培美曲塞序贯吉非替尼对A549细胞凋亡的影响

The effects of pemetrexed＋gefitinib-gefitinib sequence on apoptosis of A549 cells

目的 探讨培美曲塞序贯吉非替尼对A549细胞的疗效,及其对A549细胞增殖、凋亡及凋亡相关蛋白表达的影响.方法 采用MTT[3-（4,5-二甲基噻唑-2）-2,5-二苯基四氮唑溴盐]比色法检测培美曲塞、吉非替尼、培美曲塞联合吉非替尼及培美曲塞序贯吉非替尼对A549细胞增殖的影响;Western blot 检测凋亡相关蛋白的表达.DAPI核染色法检测A549细胞凋亡的形态学变化.结果 培美曲塞序贯吉非替尼对A549细胞增殖的抑制作用较单药及两药联合治疗更明显,A549细胞中培美曲塞、吉非替尼作用72 h的半数抑制浓度IC50分别为0.84 μmol/L和3.35 μmol/L.Western blot检测到培美曲塞序贯吉非替尼组与培美曲塞、吉非替尼单药治疗组比较,Bcl-2、Caspase3、Caspase8及PARP蛋白表达明显降低,Bax、p-H2AX蛋白表达明显升高,差异有统计学意义（P〈0.05）.DAPI核染色后可观察到培美曲塞序贯吉非替尼诱导A549细胞72 h后凋亡明显增强.结论 培美曲塞、吉非替尼均可明显抑制A549细胞增殖,诱导A549细胞凋亡,培美曲塞序贯吉非替尼作用更明显.

Objective To investigate the effects of pemetrexed ＋ gefitinib - gefitinib sequence on the proliferation and apoptosis of human non - small cell lung cancer （NSCLC） A549 cells. Methods The MTF method was used to de- tect the effect of pemetrexed （ pemetrexed group） , gefitinib （ gefitinib group）, combination of pemetrexed and gefitinib （pemetrexed ＋ gefitinib group ）, and pemetrexed ＋ gefitinib - gefitinib sequence （pemetrexed ＋ gefitinib - gefitinib group） on proliferation of A549 cells. Western blot was performed to analyze the expression of apoptosis - related proteins and the morphological changes of apoptosis were detected by DAPI staining. Results The inhibitory effects of peme- trexed or gefitinib alone and pemetrexed combined with gefitinib on A549 cells was observed. While the inhibitory effects of pemetrexed ＋ gefitinib - gefitinib sequence were more obvious compared with pemetrexed or gefitinib alone and peme- trexed combined with gefitinib. The ICs0 （ inhibitory concentration 50% ） values of pemetrexed and gefitinib on A549 cells for 72 h were 0.84 p.mol/L and 3.35 p, mol/L, respectively. Western blot showed that compared with pemetrexed group and gefitinib group, the expression of Bel -2, caspase 3, caspase 8 and procyclic acidic repetitive proteins （PARP） in pemetrexed ＋ gefitinib - gefitinib group was decreased, but Bax and p - H2AX was increased （P 〈 0.05 ）. DAPI staining showed that the apoptosis rate of pemetrexed ＋ gefitinib - gefitinib group on A549 cells for 72 h was the highest. Conclu- sion Both pemetrexed and gefitinib have obvious inhibitory and apoptosis effect on A549 cells, with the most prominent effect in the pemetrexed ＋ gefitinib - gefitinib group.