丙泊酚通过上调纤维细胞生长因子的表达激活磷酸肌醇3激酶-蛋白激酶B和磷酸化细胞外信号调节激酶1/2信号通路抑制体外培养神经元缺血/再灌注损伤

摘要

Objective To study the effect of propofol on cerebral ischemia/reperfusion injury (I/RI) and the relative mechanism.Methods The cerebral ischemia/reperfusion model was produced by oxygen glucose deprivation and reperfusion in vitro,methyl thiazolyl tetrazolium (MTT) assay was used to detect the proliferative effect of propofol.Apoptotic cells were detected with Annexin V staining.The expression level of basic fibroblast growth factor (bFGF) mRNA was measured by RT-PCR.The expression levels of bFGF,phosphorylated protein kinase B (pAkt) and phosphorylated extracellular signal-regulated kinase 1/2 (pERK1/2) protein expression were detected by Western blot.Silence of bFGF in cortical neurons by small interfering RNA.Results The apoptosis rate of neurons in oxygen-glucose deprivation/reperfusion (OGD/RP) group was 43.2％.After treated with 10 mg/L propofol,the apoptosis rate of neurons was reduced to 19.5％.The expression levels of bFGF protein in OGD treated group was significantly lower than the control group(P＜0.05),and the bFGF level was markedly upregulated in propofol treated group(P＜0.05).Propofol could upregulate the expression of pAkt and pERK1/2,and activate the two signaling pathways.Silence the expression of bFGF or treatment with the inhibitor of phosphotylinosital 3 kinase-protein kinase B (PI3K-Akt) and ERK1/2 signal pathway both resulted in the decrease of neurons viability (P＜0.05),and the inhibition of PI3K-Akt and ERK1/2 activation.Conclusions Propofol could activate PI3K-Akt and ERK1/2 signal pathway via upregulating the expression of bFGF,and promote the survival of cortical neurons.%目的 探讨丙泊酚在脑缺血/再灌注损伤(ischemia/reperfusion injury,I/RI)中发挥的作用及其具体机制. 方法 采用氧糖剥夺再灌注(oxygen-glucose deprivation/reperfusion,OGD/RP)法体外构建缺血/再灌注细胞模型,将细胞分为对照组、OGD/RP组、丙泊酚+OGD/RP组.采用甲基噻唑基四唑(methyl thiazolyl tetrazolium,MTT)法检测皮质神经细胞存活率,Annexin V-PI检测细胞凋亡情况,即时聚合酶链式反应(real-time polymerase chain reaction,RT-PCR)方法检测碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)的mRNA表达情况,免疫印迹法(Western blot)检测丙泊酚对皮质神经细胞内bFGF,磷酸化蛋白激酶B(phosphorylated protein kinase B,pAkt)以及磷酸化细胞外信号调节激酶1/2 (phosphorylated extr acellular signal-regulated kinase 1/2,pERK 1/2)蛋白表达的影响;采用小干扰RNA构建bFGF沉默的细胞. 结果 OGD/RP处理组神经细胞凋亡率为43.2％,经10 mg/L的丙泊酚预处理后,细胞的凋亡率降为19.5％.与对照组比较,OGD处理后,细胞中bFGF的含量显著下调(P＜0.05),丙泊酚处理的皮质神经元中bFGF含量显著高于OGD处理组(P＜0.05).丙泊酚能够上调pAKT以及pERK1/2的表达,激活这两条信号通路.沉默bFGF或者施加磷酸肌醇3激酶-蛋白激酶B(phosphotylinosital 3 kinase-protein kinase B,PI3K-Akt)以及pERK1/2信号通路抑制剂都会导致细胞存活率显著下降(P＜0.05),抑制PI3K-Akt以及pERK1/2的激活. 结论 丙白酚可以通过上调bFGF的表达,激活PI3K-Akt和ERK 1/2信号通路,增加皮质神经元的存活.