[关键词]
[摘要]
目的 本研究通过去甲斑蝥素(NCTD)作用于肺癌顺铂(DDP)耐药细胞A549/DDP,探讨其对DDP耐药的逆转作用及其相关机制。方法 应用CCK-8法检测NCTD对A549和A549/DDP细胞活力的影响、DDP对A549细胞和A549/DDP的半数抑制浓度(IC50)、NCTD联合不同浓度DDP对A549/DDP细胞DDP耐药性的影响;细胞克隆形成实验检测NCTD、DDP单给药及NCTD与DDP联合用药对A549/DDP细胞克隆形成率的影响;Western blotting法检测A549/DDP和A549细胞中c-met表达,以及NCTD、DDP单给药及NCTD与DDP联合用药对A549/DDP细胞内c-met蛋白表达的影响。结果 NCTD剂量相关性地抑制A549和A549/DDP细胞的活力,选择抑制率小于10%的最大NCTD浓度即10 μmol/L作为逆转浓度;DDP作用于A549/DDP和A549细胞的IC50分别为15.11和5.04 μmol/L,差异有统计学意义(P<0.01);A549/DDP细胞中c-met表达明显高于A549细胞;用NCTD(10 μmol/L)处理后,DDP对A549/DDP细胞的IC50从15.11 μmol/L减少到8.06 μmol/L,差异显著(P<0.01);与DDP单给药组比较,NCTD联合DDP作用明显降低A549/DDP细胞克隆形成率(P<0.01);Westernblotting结果显示,NCTD联合DDP作用明显抑制顺铂耐药细胞A549/DDP细胞中c-met蛋白的表达,单用DDP或NCTD与对照组比较差异不明显。结论 NCTD与DDP联用可逆转A549/DDP细胞对DDP耐药,其机制可能与抑制c-met蛋白表达有关。
[Key word]
[Abstract]
Objective To investigate the anti-resistance effects of norcantharidin (NCTD) on cisplatin (DDP) resistance A549/DDP cells and its reversal mechanism. Methods A549 and A549/DDP cells were used as a model for investigating the function of NCTD and DDP in this study.CCK-8 method was used to detect the effect of NCTD on the activity of A549 and A549/DDP cells, IC50 of DDP on A549 and A549/DDP cells, and the effect of NCTD combined with DDP of different concentrations on DDP resistance of A549/DDP cells. Cell clonogenesis assay was used to detect the clonogenesis rate of A549/DDP cells treated with NCTD, DDP alone and the combination of NCTD and DDP; Western blotting was used to detect the expression of c-Met in A549/DDP and A549 cells, and the effect of NCTD, DDP alone and the combination of NCTD and DDP on the expression of c-met protein in A549/DDP cells. Results The cell activity of the A549 and A549/DDP cells were significantly inhibited by NCTD in a dose dependent manner. The concentration of 10 μmol/L was selected to be the reversal concentration, because its maximum rate was less than 10%. The IC50 of DDP on A549/DDP and A549 cells was 15.11 and 5.04 μmol/L respectively, the difference was statistically significant (P < 0.01); The expression of c-Met in A549/DDP cells was significantly higher than that in A549 cells; After treatment with NCTD (10 μmol/ L), the IC50 of DDP on A549/DDP cells decreased from 15.11 to 8.06 μmol/L, the difference was significant (P < 0.01); Compared with DDP alone group, the effect of NCTD combined with DDP significantly reduced the formation of A549/DDP cell clone (P < 0.01). Western blotting results showed that the combination of NCTD and DDP significantly inhibited the expression of c-met protein in cisplatin resistant A549/DDP cells. Conclusion NCTD compared with DDP can reverse the DDPresistance of A549/DDP cells, and its potential mechanism may be ralated to the down-regulation of c-met expression.
[中图分类号]
R965
[基金项目]
山东省医学科学院院级科技计划(2016-41)