TIGAR通过SIRT1-PGC1α途径抑制缺血低氧再灌注神经元细胞凋亡

TIGAR inhibits neuronal apoptosis after ischemia-reperfusion via SIRT1-PGC1αpathway

目的探究TP53诱导的糖酵解和凋亡调节因子(TP53-inducible glycolysis and apoptosis regulator,TIGAR)对缺血性脑卒中的作用及其可能的作用机制。方法神经生长因子(nerve growth factor,NGF)诱导大鼠肾上腺嗜铬细胞瘤细胞(rat adrenal pheochromocytoma cell,PC12)神经元分化,氧糖剥夺再灌注(oxygen and glucose deprivation and reperfusion,OG D/R)建立缺血性脑卒中体外模型。免疫荧光染色检测PC12细胞神经元分化潜能;CCK-8检测细胞活力;RT-PCR检测TIGAR mRNA表达;Western blot检测TIGAR蛋白表达;流式细胞术检测细胞凋亡。结果NGF诱导后PC12细胞中神经元特异性标志物NSE呈阳性表达。与siRNA阴性对照(siRNA negative control,si-NC)组相比,TIGAR特异性siRNA序列(TIGAR specific siRNA sequence,si-TIGAR)组细胞中TIGAR mRNA表达、TIGAR、SIRT1和PGC1α蛋白表达量和细胞活力显著降低(P<0.05),细胞凋亡率显著升高(P<0.05);与NC组相比,TIGAR组细胞中TIGAR、SIRT1和PGC1α蛋白表达量和细胞活力显著升高(P<0.05),细胞凋亡率显著降低(P<0.05);与TIGAR组相比,TIGAR+EX 527组细胞中SIRT1和PGC1α蛋白表达及细胞活力显著降低(P<0.05),细胞凋亡率显著升高(P<0.05),TIGAR蛋白表达差异无统计学意义(P>0.05)。结论TIGAR通过激活SIRT1-PGC1α途径在OGD/R神经元细胞中发挥保护作用。

Objective To explore the effect of TP53-inducible glycolysis and apoptosis regulator(TIGAR)on ischemic stroke and its possible mechanism.Methods Nerve growth factor(NGF)induced neuronal differentiation in rat adrenal pheochromocytoma cells(PC 12),and oxygen and glucose deprivation and reperfusion(OGD/R)were used to establish an in vitro model of ischemic stroke.Immunofluorescence staining was used to detect the neuronal differentiation potential of PC12 cells;CCK-8 was used to detect cell viability;RT-PCR was used to detect TIGAR mRNA expression;Western blot was used to detect TIGAR protein expression;flow cytometry was used to detect cell apoptosis.Results The neuron-specific marker NSE was positively expressed in PC 12 cells after NGF induction.Compared with the siRNA negative control(si-NC)group,the mRNA expression of TIGAR,protein expression of TIGAR,SIRT1 and PGC1αand cell viability in the cells of the TIGAR specific siRNA sequence(si-TIGAR)group were significantly reduced(P<0.05),and the apoptosis rate was significantly increased(P<0.05);compared with the NC group,the mRNA expression of TIGAR,protein expression of TIGAR,SIRT1 and PGC1αand cell viability in the TIGAR group were significantly increased(P<0.05),and the apoptosis rate was significantly reduced(P<0.05);compared with TIGAR group,the protein expression SIRT1 and PGC1αand cell viability in the TIGAR+EX 527 group were significantly reduced(P<0.05),cell apoptosis rate was significantly increased(P<0.05),and there was no statistically significant difference in TIGAR protein expression(P>0.05).Conclusion TIGAR plays a protective role in OGD/R neuronal cells by activating the SIRT1-PGC1αpathway.