Search

Insulin signalling declines with increasing age and impacts skeletal muscle function and longevity in animal models. Our aim was to assess the relationships between insulin resistance (IR) and frailty and mortality in a unique community-dwelling cohort of older people. 991 non-diabetic subjects from the Toledo Study of Healthy Ageing (TSHA) cohort were included. IR was estimated by the homeostasis model assessment index (HOMA-IR) at baseline while frailty was determined by frailty phenotype (FP) and Frailty Trait Scale (FTS) at baseline and after 5-year follow-up. Deaths were also determined. Multivariate regression models were used to analyze the effects of HOMA-IR on outcomes. Age, gender, BMI, education level, cardio- and cerebro-vascular disease, glomerular filtration rate, and disability were included as potential confounding variables in progressive adjustment models. IR determined as increasing log HOMA-IR was inversely associated with risk of mortality. The association remained significant for all adjustment models (HR: 0.64–0.69). When we analyzed survival curves, the higher the HOMA-IR tertile, the lower the mortality rate (highest vs lowest tertile, p = 0.0082). In contrast, IR increased the risk of incident frailty determined by FP (OR 1.81 [1.14–2.87]) as well as deterioration of frailty status determined by worsening in FTS score (OR 1.28 [1.01–1.63]) at 5-year follow-up. In non-diabetic older subjects, IR significantly increases the risk for frailty and functional decline but decreased the risk of death at 5-year follow-up. This finding raises the need of assessing the effect of biomarkers on different outcomes before establishing their role as biomarkers of aging.

Background Store-operated calcium entry and its key players, stromal interaction molecule (STIM) and Orai calcium channels, have been proposed as emergent therapeutic targets in cardiovascular pathophysiology. We hypothesize alteration of STIM/Orai signaling in erectile dysfunction (ED). Aim To evaluate the contribution of STIM/Orai to human penile tissue contraction and to analyze the influence of ED on STIM/Orai signaling at functional and expression levels in human penile vascular tissues. Methods Human penile resistance arteries (HPRA) and human corpus cavernosum (HCC) were dissected from cavernosal specimens from 30 organ donors without history of ED (No ED) and from 48 patients with ED undergoing penile prosthesis insertion and functionally evaluated in wire myographs and organ chambers, respectively. Expression of STIM-1, Orai1, and Orai3 in HCC was localized and quantified by immunofluorescence. Main Outcome Measures The main outcome measures are functional responses in HCC and HPRA and STIM/Orai channel protein expression in human cavernosal tissue. Results Inhibition of Orai channels with YM-58483 (20 μM) significantly reduced norepinephrine–induced contractions in both HCC and HPRA from either No ED or ED subjects, but the effects were more marked in ED (−20.1 ± 5.9% vs −45.5 ± 13.2% and −15.9 ± 4.0% vs −31.4 ± 6.9% reduction in Emax to norepinephrine in HCC and HPRA, respectively). Thromboxane-induced contractions were reduced and neurogenic contractile and relaxant responses modulated by Orai inhibition in penile tissues from patients with ED. In fact, addition of YM-58483 concentration dependently relaxed precontracted HPRA and HCC. These relaxations were significantly more pronounced in tissues from patients with ED (EC50 7.5 vs 1.3 μM and 10.5 vs 1.3 μM, for HCC and HPRA, respectively). All HCC specimens displayed expression of STIM-1, Orai1, and Orai3. Significantly increased expression of Orai1 and Orai3 but not STIM-1 was observed in patients with ED. Clinical Translation Inhibition of enhanced Orai activity in human penile vascular tissue could facilitate erectile responses, alleviating ED. Strengths and Limitations Enhanced STIM/Orai activity contribution to penile smooth muscle tone in ED is demonstrated at functional and structural levels in human tissues from a representative sample of patients with ED and in comparison with healthy tissue. We cannot differentiate the specific contribution of risk factors associated with ED to hyperactivity of the Orai system. Conclusions Orai channels significantly contribute to human penile smooth muscle contraction. Orai contribution to penile smooth muscle tone is functionally enhanced in ED accompanied by increased expression of Orai channels in cavernosal tissue. Orai inhibition could be a potential therapeutic strategy to reduce penile smooth muscle contraction in ED.

Oxidative stress plays a key role in the aging process. Lifestyle behaviours including low physical activity and inadequate nutritional habits in addition to genetic susceptibility and some chronic diseases compromise physiological response to free radicals and promote oxidative damage. Reduced resilience (referred to the ability to respond to stressors or adverse conditions) or functional reserve in isolated organs or systems determines clinical manifestations as the age-related chronic diseases while multisystemic dysfunction results in the frailty phenotype. In older adults, frailty, but not age, is associated with elevation of oxidative stress markers and reduction of antioxidant parameters. Mitochondrial dysfunction related to oxidative stress plays a prominent role in this process affecting not only skeletal muscle but also other potential tissues and organs. Increasing endogenous antioxidant capacity in different systems by exercise outstand among therapeutic interventions with potential ability to prevent or delay frailty phenotype and to promote healthy aging.

The role of STIM/Orai calcium entry system on vascular ageing has not been elucidated. We aimed to evaluate the influence of ageing on STIM/Orai signalling and its role on ageing-induced alterations of contractile function in rat corpus cavernosum (RCC) and human penile resistance arteries (HPRA) and corpus cavernosum (HCC). RCC was obtained from 3 months-old and 20 months-old animals. HPRA and HCC were obtained from organ donors of varied ages without history of erectile dysfunction. Aging was associated with enhanced norepinephrine (NE)- and thromboxane analogue (U46619)-induced contractions in RCC which were significantly inhibited by the STIM/Orai inhibitor, YM-58483 (20 μM). Other STIM/Orai inhibitor, 2-aminoethyldiphenylborate also reduced NE-induced contractions in RCC from aged rats. YM-58483 significantly reduced neurogenic contractions and potentiated neurogenic relaxations in RCC from aged rats. In HCC and HPRA, NE-induced contractions were significantly enhanced in older subjects (>65 years-old) but YM-58483 completely reversed ageing-related hypercontractility. Ageing did not modify STIM-1 and Orai1 protein expressions but Orai3 was significantly overexpressed in cavernosal tissue from old rats and older subjects. Contribution of STIM/Orai to cavernosal contraction increases with ageing together with increased expression of Orai3. Orai inhibition could be a potential therapeutic strategy to reduce ageing-related impact on vascular/erectile function.

