HMG-CoA reductase inhibition improves endothelial cell function and inhibits smooth muscle cell proliferation in human saphenous veins
Zhihong Yang, MD*  ,
Toshiyoki Kozai, MD* ,
Bernd van de Loo, MD* ,
Hema Viswambharan, MSc* ,
Mario Lachat, MD ,
Marko I. Turina, MD,
Tadeusz Malinski, PhD and
Thomas F. Lüscher, MD, FRCP, FACC*
* Department of Cardiovascular Research, Institute of Physiology, University Zürich-Irchel, Zürich, Switzerland
Department of Cardiology, University Hospital, Zürich, Switzerland
Clinic for Cardiovascular Surgery, University Hospital, Zürich, Switzerland
Department of Chemistry, Institut of Biotechnology, Oakland University, Rochester, Michigan, USA
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Figure 1 Effects of cerivastatin on eNOS expression in human saphenous vein endothelial cells: Immunoblotting demonstrated up-regulation of eNOS by the HMG-CoA reductase inhibitor cerivastatin (Ceri) in a concentration-dependent manner. Densitometry showed about two-fold increase in eNOS protein level (n = 3).
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Figure 2 Cerivastatin up-regulates eNOS protein level and enhances NO release via inhibition HMG-CoA reductase: (A) Immunoblotting demonstrated that in cultured human saphenous vein endothelial cells, cerivastatin (10–6 mol/liter, 24 h) enhanced eNOS protein level, which was fully reversed by the HMG-CoA product mevalonate (Meva, 2 x 10–4 mol/liter). (B) Quantification of the experiments above by densitometry (n = 3). (C) In cells treated with cerivastatin (10–6 mol/liter, 24 h), NO release stimulated by Ca2+ ionophore (10–6 mol/liter) as measured by porphyrinic microsensor was significantly enhanced. This enhancement of NO release was fully reversed by mevalonate (2 x 10–4 mol/liter; n = 3). * = p < 0.01 vs. control; = p < 0.01 vs. cerivastatin plus mevalonate.
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Figure 4 Effects of cerivastatin on p42mapk and p70S6K: In cultured human saphenous vein, SMC PDGF-BB (10 ng/ml, 10 min) markedly stimulated (or phosphorylated) p42mapk as demonstrated by the slower mobility of the enzyme on Western blots. The activation of p42mapk was not inhibited by cerivastatin (10–6 mol/liter), but was markedly reduced by the specific inhibitor of MAPK kinase (MEK), PD98059 (5 x 10–5 mol/liter), while the PI3K inhibitor (wortmannin, 10–5 mol/liter) or the p70S6K inhibitor (rapamycin, 10–5 mol/liter) had no effects on p42mapk activation. The same blots were used for analysis of p70S6K. PDGF-BB stimulated p70S6K activation (or phosphorylation) as demonstrated by the slower mobility of the enzyme on Western blots, which was not influenced by cerivastatin or by PD98095, the specific inhibitor of MEK. In contrast, activation of p70S6K was completely prevented by wortmannin or rapamycin. The results were repeated with three independent experiments.
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Figure 5 Effects of cerivastatin on cell-cycle regulatory proteins: Western blotting demonstrated that stimulation of human vascular smooth muscle cells with PDGF-BB (10 ng/ml) for 24 h induced activation of Cdk2, phosphorylation of pRb and down-regulation of p27Kip1, but not p21Cip1. Cerivastatin (10–6 mol/liter) inhibited Cdk2 activation and pRb phosphorylation, but did not prevent the down-regulation of p27Kip1. The p21Cip1 was also not influenced by cerivastatin. The effects of cerivastatin on Cdk2 and Rb were reversed by the HMG-CoA product mevalonate (2 x 10–4 mol/liter). The results were repeated with three independent experiments.
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