Age-related Vascular Dysfunction Research Articles

Medin aggregation causes cerebrovascular dysfunction in aging wild-type mice

Medin is the most common amyloid known in humans, as it can be found in blood vessels of the upper body in virtually everybody over 50 years of age. However, it remains unknown whether deposition of Medin plays a causal role in age-related vascular dysfunction. We now report that aggregates of Medin also develop in the aorta and brain vasculature of wild-type mice in an age-dependent manner. Strikingly, genetic deficiency of the Medin precursor protein, MFG-E8, eliminates not only vascular aggregates but also prevents age-associated decline of cerebrovascular function in mice. Given the prevalence of Medin aggregates in the general population and its role in vascular dysfunction with aging, targeting Medin may become a novel approach to sustain healthy aging.

Trimethylamine-N-Oxide Promotes Age-Related Vascular Oxidative Stress and Endothelial Dysfunction in Mice and Healthy Humans.

Age-related vascular endothelial dysfunction is a major antecedent to cardiovascular diseases. We investigated whether increased circulating levels of the gut microbiome-generated metabolite trimethylamine-N-oxide induces endothelial dysfunction with aging. In healthy humans, plasma trimethylamine-N-oxide was higher in middle-aged/older (64±7 years) versus young (22±2 years) adults (6.5±0.7 versus 1.6±0.2 µmol/L) and inversely related to brachial artery flow-mediated dilation (r2=0.29, P<0.00001). In young mice, 6 months of dietary supplementation with trimethylamine-N-oxide induced an aging-like impairment in carotid artery endothelium-dependent dilation to acetylcholine versus control feeding (peak dilation: 79±3% versus 95±3%, P<0.01). This impairment was accompanied by increased vascular nitrotyrosine, a marker of oxidative stress, and reversed by the superoxide dismutase mimetic 4-hydroxy-2,2,6,6-tetramethylpiperidin-1-oxyl. Trimethylamine-N-oxide supplementation also reduced activation of endothelial nitric oxide synthase and impaired nitric oxide-mediated dilation, as assessed with the nitric oxide synthase inhibitor L-NAME (NG-nitro-L-arginine methyl ester). Acute incubation of carotid arteries with trimethylamine-N-oxide recapitulated these events. Next, treatment with 3,3-dimethyl-1-butanol for 8 to 10 weeks to suppress trimethylamine-N-oxide selectively improved endothelium-dependent dilation in old mice to young levels (peak: 90±2%) by normalizing vascular superoxide production, restoring nitric oxide-mediated dilation, and ameliorating superoxide-related suppression of endothelium-dependent dilation. Lastly, among healthy middle-aged/older adults, higher plasma trimethylamine-N-oxide was associated with greater nitrotyrosine abundance in biopsied endothelial cells, and infusion of the antioxidant ascorbic acid restored flow-mediated dilation to young levels, indicating tonic oxidative stress-related suppression of endothelial function with higher circulating trimethylamine-N-oxide. Using multiple experimental approaches in mice and humans, we demonstrate a clear role of trimethylamine-N-oxide in promoting age-related endothelial dysfunction via oxidative stress, which may have implications for prevention of cardiovascular diseases.

Initiation of the Gut Microbiome Targeted Compound 3,3‐Dimethyl‐1‐butanol at Mid‐life Prevents Age‐related Vascular Dysfunction

Aging induces vascular dysfunction (arterial stiffening and endothelial dysfunction) mediated by superoxide‐driven oxidative stress and reduced nitric oxide (NO) bioavailability, which increases risk of cardiovascular (CV) diseases. A potential intervention target is the gut microbiome‐derived metabolite trimethylamine N‐oxide (TMAO), which increases in circulation with advancing age, is associated with higher CV risk, and can be inhibited by the food‐derived compound 3,3‐dimethyl‐1‐butanol (DMB).PurposeTo determine if DMB initiated at mid‐life (i.e., prior to the onset of dysfunction) prevents the development of age‐related vascular dysfunction.MethodsMiddle‐aged (18 mo) male C57BL/6 mice received drinking water with (n=24) or without (control: n=27) 1% DMB until sacrifice at 21, 24, or 27 mo of age (n=7–13/age/treatment). Arterial stiffness was assessed serially in vivo by aortic pulse wave velocity (aPWV). Endothelial function (ex vivo carotid artery endothelium‐dependent dilation [EDD] to acetylcholine), plasma TMAO (LC‐MS) and aortic superoxide (electron paramagnetic resonance spectroscopy) were assessed post‐sacrifice. To determine the prevention of age‐related vascular dysfunction with DMB, functional measures were made in a reference group of young (5 mo) male C57BL/6 mice (YC, n=12).ResultsPlasma TMAO increased with age vs YC (3.1±0.5 μM), but tended to be lower in DMB (5.6±0.5 μM) vs control (6.9±0.8 μM) mice at 24 and 27 mo (p=0.15). In control mice, aPWV was similar to YC (356±14 cm/s) at 18/21 mo (359±12/368±11 cm/s, both p&gt;0.54), but increased with age (24 mo: 386±12 cm/s, p=0.06 vs 18 mo; 27 mo: 432±10 cm/s, p&lt;0.001 vs 18 mo); this increase was attenuated by ~50% in DMB mice (aPWV at 24 mo: 359±9 cm/s; 27 mo: 388±9 cm/s, both p=0.01 vs control). Age‐related declines in peak EDD were apparent in control mice at 24 mo (79±3 vs YC 93±2%, p&lt;0.001) and 27 mo (76±3%, p&lt;0.0001 vs YC) and were fully prevented by DMB (24 mo: 91±3%; 27 mo: 93±1%; both p&lt;0.05 vs control). The effects of DMB at 24 and 27 mo (data combined across these ages) were mediated by reduced aortic superoxide production (control 4.7±0.5 vs DMB 3.1±0.4 AU, p=0.01), which reduced tonic superoxide‐associated suppression of EDD (normalization of EDD with superoxide dismutase mimetic TEMPOL) and restored NO‐mediated dilation (the difference in peak EDD in the absence vs presence of the NO synthase inhibitor L‐NAME: control 45±6 vs DMB 69±4%, p&lt;0.01). There were no differences in smooth muscle sensitivity to NO (peak dilation to sodium nitroprusside; 96±1 vs 98±1%, p=0.12), indicating improvements in EDD were endothelium‐specific.ConclusionSupplementation with DMB from mid‐life through old age in mice prevented the development of arterial stiffening and endothelial dysfunction. DMB may be a promising therapeutic strategy for preventing age‐related vascular dysfunction and thereby reducing CV risk with aging in humans.Support or Funding InformationR01 HL1348870‐02S1, R21 AG060884, F32 HL140875

