目的：探讨下调中介体（mediator，Med）19表达是否增加紫杉醇(pacliatxel, PTX)对p53野生型和突变型乳腺癌细胞化疗的疗效。方法：构建Med19RNA干扰（RNAi）慢病毒，分别感染p53野生型的乳腺癌细胞MCF-7和p53突变型的乳腺癌细胞MDA-MB-231，两种细胞均分为Med19基因敲减组（KD组：细胞感染pGcscil-Med19-siRNA-GFP ）、空载体感染组（NC 组：细胞感染pGcscil-Med19-NC-GFP）、对照组（CON组：未感染慢病毒的乳腺癌细胞），荧光显微镜观察慢病毒感染效率。Western blotting检测Med19基因沉默后各组细胞内P53、磷酸化P53（pP53）、P21蛋白表达水平的变化。紫杉醇分别作用于Med19基因沉默前后的MCF-7和MDA-MB-231细胞，MTT法检测细胞增殖抑制率的变化，流式细胞术检测细胞周期和凋亡率的变化。结果：KD组和NC组细胞的荧光感染效率均大于90%，表明获得了满意的感染效率。与CON组和NC组相比，两种细胞KD组的Med19蛋白表达水平均下降，差异有统计学意义（均P<0.05）。MCF-7细胞KD组P53、pP53、P21蛋白表达上调，差异有统计意义（均P<0.05）。在0.01～50 μg/ml 范围内，紫杉醇抑制两种乳腺癌细胞的生长增殖并呈浓度依赖性（P<0.05），对KD组的抑制率和细胞凋亡率均高于NC组及CON组，差异有统计学意义（均P<0.05）；但MDA-MB-231细胞各组间抑制率差异无统计学意义（均P>0.05）。10 μg/ml紫杉醇导致两种细胞的KD组和NC组均出现G2/M期阻滞，并且MCF-7细胞KD组G0/G1期细胞较NC组显著增加（P<0.05）。结论：Med19基因沉默增强紫杉醇对p53野生型乳腺癌MCF-7细胞化疗的疗效，其机制可能为增强了p53基因的表达活化，通过其下游基因p21 调节细胞周期、促进细胞凋亡。

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 G2/M cell cycle block when treated with 10 μg/ml of paclitaxel, cycle of G0/G1 phase in KD group of MCF-7 cells more significantly increased than that in their NC group (P<005). Conclusion: The therapeutic effect of paclitaxel on MCF-7 cells( p53wild 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.

国家自然科学基金面上项目资助（No. 81472485）；江苏省“六大人才高峰”项目资助（No. WSW-014）；无锡市医院管理中心项目资助（No. YGZXQ1311）