Large Elastic Artery Stiffness Research Articles

Inorganic nitrite supplementation for healthy arterial aging.

Aging is the major risk factor for cardiovascular diseases (CVD). This is attributable primarily to adverse changes in arteries, notably, increases in large elastic artery stiffness and endothelial dysfunction mediated by inadequate concentrations of the vascular-protective molecule, nitric oxide (NO), and higher levels of oxidative stress and inflammation. Inorganic nitrite is a promising precursor molecule for augmenting circulating and tissue NO bioavailability because it requires only a one-step reduction to NO. Nitrite also acts as an independent signaling molecule, exerting many of the effects previously attributed to NO. Results of recent studies indicate that nitrite may be effective in the treatment of vascular aging. In old mice, short-term oral sodium nitrite supplementation reduces aortic pulse wave velocity, the gold-standard measure of large elastic artery stiffness, and ameliorates endothelial dysfunction, as indicated by normalization of NO-mediated endothelium-dependent dilation. These improvements in age-related vascular dysfunction with nitrite are mediated by reductions in oxidative stress and inflammation, and may be linked to increases in mitochondrial biogenesis and health. Increasing nitrite levels via dietary intake of nitrate appears to have similarly beneficial effects in many of the same physiological and clinical settings. Several clinical trials are being performed to determine the broad therapeutic potential of increasing nitrite bioavailability on human health and disease, including studies related to vascular aging. In summary, inorganic nitrite, as well as dietary nitrate supplementation, represents a promising therapy for treatment of arterial aging and prevention of age-associated CVD in humans.

Editorial: Do We Have Effective Means to Treat Arterial Stiffness and High Central Aortic Blood Pressure in Patients with and without Hypertension?

The reduction or loss of arterial elasticity or distensibility leads to arterial stiffness (AS), which has a substantial predictive value for all-cause and cardiovascular disease (CVD) mortality, as well as for non-fatal CVD events [1]. A plethora of evidence consistently showed the prognostic value of aortic stiffness for fatal and nonfatal CVD events in various populations at different levels of CVD risk, including the general population, elderly subjects and patients with hypertension, type 2 diabetes mellitus (T2DM) and end-stage renal disease (ESRD) [2]. It has been reported that 1-SD increase in pulse wave velocity (PWV) is associated with a 47% increase in the risk for total mortality [95% confidence interval (CI), 1.31-1.64] and a similar 47% increase in the risk for CVD mortality (95% CI, 1.29-1.66) [2].

Vascular calcification in end‐stage renal disease

Vascular calcification is highly prevalent in end-stage renal disease and independently predictive of future cardiovascular events and mortality. Calcification can occur in both the intimal and medial layers of vasculature, but medial calcification is the major form in end-stage renal disease. Medial calcification increases large elastic artery stiffness and pulse-pressure, promotes left ventricular hypertrophy, reduces perfusion of the coronary arteries, and ultimately promotes increased cardiovascular mortality via increased risk of myocardial infarction and heart failure. It results not from a passive deposition of calcium and phosphate due to increased circulating levels, but rather is an active cell-mediated process involving vascular smooth muscle cell apoptosis and vesicle release, a shift in the balance of inhibitors and promoters of vascular calcification, and vascular smooth muscle cell differentiation from a contractile to osteochondrogenic phenotype. This phenotypic shift requires phosphate, as well as the uptake of phosphate by the sodium-dependent phosphate cotransporter PiT-1, which is upregulated by proinflammatory cytokines and the uremic milieu. Further research is needed to determine if targeting these processes can ultimately reduce vascular calcification in this high cardiovascular risk population.