Vascular age in patients with arterial hypertension

Ageing is considered to be the major and non-modifiable risk factor for the development of hypertension and cardiovascular diseases. During ageing, the vascular system undergoes structural and functional alterations, including endothelial dysfunction, thickening of the vascular wall, reduced distensibility and increased arterial stiffness. Vascular rigidity results from fibrosis and remodelling of the extracellular matrix, processes that are associated with ageing and are amplified in hypertension. These events may be induced by vasoactive agents, such as angiotensin II, endothelin-1, and aldosterone, which are increased in the vasculature during aging and hypertension. Complex interaction between the process of ageing and prohypertensive factors results in accelerated vascular remodelling and fibrosis, as well as increased arterial stiffness. Hypertension accelerates and augments age-related vascular remodelling and dysfunction, and ageing may impact on the severity of vascular damage in hypertension, thus strongly suggesting close interactions between biological ageing and blood pressure elevation. Molecular and cellular mechanisms underlying vascular alterations in ageing and hypertension are common and include aberrant signal transduction, oxidative stress and activation of pro-inflammatory and pro-fibrotic transcription factors. Strategies to suppress age-associated vascular changes can ameliorate vascular damage associated with hypertension. This article looks into vascular alterations occurring during ageing and hypertension, focussing particularly on arterial stiffness and vascular remodelling, also emphasizing the importance of diagnostic methods.

Co-expression network analysis identified hub genes critical to triglyceride and free fatty acid metabolism as key regulators of age-related vascular dysfunction in mice.

Background: Aging has often been linked to age-related vascular disorders. The elucidation of the putative genes and pathways underlying vascular aging likely provides useful insights into vascular diseases at advanced ages. Transcriptional regulatory network analysis is the key to describing genetic interactions between molecular regulators and their target gene transcriptionally changed during vascular aging.Results: A total of 469 differentially expressed genes were parsed into 6 modules. Among the incorporated sample traits, the most significant module related to vascular aging was associated with triglyceride and enriched with biological terms like proteolysis, blood circulation, and circulatory system process. The module associated with triglyceride was preserved in an independent microarray dataset, indicating the robustness of the identified vascular aging-related subnetwork. Additionally, Enpp5, Fez1, Kif1a, F3, H2-Q7, and their interacting miRNAs mmu-miR-449a, mmu-miR-449c, mmu-miR-34c, mmu-miR-34b-5p, mmu-miR-15a, and mmu-let-7, exhibited the most connectivity with external lipid-related traits. Transcriptional alterations of the hub genes Enpp5, Fez1, Kif1a, and F3, and the interacting microRNAs mmu-miR-34c, mmu-miR-34b-5p, mmu-let-7, mmu-miR-449a, and mmu-miR-449c were confirmed.Conclusion: Our findings demonstrate that triglyceride and free fatty acid-related genes are key regulators of age-related vascular dysfunction in mice and show that the hub genes for Enpp5, Fez1, Kif1a, and F3 as well as their interacting miRNAs mmu-miR-34c, mmu-miR-34b-5p, mmu-let-7, mmu-miR-449a, and mmu-miR-449c, could serve as potential biomarkers in vascular aging.Methods: The microarray gene expression profiles of aorta samples from 6-month old mice (n=6) and 20-month old mice (n=6) were processed to identify nominal differentially expressed genes. These nominal differentially expressed genes were subjected to a weighted gene co-expression network analysis. A network-driven integrative analysis with microRNAs and transcription factors was performed to define significant modules and underlying regulatory pathways associated with vascular aging, and module preservation test was conducted to validate the age-related modules based on an independent microarray gene expression dataset in mice aorta samples including three 32-week old wild-type mice (around 6-month old) and three 78-week old wild-type mice (around 20-month old). Gene ontology and protein-protein interaction analyses were conducted to determine the hub genes as potential biomarkers in the progress of vascular aging. The hub genes were further validated with quantitative real-time polymerase chain reaction in aorta samples from 20 young (6-month old) mice and 20 old (20-month old) mice.

