七氟烷对甲状腺癌细胞AMPK-SIRT1-PGC-1α信号通路的调控作用研究

Study on the regulation effect of sevoflurane on AMPK-SIRT1-PGC-1αsignal pathway in thyroid cancer cell

目的探究七氟烷对甲状腺癌细胞能量代谢信号通路AMPK-SIRT1-PGC-1α的调控作用。方法将人甲状腺癌细胞株(TPC-1)分为对照组、NC组、七氟烷组和七氟烷+AICAR组,MTT检测各组TPC-1细胞在24、48、72 h细胞增殖情况,流式细胞仪检测细胞存活、早期凋亡及晚期凋亡水平,q PCR检测细胞AMP依赖蛋白激酶(adenosine5’-monophosphate(AMP)-activated protein kinase,AMPK)、AMPK/AD-依赖性去乙酰化酶Sirtuin-1(NAD-dependent deacetylase sirtuin-1,SIRT1)和过氧化物酶体增殖物激活受体γ辅激活因子(peroxisome proliferator-activated receptorgamma coactivator,PGC-1α)mRNA表达水平,Western blot检测细胞AMPK、SIRT1和PGC-1α蛋白表达水平。结果与对照组相比,七氟烷组细胞在24、48、72 h细胞增殖水平显著下降,分别为70.42±1.51、62.16±1.73、55.21±1.22,且有时间依赖效应,差异有统计学意义(P<0.01);七氟烷组细胞存活比例显著降低,早期凋亡和晚期凋亡比例显著增加,分别为(62.71±2.73)%、(16.21±0.22)%、(22.49±2.32)%,差异有统计学意义(P<0.01);q PCR和Western blot实验表明七氟烷组AMPK、SIRT1、PGC-1αmRNA和蛋白表达水平均显著降低,差异有统计学意义(P<0.01);NC组与对照组比较,差异无统计学意义(P>0.05)。与七氟烷组相比,七氟烷+AICAR组细胞在24、48、72 h细胞增殖显著升高,分别为86.11±1.33、78.28±1.41、62.24±1.24,差异有统计学意义(P<0.01);七氟烷+AICAR组细胞存活比例显著升高,早期凋亡和晚期凋亡比例显著降低,分别为(72.25±2.33)%、(12.21±1.01)%、(14.21±2.01)%,差异有统计学意义(P<0.01);q PCR和Western blot实验表明七氟烷+AICAR组AMPK、SIRT1、PGC-1αmRNA和蛋白表达水平均显著升高,差异有统计学意义(P<0.01)。结论七氟烷可以抑制甲状腺癌细胞的增殖,诱导细胞凋亡,此过程与抑制能量信号通路AMPK-SIRT1-PGC-1α的激活有关。

OBJECTIVE To investigate the regulation effect of sevoflurane on energy metabolism signal pathway AMPK-SIRT1-PGC-1αin thyroid cancer cells.METHODS Human thyroid cancer cell line TPC-1 was divided into four groups:control group,NC group(DMSO),sevoflurane group(sevoflurane 3%)and sevoflurane+AICAR(AMPK activator).The cell viability at 24h,48h and 72h was detected by MTT,the cell survival,early apoptosis and late apoptosis were detected by flow cytometry,and AMPK,SIRT1 and PGC-1αmRNA were detected by qPCR,protein expression level of AMPK,SIRT1 and PGC1 were detected by Western blot.RESULTS Compared with the control group,the proliferation of cells in sevoflurane group decreased significantly at 24,48 and 72 hours,with a time-dependent effect of 70.42±1.51,62.16±1.73 and 55.2±1.22,respectively(P<0.01);The proportion of cell survival decreased significantly,and the proportion of early apoptosis and late apoptosis increased significantly,which were(62.71±2.73)%,(16.21±0.22)%and(22.49±2.32)%respectively(P<0.01);QPCR and Western blot showed that AMPK,SIRT1 and PGC-1αmRNA and protein expression levels in sevoflurane group were significantly decreased(P<0.01),and there was no significant difference between NC group and control group(P<0.05);Compared with sevoflurane group,the proliferation of cells in sevoflurane+AICAR group was significantly increased at 24 h,48h and 72h,which were 86.11±1.33,78.28±1.41 and 62.24±1.24,respectively.The proportion of cell survival was significantly increased,and the proportion of early apoptosis and late apoptosis was significantly decreased,which were(72.25±2.33)%,(12.21±1.01)%and(14.21±2.01)%,respectively(P<0.01).qPCR and Western blot tests showed that AMPK,SIRT1 and PGC-1αmRNA and protein expression levels increased significantly.CONCLUSION Sevoflurane can inhibit the proliferation of thyroid cancer cells,induce apoptosis,and inhibit the activation of energy signal pathway AMPK-SIRT1-PGC-1α.