Lipid Metabolic Effect And Mechanism Of Five Flavonoids In Fat-accumulation HepG2 Cells

Objective:Nonalcoholic fatty liver disease(NAFLD)is the most common chronic liver disease in the world,and it is the manifestation of metabolic syndrome(MetS)in the liver.NAFLD refers to a condition in which more than 5%of liver cells undergo fat-accumulation and degeneration in the absence of excessive intake of alcohol.In this study,HepG2 was treated with sodium oleate and sodium palmitate solution to establish a cell model of fat-accumulation HepG2 cells.To study the regulatory effect of quercetin,apigenin,luteolin,chrysin and vitexin on lipid metabolism of the fat-accumulation HepG2 cells,and to explore the possible mechanism.Methods:1.The effect of different concentrations of quercetin,apigenin,luteolin,chrysin,vitexin on the survival rates of HepG2 cells were determined by CCK-8 method,and the optimal drug concentrations were selected.CCK-8 method was used to determine different concentrations of sodium oleate-sodium palmitate solution on the survival rates of HepG2 cells,to choose the optimal modeling concentration of the fat-accumulation HepG2 cell model.2.The effect of quercetin,apigenin,luteolin,chrysin,vitexin on the content of triglyceride and cholesterol in fat-accumulation HepG2 cells was determined,and the effect of five flavonoids on lipid metabolism was compared.Stronger flavonoids were used for subsequent experiments.3.Oil red O staining was used to observe the effect of quercetin and apigenin on lipid droplet formation in fat-accumulation HepG2 cells.To determine the antioxidant activity of quercetin and apigenin by measured the contents of superoxide dismutase(SOD),malondialdehyde(MDA)and glutathione(GSH)in fat-accumulation HepG2 cells.4.The effect of quercetin and apigenin on the mRNA and protein expression levels of SREBP-1c,LDLR and PCSK9 in fat-accumulation HepG2 cells was detected by qRT-PCR and WB assay.The mechanism of quercetin and apigenin on lipid metabolism was explored.Results:1.When the concentrations of five flavonoids were 2.5～10μg/mL,the survival rates of HepG2 cells were not significantly different from that of the blank control group(P>0.05),so the concentrations of these three drugs were selected,which were 2.5μg/mL,5μg/mL,10μg/mL.The survival rates of HepG2 cells in the concentrations range of 0.2-0.1 to 0.8-0.4 mM of sodium oleate-sodium palmitate were not significantly different from that of the blank control group(P>0.05),so 0.8-0.4 mM was selected as the modeling concentration of the fat-accumulation HepG2 cell model.2.Among the five flavonoids,quercetin and apigenin were more significant,and the concentrations of triglyceride and cholesterol were significantly lower at 5μg/mL and 10μg/mL when they were compared with the model control group,the difference was statistically significant(P<0.05).3.Compared with the model group,quercetin and apigenin can reduce the accumulation of lipid droplets in fat-accumulation HepG2 cells.When the concentrations of quercetin and apigenin were 2.5μg/mL,5μg/mL and 10μg/mL,the levels of SOD and GSH were significantly up-regulated compared with the model control group,the difference was statistically significant(P<0.05);and the MDA was significantly down-regulated,the difference was statistically significant(P<0.05).The results showed that quercetin and apigenin have good antioxidant activity,they can effectively inhibit the oxidative stress level of fat-accumulation HepG2 cells.4.Compared with the model control group,quercetin and apigenin significantly down-regulated the mRNA and protein expression levels of SREBP-1c and PCSK9 at 5μg/mL and 10μg/mL,and the difference was statistically significant(P<0.05).The mRNA and protein expression levels of LDLR were up-regulated,and the difference was statistically significant(P<0.05).The regulation mechanism of quercetin and apigenin on lipid metabolism in fat-accumulation HepG2 cells was revealed.Conclusion:In summary,all the five flavonoids can down-regulate the triglyceride and cholesterol levels in fat-accumulation HepG2 cells to varying degrees.Among them,quercetin and apigenin are the more significant ones.These two flavonoids can reduce fat droplet accumulation in fat-deposited HepG2 cells,can effectively inhibit the oxidative stress level of fat-accumulation HepG2 cells,and by adjusting the mRNA and protein expression levels of SREBP-1c,LDLR and PCSK9 to regulating lipid metabolism.