The Role Of Gallic Acid In Angiotensin Ⅱ-induced Hypertension And The Underlying Molecular Mechanism

BackgroundHypertension is a major risk factor for damageof its target organs including theheart,brain,kidneys and blood vessels.It is now believed that increased sympathetic-nervous system activity,activation of the renin-angiotensin-aldosterone system,oxidative stress,impaired endothelial cell function,and vascular remodeling play critical roles in the development of hypertension.Gallic acid(GA)is a polyphenolic plant compound,which has anti-inflammatory and antioxidant effects.Our recent study revealed that GA exerts protective effects on pressure overload-induced cardiac hypertrophy and dysfunction.However,the role of GA in angiotensin Ⅱ(AngⅡ)-induced hypertension and vascular remodeling remains unknown.AimsTo elucidate the role of GA in AngⅡ-induced hypertension and vascular remodeling and the molecular mechanism,and to will provide newexperimental evidence that GA may be a potential candidate for the treatment of hypertension..Methods:1.Animal ModelC57BL/6 mice(male,8 – 12 weeks)were selected and randomly divided into four groups,the saline control group,the AngⅡ treatment group,the AngⅡ+GA(5 mg/kg BW)group,and the AngⅡ+GA(20 mg/kg BW)group.AngⅡ(490 ng/kg/min)was infused with the osmotic mini-pumps subcutaneously AngⅡ(490 ng/kg/min)or saline for 2 weeks to establish an AngⅡ-induced hypertension model.2.Methods(1)Blood Pressure measurement:Systolic blood pressure was measured using a tail-cuff system by BP-2010.The systolic blood pressure and heart rate of mice in each group were monitored every other day after implante the pumps.We measured each mouse no less than 8-12 times and calculated the average value and observe the dynamic change trend of blood pressure.(2)Histological and functional examinations:The morphological changes of aortas was eamined with Hematoxylin & Eosin(H&E)staining;was used to examine Aortic fibrosis was detected by Masson staining;aortic infiltration of macrophages was evaluated by immunohistochemical staining;dihydroethidium(DHE)staining was used to analyze the ROS level of aortas was measured with dihydroethidium(DHE)staining.Aortic vasodilatory function was analyzed with aortic ring assay.Nitric oxide(NO)level in aortas was also measured.(3)Analysis of the mRNA and protein levels int ehe aortas:The expression levels of markers related to inflammation,fibrosis and oxidative stress were detected by real-time quantitative PCR analysis.The protein expression levels of endothelial nitric oxide synthase(eNOS)and the immunoproteasome catalytic subunits were detected by Western blot analysis.3.Statistical analysisAll results are presented as mean ± standard error of the mean(SEM).The student t test was used to compare the significant difference between two groups in normal distribution.P<0.05 was considered statistically significant.Results1.GA administration AngⅡ-induced hypertension significantlyAngⅡ infusion for 2 weeks significantly increased SBP in mice,whereas this increase was dose-dependently reduced by GA(5 or 20 mg/kg BW).2.GA administration inhibits AngⅡ-induced aortic thickness,fibrosis,inflammation and oxidative stressAngⅡ infusion significantly increased aortic wall thickness,fibrotic area,accumulation of Mac-2-positive macrophages and ROS level as well as the mRNA levels of fibrotic,proinflammatory and oxidative stress gene markers(α-SMA,Collagen I/ⅡI,IL-1β,IL-6,TNF-α,MCP-1,NOX1,NOX2,NOX4 and P22phox),but these effctswere dose-dependently reduced in GA-treated mice.3.GA treatment improves AngⅡ-induced vascular dysfunction and increases eNOS protein expression and NO levelAngⅡ infusion markedly impaired vascular endothelial diastolic function,reduced eNOS protein expression,aortic and serum NO levels,and this effect was dose-dependently reversed in GA-treated mice.4.GA treatment attenuates AngⅡ-induced increase in the activity and expression of the immunoproteasome catalytic subunitsAngⅡ infusion for 2 weeks significantly induced increase of the activity of the proteasome activities and the expression of the immunoproteasome catalytic subunits in the aorta,and the increase was abolished by GA treatment.ConclusionsThis study identifies that GA plays a critical role in AngⅡ-infused hypertension and vascular remodeling.Mechanistically,GA treatment inhibits AngⅡ-induced upregulation of the immunoproteasome catalytic subunits(β2i and β5i)expression and activities,,which suppresses degradation of eNOS,leading to increase of NO level and improvement of arterial function.These results indicate that GA may be a potential therapeutic agent for hypertension and vascular dysfunction.