Novel insights into the effect of Xiaoyao san on corticosterone-induced hepatic steatosis:inhibition of glucocorticoid receptor/perilipin-2 signaling pathway

Objective:Xiaoyao san(XYS)is a classic traditional Chinese medicinal formula.It has been clinically administered to regulate liver function.However,its mechanisms in glucocorticoid-induced hepatic steatosis are unknown.This study aimed to investigate whether XYS protects against corticosterone(CORT)-induced hepatic steatosis,and to explore its mechanism.Methods:High-fat diet mice induced with hepatic steatosis by 2mg/kg CORT were administered 2.56 g/kg or 5.12 g/kg XYS daily for 7 weeks.The effects of XYS on hepatic steatosis in mice were evaluated by H&E and Oil Red O staining and by measuring their plasma lipids(triglyceride,total cholesterol,and free fatty acids).The mechanism of XYS against hepatic steatosis was investigated by network pharmacology,immunohistochemistry,western blotting,and gain-of-function/loss-offunction experiments.Results:XYS alleviated CORT-induced steatosis,decreased plasma lipids,and inhibited glucocorticoid receptor(GR)activation in the liver.Network pharmacology data indicated that XYS may have mitigated hepatic steatosis via GR which mediated adipose differentiation-related protein(ADFP).Gain-of-function/loss-of-function experiments in vitro confirmed that GR positively regulated ADFP expression.Conclusions:XYS ameliorated CORT-induced hepatic steatosis by downregulating the GR/ADFP axis and inhibiting lipid metabolism.Our studies implicate that XYS is promising as a therapy for CORT-induced hepatic steatosis,and lay the foundation for designing novel prophylactic and therapeutic strategies on CORT-induced hepatic steatosis.

Active compounds in RenShenJian decoction ameliorate insulin resistance in vitro

Background:RenShenJian decoction,a combination of Pueraria lobata(Willd.)Ohwi and Panax ginseng C.A.Mey,has been used in China since the Song Dynasty(960-1279 C.E.)to relieve symptoms of diabetes mellitus.However,the key compounds in RenShenJian that ameliorate insulin resistance remain unclear.This study identified the anti-diabetic compounds in RenShenJian by rescuing the decreased function of adenosine 5’-monophosphate-activated protein kinase(AMPK),sirtuin 3(SIRT3),or glucose transporter isoform 4(GLUT4).Methods:After streptozotocin-induced diabetic mice were treated with RenShenJian,fasting blood glucose levels and protein expression of SIRT3,p-AMPK,and AMPK were determined.Compounds from RenShenJian in plasma were monitored using multiple responses by liquid chromatography-mass spectrometry.Additionally,two insulin-resistant cell models were incubated with compounds identified in RenShenJian in the blood.Glucose uptake was determined using the fluorescent analog 2-(N-(7-nitrobenz-2-oxa-1,3-dia-xol-4-yl)amino)-2-deoxyglucose.Protein expression levels of p-AMPK,AMPK,SIRT3,and GLUT4 were detected by western blotting.Results:RenShenJian decreased FBG levels and upregulated SIRT3 expression and AMPK phosphorylation in diabetic mice.Thirteen RenShenJian extracts were identified in the blood,11 of which increased the ratios of 2-(N-(7-nitrobenz-2-oxa-1,3-dia-xol 4-yl)amino)-2-deoxyglucose uptake in two insulin-resistant cell models.Nine extracts increased AMPK phosphorylation,nine increased SIRT3 expression,and six elevated GLUT4 expression in palmitate-induced HepG2 cells.Five extracts-puerarin,puerarin 6″-O-xyloside,genistein,ginsenoside Rb1,and ginsenoside Rd-simultaneously activated AMPK,SIRT3 and GLUT4.Conclusion:A series of compounds in RenShenJian that target AMPK,SIRT3,and/or GLUT4 was confirmed and indicate the chemical material basis of amelioration of insulin resistance by RenShenJian.