Patrick Duriez

Peroxisome Proliferator-Activated Receptor Activators Inhibit Thrombin-Induced Endothelin-1 Production in Human Vascular Endothelial Cells by Inhibiting the Activator Protein-1 Signaling Pathway

Endothelin-1 (ET-1), a 21-amino acid vasoactive peptide mainly produced by vascular endothelial cells, is involved in the regulation of vascular tone and smooth muscle cell proliferation. Peroxisome proliferator-activated receptors (PPARs), key players in lipid and glucose metabolism, have been implicated in metabolic disorders that are predisposing to atherosclerosis. Because of the potential role of ET-1 in vascular disorders such as hypertension and atherosclerosis, we investigated the regulation of ET-1 expression by PPAR activators. Western blot and reverse transcription-polymerase chain reaction analyses demonstrated that both PPARalpha and PPARgamma are expressed in human coronary artery endothelial cells as well as in endothelial cell lines such as HMEC-1 and ECV304. In bovine aortic endothelial cells and HMEC-1 cells, both PPARalpha and PPARgamma ligands inhibited thrombin-induced ET-1 secretion, whereas basal ET-1 secretion was only slightly suppressed. Reverse transcription-polymerase chain reaction experiments showed that this inhibition of ET-1 production occurs at the gene expression level. Using transient transfection assays, we demonstrated that PPARs downregulate thrombin-activated transcription of the human ET-1 promoter. Transactivation studies with c-Jun and c-Fos expression plasmids indicated that PPARs negatively interfere with the activator protein-1 signaling pathway, which mediates thrombin activation of ET-1 gene transcription. Furthermore, electrophoretic mobility shift assays demonstrated that PPAR activators reduce the thrombin-stimulated binding activity of bovine aortic endothelial cell nuclear extracts as well as c-Jun binding to an activator protein-1 consensus site. Taken together, these data indicate that (1) both PPARalpha and PPARgamma are expressed in human vascular endothelial cells and (2) PPAR activators inhibit thrombin-induced ET-1 biosynthesis, indicating a novel role for PPARs in vascular endothelial function.

Peroxisome proliterator-activated receptor-alpha activators regulate genes governing lipoprotein metabolism, vascular inflammation and atherosclerosis

The peroxisome proliferator-activated receptors (PPARs) [alpha, delta (beta) and gamma] form a subfamily of the nuclear receptor gene family. All PPARs are, albeit to different extents, activated by fatty acids and derivatives; PPAR-alpha binds the hypolipidemic fibrates whereas antidiabetic glitazo

Atherogenic Lipoprotein Particles in Atherosclerosis

The importance of low-density lipoprotein (LDL) cholesterol in the development of atherosclerosis has long been recognized, and LDL cholesterol remains the primary target of therapy for the prevention of coronary heart disease. Nevertheless, increasing research attention over the past decade has been devoted to the heterogeneity of LDL particles and the atherogenicity of lipids and lipoproteins other than LDL. Particularly atherogenic forms of LDL include small, dense LDL particles and oxidized LDL. All lipoproteins that contain apolipoprotein B, such as LDL, very-low-density lipoprotein, and intermediate-density lipoprotein, tend to promote atherosclerosis; however, these particles differ in their apolipoprotein and triglyceride content. High levels of plasma triglycerides increase the risk of acute coronary events. Lipoprotein(a) is now considered an independent risk factor in both men and women. Ultimately, better understanding of the roles of these lipid particles and subfractions in the initiation and progression of atherosclerosis may affect treatment decisions.

New Risk Factors for Atherosclerosis and Patient Risk Assessment

Advances in our understanding of the ways in which the traditional cardiovascular risk factors, including standard lipid (eg, total cholesterol, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol) and nonlipid (eg, hypertension) risk factors, interact to initiate atherosclerosis and promote the development of cardiovascular disease have enhanced our ability to assess risk in the individual patient. In addition, the ongoing identification and understanding of so-called novel risk factors may further improve our ability to predict future risk when these are included along with the classic risk factors in assessing the global risk profile. This review briefly summarizes the evidence that some newer risk factors, including impaired fasting glucose, triglycerides and triglyceride-rich lipoprotein remnants, lipoprotein(a), homocysteine, and high-sensitivity C-reactive protein, contribute to an increased risk of coronary and cardiovascular diseases.

Peroxisome proliferator-activated receptor activators inhibit oxidized low-density lipoprotein-induced endothelin-1 secretion in endothelial cells.

