心脑舒通胶囊对氧糖剥夺/复氧损伤原代神经元的保护作用

目的]探讨心脑舒通胶囊对氧糖剥夺/复氧(OGD/R)损伤的原代神经元细胞活力及微管相关蛋白2(MAP-2)表达的影响。方法]取新生24 h内SD大鼠的大脑皮质进行神经元原代培养,并通过免疫荧光法进行MAP-2目的蛋白鉴定;建立神经元OGD/R损伤的体外模型,CCK-8检测不同浓度的心脑舒通胶囊对体外正常培养和OGD/R损伤后神经元细胞活力的影响;选取最佳浓度的心脑舒通胶囊进行MAP-2的荧光表达强度实验,通过倒置荧光显微镜进行拍照处理,并通过软件对神经元突触长度进行测量及统计处理,Western blot法检测MAP-2的蛋白表达情况。结果]神经元原代培养成功,且MAP-2染色阳性。CCK-8结果显示心脑舒通胶囊在10、25、50μg/mL浓度对正常培养神经元细胞有细胞毒作用(P<0.05),因此选取低于10μg/mL浓度的心脑舒通胶囊进行后续实验。而在OGD/R损伤条件下,与模型组比较,心脑舒通胶囊在3.125、6.25μg/mL浓度能显著增加OGD/R损伤神经元的细胞活力(P<0.05);免疫荧光实验结果显示,与对照组比较,模型组MAP-2荧光表达强度及神经元突触长度明显降低(P<0.05);与模型组比较,心脑舒通胶囊在3.125μg/mL浓度能显著增加MAP-2的荧光表达强度及神经元突触长度(P<0.05)。Western blot实验结果显示,与对照组比较,模型组MAP-2蛋白表达显著降低(P<0.05),与模型组比较,心脑舒通胶囊能显著增加MAP-2的蛋白表达(P<0.05)。结论]心脑舒通胶囊能促进OGD/R损伤神经元活力,其作用机制与促进细胞骨架蛋白MAP-2表达及突触稳定性有关。

Protective effect of Xinnao Shutong Capsule on primary neurons injured by oxygen-glucose deprivation/reoxygenation

Objective]To investigate the effects of Xinnao Shutong(XNST)Capsules on the viability of primary neurons and the expression of microtubule-associated protein-2(MAP-2)in oxygen-glucose deprivation/reoxygenation cells.Methods]Newborn SD rats were used to obtain cerebral cortical neurons for primary culture and MAP-2 immunofluorescence identification.Oxygen-glucose deprivation/reoxygenation(OGD/R)injury model was established and CCK-8 was used to detect different concentrations of XNST Capsules on the cell vitality.Immunofluorescence method was used to observe the fluorescence intensity of MAP-2,and the neuronal synaptic length was measured by Image J software.Western blot method to detect MAP-2 protein expression.Results]The primary culture of neurons was successful,and MAP-2 staining was positive.CCK-8 results showed that XNST Capsules had cytotoxic effects on normal cultured neurons at concentrations of 10,25,and 50μg/mL(P<0.05).Compared with the OGD/R group,XNST Capsules at 3.125 and 6.25μg/mL concentration can significantly increase the cell viability(P<0.05).Compared with the control group,the MAP-2 fluorescence intensity and neuron synaptic length in the OGD/R group were significantly reduced(P<0.05).Compared with the OGD/R group,3.125μg/mL XNST Capsules could significantly increase MAP-2 fluorescence intensity expression and neuronal synaptic length(P<0.05).Western blot results showed that compared with the control group,the MAP-2 protein expression of the model group was significantly reduced(P<0.05).Compared with the model group,XNST Capsules can significantly increase the expression of MAP-2 protein(P<0.05).Conclusion]XNST Capsules can promote the neuron viability induced by OGD/R,and its mechanism is related to the promotion of cytoskeletal protein MAP-2 expression and synaptic stability.