Abstract-To seek evidence that the nonhuman primate arterial wall, as it ages in the absence of atherosclerosis, exhibits alterations in pathways that are involved in the pathogenesis of experimental atherosclerosis, we assessed aortic matrix metalloproteinase-2 (MMP-2) and its regulators, ie, membrane type-1 of matrix metalloproteinase (MT1-MMP) and tissue inhibitor of matrix metalloproteinase-2 (TIMP-2), and the expression of angiotensin II (Ang II), angiotensinconverting enzyme (ACE), and chymase in young (6.4Ϯ0.7 years) and old (20.0Ϯ1.9 years) male monkeys. With advancing age, (1) the intimal thickness increased 3-fold and contained numerous vascular smooth muscle cells and matrix, but no inflammatory cells; (2) the intimal MMP-2 antibody-staining fraction increased by 80% (PϽ0.01); (3) in situ zymography showed that MMP-2 activity, mainly confined to the intima, increased 3-fold (PϽ0.01); (4) the MT1-MMP antibody-staining fraction increased by 150% (PϽ0.001), but the TIMP-2 antibody-staining fraction did not significantly change; (5) steady levels of the mRNA-staining fraction (via in situ hybridization) for MMP-2 increased 7-fold, for MT1-MMP increased 9-fold, and for TIMP-2 increased 2-fold (all PϽ0.001); and (6) intimal Ang II and ACE immunofluorescence were increased 5-fold and 5.6-fold, respectively, and colocalized with MMP-2. Thus, age-associated arterial remodeling and the development and progression of experimental atherosclerosis in young animals share common mechanisms, ie, MMP-2 activation and increased Ang II signaling. This might explain, in part, the dramatically exaggerated prevalence and severity of vascular diseases with aging.

The sympathetic nervous system is designed to respond to stress. Adenylyl cyclase (AC) is the keystone of sympathetic transmission, yet its role in response to acute overload in the heart or in the pathogenesis of heart failure is controversial. We examined the effects of pressure overload, induced by thoracic aortic banding, in mice in which type 5 AC, a major cardiac AC isoform, was disrupted (AC5 ؊/؊ ). Left ventricular weight͞tibial length ratio (LVW͞TL) was not different between the WT and AC5 ؊/؊ at baseline and increased progressively and similarly in both groups at 1 and 3 wk after aortic banding. However, LV ejection fraction (LVEF) fell in WT at 3 wk after banding (from 70 ؎ 2.8 to 57 ؎ 3.9%, P < 0.05), and this decrease was associated with LV dilatation, indicating incipient cardiac failure. In contrast, AC5 ؊/؊ mice did not exhibit a fall in LVEF from 74 ؎ 2.2%. The number of apoptotic myocytes was similar at baseline, but it increased roughly 4-fold in WT at both 1 and 3 wk after banding, and significantly less, P < 0.05, in AC5 ؊/؊ . Importantly, the increase in apoptosis occurred before the decline in LVEF in WT. The protective mechanism seems to involve Bcl-2, which was up-regulated significantly more in AC5 ؊/؊ mice with pressure overload. Our findings suggest that limiting type 5 AC plays a protective role in response to pressure overload and the development of heart failure, potentially through limiting the incidence of myocardial apoptosis.

Abstract-To determine the effects of aging on vasoactivity in a primate model (Macaca fascicularis), 13 young male monkeys (aged 7.1Ϯ0.4 years) and 9 old male monkeys (aged 19.8Ϯ0.6 years) were chronically instrumented for measurement of left ventricular and aortic pressures and cardiac output. Total cholesterol, triglyceride, and fasting blood sugar levels were not different between the 2 groups. There were no significant differences in baseline mean aortic pressure and total peripheral resistance (TPR) in the young monkeys versus the old monkeys. TPR fell less (PϽ0.05) with acetylcholine (1 g/kg) in old monkeys (Ϫ25Ϯ1%) than in young monkeys (Ϫ34Ϯ2%), whereas decreases in TPR with sodium nitroprusside were similar in old and young monkeys. There was no evidence of atherosclerosis, but apoptosis of endothelial cells was enhanced (PϽ0.05) in the aortas and femoral arteries, but not in the media, of the old monkeys. There was a relationship (rϭ0.62, Pϭ0.013) between the incidence of terminal deoxynucleotidyl transferasemediated dUTP nick end-labeling (TUNEL)-positive endothelial cells and endothelial cell density in the femoral artery. The reduced endothelial cell density was also correlated (rϭ0.82, PϽ0.01) with depressed TPR responses to acetylcholine. Thus, vascular endothelial dysfunction was present in old monkeys without evidence of atherosclerosis, which may be due to endothelial apoptosis and reduced endothelial cell density. (Arterioscler Thromb Vasc Biol.

