MS-275对食管鳞癌KYSE-70细胞存活、细胞周期、凋亡及迁移的影响
Effects of MS-275 on survival,cell cycle,apoptosis and migration of human esophageal squamous cancer KYSE-70 cells

目的:观察MS-275对食管鳞癌KYSE-70细胞存活、细胞周期、凋亡以及迁移的影响,并分析其对PI3K/Akt/mTOR信号通路相关蛋白p-Akt1和p-mTOR的影响。方法:采用qRT-PCR和Western blot检测KYSE-70细胞和正常食管上皮Het-1A细胞中HDAC1 mRNA及蛋白的表达; CCK-8法检测不同浓度(0.25、0.50、1.00、2.00、4.00、8.00 μmol/L)MS-275对KYSE-70细胞存活的影响; 以0.00、0.25、0.50、1.00和2.00 μmol/L MS-275处理KYSE-70细胞, 48 h后流式细胞仪检测细胞周期,Annexin V/PI染色检测细胞凋亡,划痕实验检测细胞迁移情况,Western blot 检测细胞中Cyclin D1、Cleaved caspase-3、E-cadherin、p-Akt1和p-mTOR蛋白的表达情况。结果:与Het-1A细胞相比,KYSE-70细胞中HDAC1 mRNA和蛋白表达显著增加(P<0.05); MS-275对KYSE-70细胞存活的影响具有时间和剂量依赖性(P<0.05); 随着MS-275处理浓度的增加, KYSE-70 G0/G1期细胞增加、S期细胞降低,细胞凋亡率提高,划痕愈合率降低(P<0.05)。MS-275可提高Cleaved caspase-3和E-cadherin蛋白的表达,降低Cyclin D1、p-Akt1和p-mTOR蛋白的表达(P<0.05)。结论:MS-275可降低KYSE-70细胞存活率,有效抑制细胞迁移,阻滞细胞于G0/G1期,促进细胞凋亡,其作用可能与PI3K/Akt/mTOR信号通路的抑制有关。
Aim:To investigate the effects of MS-275 on the cell survival, apoptosis,cell cycle and migration of human esophageal squamous cancer KYSE-70 cells and analyze PI3K/Akt/mTOR signaling pathway related protein expression.Methods: The expression levels of HDAC1 mRNA and protein in KYSE-70 cells and normal esophageal epithelial cells(Het-1A)were respectively detected by qRT-PCR and Western blot; the survival of KYSE-70 cells was detected by CCK-8 to determine the optimal time and concentration; KYSE-70 cells were treated with 0.00,0.25,0.50,1.00,2.00 μmol/L MS-275 for 48 h,then the apoptosis and the change of cell cycle were detected by flow cytometry; cell migration ability was detected by cell scratches experiment; Cyclin D1,Cleaved caspase-3,E-cadherin and the key proteins(p-Akt1 and p-mTOR)of PI3K/Akt/mTOR signaling pathway were detected by Western blot.Results:HDAC1 mRNA and protein were overexpressed in KYSE-70 cells(P<0.05)compared with Het-1A cells; CCK-8 assay showed that the survival of KYSE-70 cells were inhibited by MS-275 in a dose-and time-dependent manner(P<0.05); with the increase of the concentration of MS-275, the apoptosis rate of KYSE-70 cells was significantly increased,the proportion of cells in G0/G1 phase was increased,that of S phase cells was decreased,and the number of migrating cells was significantly decreased(P<0.05); the relative contents of Cleaved caspase-3 and E-cadherin protein were increased,but those of Cyclin D1,p-Akt1 and p-mTOR protein were reduced by MS-275(P<0.05).Conclusion:MS-275 can effectively inhibit survival and migration,induce apoptosis,arrest cycle of KYSE-70 cells,which might be related with the inhibition of PI3K/Akt/mTOR signaling pathway