Abstract P122: Downregulation and Inhibition of PKCα Causes Increased Ca 2+ -Sensing Receptor-Mediated Vascular Relaxation in Aged Dahl Salt-Sensitive Rats

The G protein-coupled Ca 2+ -Sensing Receptor (CaSR) mediates extracellular calcium (Ca 2+ e ) - induced relaxation of mesenteric arteries under tension and may therefore play a role in blood pressure regulation. This presents CaSR as promising target for therapeutic drug design for cardiovascular pathologies such as hypertension. Protein kinase Cα (PKCα) is an important regulator of CaSR signaling and can phosphorylate the receptor leading to desensitization. In this study, we investigated the effects of CaSR mutation and downregulation of PKCα in CaSR-mediated relaxation of phenylephrine (PE)-contracted mesenteric arteries from aged Dahl salt-sensitive rats, fed a low salt (0.1% NaCl) diet, using automated wire myography. We have observed in our laboratory, that there is significant decrease in relaxation response of PE-contracted mesenteric arteries to Ca 2+ e . as animals get older. The results show minimal relaxation responses of arteries to Ca 2+ e and that downregulation of CaSR expression had no significant effect. PKCα downregulation (siRNA) and inhibition of PKC (Gö 6976) showed greater than 80% increase in maximum Ca 2+ -induced relaxation, with significant decreases in EC 50 values compared to untreated controls. The results indicate that PKCα plays an important role in the regulation of CaSR -mediated relaxation of mesenteric arteries. Therefore, inhibition of PKC may lead to increased CaSR sensitivity to Ca 2+ indicating a possible therapeutic mechanism for intervention in age-related vascular dysfunction.

Protein Kinase C Downregulation Enhanced Extracellular Ca2+-Induced Relaxation of Isolated Mesenteric Arteries from Aged Dahl Salt-Sensitive Rats.

The Ca2+-sensing receptor (CaSR) detects small changes in extracellular calcium (Ca2+ e) concentration ([Ca2+]e) and transduces the signal into modulation of various signaling pathways. Ca2+-induced relaxation of isolated phenylephrine-contracted mesenteric arteries is mediated by the CaSR of the perivascular nerve. Elucidation of the regulatory mechanisms involved in vascular CaSR signaling may provide insights into the physiologic functions of the receptor and identify targets for the development of new treatments for cardiovascular pathologies such as hypertension. Protein kinase Cα (PKCα) is a critical regulator of multiple signaling pathways and can phosphorylate the CaSR leading to receptor desensitization. In this study, we used automated wire myography to investigate the effects of CaSR mutation and small-interfering RNA downregulation of PKCα on CaSR-mediated relaxation of phenylephrine-contracted mesenteric arteries from aged Dahl salt-sensitive (SS) rats on a low-salt diet. The data showed minimal relaxation responses of arteries to Ca2+ e in wild-type (SS) and CaSR mutant (SS-Casrem1Mcwi) rats. Mutation of the CaSR gene had no significant effect on relaxation. PKCα expression was similar in wild-type and mutant rats, and small-interfering RNA downregulation of PKCα and/or inhibition of PKC with the Ca2+-sensitive Gӧ 6976 resulted in a >80% increase in relaxation. Significant differences in EC50 values were observed between treated and untreated controls (P < 0.05 analysis of variance). The results indicate that PKCα plays an important role in the regulation of CaSR-mediated relaxation of mesenteric arteries, and its downregulation or pharmacological inhibition may lead to an increased Ca2+ sensitivity of the receptor and reversal of age-related changes in vascular tone. SIGNIFICANCE STATEMENT: G protein-coupled CaSR signaling leads to the regulation of vascular tone and may, therefore, play a vital role in blood pressure regulation. The receptor has several PKC phosphorylation sites in the C-terminal intracellular tail that mediate desensitization. We have previously shown that activation of the CaSR in neuronal cells leads to PKC phosphorylation, indicating that protein kinase C is an important regulator of CaSR function. Therefore, PKC in the CaSR signaling pathway in mesenteric arteries is a potential target for the development of new therapeutic approaches to treat hypertension and age-related vascular dysfunction. The present studies show that small-interfering RNA downregulation of PKCα and pharmacological inhibition of PKC enhanced CaSR-mediated relaxation of phenylephrine-contracted mesenteric arteries from aged Dahl salt-sensitive rats.

Age-Related Vascular Dysfunction: What Registered Dietitian Nutritionists Need to Know

