论文部分内容阅读
[目的]研究西妥昔单抗(Cetuximab,C225)联合顺铂(Cisplatin,DDP)对人鼻咽癌(nasopharyngeal carcinoma,NPC)细胞增殖和凋亡的影响及其分子机制。[方法 ]采用人鼻咽癌HNE1和CNE2细胞株,RT-q PCR检测表皮生长因子受体(epidermal growth factor receptor,EGFR)m RNA的差异性表达。实验分为对照组、西妥昔单抗组、顺铂组、两药联合组,采用CCK-8法、平板克隆形成实验检测细胞增殖能力,并评价药物的协同效应。流式细胞术检测各组细胞的凋亡情况。Western Blot检测AKT、p-AKT以及凋亡相关蛋白Bax、caspase-3、cleaved caspase-3的表达水平。[结果 ]药物持续作用48h后,两株细胞中两药联合组与单药组相比,均不同程度地提高对细胞的抑制率。在HNE1和CNE2细胞株中,两药协同指数分别为0.6和0.9。HNE1和CNE2细胞中EGFR m RNA分别呈现相对高表达和相对低表达状态。流式细胞术结果显示:HNE1细胞中,联合组的凋亡细胞比率显著性高于单药组,另外,两药联合作用更有效地下调了HNE1中凋亡蛋白Bax、caspase-3、cleaved caspase-3的表达,同时下调了EGFR/AKT信号通路关键蛋白AKT磷酸化表达。[结论 ]西妥昔单抗可能通过抑制顺铂介导的EGFR/AKT通路的磷酸化活化,产生协同抑制HNE1细胞增殖效应。两药联合在HNE1和CNE2细胞中的差异性效应为鼻咽癌的临床个体化治疗研究提供依据。
[Objective] To investigate the effects of Cetuximab (C225) combined with cisplatin (DDP) on the proliferation and apoptosis of human nasopharyngeal carcinoma (NPC) cells and its molecular mechanism. [Methods] Human nasopharyngeal carcinoma HNE1 and CNE2 cell lines were used to detect the differential expression of epidermal growth factor receptor (EGFR) m RNA by RT-q PCR. The experiment was divided into control group, cetuximab group, cisplatin group and two drugs combined group. The cell proliferation ability was detected by CCK-8 method and plate clone formation assay, and the synergistic effect of drugs was evaluated. Flow cytometry was used to detect the apoptosis of each group of cells. Western Blot was used to detect the expression of AKT, p-AKT and apoptosis-related proteins Bax, caspase-3 and cleaved caspase-3. [Result] Compared with single drug group, the inhibitory rate of the cells in the two cell lines increased to different degrees after 48h of continuous drug treatment. In HNE1 and CNE2 cell lines, the synergistic index of the two drugs were 0.6 and 0.9, respectively. EGFR m RNA in HNE1 and CNE2 cells showed relatively high expression and relatively low expression respectively. The results of flow cytometry showed that the ratio of apoptotic cells in HNE1 cells was significantly higher than that in single drug group. In addition, the combination of the two drugs reduced the apoptosis protein Bax, caspase-3, cleaved caspase -3, and down-regulated phosphorylation of AKT, a key protein of EGFR / AKT signaling pathway. [Conclusion] Cetuximab may inhibit the proliferation of HNE1 cells by inhibiting the phosphorylation of EGFR / AKT pathway induced by cisplatin. The difference between the two drugs combined in HNE1 and CNE2 cells provide the basis for clinical individualized treatment of nasopharyngeal carcinoma.