The Effects Of Pentamethylquercetin On The Cardiac Remodeling Induced By Pressure Overload In Rat And Its Underlying Mechanism

Cardiac hypertrophy is an adaptive respose to mechanical or neurohormonal stimulation. Although initially, cardiac hypertrophy plays role as a compensatory response that tries to normalize cardiac pump function. Prolonged hypertrophy may eventually lead to heart failure and even sudden death. Clinical studies have shown that left ventricular hypertrophy is an independent risk for myocardial ischemia, arrhythmia and sudden death. Therefore, finding an effective way to prevent or reverse the cardiac hypertrophy is essential.Polymethoxy flavonoids (PMFs) are commonly found in various plants. It has been reported that PMFs play important roles in a number of biological functions. They have been reported to possess anticancer, antimutagenic activity and antiinflammatory properties. The PMFs also have been reported to positively regulate adiponectin expresion and secretion in vitro. However, there is no breakthrough in the extraction technology of natural polymethoxyl flavonoids up to now. The yield of natural polymethoxyl flavonoids is limited and the price is very high, which hindered the research.We carried out the methylation of quercetin transformation, to obtain high purity pentamethylquercetin (PMQ) and overcome the bottleneck of drug sources. Our previous experimental studies have demonstrated that PMQ could lose weight, improve type 2 diabetes, abnormal glucose and lipid metabolism of animals, up-regulate the PPARs expression in adipocytes, up-regulate the expression of PPARy-dependent adiponectin expression and secretion, inhibit of inflammatory cytokines TNF-a and IL-6 expression. The effects of PMQ in cardiovascular include:relaxing blood vessels, improvement endothelial function, improvement myocardial ischemic reperfusion injury and anti-oxidative damage.In this study, the effects of PMQ on cardiac hypertrophy and its underlying mechanisms were investigated. Part I The effects of PMQ on pressure overload cardiac hypertrophy in ratsObjective:To study the influence of PMQ on pressure overload cardiac hypertrophy in rats.Methods:Pressure overload cardiac hypertrophy was induced by abdominal aorta constriction (AAC) in rats. The day before surgery treated with PMQ (2.5,5, and 10 mg·kg·day) or the same volume of solvent for consecutive 7 weeks. Monitor the weight and blood pressure weekly. At the end of the experiment, general view of the heart were observed, the heart, lung, kindney weight and its organ index were measured. At last, hemodynamic, morphology, histological and biochemical measurements were performed.Results:(1) The SBP was significantly increased in model group (P<0.01 vs sham-operated group). PMQ treatment of 2.5,5 and 10 mg/kg could significantly decrease the SBP (P<0.01, P<0.01, P<0.05 vs AAC+vehicle group). (2) Cardiac hypertrophy model rats demonstrated an significant systolic and diastolic dysfunction. PMQ could improve the cardiac function in a dose dependent manner. (3) The cardiac index was higher in cardiac hypertrophy model group compared with sham-operated group (P<0.01), howerer, the cardiac index were lower in PMQ treatment of 5 and 10 mg/kg groups compared with the vehicle group (P<0.01, P<0.01); (4) Myofiber disarray, necrotic area increased and myocyte cross-sectional area increased were obseved in cardiac hypertrophy model group. PMQ treatment of all doses could reverse these pathological changes. (5) The Lung index and BNP mRNA expression were significantly increased in model group (P<0.01, P<0.01 vs Sham-operated group). PMQ treatment could reduce the Lung index and BNP mRNA expression in a dose dependent manner.Conclusions:PMQ exerted the anti-cardiac hypertrophy effects through decrease the blood pressure and HW/BW ratio, improvement cardiac function and myocardial pathological changes. At the same time, PMQ Prevented the progression of hypertrophy to cardiac failure through decease the LW/BW ratio and BNP mRNA expression. Part II The effects of PMQ on pressure overload cardiac fibrosis in ratsObjective：To study the influence of PMQ on pressure