Cardiovascular disease (CVD) is an age-related chronic disease, where advancing age is the primary risk factor largely due to age-related vascular dysfunction. 1 Benjamin E.J. Blaha M.J. Chiuve S.E. et al. Heart Disease and Stroke Statistics-2017 update: A report from the American Heart Association. Circulation. 2017; 135: e146-e603 Crossref PubMed Scopus (5194) Google Scholar , 2 LaRocca T.J. Martens C.R. Seals D.R. Nutrition and other lifestyle influences on arterial aging. Ageing Res Rev. 2017; 39: 106-119 Crossref PubMed Scopus (32) Google Scholar , 3 Lakatta E.G. Levy D. Arterial and cardiac aging: Major shareholders in cardiovascular disease enterprises: Part I: Aging arteries: A “set up” for vascular disease. Circulation. 2003; 107: 139-146 Crossref PubMed Scopus (1507) Google Scholar During aging, several adverse structural and functional changes occur throughout the entire vascular system (ie, heart, coronary arteries, peripheral arteries, and microvasculature); however, two prominent changes to the arteries are emerging as the most clinically important due to their striking effect on age-related CVD risk: stiffening of the large elastic arteries (carotid and aorta) and vascular endothelial dysfunction (Figure 1). 2 LaRocca T.J. Martens C.R. Seals D.R. Nutrition and other lifestyle influences on arterial aging. Ageing Res Rev. 2017; 39: 106-119 Crossref PubMed Scopus (32) Google Scholar , 4 Seals D.R. Brunt V.E. Rossman M.J. Keynote lecture: strategies for optimal cardiovascular aging. Am J Physiol Heart Circ Physiol. 2018; 315: H183-H188 Crossref PubMed Scopus (28) Google Scholar , 5 Rossman M.J. LaRocca T.J. Martens C.R. Seals D.R. Healthy lifestyle-based approaches for successful vascular aging. J Appl Physiol (1985). 2018; 125: 1888-1900 Crossref PubMed Scopus (16) Google Scholar These arterial changes, referred to together as vascular dysfunction, vascular aging, arterial dysfunction, and/or arterial aging (referred to herein as vascular dysfunction), contribute to the development of atherosclerosis and hypertension (isolated systolic hypertension in particular), and can lead to stroke, myocardial infarction, and other adverse events including death. Mechanisms underlying vascular dysfunction with aging are complex and multifactorial, but excessive oxidative stress and chronic inflammation are believed to be the major central contributors. 4 Seals D.R. Brunt V.E. Rossman M.J. Keynote lecture: strategies for optimal cardiovascular aging. Am J Physiol Heart Circ Physiol. 2018; 315: H183-H188 Crossref PubMed Scopus (28) Google Scholar , 5 Rossman M.J. LaRocca T.J. Martens C.R. Seals D.R. Healthy lifestyle-based approaches for successful vascular aging. J Appl Physiol (1985). 2018; 125: 1888-1900 Crossref PubMed Scopus (16) Google Scholar Numerous factors can accelerate these processes and therefore vascular dysfunction in aging individuals, or cause premature development in others. 6 Gimbrone Jr., M.A. Garcia-Cardena G. Endothelial cell dysfunction and the pathobiology of atherosclerosis. Circ Res. 2016; 118: 620-636 Crossref PubMed Scopus (1043) Google Scholar For instance, menopause is associated with accelerated vascular dysfunction in women, 7 Moreau K.L. Hildreth K.L. Vascular aging across the menopause transition in healthy women. Adv Vasc Med. 2014; 2014 PubMed Google Scholar whereas obesity and related metabolic disorders can also induce and accelerate vascular dysfunction with aging. 3 Lakatta E.G. Levy D. Arterial and cardiac aging: Major shareholders in cardiovascular disease enterprises: Part I: Aging arteries: A “set up” for vascular disease. Circulation. 2003; 107: 139-146 Crossref PubMed Scopus (1507) Google Scholar Certain lifestyle behaviors such as regular physical activity, healthy dietary patterns, and phytochemical-rich food consumption can help attenuate and in some cases reverse these processes and reduce age-related CVD risk. 4 Seals D.R. Brunt V.E. Rossman M.J. Keynote lecture: strategies for optimal cardiovascular aging. Am J Physiol Heart Circ Physiol. 2018; 315: H183-H188 Crossref PubMed Scopus (28) Google Scholar , 5 Rossman M.J. LaRocca T.J. Martens C.R. Seals D.R. Healthy lifestyle-based approaches for successful vascular aging. J Appl Physiol (1985). 2018; 125: 1888-1900 Crossref PubMed Scopus (16) Google Scholar

Endothelial PPARγ Is Crucial for Averting Age-Related Vascular Dysfunction by Stalling Oxidative Stress and ROCK.

Aging plays a significant role in the progression of vascular diseases and vascular dysfunction. Activation of the ADP-ribosylation factor 6 and small GTPases by inflammatory signals may cause vascular permeability and endothelial leakage. Pro-inflammatory molecules have a significant effect on smooth muscle cells (SMC). The migration and proliferation of SMC can be promoted by tumor necrosis factor alpha (TNF-α). TNF-α can also increase oxidative stress in SMCs, which has been identified to persuade DNA damage resulting in apoptosis and cellular senescence. Peroxisome proliferator-activated receptor (PPAR) acts as a ligand-dependent transcription factor and a member of the nuclear receptor superfamily. They play key roles in a wide range of biological processes, including cell differentiation and proliferation, bone formation, cell metabolism, tissue remodeling, insulin sensitivity, and eicosanoid signaling. The PPARγ activation regulates inflammatory responses, which can exert protective effects in the vasculature. In addition, loss of function of PPARγ enhances cardiovascular events and atherosclerosis in the vascular endothelium. This appraisal, therefore, discusses the critical linkage of PPARγ in the inflammatory process and highlights a crucial defensive role for endothelial PPARγ in vascular dysfunction and disease, as well as therapy for vascular aging.

