顺铂诱导人成骨肉瘤耐药细胞系的建立及生物学特性研究

目的：建立顺铂诱导的人成骨肉瘤多药耐药细胞系MG63／R并观察其生物学特性。方法：以顺铂为诱导剂,采用大剂量冲击与逐步增加剂量相结合的方法诱导人成骨肉瘤MG63细胞,建立多药耐药系MG63／R。MTF法检测药物敏感性：光镜、透射电镜观察MG63、MG63／R细胞形态及超微结构变化;生长曲线、克隆形成试验检测细胞增殖能力;流式细胞术检测细胞周期、凋亡指数、P—pg、bcl-2、p53蛋白表达。结果：经顺铂186d的诱导建立了MG63／R,MG63／R对顺铂的耐药指数为83．557,对阿霉素、长春新碱、氨甲蝶呤、环磷酰氨亦产生不同程度的交叉耐药;光镜观察可见MG63／R细胞排列不规则,形态呈三角形、多角形及多核现象;透射电镜显示MG63／R细胞表面突起增加,粗面内质网丰富;细胞周期分析显示细胞增殖能力明显增加而细胞凋亡明显降低;MG63／R细胞P—pg、bcl-2阳性表达较MG63细胞明显增加,p53表达则明显降低。结论：本研究建立了稳定的人成骨肉瘤多药耐药细胞系（MG63／R）,为进一步研究顺铂的多药耐药机制和耐药治疗提供了一种新的实验模犁。

Establishment And Characterization of A Cisplafin-multidrug Resistance Human Osteosarcoma Cell Line

Objective： To establish a muhidrug resistance ceil line from human osteosarcoma MG63 cell line and study characterization of the cell lines. Methods： The human osteosarcoma MG63 cells were exposed to high and gradually increasing dose of cisplatin for 186 days to establish muhidrug resistance cell line MG63/R. The sensitivity of muhidrug resistance was measured by M3T assay. Morphology and uhramicrostructure were observed by optical microscopy and transmission electron microscopy. The ability of proliferation was measured by growth curve and colony-forming assay. Cell cycle and apoptosis were analyzed by flow cytometry. P-pg, bcl-2, p53 expression of MG63 and MG63/R were analyzed by flow cytometry. Results： The resistance index of MG63/R cells to cisplatin is 83.557, and the cells showed various resistance to doxorubicin, vincristin, methotrexate and cyclophosphamide. The microscopy observation showed that arrangement of MG63/R was disordered, and its shape presented triangle, polygon and polynucleation, the granular endoplasmic reticulums and apophyses increased. The ability of proliferation of MG63/R clearly increased and apoptosis index decrease. The expression of P-pg and bcl-2 in MG63/R increased and p53 decreased. Conclusion： A muhidrug resistance cell line （MG63/R） from human osteosarcoma is established. It will provide a new experimental model for further study of mechanism of muhidrugresistance and therapy.