Frailty is considered a clinical marker of functional ageing. Telomere length (TL) has been proposed as a biomarker of biological age but its role in human ageing is controversial. The main aim of the study was to evaluate the longitudinal association of TL with incident frailty and mortality in two cohorts of Spanish community–dwelling older adults. TL was determined at baseline in blood samples from older adults included in Toledo Study for Healthy Aging and ENRICA cohorts while frailty was determined by frailty phenotype (FP) at baseline and at follow-up (3.5 years). Deaths occurring during follow-up were also recorded. Associations of TL with frailty and mortality were analysed by logistic regression with progressive adjustment. Data were separately analysed in the two cohorts and in all subjects by performing a meta-analysis. TL was not different between frail and non-frail subjects. Longer telomeres were not associated with lower risk of prevalent frailty. Similarly, TL at baseline failed to predict incident frailty (OR: 1.04 [0.88–1.23]) or even the development of a new FP criterion (OR: 0.97 [0.90–1.05]) at follow-up. Lack of association was also observed when analysing the development of specific FP criteria. Finally, while frailty at baseline was significantly associated with higher risk of death at follow-up (OR: 4.08 [1.97–8.43], p

Frailty, a consequence of the interaction of the aging process and certain chronic diseases, compromises functional outcomes in the elderly and substantially increases their risk for developing disabilities and other adverse outcomes. Frailty follows from the combination of several impaired physiological mechanisms affecting multiple organs and systems. And, though frailty and sarcopenia are related, they are two different conditions. Thus, strategies to preserve or improve functional status should consider systemic function in addition to muscle conditioning. Physical activity/exercise is considered one of the main strategies to counteract frailty-related physical impairment in the elderly. Exercise reduces age-related oxidative damage and chronic inflammation, increases autophagy, and improves mitochondrial function, myokine profile, insulin-like growth factor-1 (IGF-1) signaling pathway, and insulin sensitivity. Exercise interventions target resistance (strength and power), aerobic, balance, and flexibility work. Each type improves different aspects of physical functioning, though they could be combined according to need and prescribed as a multicomponent intervention. Therefore, exercise intervention programs should be prescribed based on an individual's physical functioning and adapted to the ensuing response., Highlights • Intrinsic capacity close to disability threshold and low potential recovery due to low functional reserve lead to frailty. • Frailty follows from the combination of several impaired physiological mechanisms affecting multiple organs and systems. • Exercise reduces age-related oxidative damage and inflammation and improves mitochondrial function among other benefits. • Multicomponent intervention should be considered in order to impact different aspects of physical function in the elderly. • Exercise interventions should be prescribed based on individual’s physical functioning and adapted to the ensuing response.

Background and Aims Patients with chronic kidney disease (CKD) present a high rate of cardiovascular mortality mainly associated with endothelial dysfunction, which causes more cardiovascular events in presence of atherosclerosis. Atherosclerosis is characterized by a significant increase of low density lipoproteins (LDL), reactive oxygen species (ROS) and inflammation. ROS can oxidize LDL generating oxidized-LDL (oxLDL) that promotes the development of cardiovascular pathologies. The aim of this study was to evaluate whether oxLDL induce endothelial dysfunction analysing the involvement of vascular fibrosis. Method The model used for in vivo studies was the Knockout apolipoprotein E (KO-apoE) mice, which resemble human atherosclerosis and shown high levels of cholesterol (LDL) that can be oxidized to oxLDL. In mice, blood pressure was registered before sacrificed them. After that, we measured different parameters as serum cholesterol levels, vascular function by vascular reactivity in mesenteric arteries and vascular fibrosis in aorta by Sirius Red staining and by the protein expression of fibronectin and collagen-I by immunohistochemistry. In order to investigate the mechanism of action of oxLDL, in vitro studios were performed on human smooth muscle cells (SMC) incubated with oxLDL at different times. Fibrosis was evaluated by the expression of TGF- β and extracellular matrix proteins such as fibronectin and collagen-I by Western blot and by immunofluorescence. ROS production was also measured by fluorescence confocal microscopy, using the CellROX Deep Red probe. Results KO-apoE mice shown higher levels of serum cholesterol and blood pressure than WT animals. Moreover, KO-apoE mice showed endothelial dysfunction since their arteries were less relaxed and more contracted. In addition, these mice presented thickening of vascular wall (SMC layer), more fibrosis showing intense Sirius Red staining and less expression of elastin, all compatible with their vascular dysfunction compared to WT mice. Furthermore, aortas from KO-apoE mice showed a slight increase in fibronectin and collagen-I expression assessed by immunohistochemistry. In vivo studies were confirmed in vitro after treating SMC with oxLDL. oxLDL induced fibrosis in SMC by increasing TGF-β, fibronectin and collagen-I protein expressions evaluated by Western blot and immunofluorescence assays. Treatment with oxLDL also increased ROS production, which seem to be responsible of oxLDL-induced fibrosis in human SMC, as it was blocked in the presence of the antioxidant N-Acetyl-cysteine. Conclusion In summary, these results point to endothelial dysfunction associated to atherosclerosis (oxLDL) could be mediated by an increase in the development of vascular fibrosis where ROS could play an important role. Therefore, the endothelial dysfunction typical of CKD patients could impair with more vascular fibrosis when atherosclerosis is also present.

Etamsylate is indicated for several anti-hemorrhagic indications in human and veterinary medicine. However, etamsylate has been shown to be effective only in specific hemorrhagic situations. Furthermore, mechanism of action of etamsylate is not known but recent research has shown its ability to inhibit heparin binding to several growth factors. We have evaluated the ability of etamsylate to interfere with the activities of heparin. Effects of etamsylate on vasodilatory activity of heparin were evaluated in rat aortic segments. Influence of etamsylate on anticoagulant activity of heparin was evaluated in vitro by determining prothrombin (PT) time and activated partial thromboplastin time (aPTT) in dog blood and in vivo by determining the interference of systemic and topical etamsylate on heparin-induced extension in bleeding time (BT) in rats. Despite failing to inhibit heparin-induced vasodilation of rat aorta, etamsylate significantly reduced the increase in aPTT caused by heparin (+30.4 ± 6.7% vs. +15.0 ± 2.8% for etamsylate at 100 µM, P < 0.05). Etamsylate also antagonized the anticoagulant effects driven by heparin in vivo since prevented the heparin-induced increase in BT when systemically (i.p.) administered (+94.6 ± 7.5% vs. +57.9 ± 9.2% at 10 mg/kg, P < 0.05, vs. +22.2 ± 16.8% at 30 mg/kg, P < 0.01). Additionally, topically applied etamsylate (125 mg/ml) significantly reduced heparin-induced BT increase (+102.5 ± 3.2% vs. +54.0 ± 5.8%, P < 0.01). These evidences show a pharmacological interference by etamsylate on heparin activities antagonizing pro-hemorrhagic effects of heparin in vitro and in vivo without inhibiting its vasodilatory properties. This ability could help to explain pharmacological effects of etamsylate and proposes its role for reversing pro-hemorrhagic states.

