Protective Effect Of Astragaloside Iv On H₂O₂-induced Damage In PC12Cells

Obiective Through the establishment of PC12cells in vitro model oxidative stress, toinvestigate the effect of Astragaloside IV on t-Akt、p-Akt、t-eNOS p-eNOS in H2O2-induced PC12cell, we explored whether AsIV restrained H2O2-induced PC12celldamage through activating PI3K-Akt-eNOS signal pathway. The findings provide anbeneficial experimental evidence for Astragaloside IV of neuroprotective mechanism ofaction and the clinical on antioxidant therapy strategy.Methods The viability rates of PC12cells induced by different concentrations andtreatment time of H2O2was observed using MTT assay, and a proper concentration andtreatment time were chosen to make the cell injury model. The viability rates of PC12cells on different concentrations of astragaloside IV alone and in the oxidation modelrespectively, to screen the suitable concentration. Then PC12cells were assigned to fourgroups: control group, H2O2model group, AsIV+H2O2group, LY294002+AsIV+H2O2group. LDH was to detected of LDH kit, and The western blot method was used tomeasure the expression protein of Caspase-3, t-Akt, p-Akt, t-eNOS, p-eNOS.Results1The results of cell viability detecting by MTT method: Compared with thecontrol group, Astragaloside IV concentrations in the10~400nmol/L group alone inPC12cells, no significant effect on cell survival rate. The cell activity of PC12ispositively related to H2O2concentration and its time effect. The cell viability of PC12decreased with the increase of H2O2concentration and time. In the300μmol/L H2O2group, when the time reached4h, the cell viability of PC12decreased about53%,reaching PC12median lethal dose, so the final choice of PC12cells induced by300umol/L H2O2and4h to establish model of oxidative stress. Compared to the H2O2model group, PC12cells treated with different concentrations of Astragaloside IV(100nmol/L,200nmol/L,400nmol/L) showed increase the cell activity,100nmol/L wasthe most effective concentration (P＜0.05).2The resolt of LDH released: Compared tothe control group, in the H2O2model group, the content of LDH was significantlyincreased (P＜0.05), which has obvious effect on cell injury. Compared to the H2O2model group, in AsIV+H2O2group, astragaloside IV preconditioning can reduce thecontent of LDH in H2O2induced PC12cell supernatant(P＜0.05), reduce cell damage. Compared with Astragaloside IV preconditioning group (AsIV+H2O2group), inLY+AsIV+H2O2group, Astragaloside IV pretreatment after treatment with LY294002can increase LDH content of supernatant of cells significantly(P＜0.05).3Detectionusing Western blot method, Compared with the control group, the expression of apoptosisCaspase-3significantly increased (P＜0.05), promoted apoptosis. Compared with theH2O2model group, in AsIV+H2O2group, Astragaloside IV preconditioning cansignificantly reduce the expression of apoptosis Caspase-3protein in H2O2-induced PC12cells, inhibit progression of apoptosis(P＜0.05), and can increse the expression of p-Aktand p-eNOS(P＜0.05). Compared with Astragaloside IV preconditioning group(AsIV+H2O2group), in LY+AsIV+H2O2group，Astragaloside IV pretreatment aftertreatment with LY294002can inhibite the expression of p-Akt and p-eNOS(P＜0.05).The study found that astragaloside IV might play a role through the activation ofPI3K-Akt-eNOS signal pathway.Conclusion Astragaioside IV protects the PC12cells from oxidative stress induced byH2O2via stimulating the signal pathway of PI3K-Akt-eNOS.