摘要
目的 :利用氯喹(chloroquine,CQ)及顺铂(cisplatin,DDP)作用于CAL-27细胞,探讨自噬在口腔癌化疗中的作用及机制,为临床增强口腔癌化疗敏感性提供理论依据。方法:应用四甲基偶氮唑蓝(MTT)比色法比较药物对CAL-27细胞的生长抑制作用,采用激光共聚焦显微镜观察自噬特异性蛋白LC3-Ⅱ的表达,Annexin V/PI双染色流式细胞仪检测细胞凋亡,流式细胞术观察细胞周期的分布。采用SPSS17.0软件包对数据进行统计学分析。结果:不同浓度氯喹及顺铂处理CAL-27细胞不同时间后,细胞生存率逐渐降低,呈浓度和时间依赖性。5 mg/L氯喹联合顺铂在IC50浓度(5 mg/L)处理后,与单独顺铂处理相比,显著降低了CAL-27细胞的生存率(P<0.05)。氯喹或顺铂作用于CAL-27细胞48 h后,自噬在细胞中分布清晰,顺铂组(DDP组)细胞平均荧光强度高于对照组、氯喹处理组(CQ组)及氯喹与顺铂联合处理组(CQ+DDP组)(P<0.05);氯喹组细胞平均荧光强度显著低于其他3组(P<0.05)。氯喹或顺铂作用于CAL-27细胞48 h后,与对照组相比,DDP组和CQ+DDP组细胞凋亡率显著提高(P<0.05);与顺铂单独作用相比,CQ+DDP组细胞凋亡率显著提高(P<0.05)。氯喹和顺铂作用于CAL-27细胞48 h后,DDP组和CQ+DDP组G1期细胞明显增多,出现G1期阻滞,CQ+DDP组G1期细胞数显著高于DDP组(P<0.05)。结论:抑制自噬能够提高CAL-27细胞对顺铂的化疗敏感性,CAL-27细胞本身的自噬是产生化疗耐药的重要机制。自噬抑制剂有望成为口腔癌化疗的增敏剂。
PURPOSE： To study the role and mechanism of autophagy in chemotherapy of oral squamous cell carcinoma,and provide theoretical evidence to improve chemotherapeutic efficacy of oral squamous cell carcinoma patients.METHODS： The cell survival rate changes induced by cisplatin（DDP） and chloroquine（CQ） in CAL-27 cells were assayed by methyl thiazolyl tetrazolium method（MTT）. The LC3- Ⅱ expression level was detected by laser scanning confocal microscope; The apoptotic rate was determined by flow cytometry. SPSS17.0 software package was used for statistical analysis. RESULTS： MTT results showed that compared with the control group, the cell survival rate reduced with the increasing time of DDP and CQ treatment; The optimal concentration of CAL-27 cells was 5 mg/L after treatment with CQ. IC50 of the CAL-27 cells was 5 mg/L after treatment with DDP; MTT results showed that the cell survival rate of CQ＋DDP group was significantly lower than control group, CQ group and DDP group（P0.05）. With the action of CQ and DDP to CAL-27 cells for 48 hours, immunofluorescence results showed that the average fluorescence intensity of DDP group was significantly higher than the other 3 groups（P 0.05）, while it was significantly lower in CQ group than the other 3 groups（P0.05）. With the action of CQ and DDP to CAL-27 cells for 48 hours, flow cytometry results showed that the cell apoptosis rate of DDP group and CQ＋DDP group were significantly higher than control group and CQ group. The cell apoptosis rate of CQ＋DDP group was significantly higher than DDP group（P0.05）. With the action of CQ and DDP to CAL-27 cells for 48 hours, cells in G1 phase of DDP group and CQ ＋DDP group increased, indicating G1 phase blockage. The cell count in G1 phase of CQ ＋DDP group was significantly higher than DDP group（P 0.05）.CONCLUSIONS： Inhibition of autophagy can enhance the chemotherapeutic sensitivity of DDP in CAL-27 cells. Autophagy in CAL-27 cells is an important mechanism for chemotherapy resistanc
出处
《上海口腔医学》
CAS
CSCD
2015年第1期30-36,共7页
Shanghai Journal of Stomatology
基金
吉林省科技厅基金(201105101)
吉林省卫生厅基金(20132004)~~
关键词
细胞自噬
顺铂
氯喹
细胞凋亡
口腔鳞状细胞癌
Cell autophagy
Cisplatin
Chloroquine
Cell apoptosis
Oral squamous cell carcinoma
