Carmine Vecchione

Melusin, a muscle-specific integrin β1-interacting protein, is required to prevent cardiac failure in response to chronic pressure overload

Cardiac hypertrophy is an adaptive response to a variety of mechanical and hormonal stimuli, and represents an early event in the clinical course leading to heart failure. By gene inactivation, we demonstrate here a crucial role of melusin, a muscle-specific protein that interacts with the integrin β1 cytoplasmic domain, in the hypertrophic response to mechanical overload. Melusin-null mice showed normal cardiac structure and function in physiological conditions, but when subjected to pressure overload—a condition that induces a hypertrophic response in wild-type controls—they developed an abnormal cardiac remodeling that evolved into dilated cardiomyopathy and contractile dysfunction. In contrast, the hypertrophic response was identical in wild-type and melusin-null mice after chronic administration of angiotensin II or phenylephrine at doses that do not increase blood pressure—that is, in the absence of cardiac biomechanical stress. Analysis of intracellular signaling events induced by pressure overload indicated that phosphorylation of glycogen synthase kinase-3β (GSK-3β) was specifically blunted in melusin-null hearts. Thus, melusin prevents cardiac dilation during chronic pressure overload by specifically sensing mechanical stress.

Emilin1 links TGF-β maturation to blood pressure homeostasis

TGF-beta proteins are main regulators of blood vessel development and maintenance. Here, we report an unprecedented link between TGF-beta signaling and arterial hypertension based on the analysis of mice mutant for Emilin1, a cysteine-rich secreted glycoprotein expressed in the vascular tree. Emilin1 knockout animals display increased blood pressure, increased peripheral vascular resistance, and reduced vessel size. Mechanistically, we found that Emilin1 inhibits TGF-beta signaling by binding specifically to the proTGF-beta precursor and preventing its maturation by furin convertases in the extracellular space. In support of these findings, genetic inactivation of Emilin1 causes increased TGF-beta signaling in the vascular wall. Strikingly, high blood pressure observed in Emilin1 mutants is rescued to normal levels upon inactivation of a single TGF-beta1 allele. This study highlights the importance of modulation of TGF-beta availability in the pathogenesis of hypertension.

A Common Variant of Endothelial Nitric Oxide Synthase (Glu298Asp) Is an Independent Risk Factor for Carotid Atherosclerosis

Background and Purpose— Endothelium-derived NO is formed from l-arginine by endothelial NO synthase (eNOS) encoded by the NOS 3 gene on chromosome 7. Because several studies have indicated that NO plays a key role in the development of the atherosclerotic process, we investigated whether common variants in the eNOS gene are associated with an increased risk of plaque on carotid arteries. Methods— We studied 375 subjects attending the hypertension center of our institution to be screened for arterial hypertension. The examined subjects were classified according to the presence of carotid plaques (intima-media thickness ≥1.5 mm), and 2 intronic (CA and 27-bp repeats) polymorphisms and 1 exonic (Glu298Asp) polymorphism of the eNOS gene were explored. Results— Only the Glu298Asp polymorphism of eNOS was associated with the presence of carotid plaques (P <0.05). In particular, there was an excess of homozygotes for the Asp298 variant among subjects with carotid plaques, whereas the number of subjects who had the Glu298 allele in exon 7 of the eNOS gene was equally distributed in both study groups. Interestingly, the risk of having carotid plaques was increased ≈3 times in subjects who were homozygotic for the Asp298 variant compared with subjects who were homozygotic for the Glu298 variant and was independent of the other common risk factors (age, blood pressure, and smoking). Conclusions— Homozygosity for Asp298, a common variant of the eNOS gene, is an independent risk factor for carotid atherosclerosis in this study population.

Protection from angiotensin II–mediated vasculotoxic and hypertensive response in mice lacking PI3Kγ

Hypertension affects nearly 20% of the population in Western countries and strongly increases the risk for cardiovascular diseases. In the pathogenesis of hypertension, the vasoactive peptide of the renin-angiotensin system, angiotensin II and its G protein–coupled receptors (GPCRs), play a crucial role by eliciting reactive oxygen species (ROS) and mediating vessel contractility. Here we show that mice lacking the GPCR-activated phosphoinositide 3-kinase (PI3K)γ are protected from hypertension that is induced by administration of angiotensin II in vivo. PI3Kγ was found to play a role in angiotensin II–evoked smooth muscle contraction in two crucial, distinct signaling pathways. In response to angiotensin II, PI3Kγ was required for the activation of Rac and the subsequent triggering of ROS production. Conversely, PI3Kγ was necessary to activate protein kinase B/Akt, which, in turn, enhanced L-type Ca2+ channel–mediated extracellular Ca2+ entry. These data indicate that PI3Kγ is a key transducer of the intracellular signals that are evoked by angiotensin II and suggest that blocking PI3Kγ function might be exploited to improve therapeutic intervention on hypertension.

