The Effect Of Icaritin On 6-OHDA-induced NLRP3,Caspase-1 And IL-1β Protein Expression In MN9D Cells

Objective: By studying the effect of Icaritin(ICT)on the expression of NLRP3,Caspase-1 and IL-1β proteins in 6-OHDA-induced MN9 D cells,Explore the protective effect of ICT on 6-OHDA-induced MN9 D cell injury model and its possible mechanism.Methods: 1)MN9D cells were selected for resuscitation and culture,and TH and NLRP3 in MN9D cells were detected by immunofluorescence;2)MN9D cells were induced by different concentrations of 6-OHDA(0,10,20,30,40,50,60,70,80 μmol/L).The viability of the cell was detected by CCK8 method,and the most suitable concentration of 6-OHDA was selected;3)MN9D cells were pretreated with different concentrations of ICT(0.001μmol/L,0.01 μmol/L,0.1 μmol/L,1 μmol/L,10 μmol/L).The viability of the cell was detected by CCK8 method,and the appropriate concentration of ICT was selected;4)According to the experimental design,the cells were divided into four groups: blank group,6-OHDA(50 μmol/L)group,6-OHDA+ICT(0.001 μmol/L)group and 6-OHDA+ICT(0.01μmol/L)group.The protein expression levels of α-syn and TH in each group were detected by Western blot;5)The protein expression levels of NLRP3,Casepase-1 and IL-1 β in each group were detected by Western blot.Results: 1)With the increase of the concentration of 6-OHDA,the cell viability decreased gradually.When the concentration of 6-OHDA was 50 μmol/L,the level of cell viability was about 50%,which was close to the level of modeling,so 50 μmol/ L was chosen as the concentration of modeling in this experiment;2)Different concentrations(0.001 μmol/L,0.01 μmol/L,0.1 μmol/L,1 μmol/L,10 μmol/L)ICT had no significant effect on the viability of MN9 D cells,and it was found that ICT at the concentration of 0.01 μmol/L had the best protective effect on the cell injury model induced by 6-OHDA(50 μmol/L);3)The expression of TH in 6-OHDA(50 μmol/L)group was significantly lower than that in blank group.After being pretreated with ICT for 1 hour,the expression of TH in 6-OHDA+ICT(0.001 μmol/L)group and 6-OHDA+ICT(0.01 μmol/L)group was significantly higher than that in 6-OHDA group(50 μmol/L);4)The expression of α-syn,NLRP3,Casepase-1 and IL-1 β in 6-OHDA(50 μmol/L)group was significantly higher than that in blank group.After being pretreated with ICT for 1 h,the expression of α-syn,NLRP3,Casepase-1 and IL-1 β in 6-OHDA+ICT group and 6-OHDA+ICT group was significantly lower than that in 6-OHDA group.Conclusion: ICT has a protective effect on 6-OHDA-induced MN9 D cell model,which related to the inhibition of the expression of NLRP3,Caspase-1,IL-1β inflammation-related proteins.