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Frontiers of Medicine

ISSN 2095-0217

ISSN 2095-0225(Online)

CN 11-5983/R

Postal Subscription Code 80-967

2018 Impact Factor: 1.847

Front. Med.    2014, Vol. 8 Issue (4) : 456-463     DOI: 10.1007/s11684-014-0353-z
RESEARCH ARTICLE |
Effects of miR-200c on the migration and invasion abilities of human prostate cancer Du145 cells and the corresponding mechanism
Runlin Shi,Haibing Xiao,Tao Yang,Lei Chang,Yuanfeng Tian,Bolin Wu,Hua Xu()
Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
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Abstract  

microRNAs (miRNAs) have played a key role in human tumorigenesis, tumor progression, and metastasis. On the one hand, miRNAs are aberrantly expressed in many types of human cancer; on the other hand, miRNAs can function as tumor suppressors or oncogenes that target many cancer-related genes. This study aimed to investigate the effects of miRNA-200c (miR-200c) on the biological behavior and mechanism of proliferation, migration, and invasion in the prostate cancer cell line Du145. In this study, Du145 cells were transfected with miR-200c mimics or negative control miR-NC by using an X-tremeGENE siRNA transfection reagent. The relative expression of miR-200c was measured by RT-PCR. The proliferation, migration, and invasion abilities of Du145 cells were detected by CCK8 assays, migration assays and invasion assays, respectively. The expressions of ZEB1, E-cadherin, and vimentin were observed by western blot. Results showed that DU145 cells exhibited a high expression of miR-200c compared with immortalized normal prostate epithelial cell RWPE-1. Du145 cells were then transfected with miR-200c mimics and displayed lower abilities of proliferation, migration, and invasion than those transfected with the negative control. The protein levels of ZEB1 and vimentin were expressed at a low extent in Du145 cells, which were transfected with miR-200c mimics; by contrast, E-cadherin was highly expressed. Hence, miR-200c could significantly inhibit the proliferation of the prostate cancer cell line Du145; likewise, miR-200c could inhibit migration and invasion by epithelial-mesenchymal transition.

Keywords miR-200c      proliferation      migration      invasion      prostate cancer      Du145 cell      ZEB1     
Corresponding Authors: Hua Xu   
Issue Date: 18 December 2014
URL:  
http://academic.hep.com.cn/fmd/EN/10.1007/s11684-014-0353-z     OR     http://academic.hep.com.cn/fmd/EN/Y2014/V8/I4/456
Fig.1  Expression of Du145 cells compared with that of human non-transformed prostate epithelial cell line RWPE-1; miR-200c expression in prostate cancer Du145 cells was reduced significantly. **P < 0.01.
Fig.2  Expressions of miR-200c and ZEB1 in the cells detected by qPCR. A reaction system (20 μl) and other systems were amplified and detected under the following conditions: 40 thermo cycles at 95 °C for 15 s and 60 °C for 20 s; these reactions were extended at 70 °C for 20 s. miR-200c may regulate ZEB1 expression on a transcriptional level, *P < 0.05 and ** P < 0.01. Difference was statistically significant.
Fig.3  CCK-8 assay revealing the proliferation of the prostate cancer cell line Du145. miR-200c could significantly suppress the proliferation of prostate cancer Du145 cells (P < 0.05; Fig. 3A). In vitro invasion assays showed that miR-200c could significantly suppress the invasion ability of prostate cancer cells Du145 (51.279 ± 11.338, * P < 0.05; Fig. 3B). In Transwell migration assay, cells were placed in a well at a density of 2×105 cells/well; Du145 cells were stably transfected with miR-200c mimics and miR-220-NC. The cells then penetrated the lower surface of the filter without or with Matrigel. Hence, prostate cancer Du145 cells were stably transfected with miR-200c mimics (34.345 ± 6.238, * P < 0.05) presented an impaired migration ability compared with those transfected with miR-NC group (the negative control group; Fig. 3C).
Fig.4  Epithelial-to-mesenchymal transition. Western blot result indicated that the expressions of ZEB1 and vimentin were reduced when miR-200c was transfected in the cells; on the contrary, the expression of E-cadherin was increased. Difference was statistically significant.
Fig.5  Prediction of target genes showing that locus 5 of the 3′ UTR in ZEB1 could be combined with miR-200c.
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