Daniel Hayoz

The CoLaus study: a population-based study to investigate the epidemiology and genetic determinants of cardiovascular risk factors and metabolic syndrome.

BackgroundCardiovascular diseases and their associated risk factors remain the main cause of mortality in western societies. In order to assess the prevalence of cardiovascular risk factors (CVRFs) in the Caucasian population of Lausanne, Switzerland, we conducted a population-based study (Colaus Study). A secondary aim of the CoLaus study will be to determine new genetic determinants associated with CVRFs.MethodsSingle-center, cross-sectional study including a random sample of 6,188 extensively phenotyped Caucasian subjects (3,251 women and 2,937 men) aged 35 to 75 years living in Lausanne, and genotyped using the 500 K Affymetrix chip technology.ResultsObesity (body mass index ≥ 30 kg/m2), smoking, hypertension (blood pressure ≥ 140/90 mmHg and/or treatment), dyslipidemia (high LDL-cholesterol and/or low HDL-cholesterol and/or high triglyceride levels) and diabetes (fasting plasma glucose ≥ 7 mmol/l and/or treatment) were present in 947 (15.7%), 1673 (27.0%), 2268 (36.7%), 2113 (34.2%) and 407 (6.6%) of the participants, respectively, and the prevalence was higher in men than in women. In both genders, the prevalence of obesity, hypertension and diabetes increased with age.ConclusionThe prevalence of major CVRFs is high in the Lausanne population in particular in men. We anticipate that given its size, the depth of the phenotypic analysis and the availability of dense genome-wide genetic data, the CoLaus Study will be a unique resource to investigate not only the epidemiology of isolated, or aggregated CVRFs like the metabolic syndrome, but can also serve as a discovery set, as well as replication set, to identify novel genes associated with these conditions.

Mental Stress Induces Prolonged Endothelial Dysfunction via Endothelin-A Receptors

Background—Mental stress is a risk factor for atherosclerosis and may precipitate myocardial ischemia and infarction. Because endothelial dysfunction is an early manifestation of atherosclerosis, we investigated the impact of mental stress on endothelial function. Methods and Results—The effects of a 3-minute mental stress task on endothelium-dependent vasodilation were studied in healthy subjects without cardiovascular risk factors. Flow-mediated (FMD) and nitroglycerin (0.4 mg sublingual)-induced vasodilation were studied before and after mental stress by high-resolution ultrasound of the radial artery. Additionally, FMD was assessed before and 10 to 45 minutes after mental stress during intraarterial infusion of a selective endothelin A receptor antagonist (BQ-123, 1 nmol/min) or saline, respectively. Endothelium-dependent vasodilation was reduced by half for about 45 minutes (8.0±1.1% versus 4.1±1.0%;P <0.002), whereas endothelium-independent vasodilation to nitroglycerin remained unaffected (15.6±1.6 versus 14.3±1.3%; NS). Intraarterial infusion of BQ-123, a selective endothelin-A receptor antagonist, but not saline prevented the impairment of endothelium-dependent vasodilation (8.6±1.2 versus 9.4±1.3%; NS). In contrast, intraarterial infusion of norepinephrine of similar duration as mental stress did not inhibit FMD. Conclusions—Mental stress induces prolonged endothelial dysfunction, which is prevented by selective endothelin-A receptor antagonism. This represents a novel and important link between mental stress and atherosclerotic vascular disease.

Nitric Oxide Synthase Expression in Endothelial Cells Exposed to Mechanical Forces

Nitric oxide (NO) has been demonstrated to play a central role in vascular biology and pathobiology. The expression of endothelial NO synthase (eNOS) is regulated in part by blood flow-induced mechanical factors. The purpose of this study was to evaluate how the expression of eNOS mRNA correlates with the activation of its promoter in both arterial and venous endothelial cells (ECs) exposed to mechanical forces, ie, shear stress and cyclic circumferential stretch. Bovine aortic ECs (BAECs) and EA hy.926, a cell line derived from human umbilical vein ECs, were grown on the inside of elastic tubes and subjected to combinations of pressure, pulsatile shear stress, and cyclic circumferential stretch for 24 hours. Two patterns of shear stress were used: unidirectional (mean of 6, ranging from 3 to 9 dyne/cm2) and oscillatory (mean of 0.3, ranging from -3 to +3 dyne/cm2). The expression of eNOS mRNA was quantified by Northern blot analysis. Activation of the promoter was assessed by luciferase activity after the cells were transiently transfected before the flow experiments with a plasmid construct containing the fully functional eNOS promoter coupled to a luciferase reporter gene. Expression of eNOS mRNA was increased and promoter activity was enhanced by unidirectional shear stress compared with static control. Oscillatory shear slightly upregulated eNOS mRNA in BAECs, whereas it downregulated eNOS mRNA in EA hy.926. In both BAECs and EA hy.926, there was a good correlation between the increase in eNOS mRNA expression and promoter activation by unidirectional shear stress. In contrast, in both BAECs and EA hy.926 cells exposed to shear stress, cyclic stretch did not change eNOS mRNA expression, but the activation of eNOS promoter was significantly lower. Moreover, when ECs were exposed to oscillatory shear stress, there was a dramatic activation of the eNOS promoter. These results demonstrate that unidirectional shear stress increases eNOS mRNA expression via a transcriptional mechanism. However, oscillatory shear stress and cyclic stretch appear to control eNOS expression through posttranscriptional regulatory events.

