自由基清除剂对癫痫大鼠海马组织Fyn的亚硝基化影响

The effect of free radical scavenger on the S-nitrosylation of Fyn in hippocampus of epileptic rats

目的研究自由基清除剂对癫痫大鼠海马组织Fyn的亚硝基化影响,并探讨其变化的可能机制。方法 SD大鼠随机分为癫痫对照组(saline组)、癫痫组(KA组)和药物对照组(KA+saline组)及MCI-186组(KA+MCI-186组)。采用脑室注射海人酸制作大鼠癫痫模型。海马组织匀浆,取蛋白样品用抗Fyn抗体作免疫沉淀,然后用抗-SNO-Cys抗体做免疫印迹,检测Fyn巯基亚硝基化。结果 Fyn的亚硝基化水平癫痫组和药物对照组增加(P<0.05),MCI-186组较药物对照组降低(P<0.05),Fyn的蛋白表达各组间差异无显著性(P>0.05)。结论自由基清除剂可能通过抑制癫痫诱导海马组织Fyn的巯基亚硝基化,调节NMDA受体NR2B亚基磷酸化,进而调节NMDA受体通道功能,对神经元起保护作用。

Objective To investigate whether tyrosine protein kinase Fyn can be S-nitrosylated and the effect of free radical scavenger on the S-nitrosylation of Fyn in kainic acid-induced epileptic rats and to elucidate its possible mechanism.Methods Adult male SD rats were allotted into 4 groups as following： control group（ saline）,seizure group（ KA）,drug control group（ KA ＋ saline） and MCI-186 group（ MCI-186 ＋ KA）. Seizures were induced by intra-cerebroventricular injection of KA dissolved in sterile saline. MCI-186 was intraperitoneally administrated to the rats 40 min before KA injection.Measurement of S-nitrosylated Fyn was performed by immunoprecipitation with anti-Fyn antibody,followed by immunoblotting with anti-SNO-Cys antibody. Results Compared with saline group,the level of the S-nitrosylated Fyn increased significantly in KA group and KA ＋ saline group（ P〈0. 05）. Compared with KA group and KA ＋ saline group,the S-nitrosylation of Fyn decreased significantly in MCI-186 ＋ KA group（ P〈0. 05）. In contrast,the protein expression showed no obvious alteration（ P〉0. 05）. Conclusion Free radical scavenger may involve in inhibiting the S-nitrosylation of Fyn,which suppressed the interactions of Fyn with NR2 B and NR2 B tyrosine phosphorylation and subsequently downregulated NMDA receptor overactivation,thus protected neurons from epileptic injury.