Abstract-Nitric oxide (NO) is a powerful angiogenic mediator acting downstream of vascular endothelial growth factor (VEGF). Both the endothelial NO synthase (eNOS) and the VEGFR-2 receptor colocalize in caveolae. Because the structural protein of these signaling platforms, caveolin, also represses eNOS activity, changes in its abundance are likely to influence the angiogenic process in various ways. In this study, we used mice deficient for the caveolin-1 gene (Cav Ϫ/Ϫ ) to examine the impact of caveolae suppression in a model of adaptive angiogenesis obtained after femoral artery resection. Evaluation of the ischemic tissue perfusion and histochemical analyses revealed that contrary to Cav ϩ/ϩ mice, Cav Ϫ/Ϫ mice failed to recover a functional vasculature and actually lost part of the ligated limbs, thereby recapitulating the effects of the NOS inhibitor L-NAME administered to operated Cav ϩ/ϩ mice. We also isolated endothelial cells (ECs) from Cav Ϫ/Ϫ aorta and showed that on VEGF stimulation, NO production and endothelial tube formation were dramatically abrogated when compared with Cav ϩ/ϩ ECs. The Ser1177 eNOS phosphorylation and Thr495 dephosphorylation but also the ERK phosphorylation were similarly altered in VEGF-treated Cav Ϫ/Ϫ ECs. Interestingly, caveolin transfection in Cav Ϫ/Ϫ ECs redirected the VEGFR-2 in caveolar membranes and restored the VEGF-induced ERK and eNOS activation. However, when high levels of recombinant caveolin were reached, VEGF exposure failed to activate ERK and eNOS. These results emphasize the critical role of caveolae in ensuring the coupling between VEGFR-2 stimulation and downstream mediators of angiogenesis. This study also provides new insights to understand the paradoxical roles of caveolin (eg, repressing basal enzyme activity but facilitating activation on agonist stimulation) in cardiovascular pathophysiology. Key Words: caveolin-1 Ⅲ nitric oxide Ⅲ vascular endothelial growth factor Ⅲ angiogenesis Ⅲ ischemia C aveolae are 50-to 100-nm cell surface invaginations playing key roles in vesicular transport and signal transduction. 1 The structural protein of these plasmalemmal microdomains, caveolin, acts as a scaffold for many caveolar residents. 2 The caveolin-1 isoform is particularly abundant in endothelial cells (ECs) where it regulates various functions including transcytosis, permeability, vascular tone, and angiogenesis. 3 Recently, Woodman et al 4 documented that in a model of tumor cell injection in caveolin-deficient mice (Cav Ϫ/Ϫ ), angiogenesis was markedly reduced in comparison with wild-type (WT) animals. Although the same authors showed that the reduction in vessel density could be reproduced in a model of Matrigel plugs supplemented with bFGF, 4 the mechanisms supporting the role of caveolin in the angiogenic response to exogenous stimuli remain poorly understood and, based on previous publications, 5-9 a matter of debate.For instance, the well-established inhibitory interaction between caveolin and the endothelial nitric oxide (NO) synthase (eNOS) 10,11 le...

Background-Decreased heart rate variability (HRV) and increased blood pressure variability (BPV), determined in part by nitric oxide (NO)-dependent endothelial dysfunction, are correlated with adverse prognosis in cardiovascular diseases. We examined potential alterations in BPV and HRV in genetically dyslipidemic, apolipoprotein (apo) E Ϫ/Ϫ , and control mice and the effect of chronic statin treatment on these parameters in relation to their NO synthase (NOS)-modifying properties. Methods and Results-BP and HR were recorded in unrestrained, nonanesthetized mice with implanted telemetry devices with or without rosuvastatin. Cardiac and aortic expression of endothelial NOS and caveolin-1 were measured by immunoblotting. Both systolic BP and HR were elevated in apoE Ϫ/Ϫ mice, with abolition of their circadian cycles. Spectral analysis showed an increase in their systolic BPV in the very-low-frequency (ϩ17%) band and a decrease in HRV in the high-frequency (Ϫ57%) band, reflecting neurohumoral and autonomic dysfunction. Decreased sensitivity to acute injection of atropine or an NOS inhibitor indicated basal alterations in both parasympathetic and NOS regulatory systems in apoE Ϫ/Ϫ mice. Aortic caveolin-1 protein, an inhibitor of endothelial NOS, was also increased in these mice by 2.0-fold and correlated positively with systolic BPV in the very-low-frequency band. Rosuvastatin treatment corrected the hemodynamic and caveolin-1 expression changes despite persisting elevated plasma cholesterol levels. Conclusions-Rosuvastatin

