耐甲氨蝶呤人骨肉瘤细胞株建立及其生物学特性  

Establishment of a methotrexate-resistant cell line of human osteosarcoma and its biological characteristics

作　　者：王建军[1] 赵晖[1] 林峰[1] 姚阳[1] 

机构地区：[1]上海交通大学附属第六人民医院肿瘤内科,上海200233

出　　处：《肿瘤》2007年第8期623-627,共5页Tumor

摘　　要：目的:诱导并建立耐甲氨蝶呤(methotrexate,MTX)的人骨肉瘤细胞株并观察耐药细胞系(Saos-2/MTX1、Saos-2/MTX2)的生物学特性。方法:以MTX为诱导剂,采用逐步增加药物剂量冲击诱导方法,诱导人成骨肉瘤原代Saos-2细胞株,建立不同药物浓度耐药系(Saos-2/MTX1、Saos-2/MTX2);MTT法检测原代与耐药细胞株对MTX、顺铂(cisplatin,DDP)、阿霉素(doxorubicin,ADM)、异环磷酰胺(ifosfamide,IFO)、表柔比星(epirubicine,EPI)、比柔比星(pirarubicin,THP)、紫杉醇(paclitaxel,PTX)药物敏感性;利用光学显微镜、透射电镜观察细胞形态及超微结构变化;利用细胞生长曲线检测细胞增殖能力;利用流式细胞仪检测细胞周期比例。结果:经178 d的诱导,建立Saos-2/MTX1,Saos-2/MTX2耐药细胞株,其对MTX的耐药指数(resisitant drug index,RI)分别为原代细胞株的4.87和12.73倍;耐药细胞株对ADM、EPI、THP、PTX产生不同程度的交叉耐药,对DDP敏感;光镜观察Saos-2/MTX1、Saos-2/MTX2细胞体积增大,多核现象较原代Saos-2细胞株明显增加;透射电镜显示Saos-2/MTX1、Saos-2/MTX2细胞株表面突起较原代Saos-2细胞株减少、且核仁增大增多;细胞生长曲线和流式细胞仪检测显示耐药细胞株增殖能力下降,S期细胞所占比例减少(P<0.05)。结论:本研究建立了耐MTX人成骨肉瘤细胞株Saos-2/MTX1、Saos-2/MTX2,并观察了耐药株细胞的生物学特性,为进一步研究甲氨蝶呤的耐药机制提供了一种实验模型。Objective： To establish a methotrexate （MTX）-resistant human osteosarcoma cell line （Saos-2/MTX1 and Saos-2/MTX2） derived from the Saos-2 cell line and observe its biological characters, nethods：Saos-2 cells were pulse-exposed to MTX with gradually increased dosage. Both MTX-resistant human osteosarcoma cell lines （Saos-2/MTX1 and Saos-2/MTX2 ） were es- tablished after 178 days. The sensitivity of the Saos-2, Saos-2/MTX1, and Saos-2/MTX2 cell lines to MTX, cisplatin （DDP） , doxoru- bicin （ADM） , ifosfamide （IFO） , epirubicine （EPI） , pirarubicin （PTH） , and paclitaxel （PTX） was measured by MTF assay. The morphological and ultra-structural changes of the cells were observed by optical microscopy and transmission electronic microscopy. The cell proliferation ability was calculated from the growth curve. The cell cycle distribution was analyzed by flow cytometry. Results：The MTX-resistant human osteosarcoma cell lines, Saos-2/MTXI and Saos-2/MTX2, were established after 178 d induction. The resistance indexes to MTX were 4.87 and 12.73 times than that of Saos-2, respectively. The two cell lines had cross resistance to ADM, EPI, PTH and TAXOL and were still sensitive to DDP （ P 〉 0. 05 ）. Optical microscopy showed the volume of Saoso2/MTX1 and Saos-2/MTX2 cells were enlarged and the number of poly-nucleus cells was increased. Transmission electronic microscopy revealed the cell process was decreased and the nucleoli were increased. Cell proliferation curve and flow cytometry demonstrated that the proliferation abilities of Saos-2/MTX l and Saos-2/MTX 2 cells were decreased significantly and the proportion of cells in S phase was reduced （P 〈 0.05）. Conclusion：We establish two MTX-resistant human osteosarcoma cell lines （Saos-2/MTX1 and Saos-2/MTX2） and observe their biological characteristics. It provides a new experimental model for further studying the mechanism of MTX-resistance.

关 键 词：骨肉瘤 抗药性 肿瘤 甲氨蝶呤 SAOS-2细胞 

分 类 号：R738.1[医药卫生—肿瘤]