Endothelin is a potent vasoconstrictor peptide isolated from endothelial cells and it induces smooth muscle cell proliferation. Endothelin-1 secretion is increased in atheroma and induces deleterious effects such as vasospasm and atherosclerosis. Oxidized low-density lipoproteins (LDLs) induce atherosclerosis in the vascular wall, as well as endothelin-1 secretion in endothelial cells and are activators of both peroxisome proliferator–activated receptor-&agr; (PPAR-&agr;) and PPAR-&ggr;. PPAR-&agr; (fibric acids) and PPAR-&ggr; (glitazones) activators are used to treat dyslipoproteinemias and type 2 diabetes, respectively. Furthermore, these drugs induce numerous pleiotropic effects, such as inhibiting thrombin-induced endothelin-1 secretion in endothelial cells. This study shows that both PPAR-&agr; (Wy 14643) and PPAR-&ggr; activation (rosiglitazone) partially inhibit oxidized LDL–induced protein kinase C activity and endothelin-1 secretion in endothelial cells at the transcriptional levels and suggests that synthetic PPAR activators are stronger PPAR activators than oxidized LDL. This study also suggests that fibrate and glitazone treatments should have beneficial effects on the vascular wall by reducing endothelin-1 secretion and the resulting vasospasm and atherosclerosis.

The PPARα activator fenofibrate slows down the progression of the left ventricular dysfunction in porcine tachycardia-induced cardiomyopathy

It has been reported that high intramyocardial peroxisome proliferator-activated receptor α (PPARα) stimulation or overexpression altered cardiac contractile function in mouse models of cardiac hypertrophy and heart failure. Nevertheless, it has never been demonstrated that clinically relevant doses of drugs stimulating PPARα activity such as fenofibrate increase the risk to develop heart failure in humans. To determine if fenofibrate accelerates the development of heart failure in large mammals, we have tested its effects on the progression of left ventricular dysfunction in pacing-induced heart failure in pigs. Fenofibrate treatment blunted reduction in left ventricular ejection fraction, reduced cardiac hypertrophy, and attenuated clinical signs of heart failure. Fenofibrate impeded the increase in atrial natriuretic peptide, brain natriuretic peptide, and endothelin-1 plasma levels. The expression of PPARα, fatty acyl-CoA-oxidase, and carnitine palmitoyltransferase-Iβ was reduced at mRNA levels in the left ventricle from untreated heart failure pigs but maintained near normal values with fenofibrate. Fenofibrate prevented heart failure-induced overexpression of TNFα mRNA and enhanced catalase activity in left ventricle compared to placebo. These data suggest that a clinically relevant dose of fenofibrate does not accelerate but slows down heart failure development in the model of pacing-induced heart failure in large mammals.

Paullinia pinnata extracts rich in polyphenols promote vascular relaxation via endothelium-dependent mechanisms

Paullinia pinnata L. (Sapindaceae) is an African tropical plant whose roots and leaves are used in traditional medicine for many purposes, especially for erectile dysfunction, but its action mechanism is unknown. P. pinnata root and leaf methanolic extracts are rich in phenolic compounds. This study shows that both extracts are highly antioxidative and induce a slight transcriptional activity of peroxisome proliferator activated receptor-α. They also increased and decreased endothelial nitric oxide synthase and endothelin-1 mRNA levels in bovine aortic endothelial cells, respectively. In this study P. pinnata methanolic extracts in cumulative doses elicited in a dose-dependent manner the relaxation of phenylephrine precontracted isolated rat aortic rings. NG-nitro-L-arginine methyl ester significantly attenuated the capacity of both extracts to induce arterial relaxation, indicating that this arterial relaxation was mediated by endothelial nitric oxide release. It could be suggested that the arterial relaxation induced by both extracts could be mainly linked to their capacities to inhibit nitric oxide oxidation through their antioxidant properties.

Alpha-Tocopherol But Not Beta-Tocopherol Inhibits Thrombin-Induced PKC Activation and Endothelin Secretion in Endothelial Cells

Introduction Production of endothelin by endothelial cells depends on protein-kinase C (PKC) stimulation which has been reported to be inhibited by alpha-tocopherol (α-Toch) but not by beta-tocopherol (β-Toch). The goal of this study was to determine whether α-Toch and β-Toch inhibit endothelin secretion by endothelial cells. Methods and results In a first set of experiments, cultured bovine aortic endothelial cells (BAEC) were incubated for 48 h with 100 μmol/l α-Toch or vehicle (0.1% ethanol), then cells were stimulated for 4 h or 20 h with thrombin. After stimulating bovine aortic endothelial cells with thrombin for 4 h, α-Toch inhibited PKC activity by 63% and endothelin secretion by 44%, whereas after 20 h of incubation with thrombin, α-Toch decreased the peptide secretion by 51%. In a second set of experiments, BAEC were incubated with increased concentrations (from 0 to 100 μmol/l) of α-Toch or β-Toch, PKC activity and endothelin secretion were measured after thrombin stimulation as previously reported. In these experiments, α-Toch strongly inhibited thrombin-induced PKC activity and endothelin secretion in a dose-dependent manner, whereas β-Toch was more than 10-fold less active than α-Toch in inhibiting these stimulations. Tocopherols (α-Toch + β-Toch) produced a proportional correlation on both PKC stimulation and endothelin secretion by inhibiting the effect of thrombin. Conclusion These data suggest that α-Toch strongly inhibits thrombin-induced endothelin secretion in vitro at least partly through PKC inhibition.

