洛匹那韦逆转LLC/cMOAT细胞多药耐药的作用

Reversal effects of lopinavir on multidrug resistant LLC/cMOAT cells

目的研究洛匹那韦对LLC/cMOAT细胞株多药耐药的逆转作用及其可能机制。方法采用四甲基偶氮唑盐方法(MTT)检测洛匹那韦对细胞株LLC/CMV和LLC/cMOAT细胞生存的影响,并检测阿霉素(ADM)、长春新碱(VCR)、环磷酰胺(CTX)、顺铂(DDP)对上述细胞株的半数抑制浓度(IC50)。应用流式细胞术检测洛匹那韦处理后细胞内化疗药物阿霉素相关的荧光强度变化。结果 LLC/cMOAT细胞对ADM、VCR、DDP有不同程度的耐药,对CTX未产生耐药;洛匹那韦在浓度小于10.0μmol.L-1时,对LLC/cMOAT细胞无明显细胞毒性作用;用2.5μmol.L-1洛匹那韦处理后,LLC/cMOAT细胞对VCR和DDP的敏感性增高,浓度提高至5.0μmol.L-1时,敏感性明显提高;用洛匹那韦处理后LLC/cMOAT细胞内ADM的蓄积增加,并表现为浓度依赖;细胞周期检测分析显示,使用洛匹那韦处理0、4、8、16 h后,G1期细胞百分比由(42.65±3.75)%分别增至(56.77±2.37)%、(68.13±4.30)%、(70.25±5.12)%,与0 h G1期细胞数比较差异显著(P<0.05);使用洛匹那韦处理细胞48 h后,凋亡率增高且呈时间和浓度依赖性。结论洛匹那韦可以部分逆转LLC-cMOAT的多药耐药,这种逆转可能与增加细胞内化疗药物蓄积,诱导细胞G1期阻滞,增强细胞凋亡有关。

Aim To study the reversal effects of lopinavir on the multidrug resistance of LLC/cMOAT cells.Methods The sensitivity of LLC/CMV and LLC/cMOAT cells to several chemotherapeutics,the cytoactivity of LLC/CMV and LLC/cMOAT cells treated with different concentrations of lopinavir and the inhibition rate of ADM,VCR,CTX and DDP treated with different concentration of lopinavir to LLC/CMV and LLC/cMOAT cells were evaluated with MTT assay.The IC50 of different agents was counted.Intracellular concentration of ADM and cellular cycle analysis were detected by flow cytometer（FCM）.Results LLC/cMOAT cells were resistant to ADM,VCR and DDP but not to CTX.When the concentration less than 10 μmol·L^-1,lopinavir was not significantly cytotoxic to LLC/CMV and LLC/cMOAT cells.After treatment with 2.5 μmol·L^-1 lopinavir,the chemo-sensitivity of LLC/cMOAT cells to VCR and DDP was enhanced and much higher when treated with 5.0 μmol·L^-1of lopinavir.When treated with 2.5 and 5.0 μmol·L^-1 lopinavir,ADM accumulation in LLC/cMOAT cells were enhanced significantly in a concentration-dependent manner.Cellular cycle analysis demonstrated that 0,4,8,16 hours after co-cultured with lopinavir the amount of cells at G0/G1 phase increased from（42.65±3.75）% at 0 h to（70.25±5.12）% at 16 h（P〈0.05）.The cell apoptosis rate was increased in a time-and concentration-dependent manner.Conclusion Lopinavir could perform reverse effects on the multidrug resistance of LLC/ cMOAT cells and the reversal effects correlate to lopinavir concentration.The underlying mechanism might involve the growth arrest at G1,the increasing of intracellular drug concentration and promoting apoptosis.