抗HPV16E6核酶增加宫颈癌CaSKi细胞对顺铂敏感性的研究  

The Effect of Anti-HPV16 E6-Ribozyme on Sensitivity of Cervical Carcinoma Cell Line to Cisplatin

作　　者：饶智国[1] 张积仁[1] 郑燕芳[1] 李鹏[1] 周惠[2] 屈良鹄[2] 

机构地区：[1]第一军医大学珠江医院肿瘤中心,广州510282 [2]中山大学生命科学学院真核生物学实验室,广州510275

出　　处：《中国肿瘤生物治疗杂志》2001年第4期264-268,共5页Chinese Journal of Cancer Biotherapy

基　　金：广东省自然科学基金 ( 96 0 5 8)资助项目

摘　　要：目的 :研究抗HPV16E6核酶 (Ribozyme)导入宫颈癌CaSKi细胞对顺铂 (DDP)敏感性的影响。 方法 :以脂质体法将抗HPV16E6 Ribozyme、空载体质粒分别导入CaSKi细胞 ,命名为CaSKi R ,CaSKi P细胞。点杂交检测核酶在细胞中的表达 ,Northern杂交检测 3种细胞中E6基因的表达。检测 3种细胞的细胞生长曲线和克隆形成实验 ,MTT法检测 3种细胞对DDP的敏感性 ,Annexine/PI双标法检测细胞凋亡率 ,流式细胞术检测Bcl 2 ,P5 3,Bax蛋白表达。结果 :点杂交证实核酶能在CaSKi R细胞中稳定表达 ,Northern杂交证实CaSKi R中表达E6较CaSKi P ,CaSKi明显降低。与CaSKi P和CaSKi细胞比较 ,CaSKi R细胞的生长速率明显减慢和克隆形成率下降 ,对DDP的敏感性明显增加 (P <0 .0 1) ,凋亡率明显增加 (P <0 .0 1) ,P5 3,Bax蛋白表达明显增加 ,Bcl 2蛋白表达明显减少 (P <0 .0 1) ,CaSKi P和CaSKi细胞比较无差异 (P >0 .0 5 )。结论 :抗HPV16E6 Ribozyme抑制了CaSKi R细胞的生长和克隆形成率 ,CaSKi R细胞对顺铂的敏感性增加。Objective: To investigate the effect of anti-HPV16 E6-ribozyme on sensitivity of cervical carcinoma cells line to cisplatin. Methods: With the method of lipofectin transfection, the anti-HPV16E6-ribozyme and the eucaryotic expressing plasmids per se were transfected into CaSKi cells, which were named CaSKi-R, CaSKi-P respectively. The expression of ribozyme in transfected cells was observed by RNA dot blot. The amounts of E6 mRNA in the three kinds of cells were detected by Northern blot. The growth curve and colony formation test of three cell types were detected. Their sensitivity to cisplatin were examined by MTT assay. The apoptosis rates of each cell and expressions of P53, Bcl-2 and Bax was determined by PI/Annexin V stained methods and flow cytometry analysis.Results: Anti-HPV16E6-ribozyme can be expressed stably in transfected CaSKi cells. Northern blot showed that E6 mRNA was less in CaSKi-R than in CaSKi. The growth rate and cloning efficiency of CaSKi-R cells were lower than that of CaSKi and CaSKi-P cells. The sensitivity and apoptotic rates of CaSKi-R cells to cisplatin were higher than that of CaSKi and CaSKi-P cells( P <0.01).The expression of P53, Bax protein was upregulated and the expression of Bcl-2 protein downregulated compared to that of CaSKi and CaSKi-P cells( P <0.01).Conclusion: Anti-HPVE6-ribozyme inhibited CaSKi-R cell growth, cloning efficiency and increased the sensitivity to cisplatin.

关 键 词：核酶 人乳头瘤病毒 顺铂 药物敏感性 子宫颈肿瘤 VPV16E6 CASKI细胞 

分 类 号：R737.33[医药卫生—肿瘤]