Chong Zhong1, Ming-Yi Li2, Zhi-Yuan Chen3, Hai-Kun Cheng1, Ming-Li Hu1, Yue-Lu Ruan1, Rong-Ping Guo4. 1. Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangzhou University of Traditional Chinese Medicine Guangzhou 510405, P. R. China. 2. Department of Radiotherapy, Cancer Center of Gangzhou Medical University Guangzhou 510095, P. R. China. 3. Department of Gastrointestinal Surgery, The Sixth Affiliated Hospital of Sun Yat-Sen University Guangzhou 510655, P. R. China. 4. Department of Hepatobiliary Surgery, Sun Yat-Sen University Cancer Center Guangzhou 510060, P. R. China.
Abstract
OBJECTIVE: Our study investigated the role of microRNA (miR)-200a and its molecular targets in hepatocellular carcinoma (HCC) cells. METHODS: An inhibitor of miR-200a was transiently transfected into the hepatocellular carcinoma cell line, MHCC-97L. The effect of this transfection on mRNA levels of epithelial-mesenchymal transition (EMT)-related genes was measured by fluorescence-based quantitative real-time polymerase chain reaction (qRT-PCR). Further, protein levels of EMT-related genes, cell proliferation and apoptosis-related markers were assessed by Western blot analysis in these transfected cells. MTT and wound-healing assay were used to evaluate the proliferation and migration of MHCC-97L cells in presence and in absence of miR-200a inhibitor. RESULTS: Compared with miR-NC control group, qRT-PCR results in anti-miR-200a group revealed a significant reduction in the mRNA levels of E-cadherin, with a concomitant increasing in vimentin mRNA level (all P < 0.05). Western blot results showed higher E-cadherin and Caspase-3 protein expressions in anti-miR-200a group compared to miR-NC group (P < 0.05). In addition, vimentin and Ki-67 protein expression was found sharply decreased in anti-miR-200a group compared to miR-NC group (P < 0.05). Consistent with this, wound-healing and MTT assay showed that migration and proliferation capacity of MHCC-97L cells in anti-miR-200a group is significantly increased compared with miR-NC group (both P < 0.05). CONCLUSION: Our study reveals an important role of miR-200a in inhibiting EMT, proliferation and migration in HCC cells, suggesting the possibility of miR-200a-based therapeutics in HCC.
OBJECTIVE: Our study investigated the role of microRNA (miR)-200a and its molecular targets in hepatocellular carcinoma (HCC) cells. METHODS: An inhibitor of miR-200a was transiently transfected into the hepatocellular carcinoma cell line, MHCC-97L. The effect of this transfection on mRNA levels of epithelial-mesenchymal transition (EMT)-related genes was measured by fluorescence-based quantitative real-time polymerase chain reaction (qRT-PCR). Further, protein levels of EMT-related genes, cell proliferation and apoptosis-related markers were assessed by Western blot analysis in these transfected cells. MTT and wound-healing assay were used to evaluate the proliferation and migration of MHCC-97L cells in presence and in absence of miR-200a inhibitor. RESULTS: Compared with miR-NC control group, qRT-PCR results in anti-miR-200a group revealed a significant reduction in the mRNA levels of E-cadherin, with a concomitant increasing in vimentin mRNA level (all P < 0.05). Western blot results showed higher E-cadherin and Caspase-3 protein expressions in anti-miR-200a group compared to miR-NC group (P < 0.05). In addition, vimentin and Ki-67 protein expression was found sharply decreased in anti-miR-200a group compared to miR-NC group (P < 0.05). Consistent with this, wound-healing and MTT assay showed that migration and proliferation capacity of MHCC-97L cells in anti-miR-200a group is significantly increased compared with miR-NC group (both P < 0.05). CONCLUSION: Our study reveals an important role of miR-200a in inhibiting EMT, proliferation and migration in HCC cells, suggesting the possibility of miR-200a-based therapeutics in HCC.
Authors: Cameron P Bracken; Philip A Gregory; Natasha Kolesnikoff; Andrew G Bert; Jun Wang; M Frances Shannon; Gregory J Goodall Journal: Cancer Res Date: 2008-10-01 Impact factor: 12.701
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