In the current treatment of patients with atherosclerotic coronary artery disease, percutaneous coronary intervention (PCI) or percutaneous transluminal coronary angioplasty (PTCA) is employed.1) However, PCI results in vascular restenosis due to intimal thickening, often originating mainly from vascular smooth muscle cell (VSMC) proliferation.
2)Although several classes of synthetic drug have been administered to prevent restenosis after PCI,3) no drug has been found to reduce the incidence of restenosis.The intimal thickening model in rat carotid artery injured by balloon endothelial denudation is considered to have pathological characteristics similar to restenosis after PCI in humans.4) Recently, we have reported the inhibitory effects of a traditional Chinese formulation, Chaihu-jia-Longgu-Mulitang in Chinese and Saiko-ka-Ryukotsu-Borei-To (SRB) in Japanese, on intimal thickening in carotid arteries injured by balloon endothelial denudation in rats fed a normal diet.
5)Hypercholesterolemia has been identified as one of the risk factors in the onset and progression of restenosis after PCI and atherosclerosis. 6) Therefore, in this study, we examined the inhibitory effects of SRB on intimal thickening in a similar experiment in cholesterol-fed rats and compared the effects with those of the hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase inhibitor simvastatin (SV).7) To clarify the mechanism of the inhibitory effect of SRB, the number of VSMCs labeled with antiproliferating cell nuclear antigen (PCNA) antibody (PCNA labeling index) in the intimal area was examined. SV was purchased from Banyu Pharmaceutical Co., Ltd. (Tokyo, Japan). PCNA monoclonal antibody (PC-10) and the biotinylated anti-mouse secondary antibody streptavidineconjugated peroxidase were purchased from Dako Co., Ltd. (Kyoto, Japan). All other chemicals and solvents were of analytical grade.
MATERIALS AND METHODS
SRB and Other ReagentsCholesterol-Fed Rats Male Wistar rats (13 weeks old, 340-360 g, Sankyo Laboratory Service, Tokyo) were fed a normal diet (Powdered CE-2, CLEA Japan, Tokyo) containing 1% cholesterol and SRB extract (375, 750, 1125 mg/kg daily, nϭ8) for 3 d before and then for 7 d after the injury. SV (0.83 mg/kg daily, nϭ8) was suspended in 0.5% carboxymethyl-cellulose sodium salt (Sigma) and administered orally though a gastric sonde during the same period. The doses of SRB 750 mg/kg and SV 0.83 mg/kg are 10-fold higher than the common human daily dose.Balloon Endothelial Denudation of Carotid Artery Balloon endothelial denudation of the left carotid artery in rats anesthetized with pentobarbital was performed according to our previous method.5) All animal experiments were carried out in accordance with the Guidelines of the Animal Care and Use Committee of Toyama Medical and Pharmaceutical University as approved by the Japanese Association of Laboratory Animal Care.Intimal Thickening, Intimal Area/Medical Area and Stenosis Ratios The left carotid artery sections 7 d after denudation were evaluated histologically using the ...