The Combined Effect And Mechanism Of PLIN1 Gene Silencing And Myricetin On Lipolysis Of 3T3-L1 Adipocytes

Objective:By using the method of intervention of Myricetin(Myric)combined with silencing Perilipin1(PLIN1)gene,observe the effects on the lipolysis of 3T3-L1 adipocyte and explore its mechanism,to provide a new direction and ideas for the prevention and treatment of obesity.Methods:1.3T3-L1 preadipocytes were routinely induced and differentiated.2.Mature 3T3-L1 adipocytes were intervened and differentiated by treated with Myric concentration gradient of 0,1,5,10,50,100 ?mol/L and time gradient of 48 h,72h,respectively,to determinate the contents of intracellular TG and glycerol.3.According to the optimal concentration of Myric combined with the high efficient transfection vector of sh-RNA,the combined intervention experiment was performed on the induced mature 3T3-L1 adipocytes.It was divided into four groups:combination group(Myric+sh-RNA),transfection group(sh-RNA),Myricetin group(Myric)and blank group,to detect the contents of intracellular TG and glycerol,the oil red O staining and the morphology of lipid droplets.4.The expression levels of ATGL,HSL,p-HSL,ERK and p-ERK,MEK and p-MEK were tested by Western blot after combined intervention.The expression levels of cAMP,PKA and p-PKC were detected by ABC-ELISA method.Results:1.When the concentration of Myric was 100 ?mol/L,and the intervening time was 72 h,the content of intracellular TG was the lowest,and the glycerol content was the highest.The difference was statistically significant(P<0.05).2.After combined intervention,the TG content in Myric+sh-RNA group was lower than that in Myric group and sh-RNA group,and the glycerol content increased.The difference was statistically significant(P<0.05).In Myric+sh-RNA group,the amount of intracellular lipid droplets decreased significantly and the morphology became smaller.3.After combined intervention,the expression of PLIN1 A protein in Myric+sh-RNA group was lower than that in Myric group and sh-RNA group.The expression levels of ATGL and HSL protein was higher than that in Myric group and sh-RNA group.The difference was statistically significant(P<0.05).The expression level of p-HSL protein in Myric+sh-RNA group and Myric group were higher than that in sh-RNA group and blank group,the difference was statistically significant(P<0.05).4.After combined intervention,the expression levels of c AMP and PKA in Myric+sh-RNA group and Myric group was lower than that in sh-RNA group and blank group.The difference was statistically significant(P<0.05).The expression levels of p-PKC and the ratio of p-ERK/ERK,p-MEK/MEK in Myric+sh-RNA group and Myric group were higher than those in sh-RNA group and blank group,the difference was statistically significant(P<0.05).Conclusion:1.The optimal intervening concentration of Myric was 100 ?mol/L,and the optimal intervening time was 72 h.2.The combination of Myric and PLIN1 silencing can greatly improve theefficiency of lipolysis.3.The combination of Myric and PLIN1 silencing can decrease the expression level of PLIN1 and increase the expression level of ATGL and HSL.4.Myric may promote lipolysis through the activation of PKC-MEK/ERK signaling pathway,and further increase expression leve of p-HSL in this pathway.Meanwhile,Myric may also inhibit the activity of related factors in c AMP/PKA signaling pathway.