Elsevier

Atherosclerosis

Volume 154, Issue 1, January 2001, Pages 87-96
Atherosclerosis

Anti-oxidative properties of fluvastatin, an HMG-CoA reductase inhibitor, contribute to prevention of atherosclerosis in cholesterol-fed rabbits

https://doi.org/10.1016/S0021-9150(00)00468-8Get rights and content

Abstract

Studies in vitro reveal that fluvastatin, an HMG-CoA reductase inhibitor, has a strong DPPH radical scavenging activity and achieves concentration-dependent inhibition of copper- and cell-induced oxidation of low-density lipoprotein (LDL). To further examine the anti-oxidative activity of fluvastatin in vivo, we elucidated the effects of chronic treatment with fluvastatin at a dose insufficient to reduce plasma cholesterol levels (2 mg/kg per day) on vasomotion and vascular oxidative stress in thoracic aortas of 0.5% cholesterol-fed rabbits. After 12 weeks of dietary treatment, aortic segments from rabbits fed cholesterol alone showed impaired endothelium-dependent relaxation responses to acetylcholine and A23187 compared to normal chow-fed rabbits in association with a significant increase in plasma total cholesterol levels. In contrast, although plasma total cholesterol levels were not different from those in control cholesterol-fed rabbits, aortic segments from fluvastatin-treated rabbits showed normal relaxation. Compared with rabbits fed cholesterol alone, fluvastatin treatment decreased susceptibility of LDL to ex vivo copper-induced oxidation, reduced vascular superoxide generation, and atheromatous plaque formation. In conclusion, the potent anti-oxidative properties of fluvastatin in addition to its cholesterol-lowering activity appear to contribute to its anti-atherosclerotic effect in vivo.

Introduction

Hypercholesterolemia has been shown to be one of the major risk factors of atherosclerosis [1], [2]. Recent clinical studies have demonstrated that cholesterol-lowering therapy with 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors can achieve a relatively large reduction in plasma cholesterol and they can not only decrease morbidity and mortality of coronary artery disease in humans[3], [4], but also induce regression of atherosclerosis [5]. It has been considered that these anti-atherosclerotic effects of HMG-Co/A reductase inhibitors are likely to mainly result from lipid-lowering properties of these drugs.

In arteries in hypercholesterolemic animals and patients, vascular superoxide production and oxidative stress are increased [6]. Oxidation of low-density lipoprotein (LDL) is considered as an important step in the development of atherosclerosis. In vitro studies have demonstrated that oxidized LDL (ox-LDL) activates endothelial cells to increase the expression of chemoattractant molecules leading to stimulate transendothelial migration of monocyte and formation of foam cells. Ox-LDL also increases production of growth factors including platelet-derived growth factor that stimulates migration and proliferation of smooth muscle cells. Furthermore, ox-LDL impairs endothelium-derived nitric oxide (NO) production [7], [8]. As reported previously, the antioxidant has the ability to prevent oxidation of LDL in vitro and in vivo. For example, dietary antioxidants such as vitamin E [9] and probucol [10] protect LDL from oxidation. In addition to inhibition of LDL oxidation, antioxidant therapy has been shown to produce beneficial effects on atherosclerosis and to prevent the progression of atherosclerosis in animal models by limiting vascular oxidative stress and superoxide production [11], [12], [13].

Endothelial dysfunction including the impaired endothelium-dependent vasodilation is observed in hypercholesterolemia and atherosclerosis. Recent studies have demonstrated that cholesterol-lowering therapy with HMG-CoA reductase inhibitors showed a beneficial effect on endothelium-dependent coronary vasomotion [14], [15], whereas others failed to show such an effect [16], [17]. Anderson et al. have reported that cholesterol-lowering therapy with lovastatin, an HMG-CoA reductase inhibitor, and cholestyramine was not able to improve endothelial dysfunction; however, combination therapy with a lipid-lowering drug and an antioxidant effectively improved endothelial function [16]. In contrast to failure of lovastatin to prevent the progression of atherosclerosis in cholesterol-fed rabbits [17], anti-oxidative drugs like probucol have been shown to have beneficial effects to prevent the development of atherosclerosis and to preserve endothelial function effectively [13].

In recent years, it becomes clear that all the clinical benefits of the HMG-CoA reductase inhibitors therapy can not be explained solely by their lipid-lowering properties because a variety of experimental data revealed that these drugs have direct anti-atherosclerotic effects that were unrelated to the lipid-lowering effect [18], [19], [20]. However, it remains unknown that these pleiotropic effects of HMG-CoA reductase inhibitors serve to improve endothelial dysfunction present in atherosclerosis. The aim of the present study was to examine anti-oxidative property of fluvastatin in vitro and in vivo. We investigated whether fluvastatin can preserve endothelial function and prevent atherosclerosis in cholesterol-fed rabbit model via its potent anti-oxidative property other than its hypocholesterolemic effect.

Section snippets

DPPH radical scavenging activity

1,1-Diphenyl-2-picrylhydrasyl (DPPH) radical scavenging activity was determined by the method of Blois [21]. In brief, HMG-CoA reductase inhibitors (final 160 μM) were incubated with 16 μM DPPH in acetate buffer (0.1 M, pH 5.5)/methanol, 2:3 (v/v) solution at room temperature. The absorbance at 523 nm of the tested sample solution was measured with a spectrophotometer (U-3300, Hitachi Ltd.).

Preparation of LDL

Human LDL (density, 1.019–1.063 g/ml) was isolated from the plasma of healthy volunteers by sequential

DPPH radical scavenging activity

First, we investigated the free radical scavenging activities of various HMG-CoA reductase inhibitors. DPPH is a very stable free radical and widely used for evaluation of anti-oxidative activities [21]. Probucol [10] and four HMG-CoA reductase inhibitors, fluvastatin, lovastatin, pravastatin, and simvastatin were tested. As shown in Fig. 1, probucol showed a very rapid and large decrease in the absorbance of DPPH solution, indicating that probucol has a potent anti-oxidative activity.

Discussion

The present in vitro study shows that fluvastatin has anti-oxidative activities as demonstrated that this drug has a radical scavenging activity and is capable of inhibiting copper ion- and VSMC-mediated oxidation of LDL. These anti-oxidative effects of fluvastatin were observed at therapeutic concentrations [28]. Although previous studies have shown that HMG-CoA reductase inhibitors, lovastatin [29], pravastatin [30], or simvastatin [31], also have inhibitory effects on LDL oxidation, these

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