长链非编码RNA RAB6C-1和ZNF704-1调控DNA甲基化在宫颈癌变进展中的作用

In the cervical precancerous tissue, lnc-RAB6C-1 and lnc-ZNF704-1 are over-expressed, but their roles in cervical cancer progression are not clear. LncRNAs (long non-coding RNAs) have been associated with carcinogenesis and progression, which can regulate DNA methylation. References and our previous researches showed close relationship between cervical carcinogenesis and DNA methylation. Based on that, it is hypothesized that lnc-RAB6C-1 and lnc-ZNF704-1 may involve in the development of cervical cancer through regulating DNA methylation. In this study, it will observe the association between lnc-RAB6C-1, gene expression and DNA methylation of RAB6C, and the relationship between lnc-ZNF704-1, gene expression and DNA methylation of ZNF704 in vivo. Furthermore, to examine such effects, over expression plasmid transfection and siRNA interference technology will be carried out in vitro and the changes of cervical cancer cell biology ,the levels of gene expression and the degree DNA methylation of RAB6C and ZNF704 will be detected. According to these results, the modulation function of lnc-RAB6C-1 and lnc-ZNF704-1 on DNA methylaition in the progression of cervical cancer will be evaluated. The results may provide a new theoretical basis for etiology research and new targets for the prevention and treatment of cervical cancer.

Lnc-RAB6C-1和lnc-ZNF704-1在宫颈癌前病变组织中异常高表达，但对宫颈癌变的作用不明。LncRNA可调控DNA甲基化，参与肿瘤发生发展过程。文献报道和课题组研究发现宫颈癌变与DNA甲基化密切相关。提示，lnc-RAB6C-1和lnc-ZNF704-1可能通过调控DNA甲基化影响宫颈癌变。本课题拟结合人群和体外实验，探讨lnc-RAB6C-1和lnc-ZNF704-1与基因RAB6C和ZNF704表达水平及甲基化的相关性，同时采用过表达质粒转染和siRNA干扰技术进行lnc-RAB6C-1和lnc-ZNF704-1过表达和功能缺失研究，比较干预前后宫颈癌细胞生物学功能、基因RAB6C和ZNF704表达水平和甲基化程度的变化。以综合评价lnc-RAB6C-1和lnc-ZNF704-1对DNA甲基化在宫颈癌变中的调控作用，为宫颈癌病因研究提供新依据，为宫颈癌防治提供新靶点。

Lnc-RAB6C-1和lnc-ZNF704-1在宫颈癌前病变组织中异常高表达，但对宫颈癌变的作用不明。LncRNA可调控DNA甲基化，参与肿瘤发生发展过程。文献报道和课题组研究发现宫颈癌变与DNA甲基化密切相关。提示，lnc-RAB6C-1和lnc-ZNF704-1可能通过调控DNA甲基化影响宫颈癌变。本课题人群研究选择正常宫颈(NC)、CINⅠ、CINⅡ、CINⅢ和宫颈癌人群为研究对象，形成正常-癌前病变-癌症的疾病自然发展阶梯，发现CINⅡ/CINⅢ和宫颈癌组织中lnc-RAB6C-1和 lnc-ZNF704-1的mRNA表达水平高于NC组和CINⅠ组，CINⅡ/CINⅢ和宫颈癌标本中RAB6C和ZNF704蛋白表达水平高于宫颈炎和CINⅠ组。体外实验发现，lnc-RAB6C-1在Caski细胞中表达量最高。Lnc-RAB6C-1干扰组RAB6C mRNA和蛋白表达明显下调，过表达组RAB6C mRNA和蛋白表达明显上调，差异有统计学意义。LncRNA-RAB6C干扰和过表达对RAB6C不同位点CpG岛甲基化有不同程度的影响,但干扰和过表达对RAB6C相同位点CpG岛甲基化影响方向一致。Lnc-RAB6C-1干扰后Caski细胞增殖和活性降低，过表达后增加。LncRNA-RAB6C干扰组细胞凋亡率增高（P<0.05），过表达组凋亡率略低，但未见统计学差异。Lnc-RAB6C-1干扰组细胞处于G0/G1期细胞比例增加，S期与G2/M期细胞比例下降，过表达组细胞处于S期与G2/M期比例增加，G0/G1期细胞比例下降。综上，lnc-RAB6C-1可能通过调控DNA甲基化在宫颈癌变中发挥调控作用，为宫颈癌病因研究提供了新依据，为宫颈癌防治提供了新靶点。