| 李静,王金铭,任琛琛,等.微小 RNA?558靶向调控叉头框转录因子 C1促进卵巢癌 SKOV3细胞的增殖和侵袭力[J].安徽医药,2020,24(10):1943-1947. |
| 微小 RNA?558靶向调控叉头框转录因子 C1促进卵巢癌 SKOV3细胞的增殖和侵袭力 |
| MicroRNA?558 promotes the proliferation and invasion of SKOV3 cells by targeting FOXC1 |
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| DOI：10.3969/j.issn.1009?6469.2020.10.007 |
| 中文关键词: 卵巢肿瘤 叉头转录因子类 微小 RNA 增殖 实时聚合酶链反应 荧光免疫测定 实时荧光定量 PCR 侵袭 |
| 英文关键词: Ovarian neoplasms Forkhead transcription factors microRNA(miR) Proliferation Real?time polymerase chain reaction Fluoroimmunoassay Quantitative real?time PCR Invasion |
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| 中文摘要: |
| 目的探讨 miR?558通过对叉头框转录因子 C1(Forkhead box protein C1，FOXC1)的调控从而对人卵巢癌腺癌细胞(SKOV3)增殖和侵袭能力的影响。方法选择 2014年 1月至 2017年 12月郑州大学第三附属医院妇产科收治的上皮性卵巢癌病人 25例、交界性卵巢上皮性肿瘤病人 25例和良性卵巢上皮性肿瘤病人 25例，三组病例均行手术治疗，取其部分经手术切除的卵巢组织标本，使用实时荧光定量 PCR技术来分析微小 RNA?558(miR?558)的表达水平。把卵巢癌 SKOV3细胞分成三组，分别为 miR?558模拟物组、抑制物组和阴性对照组。通过靶基因预测网站来预测 miR?558的靶基因(FOXC1基因)通过荧光素酶报告基因实验来验证 miR?558对 FOXC1基因表达的调控作用。通过实时荧光定量 ?PCR技术和蛋白印迹法来分析，转染后各组细胞的 miR?558和 FOXC1的表达水平。通过细胞增殖实验(Cell Counting Kit?8，CCK?8法)检测三组细胞的增殖率。通过基质胶侵袭实验检测三组细胞的侵袭能力。结果实时荧光定量 ?PCR结果显示，在上皮性卵巢癌、交界性卵巢上皮性肿瘤和良性卵巢上皮性肿瘤病人的卵巢组织中， miR?558的相对表达水平分别为(3.43±0.42)、(2.47±0.35)、(1.37±0.31)；在 miR?558模拟物组、抑制物组和阴性对照组中， SKOV3细胞 miR?558的表达水平分别为(2.37±0.17)、(0.64±0.17)、(1.14±0.11)。在 miR? 558模拟物组、抑制物组和阴性对照组中， SKOV3细胞的 FOXC1基因转录出的信使 RNA(mRNA)表达水平分别为(0.51±0.10)、(2.27±0.12)、(0.99±0.11)。荧光素酶报告基因实验结果显示， SKOV3细胞被含 miR?558的质粒以及含 FOXC1基因的重组质粒共同转染后，的荧光素酶活性下降了 49.50%(P＜0.05)。蛋白印迹结果显示，上述三组 SKOV3细胞中， FOXC1蛋白的表达水平分别为(0.83±0.07)、(2.17±0.15)、(1.47±0.21)。 CCK?8检测结果显示，不同时间 miR?558模拟物组 SKOV3细胞的增殖率明显高于阴性对照组(P＜0.05)，不同时间 miR?558抑制物组 SKOV3细胞的增殖率明显低于阴性对照组(P＜0.05)。基质胶侵袭实验结果显示，上述三组 SKOV3细胞穿透基底膜的细胞数分别为(158.33±9.45)、(67.01±10.58)、(117.67±16.86)(P＜0.05)。结论 miR?558能够通过靶向调控 FOXC1的表达，促进 SKOV3细胞的增殖能力和侵袭能力，从而参与卵巢癌的发生发展。 |
| 英文摘要: |
| Objective To clarify the role of miR?558 on the proliferation and invasion of SKOV3 cell and the relationship with FOXC1.Methods We recruited 25 patients with epithelial ovarian cancer，25 patients with borderline epithelial ovarian tumorsand 25 patients with benign epithelial ovarian tumors in Department of Obstetrics and Gynecology of the Third Affiliated Hospitalof Zhengzhou University from January 2014 to December 2017.The three groups underwent surgical treatment，and some ovarian tis? sue of patients were taken.Quantitative real?time polymerase chain reaction(qRT?PCR)was used to measure the expression of miR? 558 in ovarian tissues.SKOV3 cells were assigned to miR?558 mimics group，inhibitor group，negative control，respectively.Bioinfor? matics was used for predicting target genes for miRNA?558?5p(FOXC1)and dual?luciferase reporting system for verifying the tar?get gene.MiR?558 expression was evaluated by qRT?PCR.FOXC1 expression was evaluated by western blot and qRT?PCR.Cell pro?liferation was determined by CCK?8 assay.Invasive activities were assessed by cell Transwell invasion assay.Results The relative expression of miR?558 measured by qRT?PCR in the ovarian tissues of patients with epithelial ovarian cancer，borderline ovarian epithelial tumor and benign ovarian epithelial tumor were(3.43±0.42)，(2.47±0.35)，(1.37±0.31)，respectively.The expression ofmiR?558 in SKOV3 cell line in the miR?558 mimics group，inhibitor group and negative control group were(2.37±0.17)，(0.64±0.17)，(1.14±0.11)， respectively.The expression of FOXC1 mRNA in SKOV3 cell line in the miR?558 mimics group，inhibitor group and negative control group were(0.51±0.10)，(2.27±0.12)，(0.99±0.11)， respectively.Luciferase reporter assay resultsshowed that the luciferase activity of SKOV3 cells transfected with miR?558 containing plasmid and FOXC1 containing recombi?nant plasmid decreased by 49.50%(P＜0.05).The expression of FOXC1 protein in SKOV3 cell line in the miR?558 mimics group， inhibitor group and negative control group were(0.83±0.07)，(2.17±0.15)，(1.47±0.21)，respectively.The results of CCK?8 assayshowed that the proliferation rate of SKOV3 cells in the miR?558 mimics group at different times was higher than that in the nega?tive control group(P＜0.05)，and that of the miR?558 inhibitor group at different times was significantly lower when compared with negative control group(P＜0.05).The number of SKOV3 cell that passed through the membrane in miR?558 mimics group，in? hibitor group and negative control group was(158.33±9.45)，(67.01±10.58)，(117.67±16.86)，respectively(P＜0.05).Conclu? sions MiR?558 can promote the proliferation and invasion of SKOV3 cells by targeting FOXC1，which participates in the patho? genesis of ovarian cancer. |
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