Sodium Nitrite Supplementation Improves Vascular Endothelial Function but not Motor or Cognitive Function in Middle‐Aged and Older Adults

We previously observed improvements in vascular endothelial function as well as exploratory measures of motor and cognitive function in a pilot study of sodium nitrite supplementation in middle‐aged and older (MA/O) adults. Here, we sought to establish the efficacy of sodium nitrite for improving endothelial function (primary outcome) while also taking advantage of the opportunity to confirm our motor and cognitive function findings in a larger randomized, placebo‐controlled, double‐blind, parallel group study with 12 weeks of sodium nitrite (80 mg/day) vs. placebo in healthy MA/O adults (n = 49, 68±1 yr) with impaired endothelial function (baseline brachial artery flow‐mediated dilation [FMD] &lt;6%). Sodium nitrite increased plasma levels of nitrite acutely (10‐fold, p&lt;0.05 vs placebo) and chronically (p&lt;0.05) and was well‐tolerated over the 12‐week supplementation period. Vascular endothelial function, measured by brachial artery FMD, was increased by 28% vs. baseline (p&lt;0.05) after 12 weeks of supplementation (3.9±1.2 to 5.0±1.8%, p&lt;0.05), but unchanged with placebo (3.8±1.4 to 4.0±1.5%, p&gt;0.05). There were no differences in secondary outcomes of motor or cognitive function assessed with measures of the NIH Toolbox and other standardized tests between the sodium nitrite and placebo group after 12 weeks (all p&gt;0.05). There were no differences in subject characteristics, body composition or circulating markers of inflammation and oxidative stress with sodium nitrite or placebo (all p&gt;0.05). Collectively, these results indicate that sodium nitrite supplementation for 12 weeks improves endothelial function but does not affect motor or cognitive function in healthy, high‐functioning MA/O adults. These findings suggest therapeutic strategies to enhance nitrite levels may hold promise for treating age‐related vascular endothelial dysfunction.Clinicaltrials.gov Identifier: NCT02393742Support or Funding InformationNIH R01 AG013038, K01 DK115524 and NIH/NCATS Colorado CTSA Grant Number UL1 TR002535; American Physiology Society UGREF AwardThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Dietary Glycocalyx Precursor Supplementation Ameliorates Age‐Related Vascular Dysfunction

Large elastic artery stiffening and endothelial dysfunction, and associated reductions in nitric oxide (NO) bioavailability, are central features of vascular aging. We have recently demonstrated that the glycocalyx, a gel‐like structure that is bound to the luminal surface of the vascular endothelium, is dysfunctional in the aged vasculature. The glycocalyx has several functions that are critical for the maintenance of a healthy vasculature. We sought to determine if chronic dietary supplementation of glycocalyx precursors (glucosamine sulfate, fucoidan, superoxide dismutase, and high molecular weight hyaluronan) could restore glycocalyx function, while concomitantly ameliorating age‐related vascular dysfunction. Young (Y: 7 mo) and old (O: 30 mo) male B6D2F1 mice consumed a control (C) or glycocalyx precursor (GP: 37 mg/kg encapsulated chow provided courtesy of MicroVascular Health Solutions, LLC [U.S. Patent Serial No. 9,943,572]) diet ad libitum for 10 weeks. Glycocalyx barrier function (perfused boundary region [PBR]) was evaluated in the mesenteric microcirculation using an intravital microscope equipped with an automated capture and analysis system. PBR was ~13% higher in OC compared to YC, suggestive of an age‐related impairment in glycocalyx barrier function, and this was normalized in OGP mice (Both P&lt;0.05; Figure 1). At baseline, aortic pulse wave velocity (PWV), a measure of large artery stiffness, was higher in OC and OGP compared with YC mice (Both P&lt;0.05; Figure 2). However, after the dietary intervention, PWV decreased by ~13% in OGP (P&lt;0.05), whereas, PWV was unchanged in OC and YC mice after the 10 week period (P&gt;0.05). We assessed endothelial function by endothelium‐dependent dilation (EDD, maximal response to acetylcholine [ACh]) in the carotid artery. Carotid artery EDD was higher in YC and OGP compared to OC mice (92.5±2.4 and 90.7±2.3 vs. 69.0±4.9%, respectively, P&lt;0.05). EDD of OGP was similar to YC mice (P&gt;0.05). After incubation with the nitric oxide (NO) synthase inhibitor, L‐NAME, the dilatory response did not differ between groups (P&gt;0.05). NO bioavailability (max ACh dilation ‐ max ACh+L‐NAME dilation) was ~10–14 fold higher in YC and OGP compared to OC mice (Both P&lt;0.05; Figure 3). Endothelium‐independent dilation (vasodilation to sodium nitroprusside) was not different between groups (P&gt;0.05). In young mice, GP diet did not affect any of the aforementioned measurements (P&gt;0.05). In conclusion, 10 weeks of dietary GP supplementation in old mice restores glycocalyx barrier function that is accompanied by reduced aortic stiffness and augmented EDD and NO bioavailability, suggesting that the glycocalyx may be an effective therapeutic target for vascular dysfunction in older adults.Support or Funding InformationThis study was funded in part by grants from the National Institute of Health (R01 AG040297, R01 AG048366, K02 AG045339, K99 AT010017) and US Department of Veterans Affairs (1I01BX002151).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Role of endothelial NAD+ deficiency in age-related vascular dysfunction.