Dietary Sodium Restriction and Association with Urinary Marinobufagenin, Blood Pressure, and Aortic Stiffness

Systolic BP and large elastic artery stiffness both increase with age and are reduced by dietary sodium restriction. Production of the natriuretic hormone marinobufagenin, an endogenous α1 Na+,K+-ATPase inhibitor, is increased in salt-sensitive hypertension and contributes to the rise in systolic BP during sodium loading. The hypothesis was that dietary sodium restriction performed in middle-aged/older adults (eight men and three women; 60 ± 2 years) with moderately elevated systolic BP (139 ± 2/83 ± 2 mmHg) would reduce urinary marinobufagenin excretion as well as systolic BP and aortic pulse-wave velocity (randomized, placebo-controlled, and crossover design). This study also explored the associations among marinobufagenin excretion with systolic BP and aortic pulse-wave velocity across conditions of 5 weeks of a low-sodium (77 ± 9 mmol/d) and 5 weeks of a normal-sodium (144 ± 7 mmol/d) diet. Urinary marinobufagenin excretion (weekly measurements; 25.4 ± 1.8 versus 30.7 ± 2.1 pmol/kg per day), systolic BP (127 ± 3 versus 138 ± 5 mmHg), and aortic pulse-wave velocity (700 ± 40 versus 843 ± 36 cm/s) were lower during the low- versus normal-sodium condition (all P<0.05). Across all weeks, marinobufagenin excretion was related with systolic BP (slope=0.61, P<0.001) and sodium excretion (slope=0.46, P<0.001). These associations persisted during the normal- but not the low-sodium condition (both P<0.005). Marinobufagenin excretion also was associated with aortic pulse-wave velocity (slope=0.70, P=0.02) and endothelial cell expression of NAD(P)H oxidase-p47phox (slope=0.64, P=0.006). These results show, for the first time in humans, that dietary sodium restriction reduces urinary marinobufagenin excretion and that urinary marinobufagenin excretion is positively associated with systolic BP, aortic stiffness (aortic pulse-wave velocity), and endothelial cell expression of the oxidant enzyme NAD(P)H oxidase. Importantly, marinobufagenin excretion is positively related to systolic BP over ranges of sodium intake typical of an American diet, extending previous observations in rodents and humans fed experimentally high-sodium diets.

The autophagy enhancer spermidine reverses arterial aging

Arterial aging, characterized by stiffening of large elastic arteries and the development of arterial endothelial dysfunction, increases cardiovascular disease (CVD) risk. We tested the hypothesis that spermidine, a nutrient associated with the anti-aging process autophagy, would improve arterial aging. Aortic pulse wave velocity (aPWV), a measure of arterial stiffness, was ∼20% greater in old (O, 28 months) compared with young C57BL6 mice (Y, 4 months, P<0.05). Arterial endothelium-dependent dilation (EDD), a measure of endothelial function, was ∼25% lower in O (P<0.05 vs. Y) due to reduced nitric oxide (NO) bioavailability. These impairments were associated with greater arterial oxidative stress (nitrotyrosine), superoxide production, and protein cross-linking (advanced glycation end-products, AGEs) in O (all P<0.05). Spermidine supplementation normalized aPWV, restored NO-mediated EDD and reduced nitrotyrosine, superoxide, AGEs and collagen in O. These effects of spermidine were associated with enhanced arterial expression of autophagy markers, and in vitro experiments demonstrated that vascular protection by spermidine was autophagy-dependent. Our results indicate that spermidine exerts a potent anti-aging influence on arteries by increasing NO bioavailability, reducing oxidative stress, modifying structural factors and enhancing autophagy. Spermidine may be a promising nutraceutical treatment for arterial aging and prevention of age-associated CVD.

Perivascular adipose tissue contributes to large elastic artery stiffness with aging and is associated with greater superoxide bioavailability

We tested the hypothesis that perivascular adipose tissue (PVAT) contributes to age‐related large elastic artery stiffening and is associated with greater superoxide production. PVAT from young (Y, 6 mo, n=5) and old (O, 27 mo, n=5) donor C57BL6/N mice was transplanted directly onto the abdominal aorta of Y recipient (6 mo) mice for 8 weeks. Y recipient mice transplanted with PVAT from O compared with Y donors had greater arterial stiffness as assessed by aortic pulse wave velocity (409 ± 7 vs. 342 ± 8 cm/s, P &lt; 0.05) and ex vivo intrinsic mechanical stiffness (3197 ± 647 vs. 1889 ± 520 kPa, P &lt; 0.05). Systolic (109 ± 5 vs. 111 ± 2 mmHg), diastolic (75 ± 5 vs. 82 ± 2 mmHg) and mean (86 ± 5 vs. 91 ± 2 mmHg) blood pressure did not differ between groups (n=3–4). Additional aortic segments from Y and O mice with (+) or without (−) PVAT (n=5–6) were cultured for 72 hours followed by intrinsic mechanical testing. Aortic segments from O+PVAT (3947 ± 276 kPa, P &lt;0.05) had greater stiffness compared with Y+PVAT, Y−PVAT and O‐PVAT (2437 ± 226, 1768 ± 132 and 2668 ± 404 kPa, respectively); O‐PVAT had greater mechanical stiffening than Y−PVAT (P &lt; 0.05). PVAT superoxide production was greater in O vs. Y (13835 ± 2322 vs. 6657 ± 1720 AU, P &lt; 0.05, n=5–6). Collectively, these data demonstrate PVAT contributes to large artery stiffening with aging and is associated with increased superoxide bioavailability.

