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目的研究埃克替尼与塞来昔布联合应用对人肺腺癌A549细胞凋亡的影响以及对EGFR、COX-2表达的影响。方法埃克替尼、塞来昔布单独或联合干预细胞48h后,倒置相差显微镜观察药物干预后细胞形态学变化;四甲基偶氮唑盐(MTT)比色法测定细胞抑制率;HOECHEST33258荧光染色法和流式细胞术检测细胞凋亡和药物作用周期;免疫荧光检测EGFR和COX-2蛋白表达情况。结果埃克替尼联合塞来昔布,相比单药组明显出现大量颗粒和空泡,细胞变圆开始脱落;埃克替尼与塞来昔布对A549细胞增殖有明显抑制作用,并呈浓度及时间依赖性,二者联合作用时抑制作用更强(P<0.05);埃克替尼与塞来昔布均能诱导细胞凋亡,联合作用后细胞凋亡率更高(P<0.05);联合用药与单独用药相比,可使细胞发生明显的G1期阻滞(P<0.05),进一步下调了EGFR和COX-2蛋白的表达(P<0.05)。结论埃克替尼与塞来昔布联合应用具有协同作用,机制可能与介导细胞凋亡,阻滞细胞周期于G0/G1期及阻滞EGFR和COX-2信号途径有关。
Objective To investigate the effect of icotinib combined with celecoxib on the apoptosis of human lung adenocarcinoma A549 cells and its effect on the expression of EGFR and COX-2. Methods The cells were treated with imatinib and celecoxib for 48 h either alone or in combination. The morphological changes of the cells were observed by inverted phase contrast microscope. The inhibition rates of cells were determined by MTT colorimetric assay. The fluorescence of HOECHEST33258 Staining and flow cytometry were used to detect apoptosis and drug action cycle. The expression of EGFR and COX-2 protein were detected by immunofluorescence. Results Icitinib combined with celecoxib obviously showed a large number of granules and vacuoles compared with the single drug group. The cells turned round and began to fall off. Icitinib and celecoxib significantly inhibited the proliferation of A549 cells (P <0.05). Both icotinib and celecoxib could induce apoptosis, and the apoptotic rate was higher after combination (P <0.05) (P <0.05), and further down-regulated the expression of EGFR and COX-2 protein (P <0.05). Conclusions The combination of icotinib and celecoxib has a synergistic effect. The mechanism may be related to the induction of apoptosis, the arrest of cell cycle in G0 / G1 phase, and the block of EGFR and COX-2 signaling pathways.