丙戊酸钠联合全反式维甲酸对HeLa细胞的杀伤机制研究  

Valproate acid combined with all-trans retinoic acid kills HeLa cells

作　　者：曹振平[1] 冯定庆[2] 徐嵘嵘[2] 李彩荣[2] 周颖[2] 凌斌 

机构地区：[1]安徽医科大学附属省立医院妇产科,合肥230001 [2]安徽医科大学附属省立医院安徽省分子医学重点实验室,合肥230001

出　　处：《肿瘤》2010年第10期827-827,共6页Tumor

基　　金：国家"十一五"科技支撑计划资助项目(编号:2008BAI57B00)

摘　　要：目的:探讨丙戊酸钠(valproate acid,VPA)联合全反式维甲酸(all-trans retinoic acid,ATRA)对子宫颈癌HeLa细胞的杀伤作用及其可能机制。方法:采用3 mmol/L VPA、1μmol/L ATRA、1μg/mL顺铂(cis-diamminedichloroplatinum,DDP)单独或联合作用于子宫颈癌HeLa细胞,对照组仅加溶媒;采用MTT法检测细胞生长抑制率;Hochest33258标记法观察细胞凋亡情况;RT-PCR法检测凋亡相关基因p53、信号转导及转录激活因子(signal transducers and activators of transcription,STAT3)、p53上调凋亡调控因子(p53 up-regulated modulator of apoptosis,PUMA)及Bax、Bcl-2和Bcl-XL的mRNA表达;Western印迹法检测p53、STAT3及磷酸化STAT3(phosphorylated STAT,p-STAT3)蛋白表达的变化。结果:VPA对子宫颈癌HeLa细胞有生长抑制作用,且VPA联合ATRA优于单独用药。Hochest33258标记发现,用药后细胞出现核碎裂等典型的凋亡改变,VPA+ATRA组细胞凋亡率显著高于对照组。RT-PCR结果显示,VPA可在mRNA水平上增强p53、PUMA及Bax的表达,减弱STAT3的表达,且联合用药组效果更明显,并呈时间依赖关系;而Bcl-XL及Bcl-2的表达无明显变化。Western印迹结果显示,VPA联合ATRA显著增强子宫颈癌HeLa细胞中p53蛋白表达,降低STAT3和p-STAT3蛋白表达水平,亦呈现明显的时效关系。结论:VPA单独及联合ATRA在体外可有效杀伤子宫颈癌HeLa细胞株。推测其可能的作用机制是诱导p53表达水平上调,STAT3及p-STAT3表达水平下调。Objective:To investigate the action mechanism by which valproate acid(VPA) combined with all-trans retinoic acid(ATRA) kills HeLa cells.Methods:HeLa cells were treated with 3 mmol/L VPA,1 μmol/L ATRA,and 1 μg/mL cis-diamminedichloroplatinum(DDP) alone or in combination.The control group was treated with vehicle only.MTT assay was used to assess inhibitory rate of proliferation.The apoptoses of the cells were observed using Hochest33258 staining.The mRNA expressions of p53,signal transducers and activators of transcription(STAT-3),and p53 up-regulated modulator of apoptosis(PUMA),Bax,Bcl-2,Bcl-XL were detected using RT-PCR and the protein expressions of p53,STAT3,and phosphorylated STAT(p-STAT3) were determined using Western blotting.Results:The proliferation of HeLa cells was significantly inhibited by VPA combined with ATRA.The effect of combination treatment was stronger than individual treatment(P0.05).Hochest33258 staining showed that HeLa cells showed the typical apoptotic morphologic changes such as chromatin clumping,nuclear fragmentation,and chromatin condensation.The apoptotic rate in VPA+ATRA group was significantly higher than that in the control group.RT-PCR showed that VPA enhanced the mRNA expression of p53,PUMA and Bax,and reduced the expression of STAT3.The efficacy was time-dependent and more powerful in combination therapy group.No significant change was observed in the expressions of Bcl-XL and Bcl-2.Western blotting showed that VPA plus ATRA significantly increased the expression of p53 and decreased the expressions of STAT3 and p-STAT3 in a time-dependent manner.Conclusion:VPA alone or in combination with ATRA effectively kills cervical cancer HeLa cells in vitro,which is probably related with up-regulation of p53 and down-regulation of STAT3 and p-STAT3.

关 键 词：丙戊酸钠 全反式维甲酸对 HELA细胞 杀伤 机制研究 HeLa cells RETINOIC acid p-STAT3 子宫颈癌 蛋白表达水平 VPA ATRA p53 细胞凋亡 转导及转录激活因子 细胞生长抑制率 WESTERN印迹法 用药 时间依赖关系 生长抑制作用 

分 类 号：R73[医药卫生—肿瘤]