Circulating plasma factors contribute to age‐associated arterial stiffness

Previous findings from our laboratory showed that age‐related increases in large elastic artery stiffness in mice are associated with remodeling of the extracellular matrix (ECM), as indicated by increased collagen I expression, and oxidative stress, as indicated by increased nitrotyrosine staining. Here we tested the hypothesis that circulating factors in plasma contribute to arterial oxidative stress, ECM remodeling and stiffening. Nitrotyrosine, collagen I and elastic modulus, a measure of intrinsic mechanical stiffness, were determined in aortic rings from young (6 mo, n=4) male C57 mice cultured for 72 hours in media containing 10% plasma from either young (Yplasma) or old (Oplasma) mice. Compared with Yplasma, aortic rings incubated in Oplasma demonstrated increased NT abundance (3.11 ± 0.50 vs. 1.00 ± 0.30 AU, p&lt;0.05), collagen I (1.66 ± 0.18 vs. 1.00 ± 0.12 AU, p&lt;0.05) and elastic modulus (2780 ± 570 vs. 1768 ± 188 kPa, p=0.05). These preliminary results are consistent with the hypothesis that unknown factors in the circulating plasma induce “aging‐like” oxidative stress, ECM remodeling featuring increases in collagen I, and stiffening of the aorta. Identifying these circulating factors may lead to additional therapeutic targets for the prevention and treatment of ageassociated large elastic artery stiffening and clinical cardiovascular diseases.Supported by NIH AG013038, AG000279, HL007822

Curcumin ameliorates arterial dysfunction and oxidative stress with aging

We tested the hypothesis that curcumin supplementation would reverse arterial dysfunction and vascular oxidative stress with aging. Young (Y, 4–6months) and old (O, 26–28months) male C57BL6/N mice were given normal or curcumin supplemented (0.2%) chow for 4weeks (n=5–10/group/measure). Large elastic artery stiffness, assessed by aortic pulse wave velocity (aPWV), was greater in O (448±15 vs. 349±15cm/s) and associated with greater collagen I and advanced glycation end-products and less elastin (all P<0.05). In O, curcumin restored aPWV (386±15cm/s), collagen I and AGEs (AGEs) to levels not different vs. Y. Ex vivo carotid artery acetylcholine (ACh)-induced endothelial-dependent dilation (EDD, 79±3 vs. 94±2%), nitric oxide (NO) bioavailability and protein expression of endothelial NO synthase (eNOS) were lower in O (all P<0.05). In O, curcumin restored NO-mediated EDD (92±2%) to levels of Y. Acute ex vivo administration of the superoxide dismutase (SOD) mimetic TEMPOL normalized EDD in O control mice (93±3%), but had no effect in Y control or O curcumin treated animals. O had greater arterial nitrotyrosine abundance, superoxide production and NADPH oxidase p67 subunit expression, and lower manganese SOD (all P<0.05), all of which were reversed with curcumin. Curcumin had no effects on Y. Curcumin supplementation ameliorates age-associated large elastic artery stiffening, NO-mediated vascular endothelial dysfunction, oxidative stress and increases in collagen and AGEs in mice. Curcumin may be a novel therapy for treating arterial aging in humans.

