Inhibitory effect of 1,25(OH)2D3 on Aβ1-42-induced pyroptosis in the cell model of Alzheimer's disease
Objective To investigate the inhibitory effect of 1,25(OH)2D3 on Aβ1-42-induced pyroptosis in the cell model of Alzheimer's disease(AD)and its mechanism.Methods The PC12 cells were divided into control group,model group,Caspase-1-siRNA group,NC-siRNA group,1,25(OH)2D3 group and Caspase-1-siRNA+1,25(OH)2D3 group.The PC12 cells in control group were cultured with DMEM high glucose medium.AD cell model was established with 20 μmol/L Aβ1-42.The PC12 cells in Caspase-1-siRNA group were pretreated with 50 nmol/L Caspase-1-siRNA,and then given 20 μmol/L Aβ1-42.The PC12 cells in NC-siRNA group were treated with 20 μmol/L Aβ1-42 after intervention with 50 nmol/L NC-siRNA.The PC12 cells in 1,25(OH)2D3 group were intervened with 100 nmol/L 1,25(OH)2D3,followed by 20 μmol/L Aβ1-42.The PC12 cells in Caspase-1-siRNA+1,25(OH)2D3 group were firstly pretreated with 50 nmol/L Caspase-1-siRNA,and then added 100 nmol/L 1,25(OH)2D3,followed by 20 μmol/L Aβ1-42 Cellular immunofluorescence was used to observe the expression of apoptosis-associated speck-like protein containing a CARD(ASC).Western blot was used to detect the expressions of pyroptosis-related proteins,including NOD-like receptor thermal protein domain associated protein 3(NLRP3),pro-Caspase-1,cysteinyl aspartate specific proteinase 1(Caspase-1),N-terminal of gasdermin D(GSDMD-N),interleukin-1 β(IL-1β),pro-IL-1β,interleukin-18(IL-18)and pro-IL-18.The cell membrane permeability was detected by acridine orange/ethidine bromide(AO/EB)staining.Results Compared with control group,the fluorescence intensity of ASC protein was enhanced in model group(P＜0.01),the expressions of pyroptosis pathway-related proteins were increased(P＜0.05),and the cell membrane permeability was enhanced(P＜0.01).Compared with model group,the fluorescence intensity of ASC protein was decreased in 1,25(OH)2 D3 group and Caspase-1-siRNA group(P＜0.01),the expressions of pro-Caspase-1,Caspase-1,GSDMD-N,pro-IL-1β,IL-1β,pro-IL-18 and IL-18 were downregulated(P＜0.01),and the cell membrane permeability was reduced(P＜0.01).Compared with Caspase-1-siRNA group,the expressions of GSDMD-N and IL-18 were decreased in Caspase-1-siRNA+1,25(OH)2D3 group(P＜0.01),and the cell membrane permeability was declined(P＜0.01).Conclusion Aβ1-42 could induce the cell pyroptosis in PC12 cells.1,25(OH)2D3 exerts its anti-inflammatory neuroprotective effect in PC12 cells by inhibiting Aβ1-42-induced pyroptosis,and the mechanism is closely related to the inhibition of Caspase-1.
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