Atorvastatin Slows the Progression of Cardiac Remodeling in Mice with Pressure Overload and Inhibits Epidermal Growth Factor Receptor Activation

335 Hypertens Res Vol.31 (2008) No.2 p.335-344 Original Article Atorvastatin Slows the Progression of Cardiac Remodeling in Mice with Pressure Overload and Inhibits Epidermal Growth Factor Receptor Activation Yulin LIAO1), Hui ZHAO2), Akiko OGAI1), Hisakazu KATO2), Masanori ASAKURA1), Jiyoong KIM1), Hiroshi ASANUMA1), Tetsuo MINAMINO2), Seiji TAKASHIMA2), and Masafumi KITAKAZE1) The aim of this study was to investigate whether atorvastatin inhibits epidermal growth factor receptor (EGFR) activation in cardiomyocytes in vitro and slows the progression of cardiac remodeling induced by pressure overload in mice. Either atorvastatin (5 mg/kg/day) or vehicle was orally administered to male C57BL/6J mice with transverse aortic constriction (TAC). Physiological parameters were obtained by echocardiography or left ventricular (LV) catheterization, and morphological and molecular parameters of the heart were also examined. Furthermore, cultured neonatal rat cardiomyocytes were studied to clarify the underlying mechanisms. Four weeks after TAC, atorvastatin reduced the heart/body weight and lung/body weight ratios (8.69 ± 0.38 to 6.45 ± 0.31 mg/g (p < 0.001) and 10.89 ± 0.68 to 6.61 ± 0.39 mg/g (p < 0.01) in TAC mice with and without atorvastatin, respectively). Decrease of LV end-diastolic pressure and the time constant of relaxation, increased fractional shortening, downregulation of a disintegrin and metalloproteinase (ADAM)12, ADAM17 and heparin-binding epidermal growth factor genes, and reduction of the activity of EGFR and extracellular signal–regulated kinase (ERK) were observed in the atorvastatin group. Phenylephrine-induced protein synthesis, phosphorylation of EGFR, and activation of ERK in neonatal rat cardiomyocytes were all inhibited by atorvastatin. These findings indicated that atorvastatin ameliorates cardiac remodeling in mice with pressure overload, and its actions are associated with inhibition of the EGFR signaling pathway. (Hypertens Res 2008; 31: 335–344) Key Words: statins, epidermal growth factor receptor, heart failure, hypertrophy, extracellular signal–regulated kinase Introduction Although substantial evidence obtained by various clinical trials has demonstrated the efficacy of β-blockers, angio- tensin-converting enzyme inhibitors, angiotensin receptor blockers, aldosterone antagonists, and vasodilators for treatment of chronic heart failure (CHF) (1), the mortality and morbidity of this serious condition remain high. Therefore, investigation of novel treatments to improve the prognosis of From the 1)Cardiovascular Division of Medicine, National Cardiovascular Center, Suita, Japan; and 2)Department of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Suita, Japan. This work was supported by Grants (H13-Genome-011 and H13-21seiki (seikatsu)-23) from the Japanese Ministry of Health, Labour and Welfare. One of the authors (Y.L.) was supported by a grant from the Japan Society for the Promotion of Science (P05228). Address for Reprints: Masafumi Kitakaze, M.D., Ph.D., Cardiovascular Division of Medicine, National Cardiovascular Center, 5–7–1 Fujishirodai, Suita 565–8565, Japan. E-mail: Received April 5, 2007; Accepted in revised form September 2, 2007. 336 Hypertens Res Vol. 31, No. 2 (2008) Fig. 1. Effect of atorvastatin (Ator) on cardiac hypertrophy and fibrosis in mice with pressure overload. A: Representative pictures of the whole heart. B: Cardiomyocyte cross-sectional surface area (hematoxylin and eosin stain [H&E stain]). C: Longaxis view of cardiomyocytes (H&E stain). Myocardial fibrosis (D) and perivascular fibrosis (E) are revealed by Azan staining. The heart weight/body weight ratio (HW/BW, ANOVA p< 0.0001) (F) and cardiomyocyte cross-sectional surface area (ANOVA p< 0.0001) (G) were significantly reduced in atorvastatin-treated mice. Myocardial fibrosis (ANOVA p< 0.0001) (H) and perivascular fibrosis (ANOVA p< 0.003) (I) were also inhibited by atorvastatin. *p< 0.01, †p< 0.05 vs. TAC by post-hoc test. In F, the number of mice is 12, 5, 12 and 11 for the sham, sham + Ator, TAC, and TAC + Ator groups, respectively. In G–I, 3–5 hearts from each group were used to obtain data. Bar: 20 μm. CHF is an area of intense activity. Recent clinical studies performed by us as well as others have shown that hydroxymethylglutaryl-CoA (HMG-CoA) reductase inhibitors (statins) improve cardiac remodeling and survival in patients with either ischemic or non-ischemic CHF (2–5), suggesting that statin therapy may be a potential new approach for CHF. Randomized double-blind placebo-controlled trials that are still ongoing (6, 7) may eventually provide firm evidence about this issue. In the meantime, well-designed experimental studies will also be helpful to clarify whether statins are beneficial for both systolic and diastolic heart failure, as well as to explore the underlying mechanisms. Theoretically, several of the many actions of statins may contribute to the improvement of cardiac remodeling. Our previous study demonstrated that myocardial hypertrophy can be induced via activation of matrix metalloproteinases (MMPs), which is followed by the subsequent release of heparin binding–epidermal growth factor (HB-EGF) and phosphorylation of the epidermal growth factor receptor (EGFR), and we have shown that an MMP inhibitor can ameliorate Liao et al: Atorvastatin Inhibits Cardiac Remodeling 337 Table 1. Echocardiographic Findings at 4 Weeks after TAC or Sham Operation Parameters Sham (n=10) Sham+Ator (n=5) TAC (n=11) TAC+Ator (n=11) ANOVA p value LVEDd (mm) LVPWd (mm) LVESd (mm) LVFS (%) LVEF (%) HR (bpm) 2.85±0.05* 0.65±0.01*,† 1.20±0.06* 58±2* 89±1* 543±11 2.89±0.08# 0.61±0.02*,† 1.39±0.12* 52±3* 89±3* 540±10 3.22±0.07 0.92±0.02 2.10±0.1 35±2 65±3 490±27 2.89±0.05* 0.74±0.02* 1.15±0.09* 60±3* 89±2* 527±28 0.004 <0.0001 <0.0001 <0.0001 <0.0001 0.492 TAC, transverse aortic constriction; Ator, atorvastatin; LVEDd, left ventricular end-diastolic dimension; LVPWd, left ventricular diastolic posterior wall thickness; LVESd, left ventricular end-systolic dimension; LVFS, left ventricular fractional shortening; LVEF, left ventricular ejection fraction; HR (bpm), heart rate (beats per minute). *p<0.01, #p<0.05 compared with TAC, †p<0.01 vs. TAC+Ator. Data are mean±SEM. cardiac hypertrophy and improve heart failure (8). However, it remains unknown whether or not statins inhibit this signal pathway. Atorvastatin is the most frequently prescribed statin worldwide, but few studies have been performed to clarify its influence on the progression of non-ischemic CHF and the possible cellular mechanisms involved. Accordingly, we investigated whether atorvastatin had a beneficial effect on the morphology and function of the left ventricle in mice with pressure overload, and we also investigated whether inhibition of EGFR activation had a role in the beneficial effects of statin therapy. We found that atorvastatin slowed the progress (...truncated)