Contributions Of Nrf2 To Puerarin Prevent Cardiac Hypertrophy And Its Metabolic Enzymes Expression In Rats

?Background?Delaying hypertrophic course is vitally important,as it markedly reduces the risk of progressing heart failure and sudden death.Previous evidence suggested that puerarin may attenuate cardiac hypertrophy;however,the potential mechanisms were undetermined.Studies on the metabolism of puerarin and its regulatory mechanisms may help to understand its pharmacological effects.?Objective?The objective of this study is to evaluate the potential mechanisms underlying the attenuating efficacy of puerarin on cardiac hypertrophy as well as the metabolic mechanisms of puerarin involved.?Methods?Using a rat model of AAC-induced cardiac hypertrophy,we evaluated whether puerarin may attenuate cardiac hypertrophy and ventricular remodeling at a dose of 50 mg/kg/d administered from 1 week after the surgery.Catheter-based hemodynamic measurements were performed at the end of the study to further detect the changes of left ventricular systolic and diastolic function.To assess whether puerarin can activate the Nrf2-Keap1 pathway in rats with AAC surgery,we evaluated the transcription of Nrf2 mRNA by RT-PCR and the protein expression of NRF2 by Western blotting in liver and heart tissues from each group.To further determine whether puerarin could activate Nrf2 in the pathophysiological process of cardiac hypertrophy,we examined its effects on hypertrophic changes in NRCMs due to Ang II.The role of Nrf2 in puerarin's anti-hypertrophy property was further confirmed via siRNA.We detected Ugt1a1 and Ugt1A9 expression using RT-PCR and Western blotting in the liver and heart tissues from the rats in each group.We further identified the role of Nrf2 in puerarin-stimulated UGT1A1 and 1A9 upregulation with Nrf2 siRNA in AngII-treated cardiomyocytes.To demonstrate that Nrf2 may induce the transcription of Ugt1a1 and Ugt1A9 by binding to the promoters of the genes,we performed ChIP-qPCR in control and puerarin-treated cardiomyocytes.?Results?We confirmed that puerarin(50 mg/kg/d)significantly attenuated cardiac hypertrophy,upregulated NRF2 and decreased KEAP1 protein levels in the myocardium.Moreover,puerarin significantly promoted Nrf2 translocation to the nucleus in parallel with the upregulated downstream proteins including HO1,GSTP1 and NQO1.Similar results were obtained in neonatal rat cardiomyocytes(NRCMs)treated with angiotensin II(1 ?M)and puerarin(100 ?M),while the silencing of Nrf2 abolished the antihypertrophic effects of puerarin.The mRNA and protein levels of Ugt1a1 and 1a9,enzymes for puerarin metabolism,were significantly increased in the liver and heart tissues of AAC rats and angiotensin II-treated NRCMs.Interestingly,the silencing of Nrf2 attenuated the puerarin-induced upregulation of UGT1A1 and 1A9.The results of chromatin immunoprecipitation–qPCR indicated that the binding of Nrf2 to the promoter region of Ugt1a1 or Ugt1a9 was significantly enhanced in puerarin-treated cardiomyocytes.?Conclusion?These results suggest that Nrf2 is the key regulator of antihypertrophic effects and the upregulation of the metabolic enzymes UGT1A1 and 1A9 of puerarin.This autoregulatory circuits between puerarin and Nrf2-induced UGT1A1 and 1A9 are beneficial to attenuate adverse effects and maintain the pharmacological effects of puerarin.