A High Concentration of Melatonin Inhibits In Vitro Ldl Peroxidation But Not Oxidized Ldl Toxicity Toward Cultured Endothelial Cells

The pineal hormone, melatonin, was recently found to be a potent free scavenger for hydroxyl and peroxyl radicals. Melatonin also inhibits neuronal and thymocyte damage due to oxidative stress. Atherosclerosis development is mediated by low-density lipoprotein (LDL) oxidation and the endocytosis of oxidized LDL by resident macrophages in the subendothelial vascular wall. Furthermore, the cytotoxic effect of oxidized LDL increases atherogenicity. The goal of this study was to compare the antioxidant activities of melatonin and vitamin E against in vitro LDL oxidation and their cytoprotective actions against oxidized LDL-induced endothelial cell toxicity. An attempt at loading LDL with melatonin by incubating human plasma with an ethanolic melatonin solution gave only low protection against Cu2+-induced LDL oxidation in comparison with vitamin E and gave no detectable incorporation of melatonin into LDL, measured by high-performance liquid chromatography (HPLC) coupled to UV detection. High concentrations of melatonin (10-100 microM) added to the oxidative medium induced a clear inhibition of Cu2+-induced LDL oxidation, characterized as an increase in the lag-phase duration of conjugated diene formation and decreases in the maximal rate of the propagation phase and in the maximal amount of conjugated diene formation. Determination of the median efficacious dose (ED50) of melatonin and vitamin E by their ability to increase lag-phase duration showed that melatonin was less active than vitamin E (ED50, 79 vs. 10 microM, respectively). Melatonin was also less active than vitamin E in limiting the formation of thiobarbituric acid-reactive substances (TBARS) and LDL fluorescence intensity increase in the medium during Cu2+-induced LDL oxidation. Cu2+-induced LDL oxidation in the presence of 100 microM melatonin produced oxidized LDLs that were less recognizable for the scavenger receptors of J774 macrophages than were untreated LDLs. Vitamin E, 10 microM, was more active than 100 microM melatonin in inhibiting LDL oxidation and the resulting lipoprotein alterations leading to binding internalization and degradation by the J774 macrophages. Vitamin E, 100 microM, inhibited the pursuit of the oxidation of oxidized LDL mediated by bovine aortic endothelial cells (BAECs) in a culture medium containing Cu2+, whereas 100 microM melatonin had no antioxidant effect. Melatonin, 100 microM, as well as 100 microM vitamin E inhibited intracellular TBARS formation during the incubation of BAECs with highly oxidized LDL but had no influence on the increase in glutathione (GSH) concentration during this lengthy exposure (16 h) of BAECs to highly oxidized LDL. During this period, the same dose of vitamin E but not of melatonin tended to limit the decrease in adenosine triphosphate (ATP) concentration. Vitamin E, 100 microM, did not significantly reduce cellular lactate dehydrogenase (LDH) release in the culture medium during the incubation of oxidized LDL with BAECs, whereas 100 microM melatonin dramatically increased this release. These data show that melatonin is less active than vitamin E in inhibiting in vitro LDL oxidation and does not inhibit the cytotoxicity of oxidized LDL toward cultured endothelial cells. The concentrations necessary to inhibit LDL oxidation are far beyond those found in human plasma (100 microM vs. 100 pM). Therefore our results indicate that the pineal hormone melatonin per se appears to have little antiatherogenic property in the in vitro oxidation of LDL and the cytoprotective action against the toxicity of oxidized LDL. Nevertheless, in vivo LDL oxidation takes place in the subendothelium of the artery wall, and nothing is known about the concentration of melatonin or its catabolites in this environment.

PPARs: a potential target for a disease-modifying strategy in stroke.

Stroke is one of the major causes of mortality and disability in adults in industrialized countries. Despite numerous preclinical studies and clinical trials in the field of cerebral ischemia, no pharmacological agent has been validated in the treatment of acute ischemic, except thrombolysis. Cerebral ischemia is not only a neuronal disease but it affects the entire neurovascular unit. The therapeutic strategy in stroke should be more global and combine preventive approaches, acute phase treatment and long-term care to improve recovery and prevent or treat affective and cognitive post-stroke consequences. There is an imperative need to develop disease-modifying drugs, which should be able to induce neuroprotection, to serve as adjuvants for thrombolysis by decreasing the hemorrhagic risk and to limit the long-term post-stroke consequences. This review presents the potential effects of Peroxisome Proliferator-Activated Receptors (PPARs) and of their agonists in stroke. We focus on each PPAR receptor and detail their implication in stroke. PPARs are nuclear receptors, acting as ligand-dependent transcription factors. They are expressed in the neurovascular unit that suggests that PPARs could play a role in stroke. Indeed, it has been shown that they are able to interfere with pathways implicated in the pathophysiology of stroke. They could be an answer to this disease-modifying drug concept, being able to act on the different phases of ischemia.