Department of Thoracic Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, China.
The Central Operating Room, The First Affiliated Hospital of Harbin Medical University, Harbin, China.
Cancer Res Treat. 2018 Jul;50(3):936-949. doi: 10.4143/crt.2017.302. Epub 2017 Oct 12.
The purpose of this study was to investigate the biological role and mechanism of miR-373 targeting of TFIIB-related factor 2 (BRF2) in the regulation of non-small cell lung cancer (NSCLC) cells.
miRNA microarray chip analysis of four paired NSCLC and adjacent non-tumor tissues was performed. Quantitative real-time polymerase chain reaction (qRT-PCR) andwestern blotting were used to detect the expression levels of miR-373 and BRF2 in NSCLC tissues and cell lines. The dual-luciferase reporter method was performed to determine if BRF2 is a target of miR-373. MTT, wound-healing, Transwell, and flow cytometric assays were conducted to examine the proliferation, migration, invasion, and cell cycle progression of NSCLC A549 cells, respectively; western blotting was used to detect the expression of epithelial-mesenchymal transition (EMT)-related proteins.
The miRNA microarray chip analysis demonstrated that miR-373 was down-regulated in NSCLC tissues, and this result was confirmed by qRT-PCR. Additionally, miR-373 was confirmed to target BRF2. Moreover, miR-373 expression was inversely correlated with BRF2 expression in NSCLC tissues and cell lines; both miR-373 down-regulation and BRF2 up-regulation were strongly associated with the clinicopathological features and prognosis of NSCLC patients. In vitro, overexpression of miR-373 markedly inhibited cell proliferation, migration, and invasion; up-regulated the expression of E-cadherin; and down-regulated the expression of N-cadherin and Snail in A549 cell. Knockdown BRF2 by siRNA resulted in effects similar to those caused by overexpression of miR-373.
MiR-373 is decreased in NSCLC, and overexpression of miR-373 can suppress cell EMT, and inhibit the proliferation, migration, and invasion of NSCLC A549 cells by targeting BRF2.
本研究旨在探讨 microRNA-373（miR-373）靶向 TFIIB 相关因子 2（BRF2）在调控非小细胞肺癌（NSCLC）细胞中的生物学作用和机制。
对 4 对 NSCLC 组织及其相邻非肿瘤组织进行 miRNA 芯片分析。采用实时定量聚合酶链反应（qRT-PCR）和 Western blot 检测 NSCLC 组织和细胞系中 miR-373 和 BRF2 的表达水平。双荧光素酶报告基因检测法确定 BRF2 是否为 miR-373 的靶基因。采用 MTT、划痕愈合、Transwell 和流式细胞术分别检测 NSCLC A549 细胞的增殖、迁移、侵袭和细胞周期进展情况；Western blot 检测上皮间质转化（EMT）相关蛋白的表达。
miRNA 芯片分析显示，miR-373 在 NSCLC 组织中表达下调，qRT-PCR 结果进一步证实了这一点。此外，miR-373 被证实靶向调控 BRF2。进一步研究发现，miR-373 在 NSCLC 组织和细胞系中的表达与 BRF2 呈负相关；miR-373 下调和 BRF2 上调均与 NSCLC 患者的临床病理特征和预后密切相关。体外实验结果显示，过表达 miR-373 可显著抑制 A549 细胞的增殖、迁移和侵袭；上调 E-钙黏蛋白的表达；下调 N-钙黏蛋白和 Snail 的表达。siRNA 敲低 BRF2 的表达可产生与过表达 miR-373 相似的效果。
miR-373 在 NSCLC 中表达下调，过表达 miR-373 可通过靶向 BRF2 抑制 NSCLC A549 细胞的 EMT，并抑制其增殖、迁移和侵袭。
