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microRNA-27a-3p 通过靶向 SFRP1 调控 Wnt/β-catenin 信号通路促进口腔鳞状细胞癌干细胞上皮间质转化

MicroRNA-27a-3p Modulates the Wnt/β-Catenin Signaling Pathway to Promote Epithelial-Mesenchymal Transition in Oral Squamous Carcinoma Stem Cells by Targeting SFRP1.

机构信息

Department of Stomatology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, P. R. China.

Department of Oral and Maxillofacial Surgery, Guanghua School and Hospital of Stomatology; Guangdong Provincial Key Laboratory of Oral Diseases, Sun Yat-sen University, Guangzhou 510055, P. R. China.

出版信息

Sci Rep. 2017 Apr 20;7:44688. doi: 10.1038/srep44688.

Abstract

This study aimed to elucidate how microRNA27a-3p (miR-27a-3p) modulates the Wnt/β-catenin signaling pathway to promote the epithelial-mesenchymal transition (EMT) in oral squamous carcinoma stem cells (OSCSCs) by targeting secreted frizzled-related protein 1 (SFRP1). Flow cytometry was used to sort OSCSCs from the SCC-9 and Tca8113 cell lines. The OSCSCs were randomly assigned into the miR-27a-3p inhibitors group, the miR-27a-3p inhibitors-NC group, the si-SFRP1 group, the si-SFRP1 + miR-27a-3p inhibitors group and the blank group. A luciferase reporter, immunofluorescence and Transwell assays were performed to detect luciferase activity, SFRP1, and cell migration and invasion, respectively. The mRNA expression of miR-27a-3p, SFRP1 and EMT markers (E-cadherin, N-cadherin, vimentin and ZEB1) were detected using qRT-PCR. The protein expression of SFRP1, EMT markers and the proteins of the Wnt/β-catenin signaling pathway was detected by Western blotting. OSCSCs showed up-regulated miR-27a-3p, Wnt/β-catenin signaling pathway-related proteins, vimentin, N-cadherin and ZEB1 and down-regulated SFRP1 and E-cadherin. MiR-27a-3p targeted SFRP1. Down-regulated miR-27a-3p resulted in increased E-cadherin and SFRP1 but decreased vimentin, N-cadherin, ZEB1, the Wnt/β-catenin signaling pathway-related proteins, and invasive and migratory cells. Silenced SFRP1 reversed this effect. We found that miR-27a-3p modulated the Wnt/β-catenin signaling pathway to promote EMT in OSCSCs by down-regulating SFRP1.

摘要

本研究旨在阐明 microRNA27a-3p(miR-27a-3p)如何通过靶向分泌型卷曲相关蛋白 1(SFRP1)来调节 Wnt/β-catenin 信号通路,从而促进口腔鳞状细胞癌干细胞(OSCSCs)中的上皮-间充质转化(EMT)。流式细胞术用于从 SCC-9 和 Tca8113 细胞系中分选 OSCSCs。将 OSCSCs 随机分为 miR-27a-3p 抑制剂组、miR-27a-3p 抑制剂-NC 组、si-SFRP1 组、si-SFRP1+miR-27a-3p 抑制剂组和空白组。通过荧光素酶报告、免疫荧光和 Transwell 测定分别检测荧光素酶活性、SFRP1 以及细胞迁移和侵袭。使用 qRT-PCR 检测 miR-27a-3p、SFRP1 和 EMT 标志物(E-钙粘蛋白、N-钙粘蛋白、波形蛋白和 ZEB1)的 mRNA 表达。通过 Western blot 检测 SFRP1、EMT 标志物和 Wnt/β-catenin 信号通路蛋白的表达。OSCSCs 表现出上调的 miR-27a-3p、Wnt/β-catenin 信号通路相关蛋白、波形蛋白、N-钙粘蛋白和 ZEB1,以及下调的 SFRP1 和 E-钙粘蛋白。miR-27a-3p 靶向 SFRP1。下调 miR-27a-3p 导致 E-钙粘蛋白和 SFRP1 增加,但波形蛋白、N-钙粘蛋白、ZEB1、Wnt/β-catenin 信号通路相关蛋白以及侵袭和迁移细胞减少。沉默 SFRP1 逆转了这一效应。我们发现,miR-27a-3p 通过下调 SFRP1 来调节 Wnt/β-catenin 信号通路,从而促进 OSCSCs 中的 EMT。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/bbf1/5397903/7e4712045277/srep44688-f1.jpg

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