CAJ | 학술논문

目的探讨微小RNA(miRNA,miR)-494对乳腺癌细胞MCF-7的增殖和转移的调控作用以及机制.方法在乳腺癌细胞株MCF-7转染miR-494模拟物及抑制剂、阴性对照,然后采用噻唑蓝(MTT)方法检测miR494对MCF-7细胞活力的影响,并通过Transwell转移实验检测miR-494对MCF-7转移能力的影响.通过双荧光素酶实验检测miR-494的靶基因,并且用反转录-聚合酶链反应(RT-PCR)和Western blot进行验证.结果转染miR-494 mimics后,细胞活力明显提高,24 h细胞活力比值为1.28,48 h后为1.80,(P＜0.05),转染miR-494 inhibitor后,细胞活力显著下调,24h细胞活力比值为0.80,48 h后为0.68(P ＜0.05).对照组细胞均数为67个/视野,转染miR-494mimics后的MCF-7细胞的均数为:122个/视野(P＜0.05),转染miR-494 inhibitor后的MCF-7细胞的均数为:47个/视野(P＜0.01).野生型载体模拟物组相对荧光素酶活性:0.69(P＜0.01),抑制剂组:1.90(P＜0.01);突变型载体模拟物组相对荧光素酶活性:1.09,抑制剂组:1.19.miR-494过表达后PTEN的mRNA和蛋白水平确实出现明显下降,相反地,miR-494的抑制却导致PTEN蛋白水平和mRNA水平的上调.结论高表达miR-494可显著促进乳腺癌细胞株MCF-7的增殖与转移能力,抑制miR-494可显著降低MCF-7的增殖与转移能力,并且证实miR-494是通过靶向第10号染色体上缺失与张力蛋白同源的磷酸酯酶基因(PTEN)的3'端非编码区域(3'-UTR)来调节乳腺癌细胞株MCF-7的增殖和转移能力.
목적 탐토미소RNA(miRNA,miR)-494대유선암세포MCF-7적증식화전이적조공작용이급궤제.방법 재유선암세포주MCF-7전염miR-494모의물급억제제、음성대조,연후채용새서람(MTT)방법검측miR494대MCF-7세포활력적영향,병통과Transwell전이실험검측miR-494대MCF-7전이능력적영향.통과쌍형광소매실험검측miR-494적파기인,병차용반전록-취합매련반응(RT-PCR)화Western blot진행험증.결과 전염miR-494 mimics후,세포활력명현제고,24 h세포활력비치위1.28,48 h후위1.80,(P＜0.05),전염miR-494 inhibitor후,세포활력현저하조,24h세포활력비치위0.80,48 h후위0.68(P ＜0.05).대조조세포균수위67개/시야,전염miR-494mimics후적MCF-7세포적균수위:122개/시야(P＜0.05),전염miR-494 inhibitor후적MCF-7세포적균수위:47개/시야(P＜0.01).야생형재체모의물조상대형광소매활성:0.69(P＜0.01),억제제조:1.90(P＜0.01);돌변형재체모의물조상대형광소매활성:1.09,억제제조:1.19.miR-494과표체후PTEN적mRNA화단백수평학실출현명현하강,상반지,miR-494적억제각도치PTEN단백수평화mRNA수평적상조.결론 고표체miR-494가현저촉진유선암세포주MCF-7적증식여전이능력,억제miR-494가현저강저MCF-7적증식여전이능력,병차증실miR-494시통과파향제10호염색체상결실여장력단백동원적린산지매기인(PTEN)적3'단비편마구역(3'-UTR)래조절유선암세포주MCF-7적증식화전이능력.
Objective To investigate whether microRNA (miR)-494 mediated cell proliferation and migration and the regulatory mechanism in breast cancer cell MCF-7.Methods miR-494 mimics and inhibitor and negative control (NC) were transfected into MCF-7 cells, and cell viability was evaluated by methyl thiazol tetrazolium (MTT) assay.Cell migrasion was assessed by Transwell migrasion assay.Dual-luciferase reporter assay, reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting were used to identify the target gene of miR-494.Results After the transfection of miR-494 mimics, the cell vitality were improved obviously, cell vitality ratio after 24 h was : 1.28, after 48h was : 1.80.After the transfection of miR-494 inhibitor, the cell vitality were reduced obviously, cell vitality ratio after 24 h was: 0.80, after 48 h was : 0.68.(all P ＜ 0.05).The mean of the control group is : 67/view.The mimics group was 122/view (P ＜ 0.05) , the inhibitor group was 47/view (P ＜ 0.01).Wild type cartier relative luciferase activity : analog group : 0.69 (P ＜ 0.01), the inhibitor groups : 1.90 (P ＜ 0.01).Mutant carrier analog group relative luciferase activity: 1.09, inhibitor groups: 1.19.MiR-494 after expression of PTEN mRNA and protein levels significantly decline, in contrast, inhibition of miR-494 is lead to increase the level of PTEN protein and mRNA level.Conclusion These results suggested that miR-494 can regulate cell proliferation and migration through targeting PTEN in MCF-7 cells.