盐霉素增强吉非替尼诱导人肺腺癌细胞株A549凋亡的作用  

Salinomycin promotes the apoptosis-inducing effect of gefitinib on human lung adenocarcinoma cell line A549

作　　者：曾葭[1] 陈小宇[2] 祝爱珍[2] 刘成成[2] 谭广销[2] 刘革修[2] 

机构地区：[1]解放军第411医院呼吸内科,上海200081 [2]暨南大学医学院血液病研究所,广东广州510632

出　　处：《中国病理生理杂志》2012年第12期2147-2153,共7页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.30670902)

摘　　要：目的:探讨盐霉素联合吉非替尼诱导人肺腺癌细胞株A549凋亡的协同作用。方法:采用MTT的方法检测盐霉素对A549细胞生长的抑制作用;流式细胞术检测盐霉素对A549细胞凋亡和线粒体膜电位的影响;比色法检测caspase-3、-8和-9活性;Western bloting分析细胞色素C、Bcl-2、p-EGFR、p-Akt和p-ERK蛋白水平。结果:盐霉素与吉非替尼单用均出现不同程度的细胞增殖抑制作用和诱导细胞凋亡作用;而盐霉素与吉非替尼联合作用,能更显著地抑制细胞增殖,且凋亡细胞显著增加(P<0.05)。盐霉素单独作用A549细胞,线粒体膜电位显著下降,细胞内活性氧和Ca2+在短期内显著升高,胞浆细胞色素C含量以及caspase-3、-8和-9活性均显著增加,与对照组比较差异均有统计学意义;吉非替尼单用则主要表现为对p-EGFR、p-Akt和p-ERK蛋白表达的抑制作用,而对胞浆细胞色素C含量以及caspase-3、-8和-9活性影响较少。Western blotting检测发现,联合用药组的Bcl-2、p-EGFR、p-Akt和p-ERK蛋白表达量明显减少,但是对EGFR、Akt和ERK总蛋白水平无显著影响。结论:盐霉素与吉非替尼联用具有较好的协同作用,可能通过Bcl-2途径及线粒体凋亡途径诱导人肺腺癌A549细胞凋亡,提高A549细胞对吉非替尼的敏感性。AIM： To investigate the effect of salinomycin alone or in combination with gefitinib （ an inhibitor of epidermal growth factor receptor tyrosine kinase） on the growth and apoptosis of human non - small - cell lung cancer cell line A549. METHODS： The inhibitory effect of salinomycin on the growth of A549 cells was tested by MTT assay. The cell apoptosis and ttie level of mitochondrial membrane potential were determined by flow cytometry. The activity of caspase - 3, - 8, and - 9 was measured by the method of colorimetry. The protein levels of cytochrome C, Bcl - 2, p - EGFR, p - Akt and p - ERK were detected by Western blotting. RESULTS ： Salinomycin or gefitinib alone inhibited the growth of A549 ceils in a dose - dependent manner. Salinomycin or gefitinib also induced apoptosis of the ceils. Salinomycin com- bined with gefitinib produced stronger inhibitory effect on the cell proliferation, and a significant increase in cell apoptosis was also observed. Compared with control group, salinomycin alone significantly reduced mitochondrial membrane poten- tial, transitorily increased the levels of intracellular reactive oxygen species （ ROS）, cytoplasmic cytochrome C and Ca2＋ , and increased the activity of caspase - 3, - 8 and - 9 in A549 cells. Gefitinib alone inhibited the protein expression of p - EGFR, p - Akt and p - ERK, but no obvious effect on the release of cytochrome C and the activity of caspase - 3, - 8 and - 9 was found. The combination of salinomycin and gefitinib significantly reduced the protein levels of Bcl - 2, p - EGFR, p- Akt and p -ERK, but the protein levels of EGFR, Akt and ERK were not obviously changed. CONCLU- SION： The synergy of salinomycin and gefitinib is observed. Salinomycin inhibits the growth and induces apoptosis of hu- man lung carcinoma A549 cells through Bcl -2 pathway and mitochondrial apoptosis pathway. Salinomycin also increases the sensitivity of A549 cells to gefitinib.

关 键 词：盐霉素 吉非替尼 细胞凋亡 A549细胞 

分 类 号：R734.2[医药卫生—肿瘤]