Sodium nitrite de-stiffening of large elastic arteries with aging: Role of normalization of advanced glycation end-products

We tested the hypothesis that sodium nitrite treatment reverses large elastic artery stiffening in old mice via reductions in collagen I, increases in elastin and/or decreases in advanced glycation end products (AGEs) mediated by reduced oxidative stress. Aortic pulse wave velocity (aPWV), a measure of large elastic artery stiffness, was greater in old (26–28months) compared with young (4–6months) control animals (520±9 vs. 405±6cm/s, p<0.05), and this was reversed by 3weeks of sodium nitrite treatment (50mg/L) (435±17cm/s). Age-related increases (p<0.05) in aortic superoxide production were associated with greater total and adventitial nitrotyrosine staining, all of which were reversed by nitrite treatment. Total and adventitial transforming growth factor β and collagen I were increased, and total and medial elastin were reduced with aging (p<0.05), but were unaffected by sodium nitrite. Aorta from old mice had increased total, adventitial and medial AGEs (p<0.05 vs. young), which were normalized by sodium nitrite treatment. In aortic segments from young mice in vitro, pyrogallol (10μM), a superoxide generator, induced an “aging-like” increase in AGEs, and direct treatment with AGEs induced vascular stiffening; these effects were prevented by incubation with sodium nitrite. De-stiffening of aged large elastic arteries by short-term sodium nitrite therapy is mediated in part by normalization of AGEs secondary to amelioration of oxidative stress.

Mitochondria‐targeted antioxidant therapy reverses age‐related arterial stiffening

Aging leads to large elastic artery stiffening mediated by increases in superoxide‐dependent oxidative stress and related changes/damage to structural proteins. Mitochondria are a major source of cellular superoxide and may therefore be a therapeutic target for the treatment of arterial stiffening. We studied young (Y: 5 mo, n=5) and old (O: 27–29 mo, n=5) male C57BL6 mice, and O mice treated with the mitochondria targeted antioxidant MitoQ10 (OMQ: 28–29 mo, n=5). Aortic pulse wave velocity (aPWV), an index of arterial stiffness, was higher in O vs. Y mice (485 ± 24 vs. 405 ± 15 cm/s, p&lt;0.05) and was associated with increased aortic superoxide production (electron paramagnetic resonance, O: 7200 ± 1950 vs. Y: 2800 ± 515 AU, p&lt;0.05). O mice also had greater aortic levels of nitrotyrosine (NT), a marker of oxidative stress, and lower levels of manganese (mitochondrial) superoxide dismutase (MnSOD) (western blotting, both p&lt;0.05 vs. Y). MQ treatment (250 μM in water x 3w) normalized aortic stiffening (OMQ: 378 ± 33 cm/s), superoxide production (OMQ: 2835 ± 1131 AU), MnSOD and NT in O mice. MQ also increased expression of complex IV and the mitochondrial biogenesis marker PGC‐1α (both p&lt;0.05 vs. O). Incubation of aortas from Y mice with the mitochondrial superoxide inducer rotenone (0.5 μM, 48h) increased NT by ~300% (p&lt;0.05) and this was inhibited by coincubation with MQ, an effect associated with enhanced MnSOD expression. MQ treatment may be a novel therapy for age‐related stiffening of large elastic arteries by reducing mitochondrial oxidative stress and perhaps increasing mitochondrial biogenesis.NIH AG013038, AG039210

Polyamine supplementation enhances autophagy and reverses age‐related arterial stiffening

Aging leads to stiffening of large elastic arteries as a result of changes in structural proteins and increased macromolecular damage. Polyamines (PA) play a causal role in survival/longevity and have been reported to induce autophagy, the cellular process of recycling damaged biomolecules. However, the potential for PA to reduce age‐related arterial stiffness is unknown. We studied young (Y: 5 mo, n=7) and old (O: 28–29 mo, n=7) male C57/BL6 mice, and old mice treated with the PA spermidine (OPA: 29 mo, n=7). Aortic pulse wave velocity (aPWV), an index of arterial stiffness, was higher in O vs. Y mice (465 ± 25 vs. 383 ± 21 cm/s, p&lt;0.05). This was associated with ~50% greater aortic levels of both protein carbonyls, an index of oxidative damage, and advanced glycation end products (AGEs), a marker of protein cross‐linking (dot and western blotting, both p&lt;0.05, O vs. Y). PA supplementation (3 mM drinking water × 4w) ameliorated aortic stiffening in old mice (OPA aPWV pre: 482 ± 17 vs. OPA post: 343 ± 14 cm/s, P&lt;0.05) and reversed age‐related increases in protein carbonyls and AGEs. The effects of PA treatment were associated with enhanced autophagy as indicated by an ~20% reduction in acetylation (inhibition) of histone H3, an ~25% increase in expression of the transcription factors AMPK and FOXO3a, and a corresponding increase in LC3‐II and decrease in p62, markers of autophagy and autophagic protein aggregation/damage, respectively (western blotting, all p&lt;0.05, O vs. OPA). These results suggest that PA supplementation may be a novel strategy for reversing age‐related arterial stiffening, perhaps by enhancing autophagic clearance of damaged/aggregated proteins.NIH AG013038, AG039210, HL007822

