Down-regulation of extracellular signal-regulated kinase 1/2 activity in P-glycoprotein-mediated multidrug resistant hepatocellular carcinoma cells |
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Authors: | Feng Yan Xiao-Min Wang Chao Pan Quan-Ming Ma |
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Institution: | 1. Department of Hepatobiliary Surgery, Postdoctoral Station, Zhongshan Hospital, Xiamen University, Xiamen 361004, Fujian Province, China 2. Department of Hepatobiliary Surgery, Digestive Diseases Institute, Xiamen University, Xiamen361004, Fujian Province, China 3. Department of Pathology, Zhongshan Hospital,Xiamen University, Xiamen 361004, Fujian Province, China 4. Department of Hepatobiliary Surgery,Zhongshan Hospital, Xiamen University, Xiamen 361004,Fujian Province, China |
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Abstract: | AIM:To study the expression and phosphorylation of extracellular signal-regulated kinase (ERK) 1 and ERK2 in multidrug resistant (MDR) hepatocellular carcinoma (HCC) cells. METHODS:MDR HCC cell lines, HepG2/adriamycin (ADM) and SMMC7721/ADM, were developed by expo ing parental cells to stepwise increasing concentrations of ADM. MTT assay was used to determine drug sensitivity. Flow cytometry was employed to analyze cell cycle distribution and measure cell P-glycoprotein (P-gp) and multidrug resistant protein 1 (MRP1) expression levels. ERK1 and ERK2 mRNA expression levels were measured by quantitative real-time PCR (QRTPCR). Expression and phosphorylation of ERK1 and ERK2 were analyzed by Western blot. RESULTS:MTT assay showed that HepG2/ADM and SMMC7721/ADM were resistant not only to ADM, but also to multiple anticancer drugs. The P-gp expression was over 10-fold higher in HepG2/ADM cells than P < 0.001) and over 4-fold higher in SMMC7721/ADM was not significantly higher in HepG2/ADM and SMMC7721/ADM cells than in parental cells. In addition, the percentage of MDR HepG2/ADM and SMMC7721/ADM cells was significantly decreased in the G0/G1 phase and increased in the the S phase or G2/M phase. QRT-PCR analysis demonstrated that the ERK1 and ERK2 mRNA expression increased apparently in HepG2/ADM cells and decreased significantly in SMMC7721/ADM cells. Compared with the expression of parental cells, ERK1 and ERK2 protein expressions were markedly decreased in SMMC7721/ADM cells. However, ERK2 protein expression was markedly increased while ERK1 protein expression had no significant change in HepG2/ADM cells. Phosphorylation of ERK1 and ERK2 was markedly decreased in both HepG2/ADM and SMMC7721/ADM MDR cells. CONCLUSION:ERK1 and ERK2 activities are downregulated in P-gp-mediated MDR HCC cells. ERK1 or ERK2 might be a potential drug target for circumventing MDR HCC cells. |
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Keywords: | Multidrug resistance Extracellular signalregulatedMAP kinases Hepatocellular carcinoma P-glycoprotein Multidrug resistance-associated protein |
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