著低于NC组（50.650±3.693）％,两者与NC组比较差异均具有统计学意义（P＜0.05）;Si组细胞的凋亡率为（33.6±3.3）％,显著高于NC组的凋亡率（7.8±1.0）％,差异有统计学意义（P＜0.05）;同时随着Fer表达的抑制,细胞周期素（Cyclin） D1,B细胞淋巴瘤/白血病-2 （bcl-2）的表达显著下调,p21和活化型半胱氨酸-3的表达显著上升,与NC组比较差异均有统计学意义（P＜0.05）.结论 Fer-siRNA可抑制Fer在前列腺癌细胞中的表达,抑制前列腺癌细胞G1/S期转化,从而诱导肿瘤细胞凋亡,其机制可能与调控细胞周期及凋亡相关基因的表达有关.Objective To investigate the influence of small interference RNA （siRNA） silencing feline sarcoma-related protein （Fer） on cell cycle and apoptosis of prostate cancer cells.Methods PC-3 cells were transfected with three siRNAs targeting human Fer by liposome transfection reagent.Real-time reverse transcriptase-polymerase chain reaction （RT-qPCR） and Western blotting were used to detect the expression of Fer mRNA and protein respectively,and the most efficient siRNA was screened out for the following experiments.The cell cycle and apoptosis rate were analyzed by flow cytometry.Furthermore,we observed the expression levels of the cell cycle-and apoptosis-related genes in the transfected cells with down-regulation of the Fer expression.Results RT-qPCR showed that Fer mRNA relative expression levels of three siRNAs was 0.288,0.369 and 0.623,respectively,which were significantly lower than those in the negative control group （l,P ＜ 0.05）.Western blotting also demonstrated that the expression of Fer protein in PC-3 cells transfected with three siRNAs could significantly inhibited,and siRNA-1 was selected as the most efficient siRNA （P ＜ 0.01）.Flow cytometry revealed the percentage of G0/G1 phase cells in RNAi （Si） group was （52.340 ± 2.049） ％,significantly higher than that in negative control group [（40.230 ± 2.065） ％] （P ＜ 0.05）,and that of S phase cells was （35.040 ± 3.810） ％,significantly lower than that in the negative control group [（50.650 ± 3.693） ％] （P ＜ 0.05）.The apoptosis rate in Si group was （33.6 ± 3.3） ％,significanfly higher than that in negative control group [（7.8 ± 1.0） ％] （P ＜0.05）.Meanwhile,with the inhibition of Fer,the expression levels of Cyclin D1 and B cell lymphoma/leukemia-2 （bcl-2） were reduced obviously,and p21 and cleaved Caspase-3 upregulated obviously as compared with negative control group （P ＜ 0.05）.Conclusion Fer-siRNA could significantly inhibit the expression of Fer and the progression of

关 键 词：前列腺癌 猫肉瘤相关蛋白 小干扰RNA 细胞周期 脱噬作用 

分 类 号：R737.25[医药卫生—肿瘤]