microRNA-148a-3p 通过依赖于 Ago2 的方式抑制 SMAD2 表达抑制肝癌细胞的进展。
MicroRNA-148a-3p inhibits progression of hepatocelluar carcimoma by repressing SMAD2 expression in an Ago2 dependent manner.
机构信息
Hepatic Surgery Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan, 430030, China.
Clinical Medical Research Center of Hepatic Surgery at Hubei Province, Wuhan, China.
出版信息
J Exp Clin Cancer Res. 2020 Aug 4;39(1):150. doi: 10.1186/s13046-020-01649-0.
BACKGROUND
Hepatocellular carcinoma (HCC) is one of the most prevalent common cancer worldwide with high mortality. Transforming growth factor-β (TGF-β) signaling pathway was reported dysregulated during liver cancer formation and progression. As a key component of TGF-β signaling, the role of SMAD2 and its regulatory mechanisms in HCC remain unclear.
METHODS
SMAD2 expression in paired HCC specimens were determined by western blot and immunohistochemistry (IHC). quantitative real-time PCR (qRT-PCR) was used to measure mRNA and microRNA (miRNA) expression level. Cell migration, invasion and proliferation ability were evaluated by transwell, CCK8 and EdU assay. In silico websites were used to manifest overall survival rates of HCC patients or to predict miRNAs targeting SMAD2. Dual luciferase reporter assay and anti-Ago2 immunoprecipitation assay were performed to confirm the binding between SMAD2 mRNA and miRNA-148a-3p (miR-148a). Tumorigenesis and lung metastasis mouse model were used to explore the role of miR-148a in vivo. In situ hybridization (ISH) was conducted to determine the expression of miR-148a in liver tissues.
RESULTS
In this study, we found that SMAD2 was highly expressed in HCC and elevated SMAD2 expression predicted shorter overall survival (OS) time for HCC patients. SMAD2 promoted mobility and proliferation of HCC cells in vitro. We further revealed that the expression of miR-148a was negatively correlated with SMAD2 and found that miR-148a repressed SMAD2 expression by downregulating its mRNA through binding with Argonaute 2 (Ago2) in HCC. Transwell, CCK8 and animal experiments exhibited miR-148a inhibited metastasis and proliferation of HCC in vitro and in vivo. Moreover, the phenotype changes caused by miR-148a manipulation were recovered by rescuing SMAD2 expression in HCC cells. ISH assay indicated miR-148a was downregulated in HCC and low expression of miR-148a associated with more aggressive clinic features and poor prognosis.
CONCLUSION
miR-148a was identified as a repressor of HCC progression by downregulating SMAD2 in an Ago2 dependent manner.
背景
肝细胞癌（HCC）是全球最常见的癌症之一，死亡率很高。转化生长因子-β（TGF-β）信号通路在肝癌的形成和发展过程中被报道失调。SMAD2 作为 TGF-β信号的关键组成部分，其在 HCC 中的作用及其调节机制尚不清楚。
方法
通过 Western blot 和免疫组织化学（IHC）检测配对 HCC 标本中 SMAD2 的表达。采用定量实时 PCR（qRT-PCR）检测 mRNA 和 microRNA（miRNA）表达水平。通过 Transwell、CCK8 和 EdU 测定评估细胞迁移、侵袭和增殖能力。利用在线网站分析 HCC 患者的总生存率或预测靶向 SMAD2 的 miRNA。通过双荧光素酶报告基因检测和抗 Ago2 免疫沉淀检测证实 SMAD2 mRNA 与 miRNA-148a-3p（miR-148a）之间的结合。利用肿瘤发生和肺转移小鼠模型在体内研究 miR-148a 的作用。通过原位杂交（ISH）检测肝组织中 miR-148a 的表达。
结果
本研究发现 SMAD2 在 HCC 中高表达，SMAD2 表达升高预示 HCC 患者总生存期（OS）较短。SMAD2 体外促进 HCC 细胞的迁移和增殖。我们进一步揭示 miR-148a 的表达与 SMAD2 呈负相关，并发现 miR-148a 通过与 HCC 中的 Argonaute 2（Ago2）结合，下调其 mRNA，从而抑制 SMAD2 的表达。Transwell、CCK8 和动物实验表明 miR-148a 可抑制 HCC 细胞在体外和体内的转移和增殖。此外，在 HCC 细胞中恢复 SMAD2 表达可恢复 miR-148a 操纵引起的表型变化。ISH 检测表明 miR-148a 在 HCC 中下调，低表达 miR-148a 与侵袭性临床特征和不良预后相关。
结论
miR-148a 通过依赖 Ago2 的方式下调 SMAD2，被鉴定为 HCC 进展的抑制剂。
Zotero 插件