Anticontractile Effect Of Perivascular Adipose Tissue Research Articles

Beclin-1-Dependent Autophagy Preserves Perivascular Adipose Tissue Function and Suppresses Hypertension Development in Hyperaldosteronism

Hyperaldosteronism (HA), characterized by the independent overproduction of aldosterone (Aldo), leads to cardiovascular damage and perivascular adipose tissue (PVAT) dysfunction. Beclin-1 (Becn1) is a key protein regulating autophagy, a recycling process of unwanted components. Whether HA disturbs autophagic flux in PVAT via Becn1 is unknown. Thus, we hypothesize that Becn1-dependent autophagic flux will improve PVAT function and inhibit the genesis of hypertension (HTN) in HA. 10-12-weeks-old male C57BL/6J (WT) or Becn1 knock-in mice (Becn1Tg) mice were infused with aldosterone for 14 days (600ug/Kg/day via osmotic mini-pump). Vascular function was studied in PVAT-intact thoracic aortae and blood pressure was analyzed by radiotelemetry. Becn1Tg mice were protected from HA-induced HTN [mean blood pressure (mmHg) − before HA: 101.2±0.5 vs. after HA: 108.2±0.9]. HA disrupted PVAT autophagic flux characterized by accumulation of LC3A/B and p62 and reduction in Becn1 expression and activity. In terms of function, presence of PVAT decreased the contractile response in WT [PVAT (-): 137.2±1.9 vs. PVAT (+): 107.4±2.7, Emax (% KCl)], while HA induced loss of PVAT anti-contractile effect [PVAT (-): 159.9±2.9 vs. PVAT (+): 164.1±3.4]. Becn1Tg mice were protected from this effect. Furthermore, HA induced PVAT oxidative stress and inflammation, characterized by elevated TNF-α, IL-6, IL-1β, IL-17, and IFN-γ mRNA expression, and elevated reactive oxygen species (ROS), measured by lucigenin chemiluminescence. All these effects were prevented in Becn1Tg mice. Pharmacological induction of Becn1 activation, via TB-peptide (16mg/Kg/day for 7 days), improved PVAT function of WT mice with HA. To evaluate the molecular mechanisms, we treated brown adipocyte cells (BAC, WT1) with Aldo (100nM), which triggered a transient autophagic flux, depicted by an initial increase in LC3A/B (12 and 24 h), followed by a decrease at 48 and 96 h. Such effects were prevented by a selective mineralocorticoid receptor antagonist (Eplerenone, 1μM). Finally, Aldo induced BAC inflammation and oxidative stress, which were blunted by TB-peptide. In summary, our findings reveal that autophagy overactivation confers protection against HA-induced PVAT inflammation and dysfunction and HTN. Therefore, activation of Becn1 could be pharmacological tool to prevent HA-associated cardiovascular deleterious effects. FAPESP (2022/06639-2), NHLBI-R00 (R00HL14013903), AHA (CDA857268 and 23DIVSUP1069230), Vascular Medicine Institute, the Hemophilia Center of Western Pennsylvania Vitalant, and Children's Hospital of Pittsburgh of the UPMC Health System. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Browning of perivascular adipose tissue prevents vascular dysfunction and reduces hypertension in angiotensin II-infused mice.

Hypertension is a world-leading cause of cardiovascular disease and premature deaths. Vascular tone is in part regulated by perivascular adipose tissue (PVAT) that releases pro and anticontractile factors. In hypertension, dysfunctional PVAT is observed and studies have indicated a causal relationship between dysfunctional PVAT and vascular damage in hypertension. The phenotype of PVAT on resistance vessels is primarily white adipose tissue. The present study investigates the impact of a changed phenotype, i.e., browning of PVAT, on vascular function and the development of hypertension. Browning was induced by β3-adrenergic agonist in control and angiotensin II-induced hypertensive mice. Studied parameters included blood pressure by tail-cuff plethysmography and vascular function by wire myography. Browning was confirmed through an immunohistochemical and gene analysis approach. The anticontractile effect of PVAT is lost in untreated hypertensive mice and vascular tone and blood pressure are increased. Browning of PVAT resulted in a maintained anticontractile effect, improved endothelial function, and reduced development of hypertension. Phenotype of PVAT is a major determinant of PVAT health during hypertensive conditions. Our data clearly demonstrates that browning of PVAT, i.e. changing the phenotype of PVAT, protects the vascular function and counteract the development of hypertension. This study provides novel insights into how PVAT can be protected in pathologies and thus limit the development of hypertension.

Autophagy induction protects the perivascular adipose tissue during hyperaldosteronism-induced hypertension

