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目的研究H2S对缺血再灌注损伤后神经元存活信号转导通路ERK1/2/P~(90RSK)的影响。方法将培养7 d的海马神经元随机分为5组:正常培养组(C组)、缺血再灌注组(I/R组)、缺血再灌注+NaHS组(NaHS组)、缺血再灌注+NaHS+U-0126(ERK抑制剂)组(U组)、缺血再灌注+NaHS+Rapamycin(RS6K抑制剂)组(R组)。C组神经元按正常培养方法培养。NaHS组神经元在神经元进行缺血再灌注时加入NaHS使其终浓度为150μmol/L。U组和R组在加入150μmol/L NaHS的同时分别加入U-0126 10μmol/L或Rapamycin lO nmol/L。各组行细胞存活力、神经元凋亡、cAMP、磷酸化ERK1/2(PERK1/2)和磷酸化P~(90RSK)(PP~(90RSK))蛋白表达的检测。结果NaHS显著增加了cAMP的浓度(与I/R组比较,P<0.01)、PERK1/2蛋白(与I/R组比较,P<0.05)和PP~(90RSK)蛋白(与I/R组比较,P<0.05)表达,同时增加了神经元存活率(与I/R组比较,P<0.05)、降低了神经元凋亡率(与I/R组比较,P<0.05);U-0126抑制了PERK1/2蛋白(与NaHS组比较,P<0.05)和PP~(90RSK)蛋白(与NaHS组比较,P<0.05)表达同时使神经元存活率降低(与NaHS组比较,P<0.05)、神经元凋亡率升高(与NaHS组比较,P<0.05);Rapamycin抑制了PP~(90RSK)蛋白(与NaHS组比较,P<0.05)表达同时使神经元存活率降低(与NaHS组比较,P<0.05)、神经元凋亡率升高(与NaHS组比较,P<0.05)而不影响PERK1/2的表达(与NaHS组比较,P>0.05)。结论H_2S通过cAMP激活了ERK1/2/P~(90RSK)信号通路,在海马神经元缺血再灌注时抑制了神经元的凋亡,保护了神经元。
Objective To investigate the effect of H2S on neuronal survival signal transduction pathway ERK1 / 2 / P ~ (90RSK) after ischemia-reperfusion injury. Methods The hippocampal neurons cultured for 7 days were randomly divided into 5 groups: normal group (C group), ischemia / reperfusion group (I / R group), ischemia / reperfusion + NaHS group (NaHS group) The rats were infused with NaHS + U-0126 (ERK inhibitor) group (U), ischemia-reperfusion + NaHS + Rapamycin (R group). C group of neurons according to normal culture methods. NaHS group neurons added NaHS to ischemic-reperfusion neurons to a final concentration of 150μmol / L. U group and R group were added U-0126 10μmol / L or Rapamycin 10nmol / L respectively while adding 150μmol / L NaHS. The cell viability, neuronal apoptosis, cAMP, phosphorylated ERK1 / 2 (PERK1 / 2) and phosphorylated P ~ (90RSK) (90 ~ 90RK) protein expression in each group were measured. Results Compared with I / R group, NaHS significantly increased the concentration of cAMP (P <0.01), PERK1 / 2 protein (P <0.05 vs I / R group) and PP (90RSK) Compared with I / R group, the survival rate of neurons was increased (P <0.05 compared with I / R group, P <0.05) 0126 inhibited the expression of PERK1 / 2 protein (P <0.05 compared with NaHS group) and PP (90RSK) protein (P <0.05 compared with NaHS group), while decreased the survival rate of neurons (Compared with NaHS group, (P <0.05). Compared with NaHS group, Rapamycin inhibited the expression of PP 90RSK protein (P <0.05 compared with NaHS group) and decreased the survival rate of neurons NaHS group, P <0.05). The apoptotic rate of neurons was increased (P <0.05 compared with NaHS group) but not PERK1 / 2 (P> 0.05). Conclusion H_2S activates ERK1 / 2 / P ~ (90RSK) signaling pathway through cAMP, which inhibits neuronal apoptosis and protects neurons during hippocampal ischemia-reperfusion.