Age-related alterations in endothelium and the resulting vascular dysfunction critically contribute to a range of pathological conditions associated with old age. To develop therapies rationally that improve vascular health and thereby increase health span and life span in older adults, it will be essential to understand the cellular and molecular mechanisms contributing to vascular aging. Preclinical studies in model organisms demonstrate that NAD+ availability decreases with age in multiple tissues and that supplemental NAD+ precursors can ameliorate many age-related cellular impairments. Here, we provide a comprehensive overview of NAD+-dependent pathways [including the NAD+-using silent information regulator-2-like enzymes and poly(ADP-ribose) polymerase enzymes] and the potential consequences of endothelial NAD+ deficiency in vascular aging. The multifaceted vasoprotective effects of treatments that reverse the age-related decline in cellular NAD+ levels, as well as their potential limitations, are discussed. The preventive and therapeutic potential of NAD+ intermediates as effective, clinically relevant interventions in older adults at risk for ischemic heart disease, vascular cognitive impairment, and other common geriatric conditions and diseases that involve vascular pathologies (e.g., sarcopenia, frailty) are critically discussed. We propose that NAD+ precursors [e.g., nicotinamide (Nam) riboside, Nam mononucleotide, niacin] should be considered as critical components of combination therapies to slow the vascular aging process and increase cardiovascular health span.

Vasodilatory function in human skeletal muscle feed arteries with advancing age: the role of adropin.

The present study aimed to determine the impact of ageing on endogenous adropin levels in human skeletal muscle feed arteries (SMFAs) and the role of adropin in age-related vascular dysfunction. Adropin protein expression falls progressively with advancing age in the human peripheral vasculature. Endothelial-dependent vasodilatation, typically attenuated with age, was strongly correlated with SMFA adropin protein levels. Adropin incubation restored age-related endothelial-dependent vasodilatory dysfunction and increased the phosphorylated endothelial nitric oxide synthase (eNOS)/eNOS ratio in an age-dependent manner in the SMFAs. The role of nitric oxide bioavailability was additionally indicated by NOS blockade ablating both the positive vascular effects of adropin incubation and the relationship between endothelial function and adropin protein expression. Additional evidence of a mechanistic link between declining adropin and age-related endothelial dysfunction was documented by a progressively increasing magnitude of effect of adropin-induced eNOS-mediated vasodilatation with ageing. Adropin appears to be a novel therapeutic target for facilitating the restoration of endothelial function with ageing. The present study aimed to determine the impact of advancing age on endogenous adropin levels in human skeletal muscle feed arteries (SMFAs) and the role of adropin in age-related vascular dysfunction. Adropin protein expression and vasodilatory capacity was assesed in SMFAs from Young (27±2years, n=10), Middle Aged (54±2years, n=10) and Old (75±2years, n=16) subjects. Endothelial-dependent vasodilatation, with and without adropin incubation, was assessed in response to flow-induced shear stress and ACh. Both SMFA adropin protein expression and endothelial-dependent vasodilatory function exhibited a progressive, age-related, reduction (Flow: Y: 65±3%; Middle Aged: 36±3%; Old: 15±2%; ACh: Young: 63±2%, Middle Aged: 34±3%; Old: 23±3%, P<0.05). There was a strong positive correlation between SMFA adropin protein expression and both flow (r=0.81, P<0.05) and ACh (r=0.78, P<0.05). Adropin incubation in the Middle Aged and Old SMFAs restored the vasodilatory response to flow (Middle Aged+Adropin: 59±3%; Old+Adropin: 47±3%, P<0.05) and ACh (Middle Aged+Adropin: 59±3%; Old+Adropin: 49±2%, P<0.05). A mechanistic link between adropin and nitric oxide (NO) biovavailabilty was supported by (i) increased phosphorylated endothelial NO synthase (eNOS)/eNOS protein expression with adropin incubation only in the Middle Aged and Old SMFAs; (ii) eNOS blockade ablating both the positive vascular effects of adropin incubation and the relationship between endothelial function and adropin protein expression and (iii) a progressive increase in the magnitude of effect of adropin-induced eNOS-mediated vasodilatation with advancing age. Adropin could be a novel therapeutic target for facilitating the restoration of endothelial function via increased NO bioavailability, with advancing age.

Induced Trf2 deletion leads to aging vascular phenotype in mice associated with arterial telomere uncapping, senescence signaling, and oxidative stress

Age-related vascular dysfunction in large elastic and resistance arteries is associated with reductions in microvascular perfusion and elevations in blood pressure. Recent evidence indicates that telomere uncapping-induced senescence in vascular cells may be an important source of oxidative stress and vascular dysfunction in aging, but the causal relationship between these processes has yet to be elucidated. To test this important unexplored hypothesis, we measured arterial senescence signaling and oxidative stress, carotid and mesenteric artery endothelium-dependent vasodilatory capacity, markers of mesenteric microvascular perfusion and endothelial glycocalyx deterioration, and blood pressure in a novel mouse model of Cre-inducible whole body Trf2 deletion and telomere uncapping. Trf2 deletion led to a 320% increase in arterial senescence signaling (P < .05). There was a concurrent 29% and 22% reduction in peak endothelium-dependent vasodilation in carotid and mesenteric arteries, respectively, as well as a 63% reduction in mesenteric microvascular endothelial glycocalyx thickness (all P ≤ .01). Mesenteric microvascular perfusion was reduced by 8% and systolic blood pressure was increased by 9% following Trf2 deletion (both P < .05). Trf2 deletion also led to a pro-oxidative arterial phenotype characterized by increased in NADPH oxidase gene expression; a 210% increase in superoxide levels that was partly dependent on NADPH oxidase activity; and an oxidative stress mediated reduction in carotid artery vasodilation (all P ≤ .05). Collectively, our findings demonstrate that induced Trf2 deletion leads to telomere uncapping, increased senescence signaling, and oxidative stress mediated functional impairments in the vasculature similar to those seen in human aging.