Cardiac Overexpression of Melusin Protects From Dilated Cardiomyopathy Due to Long-Standing Pressure Overload

We have previously shown that genetic ablation of melusin, a muscle specific &bgr; 1 integrin interacting protein, accelerates left ventricle (LV) dilation and heart failure in response to pressure overload. Here we show that melusin expression was increased during compensated cardiac hypertrophy in mice subjected to 1 week pressure overload, but returned to basal levels in LV that have undergone dilation after 12 weeks of pressure overload. To better understand the role of melusin in cardiac remodeling, we overexpressed melusin in heart of transgenic mice. Echocardiography analysis indicated that melusin over-expression induced a mild cardiac hypertrophy in basal conditions (30% increase in interventricular septum thickness) with no obvious structural and functional alterations. After prolonged pressure overload (12 weeks), melusin overexpressing hearts underwent further hypertrophy retaining concentric LV remodeling and full contractile function, whereas wild-type LV showed pronounced chamber dilation with an impaired contractility. Analysis of signaling pathways indicated that melusin overexpression induced increased basal phosphorylation of GSK3&bgr; and ERK1/2. Moreover, AKT, GSK3&bgr; and ERK1/2 were hyper-phosphorylated on pressure overload in melusin overexpressing compared with wild-type mice. In addition, after 12 weeks of pressure overload LV of melusin overexpressing mice showed a very low level of cardiomyocyte apoptosis and stromal tissue deposition, as well as increased capillary density compared with wild-type. These results demonstrate that melusin overexpression allows prolonged concentric compensatory hypertrophy and protects against the transition toward cardiac dilation and failure in response to long-standing pressure overload.

Association and cosegregation of stroke with impaired endothelium-dependent vasorelaxation in stroke prone, spontaneously hypertensive rats.

While hypertension is a major risk factor for stroke, it is not its sole determinant. Despite similar blood pressures, spontaneously hypertensive rats (SHR) do not share the predisposition to cerebrovascular disease typical of stroke-prone spontaneously hypertensive rats (SHRSP). We investigated vascular function in male SHR and SHRSP as well as in SHRSP/SHR-F2 hybrid animals. Animals were maintained on the appropriate dietary regimen necessary for the manifestation of stroke. Among the hybrid animals, a group of stroke-prone and a group of stroke-resistant rats were selected. Blood pressure was similar in all groups. Endothelium-independent vascular reactivity tested on isolated rings of thoracic aorta and basilar artery after death showed similar contractile and dilatory responses to serotonin and nitroglycerin, respectively, in all groups. In contrast, endothelium-dependent relaxation, in response to acetylcholine or substance P, was markedly reduced in SHRSP compared with SHR. Similarly, reduced vasodilatory responses were present in aortae of F2 rats that had suffered a stroke when compared with SHR or F2 rats resistant to stroke. The observed association and cosegregation of stroke with significant and specific impairment of endothelium-dependent vasorelaxation among SHRSP and stroke-prone F2 hybrids, respectively, suggest a potential causal role of altered endothelium-dependent vascular relaxation in the pathogenesis of stroke.

