AMG-102通过调控c-Met/PI3K/Akt通路抑制喉鳞癌细胞增殖和诱导凋亡

AMG-102 inhibits proliferation and induces apoptosis of laryngeal squamous cell carcinoma cells by regulating c-Met/PI3K/Akt pathway

目的探讨c-Met抑制剂AMG-102对喉鳞癌Hep-2细胞增殖和凋亡的影响及潜在机制。方法分别以2.5、5和10μmol/L的AMG-102处理喉鳞癌Hep-2细胞,采用四甲基偶氮唑蓝(MTT)法检测AMG-102对Hep-2细胞的增殖抑制作用,流式细胞术和Hoechst染色法检测细胞凋亡情况。实时荧光定量聚合酶链反应(RT-qPCR)检测凋亡相关基因的mRNA表达,Western blot检测c-Met/PI3K/Akt通路相关蛋白的表达。结果与对照组比较,2.5、5、10μmol/L AMG-102处理Hep-2细胞24 h的增殖率分别为(89.8±1.1)%、(79.8±1.0)%和(69.1±1.2)%,处理48 h的增殖率分别为(76.8±2.0)%、(60.2±1.1)%和(49.8±1.2)%,处理72 h的增殖率分别为(50.1±2.0)%、(41.5±1.1)%和(33.6±1.0)%,差异均有统计学意义(均P<0.05)。2.5、5、10μmol/L AMG-102处理Hep-2细胞48 h的凋亡率分别为(16.09±1.53)%、(27.51±2.02)%和(36.57±1.42)%,明显高于对照组[(3.62±0.10)%],差异均有统计学意义(均P<0.05)。2.5、5、10μmol/L AMG-102处理48 h,Hep-2细胞Bcl-2 mRNA的相对表达量分别为0.58±0.13、0.38±0.12和0.20±0.13,p-Met蛋白的相对表达量分别为80.0±3.8、50.6±4.2和28.5±1.3,p-PI3K蛋白的相对表达量分别为87.1±0.9、54.2±1.2和21.0±1.2,p-AKT蛋白的相对表达量分别为98.7±5.6、56.9±3.2和32.2±4.3,均明显低于对照组(均P<0.05);Bax mRNA的相对表达量分别为1.78±0.13、2.37±0.14和3.05±0.13,caspase-3 mRNA的相对表达量分别为1.98±0.14、2.47±0.14和3.15±0.13,均明显高于对照组(均P<0.05)。结论c-Met抑制剂AMG-102通过调控c-Met/PI3K/Akt通路,可以抑制喉鳞癌Hep-2细胞增殖,诱导其凋亡。

Objective To investigate the effects of c-Met inhibitor AMG-102 on the proliferation and apoptosis of laryngeal squamous carcinoma Hep-2 cells and the underlying mechanism.Methods Laryngeal squamous carcinoma cell line Hep-2 cells were treated with 2.5,5 and 10μmol/L AMG-102,respectively.The proliferation activities of Hep-2 cells were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide(MTT).The apoptotic rate of Hep-2 cells was detected by flow cytometry analysis and Hoechst staining.The mRNA expression levels of apoptosis-related genes were detected by real-time quantitative polymerase Chain reaction(RT-qPCR),and the protein expressions of c-Met/PI3K/AKT pathway were detected by western blot.Results Compared with the control group,the proliferation rates of Hep-2 cells treated with 2.5,5 and 10μmol/L AMG-102 for 24 hours were(89.8±1.1)%,(79.8±1.0)%and(69.1±1.2)%,respectively;for 48 hours were(76.8±2.0)%,(60.2±1.1)%and(49.8±1.2)%,respectively;for 72 hours were(50.1±2.0)%,(41.5±1.1)%and(33.6±1.0),respectively,with significant differences(all P<0.05).The apoptotic rates of Hep-2 cells treated with 2.5,5 and 10μmol/L AMG-102 for 48 hours were(16.09±1.53)%,(27.51±2.02)%and(36.57±1.42)%,respectively,which were significantly higher than(3.62±0.10)%in the control group(all P<0.05).After treated with 2.5,5 and 10μmol/L AMG-102 for 48 hours,the relative expression levels of Bcl-2 mRNA in Hep-2 cells were 0.58±0.13,0.38±0.12 and 0.20±0.13,respectively;the relative protein expression of p-Met were 80.0±3.8,50.6±4.2 and 28.5±1.3,respectively;the relative protein expression of p-PI3K were 87.1±0.9,54.2±1.2 and 21.0±1.2,respectively;the relative protein expression of p-AKT were 98.7±5.6,56.9±3.2 and 32.2±4.3,respectively;which were significantly lower than those in the control group(all P<0.05).The relative expression levels of Bax mRNA were 1.78±0.13,2.37±0.14 and 3.05±0.13,respectively,and the relative expression levels of caspase-3 mRNA were 1.98±0.14,2.47±0.14 and 3.15±0.13,respectively,which were significantly higher than those in the control group(all P<0.05).Conclusion c-Met inhibitor AMG-102 could inhibit the proliferation and induce apoptosis of laryngeal squamous carcinoma Hep-2 cells by regulating the c-Met/PI3K/Akt pathway.