Healthy lifestyle-based approaches for successful vascular aging.

This review summarizes a presentation given at the 2016 Gerontological Society of America Annual Meeting as part of the Vascular Aging Workshop. The development of age-related vascular dysfunction increases the risk of cardiovascular disease as well as other chronic age-associated disorders, including chronic kidney disease and Alzheimer's disease. Healthy lifestyle behaviors, most notably regular aerobic exercise and certain dietary patterns, are considered "first-line" strategies for the prevention and/or treatment of vascular dysfunction with aging. Despite the well-established benefits of these strategies, however, many older adults do not meet the recommended guidelines for exercise or consume a healthy diet. Therefore, it is important to establish alternative and/or complementary evidence-based approaches to prevent or reverse age-related vascular dysfunction. Time-efficient forms of exercise training, hormetic exposure to mild environmental stress, fasting "mimicking" dietary paradigms, and nutraceutical/pharmaceutical approaches to favorably modulate cellular and molecular pathways activated by exercise and healthy dietary patterns may hold promise as such alternative approaches. Determining the efficacy of these novel strategies is important to provide alternatives for adults with low adherence to conventional healthy lifestyle practices for healthy vascular aging.

Vascular mitochondrial respiratory function: the impact of advancing age.

Little is known about vascular mitochondrial respiratory function and the impact of age. Therefore, skeletal muscle feed arteries were harvested from young (33 ± 7 yr, n = 10), middle-aged (54 ± 5 yr, n = 10), and old (70 ± 7 yr, n = 10) subjects, and mitochondrial respiration as well as citrate synthase (CS) activity were assessed. Complex I (CI) and complex I + II (CI+II) state 3 respiration were greater in young (CI: 10.4 ± 0.8 pmol·s-1·mg-1 and CI+II: 12.4 ± 0.8 pmol·s-1·mg-1, P < 0.05) than middle-aged (CI: 7 ± 0.6 pmol·s-1·mg-1 and CI+II: 8.3 ± 0.5 pmol·s-1·mg-1) and old (CI: 7.2 ± 0.4 pmol·s-1·mg-1 and CI+II: 7.6 ± 0.5 pmol·s-1·mg-1) subjects and, as in the case of complex II (CII) state 3 respiration, were inversely correlated with age [ r = -0.56 (CI), r = -0.7 (CI+II), and r = 0.4 (CII), P < 0.05]. In contrast, state 4 respiration and mitochondria-specific superoxide levels were not different across groups. The respiratory control ratio was greater in young (2.2 ± 0.2, P < 0.05) than middle-aged and old (1.4 ± 0.1 and 1.1 ± 0.1, respectively) subjects and inversely correlated with age ( r = -0.71, P < 0.05). As CS activity was inversely correlated with age ( r = -0.54, P < 0.05), when normalized for mitochondrial content, the age-related differences and relationships with state 3 respiration were ablated. In contrast, mitochondrion-specific state 4 respiration was now lower in young (15 ± 1.4 pmol·s-1·mg-1·U CS-1, P < 0.05) than middle-aged and old (23.4 ± 3.6 and 27.9 ± 3.4 pmol·s-1·mg-1·U CS-1, respectively) subjects and correlated with age ( r = 0.46, P < 0.05). Similarly, superoxide/CS levels were lower in young (0.07 ± 0.01) than old (0.19 ± 0.41) subjects and correlated with age ( r = 0.44, P < 0.05). Therefore, with aging, vascular mitochondrial respiratory function declines, predominantly as a consequence of falling mitochondrial content. However, per mitochondrion, aging likely results in greater mitochondrion-derived oxidative stress, which may contribute to age-related vascular dysfunction. NEW & NOTEWORTHY This study determined, for the first time, that vascular mitochondrial oxidative respiratory capacity, oxidative coupling efficiency, and mitochondrial content fell progressively with advancing age. In terms of single mitochondrion-specific respiration, the age-related differences were completely ablated and the likelihood of free radical production increased progressively with advancing age. This study reveals that vascular mitochondrial respiratory capacity declines with advancing age, as a consequence of falling mitochondrial content, as does oxidative coupling efficiency.

Endothelial PPARγ (Peroxisome Proliferator-Activated Receptor-γ) Is Essential for Preventing Endothelial Dysfunction With Aging.

