RNA干扰对宫颈癌中HPV16 E6癌基因的抑制作用

[Inhibition of HPV16 E6 oncogene in cervical cancer by RNA interference].

作者信息

Niu Xiao-yu, Peng Zhi-lan, Duan Wei-qiang, Wang He, Chen Yue-yue, Fan Yu-juan, Cheng Yi-fan, Qian Xiao-lei

机构信息

Department of Obstetrics and Gynecology, West China Second Hospital, Sichuan University, Chengdu 610041, China.

出版信息

Sichuan Da Xue Xue Bao Yi Xue Ban. 2006 Jan;37(1):14-8.

PMID:16468632

Abstract

OBJECTIVE

The efficiency of HPV16 E6 gene silenced by RNA interference in vitro and in vivo was assessed.

METHODS

The specific siRNA of HPV16 E6 was designed and transfected into CaSki cells by liposome. Cell apoptotic rates and the changes in HPV16 E6 mRNA and protein before and after transfection were measured. Cervical cancer nude mice models were set up, siRNA was injected directly into subcutaneous tumor. The function of siRNA was evaluated by the changes in tumor volume, HPV16 E6 protein expression and apoptosis of tumor cells.

RESULTS

In vitro research, the cell apoptotic rates were 7.7%, 11.8%, 37.4% and 12.6% respectively at 24 h, 48 h, 5th day and 9th day after transfection. The HPV16 E6 mRNA was reduced by 77%, 83%, 59% and 41% at 24 h, 48 h, 5th day and 9th day after transfection. The inhibition rates of E6 protein measured by Flow cytometry were 79.7%, 80.4%, 71.3% and 57.4% at 24 h, 48 h, 5th day and 9th day after transfection, which were confirmed by the results of Western blot. In vivo research, E6 siRNA administration groups had great power in inhibiting tumor growth, restraining E6 protein expression, increasing tumor necrosis and apoptosis. The result of repeated injections of siRNA was better than that of single injection.

CONCLUSION

RNA interference with HPV16 E6 is specific and highly efficient in vitro and in vivo.

摘要

目的

评估RNA干扰体外和体内沉默人乳头瘤病毒16型（HPV16）E6基因的效率。

方法

设计HPV16 E6特异性小干扰RNA（siRNA），通过脂质体转染入CaSki细胞。检测转染前后细胞凋亡率以及HPV16 E6 mRNA和蛋白的变化。建立宫颈癌裸鼠模型，将siRNA直接注射到皮下肿瘤内。通过肿瘤体积、HPV16 E6蛋白表达及肿瘤细胞凋亡的变化评估siRNA的作用。

结果

体外研究中，转染后24小时、48小时、第5天和第9天细胞凋亡率分别为7.7%、11.8%、37.4%和12.6%。转染后24小时、48小时、第5天和第9天HPV16 E6 mRNA分别降低77%、83%、59%和41%。流式细胞术检测的E6蛋白抑制率在转染后24小时、48小时、第5天和第9天分别为79.7%、80.4%、71.3%和57.4%，蛋白质免疫印迹法结果证实了该结果。体内研究中，E6 siRNA给药组在抑制肿瘤生长、抑制E6蛋白表达、增加肿瘤坏死和凋亡方面作用显著。重复注射siRNA的效果优于单次注射。