CAJ | 학술논문

目的体外构建胶质瘤耐甲磺酸伊玛替尼多药耐药细胞株，并研究其生物学特性。方法采用逐渐增加培养基中伊玛替尼药物浓度的方法诱导构建耐甲磺酸伊玛替尼的人脑胶质瘤U251细胞株(命名为U251 AR); CCK8法检测U251AR、U251对多种化疗药物的IC50和耐药指数；QRT-PCR法检测耐药相关基因ABCC1、A BCB1、A BCB4、A BCG2 mRNA的表达，流式细胞术检测ABCG2蛋白的表达。结果体外培养12个月后建立了稳定耐药的细胞株U251AR,与亲代细胞相比异形性不显著。U251和U251AR对化疗药物的IC50相比较差异均有统计学意义（P＜0.05），U251AR对甲磺酸伊玛替尼、阿霉素、顺铂的耐药指数分别为20.41、5.06、10.28，表现出多药耐药特征。QRT-PCR检测结果显示耐药细胞株U251AR中ABCC1、ABCB1、ABCB4、ABCG2 mRNA的表达明显高于亲代U251细胞，流式细胞术检测结果显示U251AR中ABCG2蛋白的表达强于亲代U251细胞，差异均有统计学意义(P＜0.05)。结论成功建立多药耐药的胶质瘤细胞株U251AR，其多药耐药与A BCC1、A BCB1、A BCB4、A BCG2 mRNA及ABCG2蛋白的表达上调相关。
목적 체외구건효질류내갑광산이마체니다약내약세포주，병연구기생물학특성。방법채용축점증가배양기중이마체니약물농도적방법유도구건내갑광산이마체니적인뇌효질류U251세포주(명명위U251 AR); CCK8법검측U251AR、U251대다충화료약물적IC50화내약지수；QRT-PCR법검측내약상관기인ABCC1、A BCB1、A BCB4、A BCG2 mRNA적표체，류식세포술검측ABCG2단백적표체。 결과 체외배양12개월후건립료은정내약적세포주U251AR,여친대세포상비이형성불현저。U251화U251AR대화료약물적IC50상비교차이균유통계학의의（P＜0.05），U251AR대갑광산이마체니、아매소、순박적내약지수분별위20.41、5.06、10.28，표현출다약내약특정。QRT-PCR검측결과현시내약세포주U251AR중ABCC1、ABCB1、ABCB4、ABCG2 mRNA적표체명현고우친대U251세포，류식세포술검측결과현시U251AR중ABCG2단백적표체강우친대U251세포，차이균유통계학의의(P＜0.05)。 결론 성공건립다약내약적효질류세포주U251AR，기다약내약여A BCC1、A BCB1、A BCB4、A BCG2 mRNA급ABCG2단백적표체상조상관。
Objective To establish the imatinib (STI-571)-resistant subline in vitro and investigate its biological characteristics. Methods Human glioblastoma multiform drug-resistant cell line (named U251AR) was established in vitro by successively increasing the concentration of imatinib in a cell culture medium. The 50％ inhibitory dose (IC50) values and the resistance indexes ([IC50U251/STI-571]/[IC50 U251]) for other chemotherapeutic agents were evaluated using cell counting kit-8 assays. Expressions of acquired multidrug resistance P-glycoprotein (MDR 1, ABCB 1; MDR3, ABCB4),breast cancer resistance protein (BCRP, ABCG2) and multidrug resistance-associated protein 1 (MRP1,ABCC1) were detected by QRT-PCR. Flow cytometry was employed to detect the protein expression of ABCG2. Results The U251AR was developed after culture for 12 months and similar morphologies of U251 and U251/STI-571 cells were determined. The resistance coefficient of U251AR cells to imatinib was 20.41 times more than that of the parent cells, and U251AR cells showed cross-resistance to many anti-tumor agents (P＜0.05). The resistance coefficients of U251AR cell line to doxorubicin and cisplatin were 5.06 and 10.28 times, respectively, more than those of U251 cells (P＜0.05). QRT-PCR indicated that the mRNA levels of MDR1, MRP1, BCRPandABCB4 (P-g4) in the U251/STI571 resistant cells were significantly higher than those in the U251 cells (P＜0.05). The protein expression of ABCG2 in U251AR cell line was significantly increased as compared with that in the parent cells (P＜0.05).Conclusion We have successfully established multidrug resistant cell line U251AR, and the drug resistance of U251/STI571 is associated with over-expressions of ABCC1, ABCB1, ABCB4, and ABCG2 mRNA, and ABCG2 protein.