大蒜素对骨肉瘤获得性耐药细胞株的作用及其机制

摘要

Objective To observe the effects of diallyl trisulfide ( DATS) on the osteosarcoma acquired drug-resist-ance cell line , and to investigate its possible mechanism .Methods We established an osteosarcoma cell line Saos-2/DOX which acquired drug-resistance to doxorubicin ( DOX) in vitro by the pulse selection .MTT assay was applied to de-termine the half maximal inhibitory concentration (IC50) of Doxorubicin (DOX) on the Saos-2 and Saos-2/DOX cells, be-sides the resistance index (RI) of Saos-2/DOX was calculated.Moreover, the DATS concentration treating Saos-2/DOX without cytotoxicity was selected by MTT assay .The cell viability was detected by MTT assay at 24, 48, and 72 h after the treatment of DOX alone or in combination with DATS on the Saos-2/DOX cells.At 48 h after the treatment of DATS (0, 10, 50, and 100 μmol/L) on the Saos-2/DOX cells, the protein expression levels of pERK 1/2, ERK1/2, pAKT1, and AKT1 were detected by Western blotting .Results The DOX-resistant osteosarcoma cell line Saos-2/DOX was successfully established and the RI of Saos-2/DOX was 6.94.The DATS concentration treating Saos-2/DOX cells without cytotoxicity was 10 μmol/L.The survival rate of Saos-2/DOX-resistant cells treated with DOX 100 μg/L and DATS 10 μmol/L for 24 h was significantly lower than that of DOXS 10μmol/L or DOX 100μg/L, and decreased over the treatment time ( all P<0.05).At 48 h after treatment of different concentrations of DATS , the protein expression level of pERK1/2 was signifi-cantly up-regulated, which was concentration dependent (P<0.05).However, no significant difference was found in the protein expression levels of ERK1/2, pAKT1, and AKT1 (all P>0.05).Conclusion DATS could reverse the acquired drug-resistance of Saos-2/DOX cells by activating the ERK 1/2 signal pathway .%目的 探讨大蒜素(DATS)对骨肉瘤获得性耐药细胞株的作用及其机制.方法 体外间歇刺激法建立对阿霉素(DOX)产生获得性耐药的骨肉瘤细胞株Saos-2/DOX,MTT法检测DOX对Saos-2及Saos-2/DOX细胞的半数抑制浓度(IC50),并计算Saos-2/DOX细胞的耐药指数(RI);MTT法筛选DATS处理Saos-2/DOX细胞的非细胞毒性浓度;DATS与DOX单独或联合干预Saos-2/DOX细胞24、48、72 h后,MTT法检测细胞生存率;不同浓度DATS(0、10、50、100μmol/L)处理Saos-2/DOX细胞48 h后,Western blotting法检测细胞中pERK1/2、ERK1/2、pAKT1和AKT1蛋白表达.结果 成功建立对DOX耐药的骨肉瘤细胞株Saos-2/DOX,细胞的RI值为6.94.DATS干预Saos-2/DOX细胞的非细胞毒性浓度为10μmol/L.DOX 100μg/L与DATS 10μmol/L联合处理Saos-2/DOX耐药细胞24 h后细胞存活率显著低于DATS 10μmol/L或DOX 100μg/L单独处理,且随处理时间延长逐渐降低(P均<0.05).不同浓度DATS处理Saos-2/DOX细胞48 h后,ERK1/2、pAKT1和AKT1蛋白表达变化差异无统计学意义(P均>0.05),而pERK1/2蛋白表达水平随DATS浓度增高显著增加(P均<0.05).结论 DATS可能通过激活ERK1/2信号通路,逆转Saos-2/DOX细胞的获得性耐药.