Abstract-Left ventricular hypertrophy (LVH) is a compensatory mechanism to cope with pressure overload. Recently, a calcineurin pathway mediating LVH and its prevention by cyclosporine was reported. We examined whether calcineurin mediates LVH due to pressure overload in mice. Pressure overload was induced by aortic banding in 53 mice (32 treated with cyclosporine [25 mg ⅐ kg], 21 treated with vehicle). There were 17 sham-operated mice (9 treated with vehicle, 8 treated with cyclosporine). At 3 weeks after surgery, LV weight to body weight was greater in the nontreatment banded group (4.39Ϯ0.16 mg/g) than in the cyclosporine-treated banded group (3.95Ϯ0.14 mg/g, PϽ0.05), with both groups being greater compared with the entire group of sham-operated mice (3.02Ϯ0.04 mg/g). The pressure gradient between the ascending and abdominal aorta was not different between the cyclosporine-treated (49.6Ϯ6.1 mm Hg) and nontreatment groups (48.7Ϯ4.6 mm Hg). Although LV systolic pressure was lower in the cyclosporine-treated banded animals, LV systolic wall stress was similar in the nontreatment banded group and in the cyclosporine-treated group. However, LV dP/dt was lower (Pϭ0.05) in the cyclosporine-treated banded group (4774Ϯ656 mm Hg/s) than in the nontreatment banded group (6604Ϯ516 mm Hg/s). During the protocol, 23 of 32 mice in the cyclosporine-treated group and 9 of 21 mice in the nontreatment group died. All deaths occurred within 10 days after surgery. Deaths caused by heart failure were 7.2-fold higher (PϽ0.05) in the cyclosporine-treated group, whereas deaths due to other causes were not different between the 2 groups. In addition, LV function of mice was assessed at 48 hours after banding; LV ejection fraction measured with echocardiography was lower (PϽ0.05) in the cyclosporine-treated banded group (66Ϯ3.0%) than in the nontreatment banded group (79Ϯ1.5%), whereas LV systolic wall stresses were similar. Calcineurin phosphatase activity was depressed similarly in both cyclosporine-treated groups compared with both nontreatment groups. Thus, cyclosporine could attenuate, but not prevent, LVH at the expense of inhibiting an important compensatory mechanism in response to pressure overload, resulting in reduced LV wall stress and function and increased susceptibility to decompensation and heart failure. 10 -12 Recently, Molkentin et al 10 reported that cyclosporine blocks this pathway in vitro and prevents hypertrophy in transgenic mice, in which the calcineurin pathway is enhanced. It is not clear whether this pathway is universal, ie, whether it mediates pressureoverload LVH as well as hypertrophy in transgenic models. An equally important question is whether LVH is salutary or deleterious, ie, if LVH is a beneficial compensatory mechanism, then blocking this mechanism may be deleterious and lead to decompensation and development of heart failure.Accordingly, the first goal of the present study was to determine whether cyclosporine prevents LVH due to aortic banding-induced pressure overload in mice. The second go...

In a genetically engineered mouse line with disruption of type 5 adenylyl cyclase (AC5-/-), a major cardiac isoform, there was no compensatory increase in other isoforms of AC in the heart. Both basal and isoproterenol (ISO)-stimulated AC activities were decreased by 30% to 40% in cardiac membranes. The reduced AC activity did not affect cardiac function (left ventricular ejection fraction [LVEF]) at baseline. However, increases in LVEF after ISO were significantly attenuated in AC5-/- (P<0.05, n=11). Paradoxically, conscious AC5-/- mice had a higher heart rate compared with wild-type (WT) mice (613+/-8 versus 523+/-11 bpm, P<0.01, n=14 to 15). Muscarinic agonists decreased AC activity, LVEF, and heart rate more in WT than in AC5-/-. In addition, baroreflex-mediated, ie, neuronally regulated, bradycardia after phenylephrine was also attenuated in AC5-/-. The carbachol-activated outward potassium current (at -40 mV) normalized to cell capacitance in AC5-/- (2.6+/-0.4 pA/pF, n=16) was similar to WT (2.9+/-0.3 pA/pF, n=27), but calcium (Ca2+)-mediated inhibition of AC activity and Ca2+ channel function were diminished in AC5-/-. Thus, AC5-/- attenuates sympathetic responsiveness and also impairs parasympathetic and Ca2+-mediated regulation of the heart, indicating that those actions are not only regulated at the level of the receptor and G-protein but also at the level of type 5 AC.