Aging impairs vascular function, but the mechanisms involved are unknown. The aim of this study was to analyze whether aging-related hyperphosphatemia is implied in this effect by elucidating the role of oxidative stress. C57BL6 mice that were aged 5 months (young) and 24 months (old), receiving a standard (0.6%) or low-phosphate (0.2%) diet, were used. Isolated mesenteric arteries from old mice showed diminished endothelium-dependent vascular relaxation by the down-regulation of NOS3 expression, increased inflammation and increased fibrosis in isolated aortas, compared to those isolated from young mice. In parallel, increased Nox4 expression and reduced Nrf2, Sod2-Mn and Gpx1 were found in the aortas from old mice, resulting in oxidant/antioxidant imbalance. The low-phosphate diet improved vascular function and oxidant/antioxidant balance in old mice. Mechanisms were analyzed in endothelial (EC) and vascular smooth muscle cells (SMCs) treated with the phosphate donor ß-glycerophosphate (BGP). In EC, BGP increased Nox4 expression and ROS production, which reduced NOS3 expression via NFκB. BGP also increased inflammation in EC. In SMC, BGP increased Collagen I and fibronectin expression by priming ROS production and NFκB activity. In conclusion, hyperphosphatemia reduced endothelium-dependent vascular relaxation and increased inflammation and vascular fibrosis through an impairment of oxidant/antioxidant balance in old mice. A low-phosphate diet achieved improvements in the vascular function in old mice.

Oxidative stress contributes to endothelial dysfunction, a key step in cardiovascular disease development. Ageing-related vascular dysfunction involves defective antioxidant response. Nuclear factor erythroid 2-like-2 (Nrf2), orchestrates cellular response to oxidative stress. We evaluated the impact of Nrf2-activation on endothelium-dependent and H2O2-mediated vasodilations in: aorta (RA), mesenteric artery (RMA), coronary artery (RCA) and corpus cavernosum (RCC) from ageing rats and in human penile arteries (HPRA) and corpus cavernosum (HCC) from erectile dysfunction (ED) patients. Relaxant responses were evaluated in organ chambers and wire myographs. Nrf2 content and heme oxygenase-1 (HO-1) were determined by ELISA. Superoxide and Nrf2 were detected by immunofluorescence. Pharmacological activation of Nrf2 with sulforaphane (SFN) improved NO- and endothelium-derived hyperpolarizing factor-mediated endothelium-dependent vasodilation and H2O2-induced relaxation in vascular beds from aging rats. SFN-induced effects were associated with increased Nrf2 (RMA, RCA) and reduced superoxide detection in RCA. Improvement of vascular function was confirmed in HPRA and HCC from ED patients and mimicked by another Nrf2 activator, oltipraz. Nrf2 increase and superoxide reduction together with HO-1 increase by Nrf2 activation was evidenced in HCC from ED patients. PDE5 inhibitor-induced relaxations of HPRA and HCC from ED patients were enhanced by SFN. Nrf2 short-term pharmacological activation attenuates age-related impairment of endothelium-dependent and reactive oxygen species (ROS)-induced vasodilation in different rat and human vascular territories by upregulation of Nrf2-related signaling and decreased oxidative stress. In ED patients target tissues, Nrf2 potentiates the functional effect of ED conventional pharmacological therapy suggesting potential therapeutic implication. Keywords: Oxidative stress, Endothelial dysfunction, Nrf2, Ageing, Human vasculature, Erectile dysfunction

Purpose of review Disability and its preceding condition, frailty, are outstanding issues for achieving healthy aging. Diabetes is a very prevalent chronic disease among older patients that favours frailty status. This review will analyse the relationship between diabetes and frailty in the elderly and summarize the current strategies to improve physical function in diabetic older patients. Recent findings We have analyzed the current knowledge providing insight on the relationship between frailty and diabetes in older people. Epidemiological evidences and potential mechanisms connecting diabetes with frailty in the aging process have been examined. Finally, the strategies to reduce frailty in aged population with diabetes were discussed. Summary Current evidence reveals the high prevalence of diabetes in frail older patients, producing an additional impairment of physical performance in this population. Insulin resistance seems to contribute to this clinical manifestation which is related to the impact of diabetes on skeletal muscle function, on vascular function, and on the hormonal milieu. Exercise, nutritional and educational interventions, and less strict glycaemic control appear as the most effective strategies to reduce frailty in diabetic older people.

