凝血酶体外促进肺癌细胞Glc-82周期进程和增殖及其分子机制

目的探讨凝血酶体外促进肺癌细胞Glc-82周期进程和增殖及其作用机制。方法采用CCK8法检测凝血酶（thrombin）对Glc-82细胞增殖活性的影响,确定其量效和时效关系;利用碘化丙锭（propidium iodide,PI）单染和AnnexinⅤ-FITC/PI双染在流式细胞仪上检测细胞周期和凋亡;以荧光实时定量PCR检测10号染色体上与张力蛋白同源缺失的磷酸酶基因（phosphatase and tensin homologue deleted on chromatosome 10,PTEN）mRNA水平的变化,免疫印迹法检测PTEN蛋白表达和AKT磷酸化水平的变化。结果凝血酶（0.5和1.0 U/ml）可以有效促进细胞的增殖（P〈0.01）。凝血酶（0.5 U/ml）促进Glc-82细胞由G1期向S期转化,可以抑制PTEN的表达,提高AKT磷酸化水平,但对细胞凋亡没有影响。结论凝血酶（0.5 U/ml）可以促进肺癌细胞Glc-82周期进程和细胞增殖,可能系通过PTEN/PI3K/AKT信号分子途径发挥作用。

Effect of thrombin on cell cycle and proliferation of lung cancer cell Glc-82 in vitro and its molecular mechanism

Objective To evaluate the effect of thrombin on cell cycle and proliferation of lung cancer cells Glc-82 in vitro and the mechanism.Methods The effect of thrombin on Glc-82 cells proliferation was determined by CCK8 assay.For cell cycle and cell apoptosis analysis,the cells were stained with propidium iodide（PI） or Annexin-Ⅴ/PI and monitored on a flow cytometer.Real-time fluorescent quantitation polymerase chain reaction was used to detect the level of phosphatase and tensin homologue deleted on chromatosome 10（PTEN） mRNA.The protein expression level of PTEN and AKT phosphorylation was detected by Western blotting assay.Results Thrombin（0.5 and 1.0 U/ml） significantly promoted Glc-82 cell proliferation（P0.01）.Thrombin（0.5 U/ml） facilitated the cell cycle process from G1 phase to S phase.Thrombin markedly down-regulated the expression level of PTEN,and increased AKT phosphorylation.Thrombin showed no effect on Glc-82 cells apoptosis.Conclusion Thrombin promotes lung cancer Glc-82 cell cycle progression and proliferation,which may be regulated by PTEN/PI3K/AKT signaling pathway.