Advanced Glycation End‐Products in Age‐Related Arterial Stiffening: Modulation by Sodium Nitrite

Aging causes large elastic artery stiffening and this is associated with oxidative stress and increased advanced glycation end‐products (AGEs). We tested the hypothesis that vascular oxidative stress with aging induces AGEs, which increase intrinsic mechanical stiffness, and that sodium nitrite prevents this stiffening. Old (O, 27 mo, n=4) vs. young (Y, 6 mo, n=4) control male C57Bl/6N mice had greater (p&lt;0.05) nitrotyrosine abundance (5.0 ± 1.1 vs. 1.0 ± 0.3 AU), a marker of oxidative stress, and AGEs (2.1 ± 0.2 vs.1.0 ± 0.1 AU) (p&lt;0.05). Sodium nitrite treatment (50 mg/L in drinking water, 3 weeks) in O reduced (p&lt;0.05) aortic nitrotyrosine (1.9 ± 0.6 AU) and AGEs (1.2 ± 0.2 AU). Stimulation of superoxide‐induced oxidative stress ex vivo with pyrogallol (10μM, 72 hr) in aortic segments from Y mice increased AGEs (2.3 ± 0.5 vs. 1.0 ± 0.1 AU, p&lt;0.05), an effect prevented by treatment with sodium nitrite (1.1 ± 0.2 AU). Aortic rings from Y mice treated with AGEs (100μg/mL, 72 hrs) had a greater elastic modulus, a measure of mechanical stiffness, compared with control (3984 ± 236 vs. 2074 ± 218 KPa, p&lt;0.05), and sodium nitrite prevented this effect (2148 ± 446 KPa). These results suggest that AGEs, induced by excessive superoxide‐associated oxidative stress, contributes to aortic stiffening with aging in mice. AGEs‐induced aortic stiffness may be preventable with sodium nitrite supplementation.

Perivascular adipose tissue in age‐associated arterial stiffening: Role of transforming growth factor beta 1

Aging causes large elastic artery stiffening associated with adventitial myofibroblast‐related increases in transforming growth factor beta 1 (TGF‐β1), collagen I and advanced glycation end‐products (AGEs). We tested the hypothesis that perivascular adipose tissue (PVAT) from older animals induces these protein changes in adventitial fibroblasts (AFB) and that the increase in TGF‐β1 causes arterial stiffening. AFB cultured from old (O, 24 mo, n=3–6) vs. young (Y, 6 mo, n=4–6) male Sprague Dawley rats had greater (p&lt;0.05) smooth muscle alpha actin (SMαA, 1.92 ± 0.21 vs. 1.00 ± 0.08), a marker of myofibroblasts, TGF‐β1 (15.77 ± 1.13 vs. 1.00 ± 0.21), collagen I (1.71 ±0.04 vs. 1.00 ± 0.14) and AGEs (4.49 ± 1.69 vs. 1.00 ± 0.20). In AFB from Y, PVAT from O rats increased (p &lt; 0.05) SMαA (1.76 ± 0.18), TGF‐β1 (7.28 ± 1.98) and collagen I (1.58 ± 0.21), but not AGEs (1.12 ± 0.38), and TGF‐β1 treatment mimicked these changes (SMαA: 1.00 ± 0.14 to 3.00 ± 0.44 and collagen I: 1.00 ± 0.04 to 1.71 ± 0.14, both p &lt; 0.05; AGEs: 1.00 ± 0.04 vs. 0.91 ± 0.22). Arterial stiffness (aortic pulse wave velocity, aPWV) was greater (p &lt; 0.05) in O (26–27 mo, n=5) vs. Y (4–6 mo, n=5) male C57Bl/6N mice (469 ± 16 vs. 351 ± 13) and 6‐week direct arterial application of recombinant TGF‐β1 to the aorta in vivo increased (p&lt;0.05) aPWV in Y (417 ± 6). These results suggest that PVAT may contribute to age‐associated arterial stiffening via TGF‐β1.NIH AG013038, HL007822

