汉防己甲素联合紫杉醇逆转脑胶质瘤C6/MDR细胞耐药及其机制研究  

Combination of tetrandrine and paclitaxel overcomes multidrug resistance on C6/MDR glioma cells

作　　者：王宏[1] 郭晓远[1] 吴国建[1] 陈陆馗[1] WANG Hong;GUO Xiao-yuan;WU Guo-jian;CHEN Lu-kui(Zhongda Hospital, Southeast University, Nanjing 210009, China)

机构地区：[1]东南大学附属中大医院,江苏南京210009

出　　处：《中草药》2018年第11期2584-2590,共7页Chinese Traditional and Herbal Drugs

基　　金：南京市科技发展计划项目(2017sc512018)

摘　　要：目的评价汉防己甲素联合紫杉醇(paclitaxel,PTX)逆转耐药型脑胶质瘤(C6/MDR)细胞多药耐药的可行性,并探讨可能的耐药逆转机制。方法通过非耐药型脑胶质瘤C6和耐药型脑胶质瘤C6/MDR细胞系作对比,以体外细胞毒性为指标,采用MTT法分别考察PTX、汉防己甲素、PTX+汉防己甲素抑制C6和C6/MDR细胞增殖作用差异;以细胞摄取能力为指标,借助HPLC和流式细胞仪技术分别测定PTX和荧光探针罗丹明123(R123)在2种细胞内的含量,评价汉防己甲素促C6/MDR细胞摄取PTX和R123的能力;以细胞凋亡率为指标,采用Annexin V-PE/7-AAD一步染色法定量PTX、汉防己甲素、PTX+汉防己甲素干预后的2种细胞凋亡率;采用P-糖蛋白(P-gp)抗体结合试剂盒和Pgp-Glo TM分析系统分别考察汉防己甲素对C6/MDR细胞P-gp表达以及P-gp ATP酶活性的影响。结果汉防己甲素+PTX对C6/MDR细胞的半数抑制浓度(IC50,以PTX浓度计)为(5.88±0.43)nmol/L;在10μmol/L汉防己甲素的干预下,C6/MDR细胞内的PTX和R123累积量相比PTX组分别提高9.4倍和12.3倍,凋亡率相应提高了2.3倍;这种汉防己甲素介导的耐药逆转机制可能与降低C6/MDR细胞P-gp表达、刺激细胞内P-gp ATP酶活性有关。结论汉防己甲素联合紫杉醇在体外具有逆转C6/MDR细胞耐药的潜力。Objective To investigate the feasibility of tetrandrine combined with paclitaxel（PTX） in multidrug resistance reversal on C6/MDR glioma cells, and explore the potential molecular mechanisms. Methods Through the comparison of non-resistant glioma C6 cells and drug-resistant glioma C6/MDR cells, the cytotoxicity of against C6/MDR（or C6） cells were assayed by MTT method. The intracellular accumulation of PTX and Rhodamine 123（R123） were determined by high performance liquid chromatography（HPLC） and flow cytometry, respectively. The cell apoptosis induction of C6/MDR（or C6） cells was detected by Annexin V-PE/7-AAD staining method after various intervention of PTX, tetrandrine, and PTX ＋ Hf A. P-glycoprotein（P-gp） expression and P-gp ATPase activity were evaluated through P-gp antibody binding assay kit and Pgp-GloTM assay systems, respectively. Results The half maximal inhibitory concentration（IC50） of tetrandrine ＋ PTX against C6/MDR cells were（5.88 ± 0.43） nmol/L. The C6/MDR intracellular accumulation of PTX and R123 were increased by 9.4-fold and 12.3-fold in the presence of 10 μmol/L tetrandrine. Accordingly, the apoptosis rate of C6/MDR cells treated with tetrandrine ＋ PTX was 2.3-fold higher than PTX treatment. The tetrandrine-mediated multidrug resistance reversal was involved with the downregulation of P-gp expression and the stimulation of P-gp ATPase activity. Conclusion The combination of tetrandrine and PTX had a potential of overcoming multidrug resistance on C6/MDR glioma cells in vitro.

关 键 词：汉防己甲素 紫杉醇 脑胶质瘤 C6/MDR细胞 逆转多药耐药 P-糖蛋白 

分 类 号：R285.5[医药卫生—中药学]