overload cardiac fibrosis in rats.Methods:Using the picrosirius red staining and Masson trichrome staining to observe the myocardial interstitial fibrosis and perivascular fibrosis, and calculate the total myocardial interstitial collagen content. By using RT-PCR method to detect left ventricular collagen type I and III (Collagen I/III) mRNA expression. Results:The heart from pressure overload rats showed severe increased perivascular fibrosis (Masson’s trichrome staining, x200) and interstitial fibrosis (picrosirius red staining, x200). Quantitative image analyses indicated a significant accumulation of interstitial collagen in cardiac hypertrophy model group(P<0.01 vs sham-operated group). It indicated that the increased pressure overload induced by abodominal aorta constriction could lead to obviously myocardial fibrosis. The collagen I/III content reflecting the level of collagen synthesis in cardiac tissue and the collagen I/III mRNA expression were significantly increased(P<0.01, P<0.01 vs sham-operated group). PMQ could improve collagen dysplasia and disordered. At the same time, PMQ could reduce the myocardial interstitial collagen content and collagen I/III mRNA expression in dose dependent manner.Conclusions:PMQ had the inhibitory effect on cardiac fibrosis induced by abdominal aorta constriction. And also, PMQ could improve the ventricular remodeling through inhibition the fibrosis. PartⅢThe effects of PMQ on TNFαIL-6, MCAD、LCAD and PPARα、PPARβmRNA expression in pressure overload rat heartsObjective:To study the influence of PMQ on TNFα、IL-6, MCAD LCAD and PPARα、PPARP mRNA expression in pressure overload rat hearts. Methods:Pressure overload cardiac hypertrophy was induced by abdominal aorta constriction in rats. Application of RT-PCR method to detect the effects of PMQ on myocardial tissue TNFα、IL-6, MCAD、LCAD and PPARα、PPARβmRNA expression.Results:(1) we found that expression of TNFa and IL-6 as pro-inflammation factors were up-regulated in AAC group rats (P<0.01, P<0.01 vs sham-operated group) and this up-regulation of inflammation factors could be inhibited by PMQ 2.5mg/Kg treatment (P<0.05, P<0.01 vs AAC+vehicle group). (2) The fatty acid oxidation rate-limiting enzyme (MCAD and LCAD) mRNA expression were decreased (P<0.01, P<0.01 vs sham-operated group), indicated that fatty acid oxidation was limited. PMQ could reverse the MCAD and LCAD mRNA expression. (3) We found that PMQ treatment of 10mg/kg enhanced the expression of PPAR-a andβwhich were both reduced in AAC group rats (P<0.01, P<0.05 vs AAC+vehicle group).Conclusions:In pressure overload cardiac hypertrophy model, the inflammatory response was increased, fatty acid oxidation and PPARa and PPARβmRNA expression were down-regulated. PMQ treatment could reduce the inflammatory response caused by pressure overload, reverse the fatty acid oxidation and up-regalate the PPARa and PPARβmRNA expression. Part IV The effects of PMQ on hypertrophy of neonatal rat cardiomyocytes induced by ET-1Objective:To observe the effects of PMQ on hypertrophy of neonatal rat cardiomyocytes induced by ET-1.Methods:Establish the neonatal rat cardiomyocytes hypertrophy model by ET-1 (0.1μM) and pretreat with PMQ 0.3,1,3,10μM or the same volume Sovent. Extraction of total cellular RNA, using RT-PCR method to detect brain natriuretic peptide (BNP), pro-inflammatory cytokines (TNF-a and IL-6), fatty acid oxidation rate-limiting enzyme (MCAD and LCAD) and peroxide proliferator-activated receptor (PPARa and PPARβ) mRNA expression.Results:TNF-a, IL-6 and BNP mRNA levels were significantly increased (P<0.01, P<0.01, P<0.01 vs control) by ET-1 treatment and these were inhibited by PMQ treatment. The MCAD, LCAD and PPARa, PPARβmRNA levels were significantly decreased by ET-1 treatment (P<0.05, P<0.01, P<0.01, P<0.01 vs control). PMQ could up-regulate the fatty acid oxidation rate-limiting enzyme and PPARa, PPARβmRNA expression. The results were consistent in vivo and in vitro.Conclusions:In ET-linduced neonatal rat cardiomyocytes hypertrophy model, also observed the protective effects of PMQ. PMQ could reduce inflammatory response and increase PPARa and PPARP mRNA expression.