Thrombin Stimulates Human Endothelial Arginase Enzymatic Activity via RhoA/ROCK Pathway Implications for Atherosclerotic Endothelial Dysfunction

Background—Arginase competes with endothelial nitric oxide synthase (eNOS) for the substrate l-arginine and decreases NO production. This study investigated regulatory mechanisms of arginase activity in endothelial cells and its role in atherosclerosis. Methods and Results—In human endothelial cells isolated from umbilical veins, thrombin concentration- and time-dependently stimulated arginase enzymatic activity, reaching a 1.9-fold increase (P<0.001) at 1 U/mL for 24 hours. The effect of thrombin was prevented by C3 exoenzyme or the HMG-CoA reductase inhibitor fluvastatin, which inhibit RhoA, or by the ROCK inhibitors Y-27632 and HA-1077. Adenoviral expression of constitutively active RhoA or ROCK mutants enhanced arginase activity (≈3-fold, P<0.001), and the effect of active RhoA mutant was inhibited by the ROCK inhibitors. Neither thrombin nor the active RhoA/ROCK mutants affected arginase II protein level, the only isozyme detectable in the cells. Moreover, a significantly higher arginase II activity (1.5-fold, not the protein level) and RhoA protein level (4-fold) were observed in atherosclerotic aortas of apoE−/− compared with wild-type mice. Interestingly, l-arginine (1 mmol/L), despite a significantly higher eNOS expression in aortas of apoE−/− mice, evoked a more pronounced contraction, which was reverted to a greater vasodilation by the arginase inhibitor l-norvaline (20 mmol/L) compared with the wild-type animals (n=5, P<0.001). Conclusions—Thrombin enhances arginase activity via RhoA/ROCK in human endothelial cells. Higher arginase enzymatic activity is involved in atherosclerotic endothelial dysfunction in apoE−/− mice. Targeting vascular arginase may represent a novel therapeutic possibility for atherosclerosis.

Local Pulse Pressure and Regression of Arterial Wall Hypertrophy During Long-Term Antihypertensive Treatment

BACKGROUND Local pulse pressure (PP) is an independent determinant of carotid artery wall thickness, stronger than mean blood pressure (BP). The present study was designed to assess whether a beta-adrenoceptor antagonist-based or an ACE inhibitor-based treatment was able to reduce carotid artery wall hypertrophy through a reduction in carotid PP rather than by lowering mean BP and whether the influence of local PP reduction could also be detected at the site of a muscular artery, the radial artery. METHODS AND RESULTS Ninety-eight essential hypertensive patients were randomized to 9 months of double-blind treatment with either celiprolol or enalapril. Arterial parameters were determined with high-resolution echo-tracking systems. PP was measured locally with applanation tonometry and independently of mean BP. After 9 months of treatment, mean BP, carotid PP, and intimal-medial thickness (IMT) decreased significantly, with no difference between the 2 groups. The reduction in carotid PP but not in mean BP was a major independent determinant of the reduction in carotid IMT. Radial artery IMT and PP decreased significantly with both treatments. However, the reduction in radial artery IMT was not related to the changes in radial artery PP. CONCLUSIONS The regression of carotid artery wall hypertrophy during long-term antihypertensive treatment was dependent on the reduction in local PP rather than on the lowering of mean BP. The effect of PP lowering on IMT reduction was observed at the site of an elastic artery but not at the site of a muscular artery.