Key points The presence of insulin resistance (IR) is determinant for endothelial dysfunction associated with obesity. Although recent studies have implicated the involvement of mitochondrial superoxide and inflammation in the defective nitric oxide (NO)-mediated responses and subsequent endothelial dysfunction in IR, other mechanisms could compromise this pathway. In the present study, we assessed the role of asymmetric dimethylarginine (ADMA) and arginase with respect to IR-induced impairment of endothelium-dependent vasodilatation in human morbid obesity and in a non-obese rat model of IR. We show that both increased ADMA and up-regulated arginase are determinant factors in the alteration of the l-arginine/NO pathway associated with IR in both models and also that acute treatment of arteries with arginase inhibitor or with l-arginine significantly alleviate endothelial dysfunction. These results help to expand our knowledge regarding the mechanisms of endothelial dysfunction that are related to obesity and IR and establish potential therapeutic targets for intervention. Abstract Insulin resistance (IR) is determinant for endothelial dysfunction in human obesity. Although we have previously reported the involvement of mitochondrial superoxide and inflammation, other mechanisms could compromise NO-mediated responses in IR. We evaluated the role of the endogenous NOS inhibitor asymmetric dimethylarginine (ADMA) and arginase with respect to IR-induced impairment of l-arginine/NO-mediated vasodilatation in human morbid obesity and in a non-obese rat model of IR. Bradykinin-induced vasodilatation was evaluated in microarteries derived from insulin-resistant morbidly obese (IR-MO) and non-insulin-resistant MO (NIR-MO) subjects. Defective endothelial vasodilatation in IR-MO was improved by l-arginine supplementation. Increased levels of ADMA were detected in serum and adipose tissue from IR-MO. Serum ADMA positively correlated with IR score and negatively with pD2 for bradykinin. Gene expression determination by RT-PCR revealed not only the decreased expression of ADMA degrading enzyme dimethylarginine dimethylaminohydrolase (DDAH)1/2 in IR-MO microarteries, but also increased expression of arginase-2. Arginase inhibition improved endothelial vasodilatation in IR-MO. Analysis of endothelial vasodilatation in a non-obese IR model (fructose-fed rat) confirmed an elevation of circulating and aortic ADMA concentrations, as well as reduced DDAH aortic content and increased aortic arginase activity in IR. Improvement of endothelial vasodilatation in IR rats by l-arginine supplementation and arginase inhibition provided functional corroboration. These results demonstrate that increased ADMA and up-regulated arginase contribute to endothelial dysfunction as determined by the presence of IR in human obesity, most probably by compromising arginine availability. The results provide novel insights regarding the mechanisms of endothelial dysfunction related to obesity and IR and establish potential therapeutic targets for intervention.

Diabetes and the ageing process independently increase the risk for cardiovascular disease (CVD). Since incidence of diabetes increases as people get older, the diabetic older adults represent the largest population of diabetic subjects. This group of patients would potentially be threatened by the development of CVD related to both ageing and diabetes. The relationship between CVD, ageing and diabetes is explained by the negative impact of these conditions on vascular function. Functional and clinical evidence supports the role of vascular inflammation induced by the ageing process and by diabetes in vascular impairment and CVD. Inflammatory mechanisms in both aged and diabetic vasculature include pro-inflammatory cytokines, vascular hyperactivation of nuclear factor-кB, increased expression of cyclooxygenase and inducible nitric oxide synthase, imbalanced expression of pro/anti-inflammatory microRNAs, and dysfunctional stress-response systems (sirtuins, Nrf2). In contrast, there are scarce data regarding the interaction of these mechanisms when ageing and diabetes co-exist and its impact on vascular function. Older diabetic animals and humans display higher vascular impairment and CVD risk than those either aged or diabetic, suggesting that chronic low-grade inflammation in ageing creates a vascular environment favouring the mechanisms of vascular damage driven by diabetes. Further research is needed to determine the specific inflammatory mechanisms responsible for exacerbated vascular impairment in older diabetic subjects in order to design effective therapeutic interventions to minimize the impact of vascular inflammation. This would help to prevent or delay CVD and the specific clinical manifestations (cognitive decline, frailty and disability) promoted by diabetes-induced vascular impairment in the elderly.

H2S signaling was proposed to participate in erectile physiology. L-cysteine (CYS)/H2S pathway stimulation causes cGMP-dependent relaxation of human corpus cavernosum (HCC) and penile arteries (HPRA). The aim was to evaluate the impact of ED on CYS/H2S pathway at functional and molecular level in human penile vascular tissues. NaHS- and CYS-induced responses were evaluated in HCC and HPRA from organ donors without ED (NoED, n = 29) and from ED patients undergoing penile prosthesis insertion (n = 45). cGMP accumulation and cystathionine β-synthase and cystathionine γ-lyase expression were also determined. NaHS-induced relaxations were slightly but significantly impaired in HCC but not in HPRA from ED patients. In contrast, CYS-induced relaxations were markedly impaired in HCC (Emax 67.6 ± 4.9% vs 46.2 ± 4.6%, P

INTRODUCTION AND OBJECTIVE:Renal function is highly dependent on adequate vascular function. This is especially relevant when considering the increasing use of older and marginal kidney donors to o...

Mechanisms contributing to pulmonary and systemic injury induced by high tidal volume (VT) mechanical ventilation are not well known. We tested the hypothesis that increased peroxynitrite formation is involved in organ injury and dysfunction induced by mechanical ventilation. Male Sprague-Dawley rats were subject to low- (VT, 9 mL/kg; positive end-expiratory pressure, 5 cmH2O) or high- (VT, 25 mL/kg; positive end-expiratory pressure, 0 cmH2O) VT mechanical ventilation for 120 min, and received 1 of 3 treatments: 3-aminobenzamide (3-AB, 10 mg/kg, intravenous, a poly adenosine diphosphate ribose polymerase [PARP] inhibitor), or the metalloporphyrin manganese(III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP, 5 mg/kg intravenous, a peroxynitrite scavenger), or no treatment (control group), 30 min before starting the mechanical ventilation protocol (n = 8 per group, 6 treatment groups). We measured mean arterial pressure, peak inspiratory airway pressure, blood chemistry, and gas exchange. Oxidation (fluorescence for oxidized dihydroethidium), protein nitration (immunofluorescence and Western blot for 3-nitrotyrosine), PARP protein (Western blot) and gene expression of the nitric oxide (NO) synthase (NOS) isoforms (quantitative real-time reverse transcription polymerase chain reaction) were measured in lung and vascular tissue. Lung injury was quantified by light microscopy. High-VT mechanical ventilation was associated with hypotension, increased peak inspiratory airway pressure, worsened oxygenation; oxidation and protein nitration in lung and aortic tissue; increased PARP protein in lung; up-regulation of NOS isoforms in lung tissue; signs of diffuse alveolar damage at histological examination. Treatment with 3AB or MnTMPyP attenuated the high-VT mechanical ventilation-induced changes in pulmonary and cardiovascular function; down-regulated the expression of NOS1, NOS2, and NOS3; decreased oxidation and nitration in lung and aortic tissue; and attenuated histological changes. Increased peroxynitrite formation is involved in mechanical ventilation-induced pulmonary and vascular dysfunction. Sin financiación 3.048 JCR (2015) Q1, 33/200 Surgery; Q2, 12/33 Critical care medicine, 29/70 Hematology, 23/63 Peripheral vascular disease UEM