Little is known about mechanisms that control vascular aging, particularly at the cell-specific level. PPARγ (peroxisome proliferator-activated receptor-γ) exerts protective effects in the vasculature when activated pharmacologically. To gain insight into the cell-specific impact of PPARγ, we examined the hypothesis that genetic interference with endothelial PPARγ would augment age-induced vascular dysfunction. We studied carotid arteries from adult (11.6±0.3 months) and old (24.7±0.6 months) mice with endothelial-specific expression of a human dominant negative mutation in PPARγ driven by the vascular cadherin promoter (E-V290M), along with age-matched, nontransgenic littermates. Acetylcholine (an endothelium-dependent agonist) produced similar relaxation in arteries from adult nontransgenic and E-V290M mice and old nontransgenic mice. In contrast, responses to acetylcholine were reduced by >50% in old male and female E-V290M mice (P<0.01). Endothelial function in old E-V290M mice was not altered by an inhibitor of COX (cyclooxygenase) but was restored to normal by a superoxide scavenger, an inhibitor of NADPH oxidase, or inhibition of ROCK (Rho kinase). Relaxation of arteries to nitroprusside, which acts directly on vascular muscle, was similar in all groups. Vascular expression of IL (interleukin)-6, Nox-2, and CDKN2A (a marker of senescence) was significantly increased in old E-V290M mice compared with controls (P<0.05). These findings provide the first evidence that age-related vascular dysfunction, inflammation, and senescence is accelerated after interference with endothelial PPARγ via mechanisms involving oxidative stress and ROCK. The finding of an essential protective role for endothelial PPARγ has implications for vascular disease and therapy for vascular aging.

Abstract 018: TET2 Loss of Function-driven Clonal Hematopoiesis Promotes Age-related Vascular and Systemic Inflammation and Metabolic Dysfunction in Mice

Background: Acquired mutations in the hematopoietic system and subsequent clonal hematopoiesis are emerging as potential new drivers of atherosclerotic cardiovascular disease. TET2 was the first gene reported to exhibit acquired mutations in blood cells of cancer-free elderly individuals and we recently reported that the clonal expansion of TET2-mutant hematopoietic cells accelerates atherogenesis in mouse models by exacerbating inflammatory responses in the atherosclerotic plaque. The objective of this study was to examine whether TET2 loss of function-driven clonal hematopoiesis also has systemic effects on age-related inflammation and metabolic function beyond its direct actions on the plaque. Methods: An adoptive transfer technique was used to introduce a small percentage of Tet2-/- hematopoietic cells in non-irradiated C57Bl/6J mice, which were monitored for 16 months following transfer. Flow cytometry techniques were used to follow the expansion of Tet2-/- cells in the blood. Results: Tet2-/- hematopoietic cells expanded progressively over time after adoptive transfer, with a mild myeloid bias. This was paralleled by increased expression of the pro-inflammatory cytokines IL-1β and IL-6 in the white adipose tissue and the aortic wall, as well as by increased serum levels of IL-6. Tet2-/- expansion also led to exacerbated systemic insulin resistance, as revealed by insulin tolerance tests. Conclusion: TET2-deficient hematopoietic cell expansion promotes vascular and systemic inflammation, as well as metabolic dysfunction in aging mice. These results support a causal role for TET2 mutation-driven clonal hematopoiesis in age-related cardiometabolic disease, and suggest that somatic TET2 mutations may contribute to atherosclerosis by promoting inflammation both systemically and locally in the atherosclerotic plaque.

Chronic Supplementation With a Mitochondrial Antioxidant (MitoQ) Improves Vascular Function in Healthy Older Adults.

URL: http://www.clinicaltrials.gov. Unique identifier: NCT02597023.

The role of adiponectin in exercise training‐induced vascular adaptation

We have previously reported that in old rats, reversal of age‐related vascular dysfunction by exercise training correlates with an increase in circulating adiponectin and its signaling within coronary vascular smooth muscle. In the current study, we investigated the effect of deletion of adiponectin on exercise training‐induced vascular adiponectin. C57BL/6 wild‐type (WT) or homozygous adiponectin knockout (AdipoKO) mice were obtained at 10–12 wks of age and underwent treadmill exercise training (EX) (12 m/min, 5° incline, 1 hr/day, 5 days/wk for 8 wks) or remained sedentary (SED) in cages. Arterioles isolated from cardiac and soleus muscle were assessed for contractile and vasodilatory function, and capillarity of the soleus muscle was evaluated. Exercise training increased flow‐induced dilation significantly in coronary arterioles of WT mice (P&lt;0.01 EX vs. SED), but decreased flow‐induced dilation in coronary arterioles from AdipoKO mice. ACh‐induced dilation was reduced in coronary arterioles from AdipoKO mice as compared to those from WT mice (P&lt;0.05 AdipoKO vs. WT). Exercise training reduced ACh‐induced dilation in arterioles from WT mice, but increased ACh‐induced dilation in arterioles from AdipoKO mice. Baseline capillarity increased in the soleus muscle of AdipoKO mice as compared to WT mice, but an exercise training‐induced increase in capillarity, detected in WT mice, was absent in AdipoKO mice. Contractile responsiveness to phenylephrine was increased in arterioles from the soleus muscle of both WT and AdipoKO mice (P&lt;0.01 EX vs. SED in both WT and AdipoKO). These data indicate that adiponectin is a critical contributor to exercise training‐induced vascular adaptations; however, locally produced adiponectin may be more critical than circulating adiponectin in mediating these adaptations.Support or Funding InformationFlorida State University College of MedicineThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.