Statins enhance postischemic hyperemia in the skin circulation of hypercholesterolemic patients: A monitoring test of endothelial dysfunction for clinical practice?☆

OBJECTIVES The present study aims to investigate whether laser Doppler flowmetry can be used to monitor improvements in vascular function during statin therapy. BACKGROUND Endothelial dysfunction is an early feature of atherosclerosis in hypercholesterolemic patients and can be improved by statins. There are several methods to assess endothelial function in vivo, none of them being feasible in everyday practice. METHODS Skin perfusion, measured by laser Doppler flowmetry, was assessed at rest and during reactive hyperemia. Nineteen hypercholesterolemic patients (age 42 to 73 years, total cholesterol 5.4 to 9.6 mmol/l) were studied before and during statin therapy. To further investigate the mechanisms, postischemic skin hyperemia was measured before and after intradermal injection of the nitric oxide synthase inhibitor L-NAME and its inactive isoform D-NAME (0.5 micromol/10 microl each). On a separate day, the healthy volunteers were reexamined before and 2 h after 1,000 mg aspirin. RESULTS Postischemic skin blood flow was markedly reduced in hypercholesterolemic patients (45 +/- 11%) compared with healthy controls (238 +/- 20%, p < 0.0001) and improved after statin therapy (113 +/- 15%, p = 0.0005 vs. pre-treatment). In the healthy volunteers, the hyperemic responses were not significantly different after L-NAME and D-NAME. Aspirin reduced hyperemia from 274 +/- 49% to 197 +/- 40% (p = 0.025). CONCLUSIONS Reactive hyperemia of the skin microcirculation can be easily and reproducibly assessed by laser Doppler flowmetry. Vasodilator prostaglandins are the major mediators of postischemic skin hyperemia, which is impaired in hypercholesterolemic patients and can be enhanced by cholesterol-lowering therapy. Thus, laser Doppler flowmetry may represent a tool to assess and monitor vascular function during therapy in everyday practice.

Isobaric compliance of the radial artery is increased in patients with essential hypertension.

Objectives: Hypertension is known to decrease arterial elasticity and systemic compliance. However, the arterial tree is not a homogeneous system, and whether a distal medium-sized artery such as the radial artery behaves like proximal arteries has not been determined. The aims of the present study were, first, to characterize non-invasively the mechanical properties of the radial artery through the determination of the pressure-diameter curve, the distensibility-pressure curve and the compliance—pressure curve, and, secondly, to compare untreated hypertensive patients with normotensive subjects. Methods: A new high-precision echo-tracking device was developed which allows the diameter of peripheral arteries to be measured continuously. By relating the changes in internal diameter (cross-sectional changes) to those in blood pressure, the cross-sectional arterial compliance could be determined. Participants: Seventy-eight untreated mild or moderate essential hypertensive patients aged 24-78 years were compared with 44 normotensive subjects aged 22-81 years. In order to increase the database and provide independent assessments of the variables examined, the cross-sectional study was performed independently using a standardized procedure in three different research centres. Results: The major finding was that diameter, distensibility and compliance of the radial artery of hypertensive patients were not significantly different from those of normotensive controls when the two populations were studied at their mean arterial pressure. Furthermore, when the two populations were compared for the same level of blood pressure, using distensibility- and compliance-pressure curves, it was clear that isobaric distensibility and compliance of hypertensives were not significantly lower than those of normotensives, being either unchanged or higher. Conclusions: These findings are in contrast with the well-known decrease in compliance of proximal large arteries due to hypertension. Whether such a difference between proximal large arteries and distal medium-sized arteries may be related to the structural vascular changes observed with long-standing hypertension is still difficult to analyse in humans, and requires further investigation.

Endogenous Angiotensin II Induces Atherosclerotic Plaque Vulnerability and Elicits a Th1 Response in ApoE−/− Mice

Rupture of vulnerable plaques is the main cause of acute cardiovascular events. However, mechanisms responsible for transforming a stable into a vulnerable plaque remain elusive. Angiotensin II, a key regulator of blood pressure homeostasis, has a potential role in atherosclerosis. To study the contribution of angiotensin II in plaque vulnerability, we generated hypertensive hypercholesterolemic ApoE−/− mice with either normal or endogenously increased angiotensin II production (renovascular hypertension models). Hypertensive high angiotensin II ApoE−/− mice developed unstable plaques, whereas in hypertensive normal angiotensin II ApoE−/− mice plaques showed a stable phenotype. Vulnerable plaques from high angiotensin II ApoE−/− mice had thinner fibrous cap (P<0.01), larger lipid core (P<0.01), and increased macrophage content (P<0.01) than even more hypertensive but normal angiotensin II ApoE−/− mice. Moreover, in mice with high angiotensin II, a skewed T helper type 1-like phenotype was observed. Splenocytes from high angiotensin II ApoE−/− mice produced significantly higher amounts of interferon (IFN)-&ggr; than those from ApoE−/− mice with normal angiotensin II; secretion of IL4 and IL10 was not different. In addition, we provide evidence for a direct stimulating effect of angiotensin II on lymphocyte IFN-&ggr; production. These findings suggest a new mechanism in plaque vulnerability demonstrating that angiotensin II, within the context of hypertension and hypercholesterolemia, independently from its hemodynamic effect behaves as a local modulator promoting the induction of vulnerable plaques probably via a T helper switch.