Aging induces vascular dysfunction, representing the major risk factor for cardiovascular disease. Our aim was to ascertain specific vulnerability of vascular territories to aging by evaluating the progressive impact of aging on vascular function in four different vascular beds: aorta, mesenteric artery (MA), coronary artery (CA), and penile corpus cavernosum (CC) from 3, 6, 9, 12, 20 or 24 months-old male rats. Contractile/relaxant responses were evaluated in organ chambers (A/CC) and wire myographs (MA/CA). Relationships of systemic biomarkers with endothelial function impairment were also determined. Although all vessels manifested aging-related impairment in endothelial vasodilation, CA was the most impacted by aging considering the onset (at 6 months) and magnitude of endothelial dysfunction (reduction by 1.5 log units in the concentration required for 50% of maximal relaxation for acetylcholine). H2O2-induced vasodilations were progressively reduced by aging in aorta, CC and CA while NO-donor-induced vasodilation was impaired by aging only in CA. Serum asymmetric dimethylarginine significantly correlated to endothelial decline in aorta, MA, and CC, while HOMA-IR was significantly associated with endothelial dysfunction in CA and MA. CA are especially vulnerable to aging-related vascular dysfunction. Correlations of vascular dysfunction with systemic biomarkers differ among vessels, further suggesting heterogeneity in aging-induced vascular impact.

Background Specific mechanisms underlying frailty syndrome are not well known. Frailty can be viewed as a loss of functional reserve resulting in increased vulnerability to stressors. We hypothesize that pathways regulating cellular response to stress are potential players in the development of frailty. The aim of this study was to evaluate the association of the expression of certain genes related to cellular response to stress with the presence of frailty in older patients. Methods A sample of 350 individuals aged 65 years or older (22% frail) was selected from the Toledo Study of Healthy Aging. RNA was extracted from blood and retro-transcribed into complementary DNA. TaqMan Low density Arrays were used for the measurement of expression of genes implicated in cellular response to oxidative stress, genes implicated in inflammation, genes implicated in vascular physiology, and genes related to hypoxia. For data analysis, a logistic regression model was used to assess the relationship of gene expression and frailty. Results Among the analyzed genes, lower expression of genes related to cellular response to hypoxia (hypoxia inducible factor-1α) or to cellular response to oxidative stress (nuclear factor erythroid 2-related factor 2 and its target genes heme oxygenase-2, thioredoxin reductase-1, and superoxide dismutase-2), but not to those related to inflammation or vascular physiology, were significantly associated with the presence of frailty after adjustment for age and sex. These associations remained significant after adjustment by type 2 diabetes and Charlson index of comorbidities. Lower expressions of genes involved in cellular response to stress were also associated with increased risk of functional impairment. Conclusions Reduced expression of several genes implicated in cellular response to oxidative stress or hypoxia is significantly associated with the presence of frailty. These results help to fill the gap of knowledge of this evolving field and provide targets for intervention to promote health and independence in the elderly.

Vascular aging, a determinant factor for cardiovascular disease and health status in the elderly, is now viewed as a modifiable risk factor. Impaired endothelial vasodilation is a early hallmark of arterial aging that precedes the clinical manifestations of vascular dysfunction, the first step to cardiovascular disease and influencing vascular outcomes in the elderly. Accordingly, the preservation of endothelial function is thought to be an essential determinant of healthy aging. With special attention on the effects of aging on the endothelial function, this review is focused on the two main mechanisms of aging-related endothelial dysfunction: oxidative stress and inflammation. Aging vasculature generates an excess of the reactive oxygen species (ROS), superoxide and hydrogen peroxide, that compromise the vasodilatory activity of nitric oxide (NO) and facilitate the formation of the deleterious radical, peroxynitrite. Main sources of ROS are mitochondrial respiratory chain and NADPH oxidases, although NOS uncoupling could also account for ROS generation. In addition, reduced antioxidant response mediated by erythroid-2-related factor-2 (Nrf2) and downregulation of mitochondrial manganese superoxide dismutase (SOD2) contributes to the establishment of chronic oxidative stress in aged vessels. This is accompanied by a chronic low-grade inflammatory phenotype that participates in defective endothelial vasodilation. The redox-sensitive transcription factor, nuclear factor-κB (NF-κB), is upregulated in vascular cells from old subjects and drives a proinflammatory shift that feedbacks oxidative stress. This chronic NF-κB activation is contributed by increased angiotensin-II signaling and downregulated sirtuins and precludes adequate cellular response to acute ROS generation. Interventions targeted to recover endogenous antioxidant capacity and cellular stress response rather than exogenous antioxidants could reverse oxidative stress-inflammation vicious cycle in vascular aging. Lifestyle attitudes such as caloric restriction and exercise training appear as effective ways to overcome defective antioxidant response and inflammation, favoring successful vascular aging and decreasing the risk for cardiovascular disease.

Frailty is a functional status that precedes disability and is characterized by decreased functional reserve and increased vulnerability. In addition to disability, the frailty phenotype predicts falls, institutionalization, hospitalization and mortality. Frailty is the consequence of the interaction between the aging process and some chronic diseases and conditions that compromise functional systems and finally produce sarcopenia. Many of the clinical manifestations of frailty are explained by sarcopenia which is closely related to poor physical performance. Reduced regenerative capacity, malperfusion, oxidative stress, mitochondrial dysfunction and inflammation compose the sarcopenic skeletal muscle alterations associated to the frailty phenotype. Inflammation appears as a common determinant for chronic diseases, sarcopenia and frailty. The strategies to prevent the frailty phenotype include an adequate amount of physical activity and exercise as well as pharmacological interventions such as myostatin inhibitors and specific androgen receptor modulators. Cell response to stress pathways such as Nrf2, sirtuins and klotho could be considered as future therapeutic interventions for the management of frailty phenotype and aging-related chronic diseases.

