Involvement of human micro-RNA in growth and response to chemotherapy in human cholangiocarcinoma cell lines

BACKGROUND & AIMS: Micro-RNA (miRNA) are endogenous regulatory RNA molecules that modulate gene expression. Alterations in miRNA expression can contribute to tumor growth by modulating the functional expression of critical genes involved in tumor cell proliferation or survival. Our aims were to identify specific miRNA involved in the regulation of cholangiocarcinoma growth and response to chemotherapy. METHODS: miRNA expression in malignant and nonmalignant human cholangiocytes was assessed using a microarray. Expression of selected miRNA and their precursors was evaluated by Northern blots and real-time polymerase chain reaction, respectively. The effect of selected miRNA on cell growth and response to chemotherapy was assessed using miRNA-specific antisense oligonucleotides to decrease miRNA expression or with precursor miRNA to increase cellular expression. RESULTS: miRNA expression was markedly different in malignant cholangiocytes, with decreased expression of many miRNA compared with nonmalignant cells. A cluster of miRNA, including miR-320, miR-200b, miR-21, miR-23a, miR-141, miR-27a, and miR-34a, were expressed in all cell lines. MiR-21, miR-141, and miR-200b were highly over-expressed in malignant cholangiocytes. Inhibition of miR-21 and miR-200b increased sensitivity to gemcitabine, whereas inhibition of miR-141 decreased cell growth. Treatment of tumor cell xenografts with systemic gemcitabine altered the expression of a significant number of miRNA. miR-21 modulates gemcitabine-induced apoptosis by phosphatase and tensin homolog deleted on chromosome 10 (PTEN)-dependent activation of PI 3-kinase signaling. Potential target genes that were modulated by selected miRNA were identified. CONCLUSIONS: Alterations in miRNA expression contribute to tumor growth and response to chemotherapy. Aberrantly expressed miRNA or their targets will provide mechanistic insight and therapeutic targets for cholangiocarcinoma.     

miR-21 targets and inhibits tumor suppressor gene PTEN to promote prostate cancer cell proliferation and invasion: An experimental study

Objective: To study whether miR-21 targets and inhibits tumor suppressor gene PTEN can promote prostate cancer cell proliferation and invasion,Methods: Prostate cancer cell lines PC-3 were cultured and divided into negative control group(NC group),miR-21 group,pc DNA3.1 group,miR-21+pc DNA3.1 group and miR-21+PTEN group that were transfected with different mi R and plasmid,respectively,After 12 h and 24 h of transfection,the cell viability and invasive cell number were determined; after 24 h of transfection,Bcl-2,Survivin,MMP2,MMP9,PTEN,PI3 K,and AKT expression in cells were determined,Results: After 12 h and 24 h of transfection,OD value and invasive cell number of miR-21 group were significantly higher than those of NC group; after 24 h of transfection,Bcl-2,Survivin,MMP2,MMP9,PI3 K and AKT expression levels were significantly higher than those of NC group while PTEN expression level was significantly lower than that of NC group; after 12 h and 24 h of transfection,OD value and invasive cell number of mi R-21+pcDNA3.1 group were significantly higher than those of pc DNA3.1 group,and the OD value and invasive cell number of mi R-21+PTEN group were significantly lower than those of mi R-21+pcDNA3.1 group; after 24 h of transfection,Bcl-2,Survivin,MMP2 and MMP9 content of mi R-21+pc DNA3.1 group were significantly higher than those of pcDNA3.1 group,and Bcl-2,Survivin,MMP2 and MMP9 content of miR-21+PTEN group were significantly lower than those of mi R-21+pcDNA3.1 group,Conclusions: miR-21 can target and inhibit tumor suppressor gene PTEN expression to promote prostate cancer cell proliferation and invasion.     

MicroRNA-21在肿瘤研究中的进展

微小RNA-21（miR-21）是由约22个核苷酸组成的参与转录后基因调控的非编码小分子RNA。miR-21在多种人类肿瘤细胞中过表达,目前研究发现其参与了细胞的分化、增殖和凋亡,与肿瘤发生密切相关。miR-21可能的靶基因有程序性细胞死亡因子4（PDCD4）、maspin、原肌球蛋白1（TPM1）、磷酸酯酶-张力蛋白同源物（PTEN）等。miR-21通过对靶基因的调控参与肿瘤细胞的生长、侵袭、转移和耐药。miR-21反义寡核苷酸与抗肿瘤药物联合应用将是肿瘤治疗的一个新方向。     

MicroRNA-194 inhibits cell invasion and migration in hepatocellular carcinoma through PRC1-mediated inhibition of Wnt/β-catenin signaling pathway

