lncRNA C5orf66-AS1促进宫颈癌细胞增殖和迁移及其机制探讨

lncRNA C5orf66-AS1 promotes the proliferation and migration of cervical cancer cells and its mechanism

目的:探讨lncRNA C5orf66-AS1对宫颈癌细胞增殖及迁移的影响及其作用机制。方法:通过TCGA筛选宫颈癌和癌旁组织中差异表达的lncRNA并采用qRT-PCR对候选lncRNA进行验证。采用qRT-PCR检测miR-149-5p和RING1在宫颈癌组织和细胞中的表达水平。使用CCK-8、克隆形成、Transwell和流式细胞术检测C5orf66-AS1对细胞增殖、迁移和凋亡的影响。通过双荧光素酶测定和Western blot评估C5orf66-AS1对miR-149-5p和RING1表达的调控作用。进一步构建裸鼠成瘤模型通过肿瘤重量和体积验证C5orf66-AS1对肿瘤生长的影响。结果:lncRNA C5orf66-AS1在宫颈癌组织和细胞中显著上调(P<0.05)。与NC组比较,C5orf66-AS1组能够促进宫颈癌细胞增殖和迁移(P<0.05)。RNA免疫沉淀(RNA immunoprecipitation,RIP)和双荧光素酶报告实验结果显示C5orf66-AS1与miR-149-5p靶向结合。过表达miR-149-5p和RING1能够逆转C5orf66-AS1介导的宫颈癌细胞增殖和迁移能力。最后裸鼠体内实验显示,与NC组比较,C5orf66-AS1组的肿瘤显著生长(P<0.05)。结论:lncRNA C5orf66-AS1作为一种竞争性内源性RNA,通过吸附miR-149-5p调节RING1对宫颈癌细胞增殖、凋亡和迁移的影响。

Objective:To investigate the effect of lncRNA C5orf66-AS1 on the proliferation and migration of cervical cancer cells and its mechanism.Methods:Differential expression of lncRNA in cervical cancer and adjacent tissues was screened by TCGA,and candidate lncRNA was verified by qRT-PCR.The expression levels of miR-149-5p and RING1 in cervical cancer tissues and cells were detected by qRT-PCR.The effects of C5orf66-AS1 on cell proliferation,migration and apoptosis were detected by CCK-8,clone formation,Transwell and flow cytometry.The regulatory effect of C5orf66-AS1 on the expression of miR-149-5p and RING1 was evaluated by double luciferase assay and Western blot.Furthermore,a nude mouse tumorigenic model was constructed to verify the effect of C5orf66-AS1 on tumor growth by tumor weight and volume.Results:lncRNA C5orf66-AS1 was significantly up-regulated in cervical cancer tissues and cells(P<0.05).Compared with NC group,C5orf66-AS1 group can promote the proliferation and migration of cervical cancer cells(P<0.05).The results of RNA immunoprecipitation(RIP)and double luciferase report showed that C5orf66-AS1 bound to miR-149-5p.Overexpression of miR-149-5p and RING1 can reverse the proliferation and migration of cervical cancer cells mediated by C5orf66-AS1.Finally,in vivo experiments in nude mice showed that,compared with NC group,the tumors in C5orf66-AS1 group grew significantly(P<0.05).Conclusion:As a competitive endogenous RNA,lncRNA C5orf66-AS1 regulates the effect of RING1 on the proliferation,apoptosis and migration of cervical cancer cells by adsorbing miR-149-5p.