Department of Pediatrics, the Affiliated Shengjing Hospital, China Medical University, Shenyang, Liaoning 110004, P.R. China.
Int J Mol Med. 2017 Nov;40(5):1435-1443. doi: 10.3892/ijmm.2017.3120. Epub 2017 Sep 6.
Trefoil factor 3 (TFF3) reconstructs the epithelial barrier by stimulating epithelial cell migration and proliferation, and significantly contributes to intestinal mucosal damage and healing. In a previous study, TFF3 was identified as a novel target of microRNA-7-5p (miR-7-5p). The aim of the present study was to investigate the roles and mechanisms of miR-7-5p in the proliferation and migration of intestinal epithelial cells. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed to determine the expression level of miR-7-5p in the experimental groups. In addition, western blot analysis was performed to examine the expression levels of TFF3, phosphoinositide 3-kinase (PI3K), Akt and phosphorylated (p)-AKT when miR-7-5p or TFF3 was overexpressed, and the effects of miR-7-5p and TFF3 on LS174T cell proliferation and migration were simultaneously investigated. miR-7-5p was demonstrated to decrease the expression level of TFF3, and inhibit LS174T cell proliferation and migration, which was accompanied by decreased expression levels of PI3K and p-Akt. miR-7-5p was decreased following combined treatment with the TFF3 plasmid and miR‑7-5p mimics, compared with treatment with miR-7-5p mimics alone, which was accompanied by increased expression levels of TFF3, PI3K and p-Akt, and enhanced LS174T cell proliferation and migration effects. The expression levels of miR-7-5p in the miRNA negative control (NC) + LY294002 group, the miR‑7-5p mimic + LY294002 group, and the miR-7-5p mimic + TFF3 plasmid + LY294002 group were higher than those in the NC group, the miR-7-5p mimic group and the miR-7-5p mimic + TFF3 plasmid group, respectively. Accordingly, the expression level of TFF3 was downregulated and the proliferation and migration ability of the cells was downregulated. The present study demonstrates that overexpressed miR-7-5p may inhibit the proliferation and migration of LS174T cells by targeting the expression of TFF3 via inhibiting the PI3K/Akt signalling pathway. The PI3K/Akt signalling pathway may exert a feedback regulation effect on miR-7-5p, inhibiting the activity of this signalling pathway, which increases the miR-7-5p expression levels and further enhances the effects of miR-7-5p on cell proliferation and migration.
三叶因子 3（TFF3）通过刺激上皮细胞迁移和增殖来重建上皮屏障，对肠道黏膜损伤和愈合有重要贡献。在之前的一项研究中，TFF3 被鉴定为 microRNA-7-5p（miR-7-5p）的一个新靶点。本研究旨在探讨 miR-7-5p 在肠上皮细胞增殖和迁移中的作用和机制。采用逆转录定量聚合酶链反应（RT-qPCR）检测实验组中 miR-7-5p 的表达水平。此外，当过表达 miR-7-5p 或 TFF3 时，通过 Western blot 分析检测 TFF3、磷酸肌醇 3-激酶（PI3K）、Akt 和磷酸化（p）-Akt 的表达水平，并同时研究 miR-7-5p 和 TFF3 对 LS174T 细胞增殖和迁移的影响。结果表明，miR-7-5p 降低了 TFF3 的表达水平，抑制了 LS174T 细胞的增殖和迁移，同时伴随着 PI3K 和 p-Akt 的表达水平降低。与单独转染 miR-7-5p 模拟物相比，转染 TFF3 质粒和 miR-7-5p 模拟物的混合物后，miR-7-5p 的表达水平降低，同时伴随着 TFF3、PI3K 和 p-Akt 的表达水平升高，以及 LS174T 细胞增殖和迁移作用增强。miRNA 阴性对照（NC）+LY294002 组、miR-7-5p 模拟物+LY294002 组和 miR-7-5p 模拟物+TFF3 质粒+LY294002 组的 miR-7-5p 表达水平均高于 NC 组、miR-7-5p 模拟物组和 miR-7-5p 模拟物+TFF3 质粒组。相应地，TFF3 的表达水平下调，细胞的增殖和迁移能力下调。本研究表明，过表达的 miR-7-5p 可能通过靶向 TFF3 的表达抑制 PI3K/Akt 信号通路，抑制 LS174T 细胞的增殖和迁移。PI3K/Akt 信号通路可能对 miR-7-5p 发挥反馈调节作用，抑制该信号通路的活性，增加 miR-7-5p 的表达水平，进一步增强 miR-7-5p 对细胞增殖和迁移的作用。
