Protective Effect Of Astragaloside Ⅳ On PC12 Cells Injured By OGD/R And Its Mechanism Of Action Research

Objective: To investigate the effect of Astragaloside Ⅳ on pheochromocytoma cell(PC12 cells) injured by oxygen-glucose deprivation which was induced by Na2S204 and its possible mechanism.Methods: Different concentrations of Na2S204(10、8、6、4、2mmol/L) diluted by glucose-free Earles solution were used on PC12 cells, we determine the best concentration of Na2S204 by CCK-8 assay; different concentrations of Nimodipine(25、5、1μmol/L) were used to pretecte the PC12 cells injured by OGD/R under the condition of best concentration of Na2S204, we determine the best concentration of Nimodipine by CCK-8 assay, detecting the effect of Astragaloside Ⅳon proliferation of PC12 cells by CCK-8 assay. The cellular morphology were observed under inverted microscope; the cell survival rate was detected by CCK-8 assay; the lactate dehydrogenase concentration was determined by LDH kit assay; detectiing the apoptosis and necrosis through Hoechst33342/PI staining by Fluorescence microscope; detectiing the changes of MMP of PC12 cells through Rh123 staining by the Fluorescence spectrophotometer; the contents of MDA, ROS, SOD as well as GSH were detected by related kits; the protein expression of the Akt, p-Akt, Bax, Bcl-2, Caspase-3 of each group were determined by western blot assay; the CCK-8 assay was used to determine the cellular proliferation of each group when we used LY294002, at the same time the western blot assay was used to the protein expression of the Akt, p-Akt, Bax, Bcl-2, Caspase-3 of each group.Results: According to the result of CCK-8, the cell survival rate was reduced to 50% after being cultured with 10mmol/L Na2S204(P<0.01), what’s more, when the concentration of Nimodipine was 5μmol/L, the best protection was presented. The result of CCK-8 also showed that Astragaloside could Ⅳpretect PC12 cells from OGD/R injury. The result of Hoechst33342/PI staining showed that fluorescence intensities were decreased following by the increasing concentration of Astragaloside Ⅳ. LDH result showed that Astragaloside could decreased the the lactate dehydrogenase concentration. The result of Rh Ⅳ123 staining by the fluorescence spectrophotometer showed that could increased the MMP of PC12 cells injured by ODG/R. The result of antioxidant kits showed that Astragaloside could increase the content Ⅳof SOD and GSH, It also could decreased the content of MDA and ROS. The result of western blot showed that Astragaloside Ⅳcould improve the protein expression of p-Akt/Akt and Bcl-2 and decrease the protein expression of Bax, Bax/Bcl-2 and Caspase-3. When LY294002 was used, the cell survival rate of adding medicine group was decreased, the protein expression of p-Akt/Akt and Bcl-2 were decreased and the protein expression of Bax, Bax/Bcl-2 and Caspase-3 were increased.Conclusion: Astragaloside can increase the the cell survival rate and MMP, it also reduces the Ⅳthe lactate dehydrogenase concentration when PC12 cells were injured by ODG/R. Astragaloside Ⅳcan increase the Antioxidant capacity of PC12 cells by increasing the content of SOD and GSH as well as decreasing the content of MDA and ROS. It can also increase the protein expression of Bcl-2 and decrease the protein expression of Bax, Bax/Bcl-2 and Caspase-3 via the activation of PI3K/Akt signaling pathway. Above all, Astragaloside can protect PC12 cells from injury by OGD/R, what’s more, Oxidative stress Ⅳinjury and PI3K/Akt/Bcl-2 signaling pathway play important roles in the protection mechanism.