The Protective Effect And The Relative Mechanism Of Swertiamarin On OGD/R-induced Injury In SH-SY5Y Cells

Objective In order to study the potential neuroprotective effect and the relative mechanism of Swertiamarin on OGD/R-induced injury in SH-SY5 Y cells,a model of OGD/R injury in SH-SY5 Y cells was established by referring to the related literature.In addition,the HPLC method of Swertiamarin was established to study the influencing factors of its solubility and stability,which provided experimental and pharmacological basis for further development and utilization of Swertiamarin.Method1.Constructed the OGD/R models of SH-SY5 Y cells and randomly divided into five groups: normal group(Control),OGD/R injury group(Vehicle)and OGD/R injury+STM group(120,60,30?M).2.Use CCK-8 to detect the cytotoxicity of SH-SY5 Y cells with different concentrations STM(30?M,60?M and 120?M)after 24 hours.And detected the activity of SH-SY5 Y cells in each group.3.Use Annexin V-FITC/PI method to determine the proportion of apoptotic cells in each group.4.Use JC-1 probe to detect the changes of mitochondrial membrane potential(MMP)in each group.5.Use DCFH-DA staining method to detect the levels of reactive oxygen species(ROS)production in each group.6.To observe the effect of STM on the OGD/R injury of SH-SY5 Y cells,and use western blot to determin the expression level of related proteins TLR4,PARP1 and NF-?B.7.Determine the solubility and stability of STM in Hank's equilibrium salt solution by HPLC,lay a foundation for Swertiamarin absorption in vivo.Result1.The results showed that there was no significant difference in cell viability between the untreated group and the STM treated group,whicshowed that the concentrations of STM(30,60 and 120?M)selected in this study were not cytotoxic;2.CCK-8 detection showed that the cell viability in OGD/R group was significantly lower than that in control group(p<0.01)and that in STM(30,60,120 ? M)pretreatment group was significantly higher than that in OGD/R group(p<0.01).3.After OGD/R injury,the cell survival rate decreased,and the pretreatment of STM(30,60,120?M)increased the viability of SH-SY5 Y cells(p<0.01).4.OGD/R decreased the ? ? m of SH-SY5 Y cells,and the pretreatment of STM(30,60,120?M)increased the ??m of cells.5.OGD/R significantly increased ROS levels in SH-SY5 Y cells.STM(30,60,120?M)pretreatment inhibited OGD/R-induced intracellular ros production and reduced apoptosis.6.OGD/R mainly induced the expression of TLR4,My D88,NF-?Bp65 and PARP1 in cells.STM pretreatment reduced the expression of TLR4,My D88,NF-?Bp65 and PARP1.7.STM is stable in HBSS solution when p H=3,p H=5 and p H=7.4,the stability is poor in the alkaline environment of p H=9 and p H=11.Light and temperature have no obvious effect on STM stability.Conclusion STM protects SH-SY5 Y cells against OGD/R-induced damage by reducing the increase of ROS levels and inhibiting apoptosis.And the stability of STM in alkaline HBSS solution is poor.