Abstract-Ischemic preconditioning confers powerful protection against myocardial infarction through pre-emptive activation of survival signaling pathways, but it remains difficult to apply to patients with ischemic heart disease, and its effects are transient. Promoting a sustained activation of preconditioning mechanisms in vivo would represent a novel approach of cardioprotection. We tested the role of the protein H11 kinase (H11K), which accumulates by 4-to 6-fold in myocardium of patients with chronic ischemic heart disease and in experimental models of ischemia. This increased expression was quantitatively reproduced in cardiac myocytes using a transgenic (TG) mouse model. After 45 minutes of coronary artery occlusion and reperfusion, hearts from TG mice showed an 82Ϯ5% reduction in infarct size compared with wild-type (WT), which was similar to the 84Ϯ4% reduction of infarct size observed in WT after a protocol of ischemic preconditioning. Hearts from TG mice showed significant activation of survival kinases participating in preconditioning, including Akt and the 5ЈAMP-activated protein kinase (AMPK). H11K directly binds to both Akt and AMPK and promotes their nuclear translocation and their association in a multiprotein complex, which results in a stimulation of survival mechanisms in cytosol and nucleus, including inhibition of proapoptotic effectors (glycogen synthase kinase-3␤, Bad, and Foxo), activation of antiapoptotic effectors (protein kinase C⑀, endothelial and inducible NO synthase isoforms, and heat shock protein 70), increased expression of the hypoxia-inducible factor-1␣, and genomic switch to glucose utilization. Therefore, activation of survival pathways by H11K preemptively triggers the antiapoptotic and metabolic response to ischemia and is sufficient to confer cardioprotection in vivo equally potent to preconditioning. (Circ Res. 2006;98:280-288.)

Background-Weight loss in obese insulin-resistant but not in insulin-sensitive persons reduces coronary heart disease risk. To what extent changes in gene expression are related to atherosclerosis and cardiovascular function is unknown. Methods and Results-We studied the effect of diet restriction-induced weight loss on gene expression in the adipose tissue, the heart, and the aortic arch and on atherosclerosis and cardiovascular function in mice with combined leptin and LDL-receptor deficiency. Obesity, hypertriglyceridemia, and insulin resistance are associated with hypertension, impaired left ventricular function, and accelerated atherosclerosis in those mice. Compared with lean mice, peroxisome proliferator-activated receptors (PPAR)-␣ and PPAR-␥ expression was downregulated in obese double-knockout mice. Diet restriction caused a 45% weight loss, an upregulation of PPAR-␣ and PPAR-␥, and a change in the expression of genes regulating glucose transport and insulin sensitivity, lipid metabolism, oxidative stress, and inflammation, most of which are under the transcriptional control of these PPARs. Changes in gene expression were associated with increased insulin sensitivity, decreased hypertriglyceridemia, reduced mean 24-hour blood pressure and heart rate, restored circadian variations of blood pressure and heart rate, increased ejection fraction, and reduced atherosclerosis. PPAR-␣ and PPAR-␥ expression was inversely related to plaque volume and to oxidized LDL content in the plaques. Conclusions-Induction of PPAR-␣ and PPAR-␥ in adipose tissue, heart, and aortic arch is a key mechanism for reducing atherosclerosis and improving cardiovascular function resulting from weight loss. Improved lipid metabolism and insulin signaling is associated with decreased tissue deposition of oxidized LDL that increases cardiovascular risk in persons with the metabolic syndrome. Key Words: atherosclerosis Ⅲ circadian rhythm Ⅲ genes Ⅲ lipoproteins Ⅲ obesity I nsulin resistance is now receiving increasing attention not only as a precursor to type 2 diabetes but also as a predictor of increased risk of cardiovascular disease. 1 Fat distributed in the abdominal region is a risk factor for type 2 diabetes and cardiovascular disease and is associated closely with insulin resistance. 2 Weight loss in insulin-resistant but not in insulinsensitive obese persons reduces their risk of coronary heart disease (CHD). 3 It is not known, however, to what extent changes in the intra-abdominal adipose gene expression profile are important for the reduction of the risk. 4 Several adipokines, and more specifically peroxisome proliferator-activated receptors (PPARs), regulate a number of the processes that contribute to the development of atherosclerosis, including dyslipidemia, arterial hypertension, endothelial dysfunction, insulin resistance, and vascular remodeling. Adipokines are preferentially expressed in intraabdominal adipose tissue, and the secretion of proinflammatory adipokines is elevated with increasing adiposity. Approaches to re...

Background-In the heart, nitric oxide synthases (NOS) modulate cardiac contraction in an isoform-specific manner, which is critically dependent on their cellular and subcellular localization. Defective NO production by NOS3 (endothelial NOS [eNOS]) in the failing heart may precipitate cardiac failure, which could be reversed by overexpression of NOS3 in the myocardium. Methods and Results-We studied the influence of NOS3 in relation to its subcellular localization on the function of cardiomyocytes isolated from transgenic mice overexpressing NOS3 under the ␣-myosin heavy chain promoter (NOS3-TG

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These data highlight the functional importance of cav-1 for the production of bioactive NO in conduit arteries and its control of central BP variability. Given the impact of the latter on target organ damage, this raises the interest for genetic, pharmacological, or molecular interventions that modulate cav-1 expression in diseases with NO-dependent endothelial dysfunction.

Beyond partial improvement in insulin sensitivity, rosuvastatin normalized BP homeostasis in obese dyslipidaemic mice independently of changes in body weight or plasma cholesterol. Upregulation of PPARgamma and SOD1 in the endothelium may be involved as a unique vasculoprotective effect of statin treatment.