Effects Of The Flavonoid Glycosides And Terpenoids In Artemisia Lavandulaefolia On The Expression Of AMPK And LPIN1 In HepG2 Cells

Objective To investigate the effects of four kinds of Flavonoid glycosides,including luteolin,naringenin,quercetin,apigenin,and Terpenoids ursolic acid in Artemisia lavandulaefolia on the expression of AMPK and LPIN1 in HepG2 cells,and to prove its active lipid-lowering ingredients at the molecular level.Method Using MTT method,the effects of studied four kinds of Flavonoid glycosides and Terpenoids ursolic acid in Artemisia lavandulaefolia on the proliferation of HepG2 cell were measured at different drug concentrations and action time.And the best drug concentration and action time were selected.Using flow cytometry,apoptosis of HepG2 cells induced by quercetin and apigenin was detected with 5-fluorouracil as a positive control.Based on the screened optimal concentration and action time,cultured cells were treated with drugs,and total RNA and protein were extracted from control group and experimental groups,respectively.mRNA levels of LPIN1,AMPK?1,AMPK?2 were measured by RT-PCR,while the protein expression levels of LPIN1,AMPK?1,AMPK?2 and p-AMPK were examined by Western blot.Results(1)All of the studied compounds could inhibit the proliferation of HepG2 cells.Drug concentration of 0.01mg/ml for 24 hours was selected as the most suitable condition,with an inhibition rate being 15%-25%.(2)Quercetin and apigenin could induce the apoptosis of HepG2 cells.For two drugs,the apoptosis rates at early stage were(9.10±0.61)%,and(7.47±1.51)%,respectively,while the apoptosis rates at later stage were(40.03 ± 3.19)%,and(38.07 ± 1.75)%,respectively.The apoptosis rates in experimental group were significantly increased than in control group,of which the apoptosis rates at early and later stages were(4.80±1.1)% and(13.7±2.87)%,respectively.Contrast difference have statistical significance(P<0.05).(3)Results of the RT-PCR indicated that no significant differences in the expression of AMPK?1 mRNA were found between the five experimental groups and control group(P>0.05).But the expression of AMPK?2 mRNA in quercetin,apigenin and ursolic acid groups was significantly increased than in control group(P<0.05).Compared withcontrol group,the expression of LPIN1 mRNA was significantly decreased in five experimental groups(P<0.01).(4)Results of the Western blot that no significant differences in the expression of AMPK?1 were found between the five experimental groups and control group(P>0.05).The expression of AMPK?2 in five experimental groups was significantly increased than in control group(P<0.05).The expression of LPIN1 in five experimental groups was significantly decreased than in control group(P<0.01).The expression of p-AMPK in five experimental groups was significantly increased than in control group,which suggested that Flavonoid glycosides and Terpenoids extreated from Artemisia lavandulaefolia by ethyl acetate could activate AMPK by phosphorylation.Conclusions(1)All of the five studied compounds extracted from Artemisia lavandulaefolia could inhibit the proliferation of HepG2 cells.The most suitable condition is at the concentration of 0.01mg/ml for 24 hours,with an inhibition rate being 15%-25%.(2)Quercetin and Apigenin can induce the apoptosis of HepG2 cells at both early and later stages.(3)In HepG2 cells,all of the five studied compounds extracted from Artemisia lavandulaefolia have no obvious effects on the expression of AMPK?1 mRNA and protein.Naringenin,quercetin and ursolic acid can increase the expression levels of AMPK?2 mRNA.All of the five studied compounds can increase the expression of AMPK?2 protein,and All of the five studied compounds can decrease the expression levels of LP1N1 mRNA and protein.All of the five studied compounds can increase the expression of p-AMPK protein,suggesting that they all can activate AMPK by phosphorylation.