ID3基因在逆转骨肉瘤顺铂耐药中的作用及其机制的研究

The role of ID3 gene in reversing cisplatin resistance of osteosarcoma and its mechanism

目的探讨ID3基因在逆转骨肉瘤顺铂耐药中的表达及对骨肉瘤细胞耐药、凋亡及RhoE、P-gp、Caspase-3表达的影响。方法 MTT法检测顺铂处理对U-2 OS及U-2 OS/DDP细胞生长功能的影响,q RT-PCR及Western blot法检测U-2 OS及U-2 OS/DDP细胞中ID3的表达;用ID3过表达载体转染U-2 OS/DDP细胞,以空脂质体转染的细胞作为对照组,各组细胞培养24 h后,用Western blot法检测细胞中RhoE、P-gp、Caspase-3蛋白表达变化;MTT检测耐药细胞耐药性的逆转作用;流式细胞仪检测细胞凋亡。结果顺铂耐药株U-2 OS/DDP细胞耐药能力明显高于U-2 OS细胞;ID3在顺铂耐药株U-2 OS/DDP细胞表达水平低于U-2 OS细胞;ID3基因过表达对骨肉瘤细胞耐药性具有逆转作用;ID3基因过表达可促进U-2 OS/DDP细胞凋亡;ID3过表达后,P-gp的表达下调,RhoE和Caspase-3的表达上调,差异有统计学意义(P﹤0.05)。结论 ID3基因在顺铂耐药株U-2 OS/DDP细胞中表达水平较低,过表达ID3基因能逆转U-2 OS/DDP细胞耐药情况,并通过调节RhoE、P-gp、Caspase-3蛋白表达促进细胞凋亡,其可能的机制与耐药蛋白的调控有关。

Objective To investigate the expression of ID3 gene in the reversal of cisplatin resistance in osteosarcoma and its effect on drug resistance, apoptosis and the expression of RhoE, P-gp and Caspase-3 in osteosarcoma cells.Method MTT assay was used to detect the growth of U-2 OS and U-2 OS/DDP cells, the expression of ID3 in U-2 OS and U-2 OS/DDP cells was detected by q RT-PCR and Western blot. Vectors with ID3 overexpression was applied to transfect U-2 OS/DDP cells, while blank liposome-transfected cells were utilized as control, all cells were cultured for 24 h,and then the expression of RhoE, P-gp and Caspase-3 protein in the cells were detected by Western blot; Reversal of drug resistance was determined by MTT; And apoptosis by flow cytometry. Result Drug resistance of resistant cells U-2 OS/DDP was significantly higher than that of U-2 OS cells, with lower expression of ID3 in the former than in the latter; the overexpression of ID3 gene could promote the apoptosis of U-2 OS/DDP cells; as ID3 overexpressed, the expression of P-gp was down-regulated while that of RhoE and Caspase-3 was up-regulated（P〈0.05）. Conclusion The expression of ID3 gene in cisplatin-resistant cells U-2 OS/DDP cells is relatively low, but overexpression of ID3 gene could significantly reverse the drug resistance of U-2 OS/DDP cells and promote the apoptosis by regulating RhoE, P-gp and Caspase-3 protein, which is possibly correlated with the regulation of drug resistance.