小RNA靶向激活p21^WAF1/CIP1基因表达对BEL-7402肝癌细胞增殖和凋亡的影响  

Activation of p21^WAF1/CIP1 by small activating RNA inhibits cell proliferation and induces apoptosis in BEL-7402 hepatoma cell

作　　者：吴志明[1] 黄欢[1] 刘卓炜[1] 郭琤琤[1] 蒋丽娟[1] 董培[1] 贾国金[2] 陈刚[2] 

机构地区：[1]中山大学肿瘤防治中心,广州510060 [2]复旦大学附属金山医院泌尿外科

出　　处：《中华医学杂志》2014年第20期1534-1538,共5页National Medical Journal of China

摘　　要：目的 探讨RNA激活技术上调p21^WAF1/CIP1基因的表达而影响人肝癌细胞株BEL-7402的增殖和凋亡能力.方法 将与p21^WAF1/CIP1基因启动子区域DNA序列互补的RNA分子（dsP21-322）转染入BEL-7402细胞中,采用qPCR法及Western印迹方法检测p21 WAF1/CIP1的表达;采用WST-1细胞增殖试剂检测细胞增殖情况;使用流式细胞仪评估肿瘤细胞凋亡情况并使用Western印迹方法检测相关分子表达水平的变化.结果 dsP21-322转染72 h后,BEL-7402细胞中p21^WAF1/CIP1表达显著上调;细胞增殖受到明显抑制,凋亡率36.86％（细胞的体积变小、变形,细胞核固缩,染色质凝聚呈深染,细胞出现皱缩、变圆、脱落）较对照组（11.51％、14.06％）相比明显升高.结论 RNA激活技术靶向上调p21^WAF1/CIP11基因表达并抑制细胞增殖、增加诱导肿瘤细胞凋亡,可作为肝细胞癌或其他恶性肿瘤基因治疗的一种有效手段.Objective To evaluate the anti neoplastic effects of p21^WAF1/CIP1 transcriptional activation induced by duplex RNAs in hepatocellular carcinoma （HCC） cell line BEL-7402.Methods Cells were treated with dsRNAs complementary to promoter sequences of p21^WAF1/CIP1.Quantitative polymerase chain reaction （qPCR） and Western blot were employed to detect the expression of p21.At various timepoints post-transfection,cell viability assay and apoptosis analysis were used to determine the effect of RNA activation.After transfection Western blot was also performed to detect the expression of BclxL,cleaved caspase-3,cleaved caspase-9 and cleaved PARP.Results DsP21-322 transfection significantly inhibited cell viability.And,at Day 5,dsP21-322 inhibited cell growth by 65.84% versus control.Flow cytometry revealed that dsP21-322 caused a significant increase of cell apoptosis.The total percent of apoptotic cells （UR ＋ LR） increased to 36.86% versus 11.51% and 14.06% in mocks and controls respectively.Such phenomena correlated with a decrease of anti-apoptotic protein Bcl-xL and an increase of cleaved caspase-3,cleaved caspase-9 and cleaved PARP.Conclusion Activation of p21 gene expression by saRNA may offer therapeutic benefits for HCC and other cancers.

关 键 词：癌 肝细胞 细胞增殖 P21^WAF1 CIP1 

分 类 号：R735.7[医药卫生—肿瘤]