The mechanisms contributing to sepsis vascular dysfunction are not well known. We tested the hypothesis that peroxynitrite scavenging ameliorates sepsis-induced macrovascular and microvascular dysfunction. Male Sprague-Dawley rats were killed 48 h after cecal ligation (n = 15) and puncture or sham procedure (n = 15). Their aortas and mesenteric vessels were mounted in organ baths for isometric tension recording. We studied contraction in resting vessels (norepinephrine 1 nM-10 μM and 10 nM-10 μM) and endothelium-dependent relaxation (acetylcholine, 10 nM-10 μM and 1 nM-10 μM) for aortas and microvessels, respectively. Vascular rings were preincubated for 30 min with the superoxide scavenger Cu-Zn-superoxide dismutase (SOD) (100 U/mL), the SOD mimetic and peroxynitrite scavenger tempol (10 M), the NO synthase inhibitor N-nitro-l-arginine methyl ester (10 M), or the peroxynitrite decomposition catalyst manganese tetrakis(4-N-methylpyridyl)porphyrin (MnTMPyP) (10 M). Fluorescence to 3-nitrotyrosine, oxidized dihydroethidium, and NOS2 was assessed in vascular tissue. Vascular NOS2, endothelial nitric oxide synthase (NOS1), NADPH-oxidase-1 (NOX-1), and SOD expression was analyzed by reverse transcription-polymerase chain reaction. Sepsis induced (i) in macrovessels, impairment of norepinephrine-induced contractions; (ii) in microvessels, impairment in norepinephrine-induced contractions and acetylcholine-induced relaxations; (iii) aortic and microvascular tissue increased reactivity to 3-nitrotyrosine, oxidized dihydroethidium, NOS2, and increased expression of NOS2, as well as increased expression of NOX-1 in microvascular tissue. Contractile responses in aortic and microvascular rings improved by ex vivo treatment with MnTMPyP and tempol, whereas vascular relaxation in microvessels improved only with MnTMPyP. Peroxynitrite scavenging protects from vascular dysfunction in sepsis.

Background: Apoptosis and inflammatory/oxidative stress have been associated with hyperglycemia in human peritoneal mesothelial cells (HPMCs) and other cell types. We and others have highlighted the role of early products of non-enzymatic protein glycation in inducing proinflammatory conditions and increasing apoptotic rates in HPMCs. Loss of HPMCs seems to be a hallmark of complications associated with peritoneal membrane dysfunction. The aim of this work is to elucidate the mechanisms by which Amadori adducts may act upon HPMC apoptosis. Methods: HPMCs isolated from different patients were exposed to different Amadori adducts, i.e. highly glycated hemoglobin (10 nM) and glycated bovine serum albumin (250 µg/ml), to study cell death and several proapoptotic markers by different experimental approaches. Results: Amadori adducts, but not their respective controls, impaired cell proliferation and cell viability by means of apoptosis in a time-dependent manner. They regulated the intrinsic mitochondrial cell death signaling pathway and modulated activation of caspases, Bax, iNOS, p53, NF-ĸB, and mitogen-activated protein kinases (p38 and JNK) through different reactive oxygen and nitrosative species. Conclusions: Our data strongly support the idea that long-term hyperglycemia could act as an inducer of apoptosis in HPMCs through Amadori adducts, involving different oxidative and nitrosative reactive species.

Summary Vascular endothelial dysfunction occurs during the human aging process, and it is considered as a crucial event in the development of many vasculopathies. We investigated the underlying mechanisms of this process, particularly those related with oxidative stress and inflammation, in the vasculature of subjects aged 18‐ 91 years without cardiovascular disease or risk factors. In isolated mesenteric microvessels from these subjects, an age-dependent impairment of the endotheliumdependent relaxations to bradykinin was observed. Similar results were observed by plethysmography in the forearm blood flow in response to acetylcholine. In microvessels from subjects aged less than 60 years, most of the bradykinin-induced relaxation was due to nitric oxide release while the rest was sensitive to cyclooxygenase (COX) blockade. In microvessels from subjects older than 60 years, this COX-derived vasodilatation was lost but a COX-derived vasoconstriction occurred. Evidence for age-related vascular oxidant and inflammatory environment was observed, which could be related to the development of endothelial dysfunction. Indeed, aged microvessels showed superoxide anions (O2 ) ) and peroxynitrite (ONOO ) ) formation, enhancement of NADPH oxidase and inducible NO synthase expression. Pharmacological interference of COX, thromboxane A2 ⁄prostaglandin H2 receptor, O2 ) , ONOO ) , inducible NO synthase, and NADPH oxidase improved the age-related endothelial dysfunction. In situ vascular nuclear factor-jB activation was enhanced with age, which correlated with endothelial dysfunction. We conclude that the age-dependent endothelial dysfunction in human vessels is due to the combined effect of oxidative stress and vascular wall

Objective To describe the time course of the changes in pulmonary and vascular function, and systemic inflammation induced by injurious mechanical ventilation.

Background and purpose: Diabetes mellitus is prevalent in the elderly population. It is also a disease causing tissue damage through several different mechanisms. Some of these mechanisms are also activated by ageing and this overlap raises questions about how diabetes induces damage in the elderly. Early products of non-enzymatic glycation of proteins (Amadori adducts), and the ageing process share the capacity to induce oxidative stress and inflammation in human peritoneal mesothelial cells (HPMCs). We have evaluated the interactions between the age of the donor of the HPMCs and the pro-inflammatory effects of Amadori adducts in those cells. Experimental approach: HPMCs were isolated from 20 individuals (age range 21–81 years) and grown in culture. Using different experimental approaches we determined NF-κB dependent transcriptional activity and different NF-κB-related pro-inflammatory gene and protein expressions in basal (or non-stimulated) conditions and after stimulation with two Amadori adducts; highly-glycated haemoglobin and glycated bovine serum albumin. Key results: Amadori-induced effects on NF-κB dependent-transcription and on the activity of NOS, COX and several NF-κB-related pro-inflammatory genes (iNOS, COX-2, TNF-α, IL-1β, and IL6) diminished as the donor's age increased, being practically absent in cells from donors more than 65 years old. Such decreased effects were inversely correlated with an increased basal expression and activity of these pro-inflammatory markers with age. Conclusions and implications: Pro-inflammatory effects of Amadori-adducts in HPMCs were strongly dependent on cell donor's age. This may have significant implications for the mechanisms underlying diabetes-induced tissue damage in patients of different ages. British Journal of Pharmacology (2006) 149, 979–987. doi:10.1038/sj.bjp.0706864

