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目的:观察西妥昔单抗联合低温热疗诱导人鼻咽癌细胞株CNE凋亡的效果,并初步探讨其可能的机制。方法:试验分对照组、单靶组、单热组及热靶组,MTT法测定西妥昔单抗对CNE细胞株48h20～30%抑制的药物浓度;以该浓度药物与低温热疗(43℃,30分钟)联合,Hoechst33258荧光染色法观察细胞凋亡形态学变化;流式细胞术检测24h、48h凋亡率;western blot检测Bax、Bcl-2、EGFR蛋白的表达。结果:以48h20～30%抑制率的药物浓度(IC20～30)为试验的工作浓度,确定西妥昔单抗对CNE细胞株的工作浓度为10ug/ml;荧光染色法观察到热靶组CNE细胞发生典型的凋亡形态学改变;流式凋亡检测显示24、48h热靶组凋亡率显著高于相应单靶组、单热组及对照组(P<0.05)。western blot检测显示各处理组较之对照组都有上调Bax、下调Bcl-2、EGFR蛋白表达的作用,以热靶组最为显著(P<0.05);热疗后EGFR蛋白表达逐渐下调。结论:西妥昔单抗联合低温热疗能显著增加CNE细胞的凋亡率,这可能与EGFR的下调与受抑,进而上调Bax、下调Bcl-2蛋白的表达促进凋亡有关。
OBJECTIVE: To observe the effect of cetuximab combined with hypothermia hyperthermia on the apoptosis of human nasopharyngeal carcinoma cell line CNE and to explore its possible mechanism. Methods: The control group, single-target group, single-heat group and hot target group were used to test the concentration of cetuximab in 48 hours from 20% to 30% inhibition of MTT assay. ℃, 30 min). Hoechst33258 fluorescence staining was used to observe the morphological changes of apoptosis. Flow cytometry was used to detect the apoptotic rate at 24h and 48h. Western blot was used to detect the expression of Bax, Bcl-2 and EGFR. Results: The concentration of cetuximab in CNE cell line was determined to be 10 ug / ml at the concentration of 20 to 30% inhibition of IC50 (IC20 ~ 30). The CNE of hot target group was observed by fluorescence staining Apoptotic morphological changes were observed in typical cells. Flow cytometry showed that the apoptotic rate of hot target group was significantly higher than that of the corresponding single target group, single heat group and control group (P <0.05) at 24,48 hours. The results of western blot showed that the expression of Bcl-2 and EGFR in all treatment groups was significantly higher than that in control group (P <0.05). The expression of EGFR protein was down-regulated after hyperthermia treatment. CONCLUSION: Cetuximab combined with hypothermia can significantly increase the apoptosis rate of CNE cells, which may be related to down-regulation and down-regulation of EGFR, up-regulation of Bax and down-regulation of Bcl-2 protein expression.