中文摘要：

目的探讨D5 Stat5a对人类前列腺癌细胞增殖的影响及其对胰岛素样生长因子结合蛋白7（IGFBP-7）表达的影响。方法常规培养人前列腺癌细胞（DU145及PC3）,并随机分为四组：对照组及三个实验组。对照组以不含外源基因的腺病毒感染,三个实验组细胞则以携带D5 Stat5a cDNA的腺病毒感染,其病毒感染增殖活性（MOI）依次为10、20及30。四组细胞均在培养液中加入催乳素（50 ng/m L）以启动细胞信号转导。运用MTS方法检测细胞增殖;实时定量PCR检测抑癌基因IGFBP-7及EZH2 mRNA水平;染色质免疫共沉淀技术（ChIP）检测IGFBP-7基因启动子区域组蛋白甲基化程度;蛋白质印迹法检测IGFBP-7蛋白表达。结果携带D5 Stat5a cDNA的腺病毒感染能够呈剂量依赖式地促进前列腺癌细胞（DU145及PC3）增殖。如PC3细胞,对照组细胞的相对活细胞数为（1.362±0.018）,三个实验组相对活细胞数分别为（1.453±0.022）（P〉0.05）、（1.649±0.020）（P〈0.05）及（1.829±0.027）（P〈0.05）。实时定量PCR及蛋白印迹证明D5 Stat5a过表达明显下调IGFBP-7的表达。DU145及PC3对照组细胞IGFBP-7 mRNA相对值分别为（0.796±0.023）及（0.871±0.046）,而感染携带D5 Stat5a cDNA的腺病毒（MOI 30）后,其IGFBP-7 mRNA相对值则分别下降到（0.148±0.019）（P〈0.01）及（0.078±0.021）（P〈0.01）。染色质免疫共沉淀分析（ChIP-Assay）证实,D5 Stat5a导致IGFBP-7基因启动子区域组蛋白甲基化（H3K27Me3）水平显著上升。DU145细胞对照组与实验组,其沉淀DNA比例分别为（0.0176±0.0030）%及（0.0650±0.0099）%,两者差异有高度统计学意义（P〈0.01）。此外,实时定量PCR检测表明,D5 Stat5a引起组蛋白甲基转移酶EZH2 mRNA大幅上升。DU145细胞对照组及D5 Stat5a感染组相对mRNA水平分别为（0.0033±0.0004）及（0.0160±0.0035）,两组比较差异有统计学意义（P〈0.05）。结论转录因子D5 Stat5a能够显

英文摘要：

Objective To clarify the effect of D5 Stat5 a on the proliferation of prostate cancer cells and the expression of insulin-like growth factor binding protein 7（IGFBP-7）.Methods Prostate cancer cells（DU145 and PC3） were randomly divided into four groups：control and three experimental groups.The cells were infected with adenovirus carrying or not carrying（Con） D5 Stat5 a cDNA at MOI 10,20 and 30 respectively for 120 min and cultured in the presence of prolactin（50 ng/m L） for further 48 h.The relative viable cell number was determined by MTS assay.Quantitative real time polymerase chain reaction（qRT-PCR） and Western blot were used to examine the expression of IGFBP-7.Chromatin immunoprecipitation（ChIP） assay was also applied to examine the effect of D5 Stat5 a on the trimethylation of histone 3 of IGFBP-7 gene promoter region.Results Adenovirus（carrying D5 Stat5a） infection induced proliferation of prostate cancer c ells in a dose-dependant manner.Taking PC3 as example,the relative viable cell number was（1.362±0.018） in control cells,while infection with adenovirus carrying D5 Stat5 a cDNA at MOI 10,20 or 30 increased the relative vialble cell number to（1.453 ±0.022）（P〈0.05）,（1.649±0.020）（P〈0.05）and（1.829±0.027）（P〈0.05）,respectively.q RT-PCR showed that the relative IGFBP-7 m RNA amounts were（0.796±0.023） and（0.871±0.046） in DU145 and PC3 control cells respectively,however,they decreased to（0.148±0.019）（P〈0.01） and（0.078±0.021）（P〈0.01） in DU145 and PC3 cells infected with virus carrying D5 Stat5 a cDNA at MOI 30,respectively.Increased trimethylation on histone 3 lysine 27（H3K27Me3） of IGFBP-7 gene promoter region and downregulated IGFBP-7 expression were also found in prostate cancer cells over-expressing D5 Stat5 a including DU145 and PC3 cells.Ch IP assay showed that the percentage of immunoprecipitated DNA in control cells and cells infected with D5 Stat5 a cDNA adenovirus at MOI 30 were（0.0176 ±0.0030）% a