摘要
目的阐明miR-106b在结直肠癌细胞放射敏感性中的作用及机制。方法构建稳定过表达和干扰miR-106b结直肠癌细胞系,通过免疫荧光和平板克隆实验探讨miR-106b对结直肠癌细胞放射敏感性的影响;Western blot检测Caspase-3和γ-H2AX的表达;生物信息学预测miR-106b下游调控的靶基因,双荧光素酶报告系统、荧光定量PCR及Western blot进一步验证。在稳定过表达miR-106b的结直肠癌细胞系中转染pCDNA3.0-PTEN,探讨细胞放射敏感性变化,明确miR-106b是否通过靶向调控PTEN增加结直肠癌细胞放射抵抗。结果在结直肠癌细胞系过表达miR-106b,经过同等条件的照射处理后,与对照组相比,过表达miR-106b组细胞存活分数升高,放射抵抗增强(P<0.05),Caspase-3及γ-H2AX表达降低。在稳定过表达miR-106b的细胞中上调PTEN后能逆转miR-106b引起的放射抵抗。结论miR-106b通过靶向抑制PTEN从而诱导结直肠癌细胞放射抵抗,预示着miR-106b-PTEN位点可能为临床提高结直肠癌放疗效果寻找相关靶点提供理论依据。
Objective To investigate the role of miR-106b in the cell radioresistance in colorectal carcinoma(CRC),and unravel the underlying mechanism.Methods The CRC cell lines stably overexpressing and interfering miR-106b were established.The effect of miR-106b upon the CRC cell radiosensitivity was evaluated by cell radiation,immunofluorescence and colony formation assay.The expression levels of Caspase-3 andγ-H2AX were detected by Western blot.The target genes of miR-106b were identified by bioinformatics prediction,which were further validated by dual luciferase assay,fluorescence quantitative PCR and Western blot.The CRC cell lines stably overexpressing miR-106b were transfected with pCDNA3.0-PTEN.The changes of CRC cell radiosensitivity were investigated.Whether miR-106b could increase the radioresistance of CRC cells by targeting PTEN was clarified.Results Compared with the control group(miR-ctr group),the cell surviving fraction was significantly elevated(P<0.05),the radioresistance(P<0.05)was considerably enhanced and the expression levels of Caspase-3 andγ-H2AX were significantly down-regulated(both P<0.05)in the miR-106b overexpression group.PTEN up-regulation in CRC cell lines stably overexpressing miR-106b could reverse the radioresistance induced by miR-106b.Conclusion miR-106b can induce CRC cell radioresistance by inhibiting PTEN,prompting that miR-106b-PTEN might provide theoretical evidence for relevant targets which can enhance the clinical efficacy of radiotherapy.
作者
张余琴
郑林
丁轶
Zhang Yuqin;Zheng Lin;Ding Yi(Department of Radiation Oncology,Nanfang Hospital,Southern Medical University,Guangzhou 510515,China;Department of Pathology,School of Basic Medical Sciences,Southern Medical University/Department of Pathology,Nanfang Hospital,Southern Medical University,Guangzhou 510515,China)
出处
《中华放射肿瘤学杂志》
CSCD
北大核心
2020年第11期991-995,共5页
Chinese Journal of Radiation Oncology
基金
国家自然科学基金(81903125)
广东省自然科学基金博士启动项目(2017A030310081)
广东省自然科学基金博士启动项目(2018A030310513)
肿瘤免疫病理学教育部重点实验室开放课题(2018jsz105)。
