论文部分内容阅读
目的 探讨百里醌(TQ)对非小细胞肺癌(NSCLC)细胞毒性作用的分子机制.方法 NSCLC鳞癌细胞SK-MES-1接种于96孔板,予20、40、60、80、100 μmol/L TQ培养24 h,观察浓度依赖性,计算TQ的IC50值;予接近IC50浓度TQ培养SK-MES-1,观察时间依赖性;予ERK抑制剂U0126和p38抑制剂SB203580分别作用于SK-MES-1、95-D,观察细胞存活率;Western blot检测U0126预处理1h,加TQ孵育SK-MES-1 30 min后测p-p38、p38,p-ERK1/2、ERK1/2,p-JNK、JNK蛋白表达.结果 ①TQ呈浓度和时间依赖性降低NSCLC细胞存活率;②30 μmol/L TQ和10 μmol/L U0126 +30μmol/L TQ比较其细胞存活率有显著差异(P =0.000),但与10 μmol/L SB203580 +30μmol/L TQ比较无显著差异(P=1.00),10 μmol/L SB203580+ 30 μmol/L TQ与10 μmol/L U0126+ 30 μmol/L TQ组比较有显著差异(P =0.000);60 μmol/L TQ、10 μmol/L U0126+60 μmol/L TQ、10 μmol/L SB203580 +60μmol/L TQ两两比较均无显著差异(P＞0.05);随TQ浓度增加,SB203580对95-D细胞的保护作用逐渐减弱,10μmol/L SB203580+ 40μmol/L TQ组与40 μmol/L TQ组比较细胞存活率仍有显著差异(P=0.033);③Western blot结果表明U0126显著抑制ERK1/2磷酸化,p38随TQ浓度增加而磷酸化增加,但ERK 1/2磷酸化减少,JNK磷酸化无明显改变.结论 TQ透过磷酸化p38途径介导NSCLC毒性作用,而不是ERK1/2途径.“,”Objective To explore the mechanism of thymoquinone (TQ) on NSCLC cytotoxicity.Methods SK-MES-1 was inoculated into 96-well plates and cultured at 20,40,60,80 and 100 μmol/L TQ for 24 h,and the IC50 of TQ was calculated.SK-MES-1 was cultured in close proximity to IC50 concentration TQ,and time-dependent was observed.The ERK inhibitor U0126 and the p38 inhibitor SB203580 were applied to SK-MES-1 and 95-D,respectively,then TQ-activated MAPK-mediated cytotoxicity were observed.The SK-MES-1 expression of p-p38,p38,p-ERK1/2,ERK1/2,pJNK and JNK protein were detected by U0126 pretreatment for 1 h and TQ-cultured for 30 min.Results ① TQ was used to mediate NSCLC cells in a concentration and time-dependent manner,and the viability of NSCLC cells was decreased.② The cell viability of 30 μmol/L TQ and 10 μmol/L U0126 +30 μmol/L TQ showed significant differences (P =0.000),but no significant difference was found when compared with 10 μmol/L SB203580 + 30 μmol/L TQ (P =1.00).10 μmol/L SB203580 + 30 μmol/L TQ was significantly different from 10 μmol/L U0126 + 30 μmol/L TQ (P =0.000).60μmol/LTQ,10μmol/LU0126 + 60μmol/LTQ,10μmol/LSB203580 + 60 μmol/L TQ showed no significant differences between every two groups (P ＞ 0.0 5).With the increase of TQ concentration,the protective effect of SB203580 on 95-D cells gradually decreased,and 10 μmol/L SB203580 +40 μmol/L TQ group was significantly different from 40 μmol/L TQ group (P =0.033).③ Western blot analysis showed that U0126 could significantly inhibit the phosphorylation of ERK1/2,phosphorylated p38 increased with the increasing of TQ concentration.However,ERK1/2 phosphorylation decreased,and JNK phosphorylation did not change significantly.Conclusion TQ can mediate NSCLC cytotoxicity through phosphorylation of p38 pathway,but not ERK1/2 pathway.