miR-155-5p调控Ndfip1表达影响乳腺癌细胞恶性生物学行为及相关机制研究

Regulation of Ndfip1 expression by miR-155-5p affects the malignant biological behavior of breast cancer cells and its related mechanisms

目的:探究miR-155-5p靶向调控Ndfip1影响乳腺癌细胞增殖、凋亡、侵袭与迁移改变及相关机制。方法:实时荧光定量聚合酶链式反应(qRT-PCR)检测本院25例有明确病理诊断的乳腺癌及对应的癌旁正常组织中miR-155-5p及Ndfip1基因表达量。选取MCF-7细胞,构建Control组、NC-inhibitor组、miR-155-5p inhibitor组、miR-155-5p inhibitor+si-NC组、miR-155-5p inhibitor+si-Ndfip1组细胞,CCK8实验、流式细胞仪、Transwell实验、划痕愈合实验分别检测细胞增殖、凋亡、侵袭、迁移能力改变,Western blot检测PI3K/AKT信号通路相关蛋白表达。Targetscan生物信息学网站预测miR-155-5p下游靶基因,双荧光素酶实验检测miR-155-5p与靶基因Ndfip1相关性。结果:与癌旁正常组织相比,miR-155-5p在乳腺癌组织中的基因表达量显著升高(P<0.05),Ndfip1在乳腺癌组织中的基因表达量明显降低(P<0.05),且乳腺癌组织中miR-155-5p和Ndfip1表达呈负相关(r=-0.969 8,P<0.001)。与Control组及NC-inhibitor组相比,miR-155-5p inhibitor组细胞增殖、侵袭及迁移能力均降低(P<0.05),细胞凋亡数量增多(P<0.05),p-PI3K和p-AKT蛋白表达量降低(P<0.05)。双荧光素酶报告基因检测证明Ndfip1是miR-155-5p的靶标。回复实验检测得出,与miR-155-5p inhibitor组及miR-155-5p inhibitor+si-NC组相比,miR-155-5p inhibitor+si-Ndfip1组细胞增殖、侵袭及迁移能力均明显提升(P<0.05),细胞凋亡数量减少(P<0.05),p-PI3K和p-AKT蛋白表达量增高(P<0.05)。结论:miR-155-5p靶向调控Ndfip1影响乳腺癌MCF-7细胞恶性生物学行为改变,这种改变可能与PI3K/AKT信号通路蛋白表达相关。

Objective:To explore the effect of miR-155-5p on the proliferation,apoptosis,invasion and migration of breast cancer cells by targeting Ndfip1 and the related mechanisms.Methods:Real-time fluorescence quantitative polymerase chain reaction(qRT-PCR)was used to detect miR-155-5p and Ndfip1 gene expressions in 25 cases of breast cancer with definite pathological diagnosis and corresponding adjacent normal tissues.MCF-7 cells were selected.Cells of Control group,NC-inhibitor group,miR-155-5p inhibitor group,miR-155-5p inhibitor+si-NC group and miR-155-5p inhibitor+si-Ndfip1 group were constructed.CCK8 assay,flow cytometry,Transwell assay and scratch assay were used to detect changes in cell proliferation,apoptosis,invasion and migration ability,respectively.Western blot assay was used to detect protein expression related to PI3K/AKT signaling pathway.The downstream target genes of miR-155-5p were predicted by Targetscan bioinformatics website,and the correlation between miR-155-5p and target gene Ndfip1 was detected by dual luciferase assay.Results:Compared with adjacent normal tissues,the gene expression of miR-155-5p in breast cancer tissues was significantly increased(P<0.05),and the gene expression of Ndfip1 in breast cancer tissues was significantly decreased ( P <0.05),and the expression of miR-155-5p and Ndfip1 in breast cancer tissues was negatively correlated ( r =-0.969 8, P <0.001).Compared with Control group and NC-inhibitor group,cell proliferation,invasion and migration of miR-155-5p inhibitor group were decreased ( P <0.05),apoptosis number was increased ( P <0.05),p-PI3K and p-AKT protein expression levels were decreased ( P <0.05).Dual luciferase reporter gene assay confirmed that Ndfip1 was the target of miR-155-5p.The recovery test showed that compared with the miR-155-5p inhibitor group and the miR-155-5p inhibitor+si-NC group,cell proliferation,invasion and migration of miR-155-5p inhibitor+si-Ndfip1 group were significantly increased ( P <0.05),the number of apoptosis decreased ( P <0.05),and the expression levels of p-PI3K and p-AKT protein increased ( P <0.05). Conclusion: The targeted regulation of Ndfip1 by miR-155-5p may affect the malignant biological behavior changes of breast cancer MCF-7 cells,which may be related to the expression of PI3K/AKT signaling pathway protein.