Systemic hypertension and coronary artery disease: The link

A direct, continuous, and independent association between blood pressure values and incidence of coronary artery disease has been well documented. However, the evidence that the reduction of blood pressure alone is not able to completely reverse the increase in the risk of coronary artery disease associated with essential hypertension suggests that the link between hypertension and coronary artery disease is a complex process including other factors beside the increase in blood pressure values. In this regard, the main determinant of coronary artery disease in hypertensive patients seems to be the development of left ventricular hypertrophy (LVH). In fact, hypertensive patients who died from sudden cardiac death showed a lesser degree of coronary atherosclerosis compared with normotensives, but a higher incidence of LVH. Several mechanisms can account for the increased coronary risk with LVH, including (1) an increase in left ventricular (LV) mass, which by itself requires more oxygen for tissue perfusion; (2) impairment of coronary flow reserve; (3) perivascular fibrosis, which then impairs oxygen supply to the myocardium; and (4) deterioration of LV diastolic function, which hampers myocardial perfusion. Recently, a study reported an impairment of endothelial function and abnormal control of the sympathetic tone in hypertensive patients, which may contribute to the risk of coronary artery disease. In particular, the impaired endothelial function resulting in a prevalence of vasoconstrictive, thrombogenic, and proliferative factors may account for the enhanced ischemic susceptibility of these patients. Furthermore, the cardiac adrenergic system plays an important role in regulating myocardial blood flow. On one hand, hypertensive patients show an exaggerated sympathetic response to physiologic stimuli, whereas on the other hand, the beta-adrenergic receptor-mediated vasodilating component of the sympathetic response is blunted in hypertension. Finally, excess body weight, dyslipidemia, glucose intolerance, and hyperinsulinemia, which are frequently interrelated, represent independent predictors of both coronary artery disease and hypertension.

Replacement of K-Ras with H-Ras supports normal embryonic development despite inducing cardiovascular pathology in adult mice

Ras proteins are highly related GTPases that have key roles in regulating growth, differentiation and tumorigenesis. Gene‐targeting experiments have shown that, out of the three mammalian ras genes, only K‐ras is essential for normal mouse embryogenesis, and that mice deprived of H‐ras and/or N‐ras show no major phenotype. We generated mice (HrasKI) in which the K‐ras gene had been modified to encode H‐Ras protein. HrasKI mice produce undetectable amounts of K‐Ras but—in contrast to mice homozygous for a null K‐ras allele—they are born at the expected mendelian frequency, indicating that H‐Ras can be substituted for K‐Ras in embryonic development. However, adult HrasKI mice show dilated cardiomyopathy associated with arterial hypertension. Our results show that K‐Ras can be replaced by H‐Ras in its essential function in embryogenesis, and indicate that K‐Ras has a unique role in cardiovascular homeostasis.

β-Amyloid deposition in brain is enhanced in mouse models of arterial hypertension

There are conflicting evidence regarding the association of hypertension with Alzheimers disease (AD), and so far it is still unexplored whether increased blood pressure levels can be mechanistically related to the pathophysiology of AD. Since the deposition of beta-amyloid (A beta) in brain represents the first pathogenetic event in the onset of AD, in this study we investigated the role of hypertension in the brain deposition of A beta. We analyzed two independent mouse models of hypertension. In both models we observed an increased permeability of blood-brain barrier in cortex and hippocampus. More interestingly, in the same areas hypertensive mice showed a marked positivity to anti-A beta antibodies and the presence of A beta-like fragments. Finally, we analyzed mice after passive immunotherapy with anti-A beta IgG. We observed that this latter approach determined a markedly reduced A beta immunopositivity in both cortex and hippocampus. Our study demonstrates that chronic hypertension determines an impairment of the blood-brain barrier permeability with deposition of A beta in brain tissue and that passive immunotherapy prevents this latter phenomenon.

Acute hypertension induces oxidative stress in brain tissues

Arterial hypertension is not only a major risk factor for cerebrovascular accidents, such as stroke and cerebral hemorrhage, but is also associated to milder forms of brain injury. One of the main causes of neurodegeneration is the increase in reactive oxygen species (ROS) that is also a common trait of hypertensive conditions, thus suggesting that such a mechanism could play a role even in the onset of hypertension-evoked brain injury. To investigate this issue, we have explored the effect of acute-induced hypertensive conditions on cerebral oxidative stress. To this aim, we have developed a mouse model of transverse aortic coarctation (TAC) between the two carotid arteries, which imposes acutely on the right brain hemisphere a dramatic increase in blood pressure. Our results show that hypertension acutely induced by aortic coarctation induces a breaking of the blood–brain barrier (BBB) and reactive astrocytosis through hyperperfusion, and evokes trigger factors of neurodegeneration such as oxidative stress and inflammation, similar to that observed in cerebral hypoperfusion. Moreover, the derived brain injury is mainly localized in selected brain areas controlling cognitive functions, such as the cortex and hippocampus, and could be a consequence of a defect in the BBB permeability. It is noteworthy to emphasize that, even if these latter events are not enough to produce ischemic/hemorrhagic injury, they are able to alter mechanisms fundamental for maintaining normal brain function, such as protein synthesis, which has a prominent role for memory formation and cortical plasticity.