BackgroundHepatocellular carcinoma (HCC) is a commonly occurring malignancy accompanied by significant mortality rates. More recently, extensive investigations into microRNA (miRNA) expression profiles have been conducted to identify their ability to inhibit tumors. Thus, this study explored the role of miR-194 in epithelial-mesenchymal transition (EMT), cell invasion and migration through Wnt/β-catenin signaling pathway by binding to protein regulator of cytokinesis 1 (PRC1) in HCC.MethodsInitially, HCC related microarray data were retrieved and analyzed, and regulatory miRNAs of PRC1 were predicted accordingly. Next, the roles of miR-194, PRC1, and Wnt/β-catenin signaling pathway in HCC were determined, with relationship between PRC1 and miR-194 being verified subsequently. The role of miR-194 in cell EMT, migration, proliferation and invasion was evaluated through gain- and loss- function studies. Finally, tumor xenograft in nude mice was induced to assess tumor growth of HCC.ResultsmiR-194 affected HCC development in Wnt/β-catenin signaling pathway with putative binding sites to PRC1. MiR-194 could target PRC1. MiR-194 was downregulated while PRC1 was upregulated in HCC tissues. Additionally, miR-194 elevation and PRC1 silencing could suppress EMT, growth, proliferation, invasion, and migration in HCC cells by inactivating Wnt/β-catenin signaling pathway.ConclusionTaken together, this study demonstrated that miR-194 inhibited EMT, cell invasion and migration through inactivation of PRC1-dependent Wnt/β-catenin signaling pathway.     

Exploration of the regulatory effect of miR-21 on breast cancer cell line proliferation and invasion as well as the downstream target genes

Objective: To study the regulatory effects of miR-21 on breast cancer cell line proliferation and invasion as well as the downstream target genes. Methods: Breast cancer cell lines MCF-7 were cultured and transfected with miR-21 mimics and the corresponding negative control mimics(NC mimics), and then MTS kits were used to detect cell viability. Transwell experiment was used to detect cell invasion ability, and fluorescence quantitative PCR was used to detect the expression of proliferation and invasion-related genes in cells. Results: 24 h after transfection of miR-21 mimics and NC mimics, cell OD value and the number of invasive cells of miR-21 group were significantly higher than those of NC group, and m RNA contents of PDCD-4, Fas L, PTEN, Rho B, Maspin, TIMP3 and RECK in cells were significantly lower than those of NC group. Conclusion: miR-21 can promote the proliferation and invasion of breast cancer cell lines, and its downstream target genes include PDCD-4, Fas L, PTEN, Rho B, Maspin, TIMP3 and RECK.     

沉默miR-107对乳腺癌细胞侵袭及上皮-间质转化的影响

目的探究miR-107对乳腺癌细胞侵袭及上皮-间质转化(EMT)的影响和机制。方法利用荧光定量PCR方法分析miR-107在正常乳腺细胞及乳腺癌细胞系中的表达差异;HS578T细胞分别转染对照miRNA和miR-107抑制剂,利用划痕实验分析细胞迁移能力,用Transwell实验分析细胞侵袭能力,用实时荧光定量PCR方法检测细胞EMT标志物的表达水平,用蛋白质免疫印迹实验检测细胞PTEN/AKT信号通路。结果与正常乳腺细胞相比,miR-107在乳腺癌细胞株中高表达;沉默miR-107的表达抑制HS578T细胞的迁移、侵袭及EMT,促进PTEN/AKT信号通路。结论miR-107在乳腺癌细胞中高表达,沉默miR-107表达通过PTEN/AKT信号通路抑制乳腺癌细胞迁移、侵袭和EMT。     

Down-Regulation of PTEN Expression Modulated by Dysregulated miR-21 Contributes to the Progression of Esophageal Cancer

Background and Aim

miR-21, a putative tumor oncomiR, is a frequently overexpressed miRNA in a variety of tumors. Because it targets tumor-suppressor genes it has been linked to tumor progression. In this study we investigated the role of miR-21 in esophageal squamous cell carcinoma (ESCC), and its possible mechanism.

Methods

Expression of miR-21 was detected by stem–loop RT-PCR in tissue from 76 invasive ESCC at stage I–IV and in their corresponding para-cancerous histological normal tissues (PCHNT). Thirty endoscopic esophageal mucosal biopsy specimens from non-tumor patients were used as controls. Expression of PTEN in 76 paired ESCC and PCHNT was investigated by real-time RT-PCR and an immunohistochemical method, respectively. Paired tumor and PCHNT specimens of 20 ESCC cases were randomly selected for western blot analysis. The effect of miR-21 on PTEN expression was assessed in the ESCC cell line with an miR-21 inhibitor to reduce miR-21 expression. Furthermore, the roles of miR-21 in cell biology were analyzed by use of miR-21 inhibitor-transfected cells.

Results

Stem–loop RT-PCR revealed miR-21 was significantly overexpressed in ESCC tissues and cell lines. Overexpression of miR-21 correlated with tumor status, lymph node metastasis, and clinical stage. We demonstrated that knockdown of miR-21 significantly increased expression of PTEN protein. Consequent PTEN expression reduced cell proliferation, invasion, and migration.

Conclusions

Our findings suggest that miR-21 could be a potential oncomiR, probably by regulation of PTEN, and a novel prognostic factor for ESCC patients.