Reduced large elastic artery stiffness in older exercising adults is associated with suppressed nuclear factor kappa B signaling

Large elastic artery stiffness increases with age and is a strong independent predictor of incident cardiovascular diseases (CVD) in healthy middle‐aged and older (MA/O) adults. Regular aerobic exercise attenuates this age‐associated stiffening. We hypothesized that suppression of the pro‐inflammatory transcription factor nuclear factor κB (NFκB) may be involved. Aortic pulse‐wave velocity (aPWV) was measured in young sedentary (YS, n=7, blood pressure [BP] 101±3/58±2 mmHg), MA/O sedentary (OS, n=11, 110±4/67±2 mmHg) and MA/O aerobic exercise‐trained (OT, n=12, 120±6/70±4 mmHg) healthy non‐hypertensive men and women. Baseline aPWV increased with age (602±25 [YS] vs. 852±38 [OS] cm/sec, p&lt;0.001) but was 20% lower in OT (686±30 cm/sec) compared with OS (p&lt;0.01). Short‐term (4 d × 2500–4500 mg) treatment with the NFκB inhibitor salsalate (randomized, cross‐over design), reduced aPWV (to 782±44 cm/sec, p&lt;0.05) without changing BP (p=0.74) in OS, but had no effect in YS (599±23) or OT (699±30). Following salsalate treatment, aPWV no longer was significantly different in OS vs. OT (p=0.14). These results support the hypothesis that reduced NFκB signaling contributes to the lower large elastic artery stiffness in MA/O adults who regularly perform aerobic exercise. NIH AG013038, AG006537, AG03114, AG033994, RR025780

Vascular health in the ageing athlete

The demographics of ageing are changing dramatically such that there will be many more older adults in the near future. This setting is projected to produce a new 'boomer-driven' epidemic of physiological dysfunction, disability and risk of chronic degenerative disorders, including cardiovascular diseases. Standing out against this dreary biomedical forecast are Masters athletes, a group of middle-aged and older adults who engage in regular vigorous physical training and competitive sport. Compared with their sedentary/less active (untrained) peers, Masters athletes who perform endurance training-based activities demonstrate a more favourable arterial function-structure phenotype, including lower large elastic artery stiffness, enhanced vascular endothelial function and less arterial wall hypertrophy. As such, they may represent an exemplary model of healthy or 'successful' vascular ageing. In contrast, Masters athletes engaged primarily/exclusively in intensive resistance training exhibit less favourable arterial function-structure than their endurance-trained peers and, in some instances, untrained adults. These different arterial properties are probably explained in large part by the different intravascular mechanical forces generated during endurance versus resistance exercise-related training activities. The more favourable arterial function-structure profile of Masters endurance athletes may contribute to their low risk of clinical cardiovascular diseases.

Superoxide‐lowering therapy with TEMPOL reverses arterial dysfunction with aging in mice

