MiR-138对MCF-7细胞端粒酶活性及端粒稳定性的调控作用

The regulation of telomerase activity and telomere stability by MiR-138 in MCF-7 cells

目的研究MiR-138通过人端粒酶反转录酶(hTERT)作为下游靶基因,对人乳腺癌MCF-7细胞端粒酶活性的调控作用及端粒稳定性的影响。方法在人乳腺癌MCF-7细胞中瞬时转染MiR-138模拟物,用MTT法检测细胞增殖活性,并于转染后48 h用实时定量RT-PCR检测端粒酶催化亚单位hTERT表达、TRAP assay检测端粒酶活性,同时对细胞进行53BP1抗体免疫荧光染色及端粒的FISH染色。结果转染后48 h,MiR-138模拟物处理的MCF-7细胞hTERT表达水平比对照细胞降低2.18倍(2-△△Ct),端粒酶活性比对照细胞降低2.69倍,53BP1聚集形成的凝集点(Foci)部分与端粒位点重合,比率达到20.62%±1.55%。结论 MiR-138以hTERT作为下游靶分子调控MCF-7细胞端粒酶活性,影响细胞端粒稳定性。

Objective To study the regulation mechanism of telomerase activity of MiR-138 by targeting hTERT as the downstream molecular,and its effect on telomere stability in breast cancer cell line MCF-7.Methods MiR-138 mimics were transiently transfected into breast cancer cell line MCF-7.MTT method was applied for detecting the effect on cell proliferation.The hTERT expression level of the cells was detected by real-time RT-PCR and telomerase activity was checked by TRAP assay 48 h after transfection.Meanwhile,the cells were simultaneously checked by 53BP1 antibody immunofluorescent staining and telomere FISH staining.Results The hTERT expression level of MCF-7 cells treated with MiR-138 was repressed by 2.18 folds（2-△△Ct） when compared with the control cells 48 h after transfection.Telomerase activity was also decreased by 2.69 folds when compare with the control cells.53BP1congregated into many Foci and co-localized with the telomere spots at a percentage of 20.62%±1.55%.Conclusion MiR-138 can regulate telomerase activity in MCF-7 cells by targeting hTERT as the downstream molecular,and affects the telomere stability as well.