The number of older patients admitted to peritoneal dialysis (PD) programmes is growing. At the same time, there is increasing data about the role of mesothelial cells in determining the functional alteration of the peritoneum during PD. However, little is known about the functional changes accompanying the ageing process in mesothelial cells. We aimed to evaluate whether the aging process is accompanied by changes in some functional characteristic of the human peritoneal mesothelial cells (HPMC), which could account for the poor prognosis observed in old patients with PD. HPMCs were isolated from patients undergoing a nonurgent, nonseptic abdominal surgical procedure, without renal, vascular or inflammatory disease. Cytokine levels (by enzyme-linked immunosorbent assay (ELISA)), nitrates+nitrites, and cyclooxygenase (COX) activity (by a chemiluminescence assay), cytokines, COX, nitric oxide synthase (NOS), and nuclear factor (NF)-kappaB1, two messenger ribonucleic acid (mRNA) gene expressions (by reverse transcriptase (RT)-Multiplex PCR), COX, and NOS promoter gene activities, and NF-kappaB-dependent transcription (by transient transfection assays) were determined. Our data show a significant increase in cytokines, COX, and NOS activities, and mRNA expression of cytokines, COX-2, inducible nitric oxide synthase (iNOS) and precursors of NF-kappaB in HPMCs from old people. This was also the case for COX-2 and iNOS promoter gene activities and NF-kappaB-dependent transcription. There was a positive correlation between the age of the donor's cell and the proinflammatory profile of the HPMCs. Such age-dependent increase (around two-three times) is partially abolished by different antioxidant or free-radical scavengers. Thus, aging is accompanied by the presence of an inflammatory state in HPMCs, which involves the participation of different reactive oxygen species.

Diabetes mellitus leads to a high incidence of several so-called complications, sharing similar pathophysiological features in several territories. Previous reports points at early nonenzymatic glycosylation products (Amadori adducts) as mediators of diabetic vascular complications. In the present study, we analysed a possible role for Amadori adducts as stimulators of proinflammatory pathways in human peritoneal mesothelial cells (HPMCs). Cultured HPMCs isolated from 13 different patients (mean age 38.7+/-16 years) were exposed to different Amadori adducts, that is, highly glycated haemoglobin (10 nM) and glycated bovine serum albumin (0.25 mg ml(-1)), as well as to their respective low glycosylation controls. Amadori adducts, but not their respective controls, elicited a marked increase of NF-kappaB activation, as determined by electromobility shift assays and transient transfection experiments. Additionally, Amadori adducts significantly increased the production of NF-kappaB-related proinflammatory molecules, including cytokines, such as TNF-alpha, IL-1beta or IL-6, and enzymes, such as cyclooxygenase-2 and inducible nitric oxide (NO) synthase, this latter leading to the release of NO by HPMCs. The effects of Amadori adducts were mediated by different reactive oxygen and nitrosative species (e.g. superoxide anions, hydroxyl radicals, and peroxynitrite), as they were blunted by coincubation with the appropriate scavengers. Furthermore, NO generated upon exposure to Amadori adducts further stimulated NF-kappaB activation, either directly or after combination with superoxide anions to form peroxynitrite. We conclude that Amadori adducts can favour peritoneal inflammation by exacerbating changes in NO synthesis pathway and triggering NF-kappaB-related proinflammatory signals in human mesothelial cells.

Background Insulin resistance (IR) is frequently associated with endothelial dysfunction and has been proposed to play a major role in cardiovascular disease (CVD). On the other hand, obesity has long been related to IR and increased CVD. However it is not known if IR is a necessary condition for endothelial dysfunction in human obesity, allowing for preserved endothelial function in obese people when absent. Therefore, the purpose of the study was to assess the relationship between IR and endothelial dysfunction in human obesity and the mechanisms involved. Methods Twenty non-insulin resistant morbid obese (NIR-MO), 32 insulin resistant morbid obese (IR-MO), and 12 healthy subjects were included. Serum concentrations of glucose, insulin, interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), resistin and adiponectin were determined. IR was evaluated by HOMA-index. Endothelium-dependent relaxation to bradykinin (BK) in mesenteric microvessels was assessed in wire myograph. Results Serum IL-6, and TNF-α levels were elevated only in IR-MO patients while resistin was elevated and adiponectin reduced in all MO individuals. Mesenteric arteries from IR-MO, but not from NIR-MO subjects displayed blunted relaxation to BK. Vasodilatation was improved in IR-MO arteries by the superoxide scavenger, superoxide dismutase (SOD) or the mitochondrial-targeted SOD mimetic, mito-TEMPO. NADPH oxidase inhibitors (apocynin and VAS2870) and the nitric oxide synthase (NOS) cofactor, tetrahydrobiopterin failed to modify BK-induced vasodilatations. Superoxide generation was higher in vessels from IR-MO subjects and reduced by mito-TEMPO. Blockade of TNF-α with infliximab, but not inhibition of inducible NOS or cyclooxygenase, improved endothelial relaxation and decreased superoxide formation. Conclusions Endothelial dysfunction is observed in human morbid obesity only when insulin resistance is present. Mechanisms involved include augmented mitochondrial superoxide generation, and increased systemic inflammation mediated by TNF-α. These findings may explain the different vascular risk of healthy vs unhealthy obesity.

OBJECTIVE: The objective of this study was to analyze the association between candidate gene polymorphisms and susceptibility to acute respiratory distress syndrome (ARDS) in patients with severe sepsis. METHODS: Patients older than 18 years admitted to the intensive care unit (ICU) with the diagnosis of severe sepsis were prospectively included. A blood sample was drawn on the first day of ICU admission, and DNA was extracted. We genotyped the insertion/deletion polymorphism of the angiotensin-converting enzyme (ACE) gene (polymerase chain reaction) and the following single-nucleotide polymorphisms (TaqMan SNP genotyping assay): tumor necrosis factor α -376 G/A, -308 G/A, and -238 G/A; interleukin 8 -251 T/A; pre-B cell colony-enhancing factor -1001 G/T; and vascular endothelial growth factor +405 C/G and +936 C/T. Polymorphisms were selected based on reports on their association with ARDS. Variables associated in univariate analysis (P < 0.1) with the diagnosis of ARDS were included in a multiple logistic regression analysis. RESULTS: We studied 149 patients, of whom 35 presented ARDS. Variables included in the maximal multivariate model were male sex, chronic alcoholism, use of ACE inhibitors or angiotensin-receptor blockers, Simplified Acute Physiology Score II score, serum glucose concentration at ICU admission, and the presence of the allele D of the ACE gene. After adjustment for those variables, the presence of the allele D of the ACE gene (odds ratio, 4.75; 95% confidence interval, 1.02-22.20; P = 0.048) was significantly associated with the diagnosis of ARDS. CONCLUSION: The presence of the allele D of the ACE gene is associated with ARDS in patients with severe sepsis. 2.732 JCR (2013) Q2, 11/27 Critical care medicine, 27/68 Hematology

