miR-146b-5p靶向ZNRF2影响IL-17a诱导乳腺癌细胞增殖、迁移和侵袭

miR-146b-5p affects proliferation,migration and invasion of breast cancer cells induced by IL-17a through regulating ZNRF2

目的:研究miR-146b-5p对IL-17a诱导的乳腺癌细胞增殖、迁移和侵袭的影响及其潜在分子机制。方法:免疫组织化学法检测20例乳腺癌组织和对应癌旁组织中IL-17a表达,qRT-PCR法检测组织中miR-146b-5p表达,Western blot检测组织中ZNRF2蛋白表达。Pearson相关性检验分析乳腺癌组织中IL-17a与miR-146b-5p及miR-146b-5p和ZNRF2表达的相关性。不同浓度的IL-17a处理乳腺癌细胞MDA-MB-231后,MTT法检测细胞增殖,Transwell检测细胞迁移和侵袭,qRT-PCR法检测细胞中ZNRF2 mRNA和miR-146b-5p表达,Western blot检测ZNRF2、MMP-2、MMP-9及STAT3信号通路蛋白STAT3和pSTAT3蛋白表达水平。双荧光素酶报告基因实验验证细胞中miR-146b-5p与ZNRF2的关系。结果:与癌旁组织相比,乳腺癌组织中IL-17a表达呈阳性(P<0.05),miR-146b-5p表达水平明显降低(P<0.05),ZNRF2蛋白表达水平明显升高(P<0.05)。IL-17a处理MDA-MB-231细胞后,细胞增殖能力升高(P<0.05),迁移和侵袭数增多(P<0.05),细胞中MMP-2、MMP-9和p-STAT3蛋白表达水平升高(P<0.05),miR-146b-5p表达水平降低(P<0.05),ZNRF2的mRNA和蛋白表达水平升高(P<0.05)。miR-146b-5p靶向负调控ZNRF2表达。过表达miR-146b-5p或沉默ZNRF2后,细胞增殖能力降低(P<0.05),迁移和侵袭数减少(P<0.05),细胞中MMP-2、MMP-9和p-STAT3蛋白表达水平降低(P<0.05)。IL-17a可逆转过表达miR-146b-5p或沉默ZNRF2对MDA-MB-231细胞增殖、迁移、侵袭及STAT3信号通路的影响。结论:miR-146b-5p通过靶向下调ZNRF2表达并抑制STAT3信号通路降低IL-17a诱导的乳腺癌MDA-MB-231细胞增殖、迁移和侵袭。

Objective:To explore the effect of miR-146b-5p on proliferation,migration and invasion of breast cancer cells induced by IL-17 a and its potential molecular mechanism.Methods:Immunohistochemical method was used to detect expression of IL-17 a in 20 breast cancer tissues and corresponding adjacent tissues,qRT-PCR was used to detect expression of miR-146b-5p,and Western blot was used to detect expression of ZNRF2 protein.Pearson correlation was used to analyze the correlation between IL-17 a and miR-146b-5p as well as miR-146b-5p and ZNRF2 in breast cancer tissues.After treating breast cancer cells MDA-MB-231 with different concentrations of IL-17 a,MTT method was used to detect cell proliferation,Transwell was used to detect cell migration and invasion,qRT-PCR was used to detect expression levels of ZNRF2 mRNA and miR-146b-5p in cells,and Western blot was used to detect expression levels of ZNRF2,MMP-2,MMP-9 and STAT3 signaling pathway proteins STAT3 and p-STAT3 protein.Dual luciferase reporter gene experiment verified the relationship between miR-146b-5p and ZNRF2 in cells.Results:Compared with adjacent tissues,expression of IL-17 a was positive in breast cancer tissues(P<0.05),expression level of miR-146b-5p was significantly reduced(P<0.05),and expression level of ZNRF2 protein was significantly increased(P<0.05).After IL-17 a treated MDA-MB-231 cells,the cell proliferation capacity increased(P<0.05),number of migration and invasion increased(P<0.05),expressions of MMP-2,MMP-9 and p-STAT3 protein increased(P<0.05),expression of miR-146b-5p decreased(P<0.05),and mRNA and protein expression levels of ZNRF2 increased(P<0.05).miR-146b-5p negatively regulated expression of ZNRF2.After overexpressing miR-146b-5p or silencing ZNRF2,the cell proliferation capacity reduced(P<0.05),number of migration and invasion reduced(P<0.05),and expression levels of MMP-2,MMP-9 and p-STAT3 protein in cells were reduced(P<0.05).IL-17 a could reverse the effects of overexpressing miR-146b-5p or silencing ZNRF2 on proliferation,migration and invasion of MDA-MB-231 cells and STAT3 signaling pathway.Conclusion:miR-146b-5p inhibits the proliferation,migration and invasion of breast cancer MDA-MB-231 cells by targeting ZNRF2 gene and inhibiting the STAT3 signaling pathway.