Mechanism Of Puerarin Inhibiting Cardiac Fibrosis By Regulating Metabolic Feedback Loop In Pressure Overload Rats

Background:Ventricular remodeling is a risk factor of cardiovascular death.Cardiac fibrosis takes part in the pathological process of ventricular remodeling.Puerarin could inhibit cardiac fibrosis.But the mechanism of it is not clear.AIM:To exploit the possible pathway between the metabolic enzymes modulated by puerarin and cardiac fibrosis focusing on the feedback loop of UDP-glucuronosyltransferase(UGT)substrate and ligand-activated transcription factors(LATFs).Method:Rats with pressure overload were developed by abdominal aotic banding(AB).Echocardiograohy was used to detect cardiac strcture and cardiac function in SD rats.Right carotid artery intubation was used to detect hemodynamic parameters in SD rats.H&E staining and masson staining were used to detect cardiac fibrosis in SD rats.Immunohistchemical and western blot were used to check the expression of collagen ?,collagen III and Nrf2 in the heart of rats.In vitro,Neonatal SD rats cardiac fibroblasts(NRCF)were treated with Ang ?so as to eastablish cell model of cardiac fibrosis.Cardiac fibroblast proliferation was checked by CCK-8.The expression of Nrf2,UGT1A1,collagen ?,collagen ?,P38 MAPK,and p-P38 MAPK were detected by western blot.The expression of Nrf2 also detected by immunofluorescence.Reactive oxygen species(ROS)were checked by fluorescence microscope and fluorometry.ChIP was used to detect the effect of puerarin which contributed Nrf2 to UGT1A1 expression.Results:?Puerarin inhibited cardiac hypertrophy and cardiac fibrosis in pressure overload rats and up-rugulate the expression of Nrf2 in heart,manifesting as decreasing cardiac weight,ventricular wall thickness,and collagen deposion(mainly collagen ?and collagen ?)and increasing expression of Nrf2.? Puerarin inhibited the proliferation of NRCF induced by Ang ? and reduced the expression of collagen ?and collagen ?.Puerarin can activate Nrf2 by increasing the expression of Nrf2 and transferring Nrf2 from cytoplasm to nucleus,and reduce the production of ROS,so as to reduce the expression of collagen ? and collagen ?.The same results were observed in the cell experiment using siRNA Nrf2,agonists and inhibitors of Nrf2.?The protein level of UGT1A1 was dramatically upregulated by puerarin or tBHQ treatment in a time-dependent manner,but opposite when treated with brusatol.The ChIP results showed that puerarin significantly increased Nrf2-associated UGT1A1 promoter activity.?Puerarin significantly inhibited the phosphorylation of P38 MAPK induced by Ang ?.SB203580(an inhibitor of P38 MAPK)inhibited the proliferation of NRCF induced by Ang ?,reduced production of ROS,and the expression of collagen ? and collagen ? in NRCF induced by Ang ?.All these effects were independent of Nrf2.Conclusion:Puerarin prevents cardiac fibrosis via activation of Nrf2 and inactivation of p38-MAPK.Nrf2 is the key regulator of anti-fibrotic effects and upregulates metabolic enzymes UGT1A1.Autoregulatory circuits between puerarin and Nrf2-regulated UGT1A1 attenuates side effects associated with treatment,but it does not weaken puerarin's pharmacological effects.