Effect Of Celastrol On Degranulation Of Substance P Pre-treated Mast Cell And Mechanism Of PI3K/AKT/GSK3? Pathway

Objective This study was designed to discuss the mechanism of pre-treated mast cell degranulation process,observe the effects of Celastrol inhibition induced by substance P in P815 sensitized mast cell degranulation and the expression of key signaling gene on PI3K/AKT/GSK3? pathway.The aim of the study was to provide a certain theoretical and experimental basis for the clinical application of Celastrol in anti-inflammatory and anti-allergy.Methods First,cell viability of substance P pre-treated cells was detected.Then based on the first experiment results,the optimal concentration and intervention time of model group was selected to detect the cell viability of of Celastrol on sensitized mast cells.q-PCR,and the relative expression of key genes in PI3K/AKT/GSK3? signaling pathway.After that,the expression of AKT2 and GSK3 was detected by flow cytometry.Results 1.cell viability:50-150?M substance P exposure induced a dose-and time-dependent reduction of cell viability(P<0.05).Treatment with Celastrol within 2?M and 1h cell did not cause obvious difference.The concentration of Celastrol was between 2?M-20pM,and the survival rate of the cells was significantly a dose-and time-dependent reduction of cell viability after more than lh(P<0.05).2?M Celastrol group after 1h and 1.5h on sensitized mast cell survival rate was restored to 83.51%?82.4%.2.q-PCR:Compared with the normal group,Gnai1,Pik3r3,Akt2,Fgr,Arrb2 of substance P model group were upregulated,and the level of Gnai1?Pik3r3?Akt2?Fgr?Arrb2 of Celastrol group with the normal group and model group were downregulated(P<0.05).3.Compared with the normal group,Adrbk2of substance P model group were increased,and the level of Adrbk2 of Celastrol group with the normal group and model group were significantly increased(P<0.05).4.Compared with the normal group,Gsk3p,Ccr6 of substance P model group were downregulated,and the level of Gsk3??Ccr6 of Celastrol group with the normal group and model group were upregulated(P<0.05).5.Compared with the normal group,Ccl24 of substance P model group were downregulated,and the level of Ccl24 of Celastrol group with the normal group and modelgroup were obviously downregulated(P<0.05).6.Flow cytometry text:Compared with the control group substance P group protein expression of Akt2 and Gsk3? was obviously increased,after Celastrol treatment,Akt2 were obviously downregulated but Gsk3? was upregulated.Conclusion 1.The cell viability of Celastrol on the degranulation of mast cells pre-treated by substance P was inhibited.2.The mechanism of substance P induced on mast cells is related to the expression of PI3K/AKT/GSK3? signaling pathway.3 Celastrol plays the guiding inhibition role through PI3K/AKT/GSK3?signaling pathway by chemokine,GPCR and main target genes of Pik3r3,Akt2,Gsk3?.