To test the hypothesis that the antioxidant enzyme superoxide dismutase (SOD) mimetic TEMPOL improves arterial aging, young (Y, 4-6 months) and old (O, 26-28 months) male C57BL6 mice received regular or TEMPOL-supplemented (1mM) drinking water for 3 weeks (n = 8 per group). Aortic superoxide was 65% greater in O (P < 0.05 vs. Y), which was normalized by TEMPOL. O had large elastic artery stiffening, as indicated by greater aortic pulse wave velocity (aPWV, 508 ± 22 vs. 418 ± 22 AU), which was associated with increased adventitial collagen I expression (P < 0.05 vs. Y). TEMPOL reversed the age-associated increases in aPWV (434 ± 21 AU) and collagen in vivo, and SOD reversed the increases in collagen I in adventitial fibroblasts from older rats in vitro. Isolated carotid arteries of O had impaired endothelial function as indicated by reduced acetylcholine-stimulated endothelium-dependent dilation (EDD) (75.6 ± 3.2 vs. 94.5 ± 2.0%) mediated by reduced nitric oxide (NO) bioavailability (L-NAME) associated with decreased endothelial NO synthase (eNOS) expression (P < 0.05 vs. Y). TEMPOL restored EDD (94.5 ± 1.4%), NO bioavailability and eNOS in O. Nitrotyrosine and expression of NADPH oxidase were ~100-200% greater, and MnSOD was ~75% lower in O (P < 0.05 vs. Y). TEMPOL normalized nitrotyrosine and NADPH oxidase in O, without affecting MnSOD. Aortic pro-inflammatory cytokines were greater in O (P < 0.05 vs. Y) and normalized by TEMPOL. Short-term treatment of excessive superoxide with TEMPOL ameliorates large elastic artery stiffening and endothelial dysfunction with aging, and this is associated with normalization of arterial collagen I, eNOS, oxidative stress, and inflammation.

Autophagy‐enhancing treatment reverses age‐associated large elastic artery stiffening and modulates arterial superoxide production, inflammation and collagen I

Aging leads to stiffening of large elastic arteries associated with increased superoxide (O2‐) production, inflammation and collagen protein. Impairments in autophagy, a process that recycles damaged cellular components and influences oxidative stress and inflammation, may contribute to age‐related arterial stiffening. We tested the hypothesis that treatment with the natural disaccharide trehalose, an autophagy inducer, would reduce aortic pulse wave velocity (aPWV), a clinically important measure of arterial stiffness, and decrease superoxide production, inflammation and collagen I in old (O) male C57/BL6 mice. Autophagy was impaired in aorta from O (28 mo, n=6) vs. young (Y: 5 mo, n=6) mice as indicated by a ~40% reduction (p&lt;0.05) in beclin‐1 and LAMP‐2a, and these markers were restored by trehalose treatment (4 weeks, 20 g/L drinking water) in O mice. aPWV was higher in O vs. Y mice (471 ± 23 vs. 405 ± 25 cm/s, p&lt;0.05), and trehalose reduced aPWV in O mice (pre: 478 ± 32 vs. post: 417 ± 18 cm/s, p&lt;0.05). Trehalose treatment reversed age‐associated increases in aortic O2‐ production and inflammatory cytokines (tumor necrosis factor α and interleukin‐1), and reduced aortic collagen I by ~50%. Our results suggest that autophagy may be a promising therapeutic target for treating age‐associated large elastic artery stiffening, possibly by reducing O2‐/inflammation modulation of collagen.

Curcumin improves large elastic artery stiffness and reverses superoxide suppression of nitric oxide‐mediated vascular endothelial function in old mice

We tested the hypothesis that curcumin, a polyphenol found in the Indian spice turmeric, improves age‐associated large elastic artery stiffness and vascular endothelial dysfunction. Old control C57 male mice (27 mo, n=8) had greater aortic pulse wave velocity (aPWV) vs. young control (6 mo n=7) (448 ± 15 vs. 349 ± 15 cm/s, p &lt; 0.05) and 4 wk of curcumin supplemented chow (0.2%) reduced aPWV in old mice (396 ± 15 cm/s, n=8). Old control mice had impaired ex‐vivo carotid artery endothelium‐dependent dilation (EDD) to acetylcholine (ACh) vs. young (79 ± 3 vs. 93 ± 2%), mediated by a smaller nitric oxide (NO) component (EDD to ACh + LNAME minus EDD to ACh alone, 39 ± 11 vs 74 ± 6%) (both p &lt; 0.05). Curcumin restored EDD in old mice (92 ± 1%) by restoring its NO component (61 ± 7%). Incubation with the superoxide dismutase mimetic, TEMPOL, selectively improved EDD in old control mice (91 ± 3%, p &lt; 0.05), but had no effect in the other groups. Endothelium‐independent dilation did not differ between groups. Superoxide production (electron paramagnetic resonance spectroscopy) from aortic rings of old mice was greater vs. young control (7748 ± 897 vs. 4820 ± 492, p &lt; 0.05), and was markedly reduced with curcumin supplementation (2486 ± 533, p &lt; 0.05). Curcumin may be a novel therapeutic nutraceutical agent to destiffen large elastic arteries, restore NO‐mediated vascular endothelial function and reduce arterial superoxide production with aging.

