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Archive > Volume 22 Issue 6 > 2015,22(6):716-723. DOI:10.3872/j.issn.1007-385X.2015.06.007 Prev Next
Basic Research
The influence of silencing Med19 on the therapeutic effect of paclitaxel in human breast cancer cells with wild-type or mutated p53
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Abstract:
Objective:To investigate the influence of downregulating Med19(mediator 19) expression on the therapeutic effect of paclitaxel in breast cancer cells with wild type or mutated p53 . Methods： Lentiviruses encoding small RNA that interferes Med19 expression were generated to infect cells of breast cancer cell lines MCF-7( p53 wild type) and MDA-MB-231 ( p53 mutation type). They were divided into Med19 knock-down groups (KD group, infected with pGcscil-Med19-siRNA-GFP), empty vector groups (NC group, infected with pGcscil-Med19-NC-GFP), and control groups (CON group, non-infected). GFP expression visualized under fluorescence microscope was used as an indicator of the efficiency of lentiviral infection. After silience of Med19 the levels of P53, phosphorylated P53 and P21 in cells of the different groups were determined by immunoblotting. The proliferation of MCF-7 and MDA-MB-231 cells under various experimental conditions was measured by the MTT assay. Flow cytometry was used to examine cell cycle and apoptosis. Results: Judged by fluorescent protein expression, more than 90% of MCF-7 cells and MDA-MB-231 cells infected with the lentiviruses in both KD and NC groups. The levels of Med19 expression in MCF-7 and MDA-MB-231 cells of the KD group were significantly lower than those in both CON and NC groups (P<0.05). The expressions of P53, phosphorylated P53, and P21 were significantly upregulated in MCF-7 cells of the KD group (P<0.05). In the 0.01-50 μg/ml range, paclitaxel induced a dose-dependent growth inhibition of both MCF-7 and MDA-MB-231 cells. For MCF-7 cells, the degree of inhibition and the number of apoptotic cells in KD group were significantly higher than those in the both NC and CON groups (P＜0.05). However, they were not significantly different between the three groups of MDA-MB-231 cells (P>0.05). While both MCF-7 and MDA-MB-231 cells underwent G2/M cell cycle block when treated with 10 μg/ml of paclitaxel, cycle of G0/G1 phase in KD group of MCF-7 cells more significantly increased than that in their NC group (P<005). Conclusion: The therapeutic effect of paclitaxel on MCF-7 cells( p53wild type) is enhanced by silencing Med19, which is likely mediated by the activation of p53 and expression of its downstream genes including p21 gene, leading to cell cycleb lock and apoptosis.
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Project Supported:
Projected supported by the National Natural Science Foundation of China (No. 81472485), the Project of “Six Major Talent Summit” of Jiangsu Province (No. WSW-014), and the Project of Hospital Management Center of Wuxi City (No. YGZXQ1311)
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