miR-139-5p对人肝癌细胞增殖和侵袭的影响  

Effect of miR-139-5p on the cell proliferation and invasion of human hepatocellular carcinoma

作　　者：袁震[1] 杨晓娣 于红梅[1] 叶涛[1] YUAN Zhen;YANG Xiaodi;YU Hongmei;YE Tao(Department of Oncology,Minhang Hospital,Fudan University,Shanghai 201199,China)

机构地区：[1]复旦大学附属闵行医院肿瘤科,上海201199

出　　处：《肝胆胰外科杂志》2023年第3期156-163,共8页Journal of Hepatopancreatobiliary Surgery

基　　金：上海市闵行区科委项目(2021MHZ100)。

摘　　要：目的 探讨微小RNA(miRNA)-139-5p在肝癌组织中的表达及其对人肝癌细胞增殖、侵袭的影响和潜在的分子机制。方法 采用生物信息学方法,从GEO数据库中下载GSE36915数据集,进行差异miRNA分析,结合TCGA-LICH数据集中的生存数据,筛选与预后显著相关的miRNA。采用实时荧光定量PCR(qRT-PCR)检测miR-139-5p在肝癌及其癌旁组织中的表达。利用CCK-8实验、Transwell实验观察miR-139-5p对人肝癌细胞SK-Hep-1和SMMC-7721的表型变化。采用高通量测序检测过表达miR-139-5p的SK-Hep-1细胞中基因表达的变化,确定miR-139-5p调控的潜在的信号通路和靶基因,并采用免疫印迹实验和报告基因实验进行验证。结果 通过对GSE36915数据集进行分析,共鉴定到50个显著差异表达的miRNA。结合TCGA-LICH数据集中的生存数据,发现在差异最显著的20个miRNA中,miR-15b-3p、miR-1180-3p、miR-139-5p、miR-139-3p与预后显著相关,最终确定miR-139-5p为进一步研究对象。miR-139-5p在100对肝癌组织的相对表达水平显著低于对应癌旁组织[(0.50±0.33) vs (1.02±0.43),P<0.001],且低水平的miR-139-5p肝癌患者中位生存期较短(25.5个月vs 43.0个月,P<0.01)。人肝癌细胞系SK-Hep-1和SMMC-7721中过表达miR-139-5p,可以显著抑制肝癌细胞的增殖和侵袭。高通量测序分析发现,在过表达miR-139-5p的SK-Hep-1细胞中,差异表达基因显著富集在Sphingolipid信号通路。经免疫印迹实验和报告基因实验验证,Sphingolipid信号通路中神经酰胺合成酶6(ceramide synthase 6,CerS6)是miR-139-5p的新的靶点。结论 miR-139-5p在肝癌组织中表达减少,导致其下游靶基因CerS6过量表达,从而激活Sphingolipid信号通路,促进肝癌发生发展。本研究提示miR-139-5p可以作为肝癌治疗的新的靶点。Objective To explore the effect of miR-139-5p on the cell proliferation and invasion of human hepatocellular carcinoma(HCC)and the underlying mechanism.Methods The differently expressed miRNAs were identified by the miRNA expression data from GSE36915,which was downloaded from GEO database.Combined with the prognostic data from the TCGA-LICH dataset,the survival-related miRNAs were screened out.Then,RT-qPCR was performed to detect the expression of miR-139-5p in HCC and adjacent normal samples.CCK-8 and transwell assays were performed to explore the effect of miR-139-5p on the cell proliferation and invasion of HCC,respectively.Finally,the miR-139-5p-regulated KEGG signaling pathways and target gene via high-throughput sequencing were investigated,followed by validation with double luciferase assay and Western blotting.Results By bioinformatic analysis of GSE36915,50 differently expression miRNAs were identified.Combined with the TCGA-LICH datasets,4 miRNAs as survival-related miRNA were screened out,including miR-15b-3p,miR-1180-3p,miR-139-5p and miR-139-3p.Furthermore,miR-139-5p was selected for validation experiments.Compared with adjacent normal tissues,the expression of miR-139-5p was significantly decreased in HCC tissues[(0.50±0.33)vs(1.02±0.43),P<0.001],and for HCC patietnts,lower expression of miR-139-5p predicted shorter median survival time(25.5 months vs 43.0 months,P<0.01).The overexpression of miR-139-5p inhibited the cell proliferation and invasion in HCC cell lines SK-Hep-1 and SMMC-7721.Finally,it was demonstrated that miR-139-5p-regulated genes were significantly enriched in Sphingolipid signaling pathway,and double luciferase assay and Western blotting results showed that ceramide synthase 6(CerS6)was a novel target gene of miR-139-5p.Conclusion The expression of miR-139-5p is reduced in HCC tissues,which leads to overexpression of its target gene CerS6.Then CerS6 activates the Sphingolipid signaling pathway,promotes the development of HCC.This study suggests that miR-139-5p is a potential no

关 键 词：肝细胞癌 miR-139-5p Sphingolipid信号通路 神经酰胺合成酶6(CerS6) 

分 类 号：R735.7[医药卫生—肿瘤]