摘要
目的分析miR-143对前列腺癌细胞PC3增殖和凋亡的影响。方法实验分为未转染的PC3细胞(PC3组)、转染miR-NC的对照组细胞(PC3/miR-NC组)和稳定表达miR-143的PC3细胞(PC3/miR-143组),通过免疫荧光和Real-time PCR进行鉴定。利用CCK-8法和流式细胞术分别分析miR-143表达水平变化是否影响PC3细胞的增殖和凋亡。再使用在线分析软件预测miR-143的靶基因,随后构建荧光素酶报告基因质粒,通过荧光素酶报告基因法分析miR-143与靶基因的靶向结合位点。结果在PC3/miR-143组中miR-143的表达水平高于PC3组(P<0.01)和PC3/miR-NC组(P<0.01)。CCK-8检测结果显示PC3/miR-143组细胞的增殖能力与PC3组(P<0.05)和PC3/miR-NC组相比下降(P<0.01)。流式细胞术检测的结果显示,PC3/miR-143组细胞的凋亡水平与PC3组和PC3/miR-NC组细胞比较增加(P<0.01)。在线分析软件预测miR-143能够靶向结合三叶因子3(TFF3)的3′-UTR;双荧光素酶报告基因的结果进一步证实miR-143能够靶向作用于TFF3的3′-UTR,Real-time PCR的结果表明在过表达miR-143能够抑制PC3细胞中TFF3的表达。在稳定表达miR-143的PC3细胞中转染TFF3真核表达质粒能够抵消miR-143的作用。结论miR-143通过靶向作用于TFF3的3′-UTR,抑制其表达,抑制PC3细胞的增殖。
Objective To investigate the effect of miR-143 on the proliferation and apoptosis of prostate cancer cell line PC3.Methods The experiment was divided into non transfected PC3 cells group(PC3 group),transfected with miR-NC group(PC3/miR-NC group)and PC3 cells stably expressing miR-143 group(PC3/miR-143 group),and identified by immunofluorescence and real-time PCR.The effect of miR-143 on the proliferation of PC3 cells by CCK-8 method.The effect of miR-143 on PC3 cells apoptosis was analyzed by flow cytometry.The online analysis software was used to predict the target genes of miR-143,and then the luciferase reporter gene detection plasmid was constructed.The targeted binding sites of miR-143 and target genes were analyzed by luciferase assay.Results The expression level of miR-143 in PC3/miR-143 group was significantly higher than that of PC3 group(P<0.01)and PC3/miR-NC group(P<0.01).CCK-8 test showed that the cell proliferation ability of PC3/miR-143 group decreased significantly compared with PC3 group(P<0.05)and PC3/miR-NC group(P<0.01).The results of flow cytometry showed that the apoptosis level of PC3/miR-143 group was significantly higher than that of PC3 group and PC3/mir-nc group(P<0.01).Online analysis software predicted that miR-143 could target 3′-UTR binding to Trefoil Factor 3(TFF3);The results of dual luciferase reporter gene further confirmed that miR-143 could target the 3′-UTR of TFF3.Real time PCR results showed that overexpression of miR-143 could significantly inhibit the expression of TFF3 in PC3 cells.Transfection of TFF3 eukaryotic expression plasmid into PC3/miR-143 group could counteract the effect of miR-143.Conclusion MiR-143 inhibits the expression of TFF3 and the proliferation of PC3 cells by targeting the 3′-UTR of TFF3.
作者
郭亮
肖峻
陶陶
Guo Liang;Xiao Jun;Tao Tao(Dept of Urinary Surgery,The Affiliated Provincial Hospital of Anhui Medical University, Hefei 230001;Dept of Urinary Surgery, The Affiliated Lu′an Hospital of Anhui Medical University, Lu′an 237005)
出处
《安徽医科大学学报》
CAS
北大核心
2022年第5期684-689,共6页
Acta Universitatis Medicinalis Anhui
基金
国家自然科学基金(编号:81702540)。