Renin Inhibition by Aliskiren Prevents Atherosclerosis Progression. Comparison With Irbesartan, Atenolol, and Amlodipine

Hypertension is associated with increased risk of cardiovascular diseases. Antihypertensive treatment, particularly blockade of the renin-angiotensin system, contributes to prevent atherosclerosis-mediated cardiovascular events. Direct comparison of different antihypertensive treatments on atherosclerosis and particularly plaque stabilization is sparse. ApoE−/− mice with vulnerable (2-kidney, 1-clip renovascular hypertension model) or stable (1-kidney, 1-clip renovascular hypertension model) atherosclerotic plaques were used. Mice were treated with aliskiren (renin inhibitor), irbesartan (angiotensin-receptor blocker), atenolol (&bgr;-blocker), or amlodipine (calcium channel blocker). Atherosclerosis characteristics were assessed. Hemodynamic and hormonal parameters were measured. Aliskiren and irbesartan significantly prevented atherosclerosis progression in 2-kidney, 1-clip mice. Indeed, compared with untreated animals, plaques showed thinner fibrous cap (P<0.05); smaller lipid core (P<0.05); decreased media degeneration, layering, and macrophage content (P<0.05); and increased smooth muscle cell content (P<0.05). Interestingly, aliskiren significantly increased the smooth muscle cell compared with irbesartan. Despite similar blood pressure lowering, only partial plaque stabilization was attained by atenolol and amlodipine. Amlodipine increased plaque smooth muscle cell content (P<0.05), whereas atenolol decreased plaque inflammation (P<0.05). This divergent effect was also observed in 1-kidney, 1-clip mice. Normalizing blood pressure by irbesartan increased the plasma renin concentration (5932±1512 ng/mL per hour) more than normalizing it by aliskiren (16085±5628 ng/mL per hour). Specific renin-angiotensin system blockade prevents atherosclerosis progression. First, evidence is provided that direct renin inhibition mediates atherosclerotic plaque stabilization. In contrast, &bgr;-blocker and calcium channel blocker treatment only partially stabilize plaques differently influencing atherogenesis. Angiotensin II decisively mediates plaque vulnerability. The plasma renin concentration measurement by an indirect method did not confirm the excessive increase of plasma renin concentration reported in the literature during aliskiren compared with irbesartan or amlodipine treatment.

High Prevalence of Peripheral Arterial Disease in HIV-Infected Persons

BACKGROUND Atherosclerosis has been assessed in human immunodeficiency virus (HIV)-infected persons by using various methods. Peripheral arterial disease (PAD) has not been evaluated, however. We studied the cross-sectional prevalence of lower limb PAD in an HIV-infected population. METHODS PAD was assessed using the Edinburgh Claudication Questionnaire and by measuring the systolic ankle-brachial blood pressure index (ABI) at rest and after exercise. Patients with PAD were further evaluated by duplex scan of lower limb arteries. RESULTS Ninety-two consecutive HIV-infected patients were evaluated (23.9% women; mean age, 49.5 years; 61.9% current smokers). Claudication was reported by 15.2% of the patients. PAD was found in 20.7% of the patients: 9.8% had an abnormal ABI (<0.90) at rest, and 10.9% had normal ABI at rest but a >25% decrease after exercise. Of the patients with PAD, 84.2% were investigated with duplex scan, all of whom had atherosclerotic occlusions or stenoses of the iliac or femoral arteries. Age, diabetes, smoking, and low CD4+ T lymphocyte counts were identified as independent predictors of PAD. CONCLUSIONS The prevalence of symptomatic and asymptomatic PAD is high in the HIV-infected population and is much higher than expected (prevalence in the general population, approximately 3% at 60 years). This study suggests the presence of an epidemic of PAD approximately 20 years earlier in the HIV-infected than in the general population. Larger epidemiological studies are needed to better define risk factors and to evaluate whether PAD is associated with increased mortality, as it is in the general population.