Study On The Protective Effect Of Butylphthalide On Parkinson's Cell Model By Activating PI3K/AKT Pathwa

Object:Some studies have demonstrated that DL-3-n-Butylphthalide(NBP)has a neuroprotective effect on Parkinson’s disease,but the specific mechanism of action is unclear.In this study,SH-SY5Y human neuroblastoma cells secreting dopamine(DA)were cultured in vitro to explore the mechanism of NBP in protecting neuronal cells by regulating PI3K/AKT signaling pathway in Parkinson’s cell model.Methods:Human neuroblastoma cells SH-SY5Y were used for in vitro culture.Different concentrations of 1-methyl-4-phenylpyridine(MPP~+)were used to treat SH-SY5Y cells to construct Parkinson’s cell model.Cell survival rate was measured by CCK8 assay to determine the optimal MPP~+modeling concentration.Different concentrations of NBP were added for pretreatment treatment.After 3 hours of culture,the optimal concentration of MPP~+was added to continue culture for 24 hours.Cell survival rate was measured by CCK8 assay to determine the optimal protective concentration of NBP.The experiment was divided into the following four groups:Control group,MPP~+group,MPP~++NBP group,MPP~++NBP+LY294002 group(LY294002 is an inhibitor of PI3K);after successful model construction,cell survival rate was measured by CCK8assay;NO kit and DCFH-DA fluorescence probe were used to detect intracellular oxidative stress level;Hoechst staining was used to detect apoptosis;Western blot was used to detect p-AKT,TH,andα-synuclein.Results:The results of cell survival measured by CCK8 assay showed that the optimal concentration of MPP~+group was 1 mmol/L,which was statistically significant(P<0.001);the optimal concentration of NBP was 10μmol/L,which was statistically significant(P<0.001).Compared with the control group,the cell survival rate was significantly decreased,the intracellular NO level was increased,the number of apoptosis was increased,the intracellular ROS production was increased,the TH and p-AKT expression levels were decreased,and theα-synuclein expression was increased in the MPP~+group,and the experiments were statistically significant(P<0.05);compared with the MPP~+group,the addition of NBP to treatment could significantly increase the cell survival rate,inhibit the intracellular NO production,reduce the number of apoptotic cells,inhibit the generation of ROS in SH-SY5Y cells,increase the protein expression levels of TH and p-AKT,and decrease the protein expression ofα-synuclein in the MPP~++NBP+LY294002 group,and the protective effects of NBP on SH-SY5Y cells disappeared after the addition of LY294002 inhibitor treatment,and the results of various experiments tended to be in the MPP~+group.Conclusions:NBP can inhibit intracellular NO and ROS production and play an antioxidant role;NBP reduces oxidative stress damage caused by MPP~+by activating the PI3K/AKT signaling pathway.