Treatment with the SIRT1 activator SRT1720 reduces large elastic artery stiffness, superoxide production and inflammation in old mice

Aortic pulse wave velocity (aPWV), the gold‐standard measure of large elastic artery stiffness, is a strong independent predictor of incident cardiovascular disease in older adults. We hypothesized that chronic activation of the protein deacetylase sirtuin‐1 (SIRT1) would reduce aPWV in old mice and that this would be associated with reduced superoxide production and inflammation. Aortic SIRT1 activity (inverse of acetyl‐p53 protein expression) was markedly reduced in old (29–31 mo, n=11) vs. young (4–7 mo, n=11) control B6D2F1 mice (5.4±1.4 vs. 1.0±0.2 AU, p&lt;0.005), and 4 weeks of treatment with SRT1720 (100 mg/kg body weight, Sirtris, a GSK Co.) in old mice (29–32 mo, n=19) restored SIRT1 activity (1.1±0.2 AU). In vivo aPWV was 14% greater in old (OC, 29 mo, n=8) vs. young control mice (442±16 vs. 387±9 cm/s, p&lt;0.05), but was reduced by 13% after vs. before SRT1720 treatment in old mice (411±12 vs. 354±12 cm/s, p&lt;0.005). Aortic superoxide production (3149±493 vs. 5038±899 AU, p&lt;0.05) and expression of inflammatory cytokines IL‐6 (4.03±0.79 vs. 6.23±1.05 pg/mL, p=0.05) and TNF‐α (1.81±0.26 vs. 2.91±0.50 pg/mL, p&lt;0.05) were lower in treated vs. control old mice. SIRT1 activation with SRT1720 may be an effective treatment for reducing aPWV and destiffening large elastic arteries in older adults, possibly by decreasing arterial superoxide production and inflammation.

Nitrite supplementation reverses vascular endothelial dysfunction and large elastic artery stiffness with aging

We tested the hypothesis that short-term nitrite therapy reverses vascular endothelial dysfunction and large elastic artery stiffening with aging, and reduces arterial oxidative stress and inflammation. Nitrite concentrations were lower (P < 0.05) in arteries, heart, and plasma of old (26-28 month) male C57BL6 control mice, and 3 weeks of sodium nitrite (50 mg L(-1) in drinking water) restored nitrite levels to or above young (4-6 month) controls. Isolated carotid arteries of old control mice had lower acetylcholine (ACh)-induced endothelium-dependent dilation (EDD) (71.7 ± 6.1% vs. 93.0 ± 2.0%) mediated by reduced nitric oxide (NO) bioavailability (P < 0.05 vs. young), and sodium nitrite restored EDD (95.5 ± 1.6%) by increasing NO bioavailability. 4-Hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPOL), a superoxide dismutase (SOD) mimetic, apocynin, a nicotinamide adenine dinucleotide phosphate-oxidase (NADPH) inhibitor, and sepiapterin (exogenous tetrahydrobiopterin) each restored EDD to ACh in old control, but had no effect in old nitrite-supplemented mice. Old control mice had increased aortic pulse wave velocity (478 ± 16 vs. 332 ± 12 AU, P < 0.05 vs. young), which nitrite supplementation lowered (384 ± 27 AU). Nitrotyrosine, superoxide production, and expression of NADPH oxidase were ∼100-300% greater and SOD activity was ∼50% lower in old control mice (all P < 0.05 vs. young), but were ameliorated by sodium nitrite treatment. Inflammatory cytokines were markedly increased in old control mice (P < 0.05), but reduced to levels of young controls with nitrite supplementation. Short-term nitrite therapy reverses age-associated vascular endothelial dysfunction, large elastic artery stiffness, oxidative stress, and inflammation. Sodium nitrite may be a novel therapy for treating arterial aging in humans.