耐阿霉素人骨肉瘤细胞株的建立及评价-第三军医大学学报.doc

人骨肉瘤耐阿霉素细胞株的建立 陈渝 邓忠良* （重庆医科大学附属第二医院骨科，重庆400010） [摘要] 目的 诱导并建立耐阿霉素(Adriamycin，ADM)的人骨肉瘤细胞株，观察耐药细胞系(143B/ADM)的生物学特性。方法 采用逐步递增ADM浓度间歇作用的方法诱导143B细胞株建立143B/ADM耐药细胞株。CCK法检测药物敏感性；冻存/复苏试验和撤药试验检测细胞稳定性；激光共聚焦显微镜观察细胞形态变化；贴壁率、生长曲线、倍增时间检测细胞增殖能力；流式细胞术检测细胞周期；双层软琼脂克隆形成试验检测细胞克隆形成能力。结果 经ADM诱导45 d建立了143B/ADM细胞株，其对ADM耐药指数为15.7，对顺铂、甲氨喋啶、异环磷酰胺、长春新碱和紫杉醇亦产生不同程度交叉耐药；冻存/复苏和撤药对143B/ADM耐药性无显著影响；激光共聚焦显微镜可见143B/ADM细胞和细胞核体积均增大；生长曲线显示143B/ADM增殖能力下降；流式细胞仪检测显示细胞周期混乱；双层软琼脂克隆形成试验显示143B/ADM克隆形成能力显著升高。结论 本研究建立了稳定的人骨肉瘤多药耐药细胞系(143B/ADM)，为进一步研究多药耐药机制和耐药治疗提供了一种新的实验模型。 [关键词]骨肉瘤；细胞；阿霉素；多药耐药 [中图分类号] R681，R738.1 [文献标识码] A Establishment of adriamycin resistance cell lines of human osteosarcoma Chen Yu，Deng Zhongliang（ Department of Orthopedics, the Second Affiliated Hospital, ChongqingMedical University, Chongqing 400010, China） [Abstract] Objective To establish a multidrug resistance cell line from human osteosarcoma 143B cell line and study characterization of the cell line. Methods The 基金项目：国家自然科学基金 Supported by the National Natural Science Foundation of China 作者简介：陈渝，男，博士，主治医师，从事骨肿瘤研究。E-mail:chenyu@ *通信作者：邓忠良，教授 E-mail: deng7586@ human osteosarcoma 143B cells were exposed to high and gradually increasing dose of adriamycin for 45 d to establish muhidrug resistance cell line 143B/ADM. The sensitivity of muhidrug resistance was measured by CCK assay. The cell stability was measured by cryopreservation/resuscitation test and withdrawal test. Morphology was observed by laser scanning confocal microscope. The ability of proliferation was measured by growth curve, adherence ratio and double time. Cell cycle was analyzed by flow cytometry. Clonality was detected by soft agar colony formation test. Results Resistant cell line(143B/ADM) was established after induction for 45 d. The resistance index of 143B/ADM cells to adriamycin was 15.7, and the cells showed various resistance to cisplatin, methotrexate, iso