山东医药2019，Vol.59Issue(2)：28-31,4.DOI:10.3969/j.issn.1002-266X.2019.02.007
miR-130b对宫颈癌细胞修复TNF-α诱导性基因组双链DNA断裂作用的影响及机制
Effect of miR-130b on repair of TNF-α-induced genomic DNA double strand breaks in cervical cancer cells
摘要
Abstract
Objective To observe the impact of miR-130 b on the repair of DNA double strand breaks in cervical cancer cells and to discuss the mechanism underlying them. Methods Siha cells were classified into 6 groups. The cells transfected with pc DNA 3. 1-CDKN1A ( cyclin-dependent kinase inhibitor 1A) , pc DNA 3. 1, miR-130 b mimics and the negative control of miR-130 b were taken as the CDKN1 A, pc DNA 3. 1, miR-130 b and NC groups in turn. The ones cotransfected with miR-130 b and pc DNA 3. 1-CDKN1 A and those with miR-130 b and pc DNA 3. 1 were taken as the CDKN1 A + miR-130 b and pc DNA 3. 1 + miR-130 b groups. Cells were stimulated with TNF-α after the transfection. DNA oliver tail moments ( OTM) were determined in comet assays and the phosphorylated H2 AX histone protein variant ( γ-H2AX) levels were measured by flow cytometry. The CDKN1 A mRNA and its protein levels were measured by using semiquantitative real-time PCR and Western blotting. pc DNA 3. 1/EGFP, pc DNA 3. 1/EGFP-CDKN1A-wtUTR ( wild type) and pc DNA 3. 1/EGFP-CDKN1A-mutUTR ( mutant type) were transfected into Siha cells with miR-130 b mimics or NC, and the expression levels of reporter genes were determined. We used TNF-α and DNA damage enhancer AZD2461 to stimulate the cells in each group, and then used flow cytometry to determine the level of apoptosis. Results Compared with the pc DNA 3. 1 group, both the oliver tail moment and the γ-H2 AX protein level reduced in the cells of pc DNA 3. 1-CDKN1 A group ( both P < 0. 05) ; compared with the NC group, the oliver tail moment and the γ-H2 AX protein level increased whereas the CDKN1 A mRNA and protein levels decreased in the cells of miR-130 b group ( all P < 0. 05) . Compared with the pc DNA 3. 1 + miR-130 b group, the oliver tail moment, γ-H2 AX protein level and apoptosis rate decreased ( all P < 0. 05) ; compared with NC cotransfection, the cotransfection with miR-130 b mimics reduced the fluorescence levels within cells, but did not change the fluorescent relative activity of the mutant cells. Conclusion The miR-130 b hampers the repair of DNA double strand breaks in cervical cancer cells by inhibiting CDKN1 A gene expression through directly targeting CDKN1 A mRNA to increase the apoptosis rates.关键词
宫颈癌/微小核糖核酸/DNA修复/细胞周期蛋白依赖性蛋白激酶抑制物1A/磷酸化组蛋白2A变异体/细胞凋亡Key words
cervical carcinoma/microR-130b/DNA repair/cyclin-dependent kinase inhibitor 1A/phosphorylated H2AX histone protein variant/apoptosis分类
医药卫生引用本文复制引用
杨磊,刘涛,段继惠,穆红..miR-130b对宫颈癌细胞修复TNF-α诱导性基因组双链DNA断裂作用的影响及机制[J].山东医药,2019,59(2):28-31,4.基金项目
国家自然科学基金资助项目(81602403). （81602403）
