吉林大学学报(医学版)

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miR-21对宫颈癌HeLa细胞辐射敏感性的影响

徐珊，燕蒙蒙，刘 扬，李鹏武，马淑梅，刘晓冬   

  1. (吉林大学公共卫生学院 卫生部放射生物学重点实验室，吉林 长春 130021)
  • 收稿日期:2014-04-11 出版日期:2014-11-28 发布日期:2015-01-18
  • 通讯作者: 马淑梅 E-mail:（Tel：0431-85619443，E-mail:shmm2001@126.com）
  • 作者简介:徐珊（1988-），女，河北省石家庄市人，在读医学硕士，主要从事放射生物学的研究。
  • 基金资助:

    国家自然科学基金资助课题（31370837）；吉林省科技厅科研基金资助课题（201205010）；吉林省科技厅科技发展计划国际合作项目资助课题（2009）；吉林大学科学前沿与科学交叉创新项目资助课题（2011）

Influence of miR-21 on radiation sensitivity of cervical cancer HeLa cells

XU Shan,YAN Meng-meng，LIU Yang，LI Peng-wu,MA Shu-mei,LIU Xiao-dong   

  1. （Key Laboratory of Radiobiology,Ministry of Health,School of Public Health,Jilin University,Changchun 130021,China）
  • Received:2014-04-11 Online:2014-11-28 Published:2015-01-18

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摘要：

目的：探讨miR-21与宫颈癌HeLa细胞辐射敏感性之间的关系，阐明miR-21对细胞凋亡和自噬改变的可能作用机制。方法：人宫颈癌HeLa细胞分为假照组和4 Gy照射组，分别转染miR-21 NC（阴性对照）、miR-21 mimics、miR-21 inhibitor NC和miR-21 inhibitor。实时荧光定量PCR检测照射前和照射后miR-21的表达水平；集落形成实验观察miR-21对HeLa细胞辐射敏感性的影响；流式细胞术检测细胞凋亡和自噬细胞百分率；采用生物信息学方法预测miR-21的靶基因；荧光素酶报告基因分析验证miR-21与靶基因3′UTR的结合；Western blotting法检测相关蛋白beclin1表达水平。结果：与假照组比较，4 Gy照射组miR-21表达水平明显增加(P<0.05）；集落形成实验，与miR-21 NC组比较，miR-21 mimics组HeLa细胞辐射敏感性无明显变化(P>0.05）；与miR-21 inhihitor NC组比较，miR-21 inhibitor 组HeLa细胞辐射敏感性无明显变化(P>0.05）；与miR-21 NC+4 Gy组比较，miR-21 mimics+4 Gy组HeLa细胞凋亡率明显增加(P<0.05）；与miR-21 inhibitor NC组比较，miR-21 inhibitor组HeLa细胞凋亡率明显减少（P<0.05）。与miR-21 NC组比较，miR-21 mimics组自噬率升高（P<0.05）。与miR-21 NC+4 Gy组比较，miR-21 mimics+4 Gy组自噬率升高（P<0.05）;与miR-21 NC+4 Gy组比较，miR-21 mimics+4 Gy组beclin1蛋白表达水平增加（P<0.05）。
结论：发现了miR-21新的靶基因beclin1。过表达miR-21促进辐射诱导的凋亡和自噬，但对于宫颈癌HeLa细胞辐射敏感性无直接影响。
［关键词］ miR-21；宫颈肿瘤；HeLa细胞；辐射敏感性；细胞凋亡；自噬

关键词: miR-21, 宫颈肿瘤, HeLa细胞, 辐射敏感性, 细胞凋亡, 自噬

Abstract:

Abstract:Objective  To research the relationship between miR-21 and radiation sensitivity of cervical cancer HeLa cells,and to elucidate the possible action mechanism of miR-21 in apoptosis and autophagy of cells.Methods The HeLa cells were divided into mock groups (transfected with miR-21 NC,mimics, inhibitor NC or inhibitor) and 4 Gy irradiation groups(transfected with miR-21 NC,mimics, inhibitor NC or inhibitor with irradiation).The miR-21 expression was detected by real-time PCR;colony-forming assay was used to detect the changes of radiation sensitivity;the apoptosis and autophagy were analyzed by FACS assay;the target gene of miR-21 was predicted by bioinformatics methods;Dual-luciferase Reporter Assay was used to demonstrate the binding of miR-21 and targets;the expression of beclin1 protein was measured by Western blotting method.Results After 4 Gy radiation,compared with mock group,the expression of miR-21 was increased remarkably (P<0.05). Colony-forming assay showed that there were no changes of radiation sensitivity of HeLa cells after overexpressing or inhibiting miR-21  compared with miR-21 NC or miR-21 inhibitor NC groups(P>0.05). Compared with miR-21 NC＋4 Gy group,the apoptotic rate of HeLa cells in　miR-21 mimics+4 Gy　group was  increased (P<0.05).After transfection with  miR-21 inhibitor,compared with miR-21 inhibitor NC  group,the apoptotic rate of HeLa cells in duced by radiation was  decreased (P<0.05).Compared with miR-21 NC  group,the autophagy rate was increased after transfection with miR-21 mimics (P<0.05).After transfection with miR-21 mimics and radiation,the expression level of beclin1 protein was increased(P<0.05) compared with miR-21 NC+4 Gy group.Conclusion Beclin1 is the target gene of miR-21 in HeLa cells.Overexpression of miR-21 can increase the radiation-induced apoptosis and autophagy,but has on influence in radiation sensitivity of HeLa cells.

Key words: miR-21, cervical neoplasms, HeLa cells, radiation sensitivity, apoptosis, autophagy

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