拉帕替尼对A549人肺癌细胞增殖抑制和凋亡机制的实验研究

Experimental Study of Lapatinib on Proliferation Inhibition and Apoptosis of A549 Human Lung Cancer Cells

目的探讨人表皮生长因子(ErbB)抑制剂拉帕替尼对肺癌细胞A549凋亡的影响及其作用机理。方法2,4,8μmol/L的拉帕替尼与A549肺癌细胞共培养48 h,MTT检测细胞增殖活力,流式细胞实验检测细胞凋亡与细胞周期,免疫印迹法(western blot)检测细胞中磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(AKT)信号通路相关蛋白表达。结果与对照组(0.563±0.056)比较,2,4,8μmol/L的拉帕替尼能显著降低A549细胞活性(0.492±0.050,0.435±0.044,0.356±0.025),差异均有统计学意义(t=2.316,4.402,8.267,均P<0.056)。与对照组比较,2,4,8μmol/L的拉帕替尼能显著提高A549细胞早期凋亡率(t=-5.387,-17.921,-19.981,均P<0.05)及晚期凋亡率(t=-16.174,-19.348,-20.767,均P<0.05),并使细胞周期G1期延长(t=-4.535,-7.276,-9.831,均P<0.05),差异均具有统计学意义;与对照组比较,2,4,8μmol/L的拉帕替尼能显著下调PI3K表达(t=2.165,5.388,17.822,均P<0.05)及CyclinD1表达(t=2.449,16.634,20.208,均P<0.05),下调p21表达(t=2.165,4.898,18.416,均P<0.05),4,8μmol/L的拉帕替尼能显著降低AKT磷酸化水平(t=2.486,18.014,均P<0.05)。结论拉帕替尼通过阻断PI3K/AKT信号通路诱导A549细胞凋亡、抑制细胞增殖、阻滞细胞于G1期。

Objective To explore effect of human epidermal growth factor recpetor(ErbB)inhibitor Lapatinib on proliferation and cell cycle in A549 cell.Methods Cell was cultured by 2,4,8μmol/L dose of Lapatinib for 48 h,cell viability was measured by MTT assay.Cell apoptotic rate and cell cycle was detected by flow cytometry.The expression of phosphatidylinositol 3 kinase(PI3K)/protein kinase B protein(AKT)related protein was measured by western blot.Results Compared with the control group(0.563±0.056),2,4,8μmol/L lapatinib significantly reduced the activity of A549 cells(0.492±0.050,0.435±0.044,0.356±0.025),with statistical significance(t=2.316,4.402,8.267,all P<0.05).Compared with the control group,2,4,8μmol/L lapatinib significantly increased the early apoptosis rates(t=-5.387,-17.921,-19.981,all P<0.05)and late apoptosis rates(t=-16.174,-19.348,-20.767,all P<0.05)of A549 lung cancer cells,made the cell cycle arrested in G1 phase(t=-4.535,-7.276,-9.831,all P<0.05),the differences were statistically significant,repectively.Down-regulated the expression of CyclinD1(t=2.449,16.634,20.208,all P<0.05)and PI3K(t=2.165,5.388,17.822,all P<0.05),down-regulated the expression of p21(t=2.165,4.898,18.416,all P<0.05).4,8μmol/L dose of lapatinib down-regulated the expression of p-AKT(t=2.486,18.014,all P<0.05).Conclusion Lapatinib could induce A549 cell apoptosis and made the cell cycle arrested in G1 phase by blocking PI3K/Akt signaling pathway.