Purpose The aim of this study was to demonstrate that candidate gene polymorphisms are associated with an increased risk of acute kidney injury (AKI). Materials and Methods Patients admitted to the intensive care unit with the diagnosis of severe sepsis and an expected intensive care unit length of stay more than 48 hours were included. Genetic polymorphisms studied included angiotensin-converting enzyme insertion/deletion (polymerase chain reaction); tumor necrosis factor α − 376, − 308, and − 238; interleukin-8 − 251; vascular endothelial growth factor (VEGF) + 405 and + 936; and pre–B-cell colony-enhancing factor − 1001 (TaqMan SNP genotyping assay, Life Technologies, Grand Island, NY). Acute kidney injury was defined as the risk, injury, and failure categories, as per the RIFLE (risk, injury, failure, loss, end-stage kidney disease) classification. Results One hundred thirty-nine patients were included, 65 of whom developed AKI. In univariate analysis, the VEGF + 936 CC and the pre–B-cell colony-enhancing factor − 1001 GG genotypes were associated with AKI. In multivariate analysis, Simplified Acute Physiology Score II score (odds ratio [95% confidence interval], 1.06 [1.03-1.09]), chronic arterial hypertension (3.15 [1.39-7.15]), and the presence of the VEGF + 936 CC genotype (3.41 [1.19-9.79]) were associated with AKI. Conclusion This is the first study demonstrating an association between the VEGF + 936 CC genotype and the risk to develop AKI in patients with severe sepsis.

Endothelial vasodilation in human vessels is impaired by aging and other cardiovascular risk factors (CVRF) but the differential impact of aging and CVRF in human endothelial function is not completely elucidated. The aim of this work was to evaluate the influence of aging on the effects of peroxisome proliferator-activated receptor (PPAR)-α and -γ subtype agonists on endothelium-dependent vasodilation of isolated human vessels from subjects with or without CVRF. Human mesenteric microarteries were dissected from omentum specimens obtained from subjects younger or older than 60 years having or not CVRF and mounted in wire myographs to evaluate endothelium-dependent relaxation to bradykinin (BK). Aging and CVRF independently reduced endothelium-dependent relaxations. An additional impairment was produced when aging and CVRF co-existed (p0.001). In vessels from adult subjects PPARγ agonist, GW1929 (1 μM) improved BK-induced responses only in those obtained from subjects with CVRF. By contrast, GW1929 improved the responses in vessels from elderly subjects having or not CVRF. PPARα agonist, GW7647 (1 μM), enhanced endothelial vasodilation in adults with CVRF (p0.001) but lack any effect in vessels from older subjects having or not CVRF. In vessels from subjects with CVRF, superoxide dismutase (SOD; 100 U/ml) improved BK-induced responses only in elderly subjects (p0.001). Vascular aging negatively impacts endothelial function independently of the presence of additional CVRF through specific molecular mechanisms involving superoxide generation. While PPARγ activation remains effective, the improving effects of PPARα agonists on endothelial responses disappear in aged human vessels.

Objective: To describe the lung pathological changes in influenza A (H1N1) viral pneumonia. We studied morphological changes, nitro-oxidative stress and the presence of viral proteins in lung tissue. Methods and patients: Light microscopy was used to examine lung tissue from 6 fatal cases of pandemic influenza A (H1N1) viral pneumonia. Fluorescence for oxidized dihydroethydium, nitrotyrosine, inducible NO synthase (NOS2) and human influenza A nucleoprotein (NP) (for analysis under confocal microscopy) was also studied in lung tissue specimens. Results: Age ranged from 15 to 50 years. Three patients were women, and 5 had preexisting medical conditions. Diffuse alveolar damage (DAD) was present in 5 cases (as evidenced by hyaline membrane formation, alveolo-capillary wall thickening and PMN infiltrates), and interstitial fibrosis in one case. In the fluorescence studies there were signs of oxygen radical generation, increased NOS2 protein and protein nitration in lung tissue samples, regardless of the duration of ICU admission. Viral NP was found in lung tissue samples from three patients. Type I pneumocytes and macrophages harbored viral NP, as evidenced by confocal immunofluorescence microscopy. Conclusions: Lung tissue from patients with pandemic influenza A (H1N1) viral pneumonia shows histological findings consistent with DAD. Prolonged nitro-oxidative stress is present despite antiviral treatment. Viral proteins may remain in lung tissue for prolonged periods of time, lodged in macrophages and type I pneumocytes.

Amylin has been linked to the development of hypertension in several pathological states related to hypertension and insulin resistance, although there is scant data regarding its potential mechanisms of action. The -132 G/A mutation located within an activator domain of the amylin gene's promoter was first identified in a small cohort of Spanish patients with type 2 diabetes.The objective of the study was to test the interference of amylin peptide with endothelium-dependent responses as an added potential mechanism for amylin-induced hypertension.A total of 384 patients with type 2 diabetes and 207 healthy controls were subjected to clinical analysis and genetic screening for the -132 G/A mutation of the amylin gene. The effect of amylin on endothelium-dependent responses was analyzed in aortic rings and mesenteric microvessels from nondiabetic rats.The prevalence of the mutation was 10.1 vs. 0.9% in the control population (P0.001). Hypertension was higher in a diabetic population carrying the mutation than in diabetic noncarriers (74 vs. 57%; P0.05). Diabetic carriers showed higher fasting amylin levels than diabetic noncarriers (11.4+/-7 vs. 8.2+/-3 pmol/liter; P0.05). Preincubation with 20 pmol/liter amylin impaired the relaxant responses induced by acetylcholine in rat aorta and mesenteric microvessels. This effect was abolished in both vascular beds in the presence of 100 micromol/liter NG-nitro-L-arginine methyl ester.We propose that amylin levels and hypertension may be linked by a novel mechanism involving the capacity of amylin to induce endothelial dysfunction by interfering with nitric oxide-mediated responses.