Hyperaldosteronism (HA), characterized by an autonomous production of aldosterone, promotes increased blood pressure and cardiovascular damage. The perivascular adipose tissue (PVAT) participates of the control of vascular function and is inflamed in HA. Autophagy is a recycling process of unnecessary or dysfunctional components and it is important for inflammatory tolerance, but whether the autophagic flux is dysregulated in the PVAT during HA, which would contribute to the inflammation and dysfunction of this tissue, is still to be elucidated. Herein, we hypothesize that treatment with autophagy inducer will rescue the autophagic unbalance, subsequently decreasing inflammation, and improving PVAT function during HA. 10-12-weeks-old male C57BL/6J mice were infused with aldosterone for 14 days (600ug/Kg/day via osmotic mini-pump). Vascular function was studied in PVAT-intact thoracic aortae and blood pressure was analyzed by the tail-cuff. Brown adipocyte cells (WT1) were used to evaluate the molecular mechanisms by which HA induces PVAT dysfunction. HA promoted an increase in systolic blood pressure [mmHg – before HA: 123.8±0.9 vs. after HA: 149.5±1.1]. Furthermore, PVAT decreased the contractile response [Emax (% KCl)] in control arteries [Control PVAT (-): 137.2±1.9 vs. Control PVAT (+): 107.4±2.7], whereas HA promoted loss of PVAT anti-contractile effect [HA PVAT (-): 159.9±2.9 vs. HA PVAT (+): 164.1±3.4]. We found that PVAT from HA mice presents an unbalance in LC3II/LC3I ratio, an important autophagy marker, inflammation, characterized by elevated TNF-α, IL-6, IL-1β, IL-17, and IFN-γ gene expression, and elevated reactive oxygen species (ROS). To induce autophagy, we treated mice with TB-peptide (16mg/Kg/day via ip for 7 days). TB-peptide partially improved the PVAT function of mice with HA [HA PVAT (+): 122.5±4.1]. On mechanistic levels, aldosterone (100nM, 12-96 h) induced an oscillatory autophagy flux in WT1 cells, characterized by an initial increase in LC3II/LC3I ratio at 12 and 24 h, followed by a decrease at 48 and 96 h. These effects were not observed in the presence of the mineralocorticoid receptor antagonist (Eplerenone, 1μM). Aldosterone also led to the expression of pro-inflammatory genes (TNF-α, IL-6, IL-1β, IL-17, and IFN-γ) and elevated ROS, such effects were prevented by inducing autophagy with TB-peptide. In summary, these findings reveal a new mechanism in vascular injury associated with HA, aldosterone impairs autophagic flux in PVAT, inducing inflammation, oxidative stress, and dysfunction. Our study places PVAT and autophagic process as attractive targets to reduce cardiovascular risk in HA. FAPESP (2022/06639-2), NHLBI-R00 (R00HL14013903), AHA-CDA (CDA857268), Vascular Medicine Institute, the Hemophilia Center of Western Pennsylvania Vitalant, and Children's Hospital of Pittsburgh of the UPMC Health System. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 14157: Effects of Follistatin on the Regulation of Vascular Function by Perivascular Adipose Tissue in a Model of Essential Hypertension

Introduction: We previously showed that follistatin (FST), a potent activin inhibitor, lowers BP in the spontaneously hypertensive rat (SHR), a model of essential HTN. This was associated with improved vessel structure (reduced collagen) and function (reduced hypercontractility and improved endothelium-dependent relaxation). FST also reduced vascular oxidative stress (ROS). Perivascular adipose tissue (PVAT) is known to regulate vascular tone. FST induces browning and thus increased thermogenic activity of peripheral adipose tissue. Transition from white to brown adipose tissue is associated with decreased risk of CVD. Here, we assess FST effects on PVAT and its regulation of SHR vascular function. Methods: SHR were treated with FST from 12-20 weeks of age. Normotensive WKY rats served as controls. Mesenteric resistance arteries were isolated with or without intact PVAT. Vessel function was assessed using wire myography. In separate studies, white mesenteric and brown aortic PVAT, or peripheral fat depots, were isolated from WKY rats. These were introduced to WKY mesenteric vessels to study PVAT-isolated function. Results: Intact PVAT augmented constriction to KCl in SHR, but not WKY arteries, an effect prevented by FST. Improvement in endothelium-dependent vasodilation in FST-treated SHR was PVAT-dependent. SHR PVAT had increased adipocyte size and ROS (by DHE), both reversed by FST. Isolated PVAT from WKY incubated with pyocyanin (30’) to induce ROS augmented the KCl response, suggesting that FST inhibition of PVAT oxidative stress in HTN restores beneficial vascular effects. Incubation of WKY vessels with brown PVAT inhibited constriction to KCl more effectively than did white PVAT. Interestingly, peripheral white adipose (inguinal) did not have any anticontractile effect. Brown adipose (interscapular) effects were intermediate. Conclusions: Improvement in anticontractile effects of PVAT by FST in resistance vessels is associated with reduced PVAT oxidative stress. Brown PVAT is more effective than white PVAT in inhibiting vessel contraction, and unlike white PVAT, peripheral white adipose does not have anticontractile effects. Future work will assess effects of FST on PVAT browning, and whether browning itself can improve BP.

Neutrophil Extracellular Traps Contribute to Perivascular Adipose Tissue Dysfunction in High‐Fat Diet Obese Mice

BackgroundObesity triggers functional changes in the perivascular adipose tissue (PVAT), favoring the release of vasoconstrictor factors and the activation of contractile mechanisms in vascular smooth muscle cells. In the PVAT of obese individuals, there is infiltration of immune cells, including neutrophils. A neutrophil defense mechanism is the formation of neutrophil extracellular traps (NETs), a cytotoxic framework that allows maintenance and expansion of inflammatory processes. We hypothesize that PVAT dysfunction in obese mice is due to increased neutrophil infiltration, NETs formation and inflammation.MethodsSix weeks‐old male Balb/C mice received control (CD) or high fat diet (HFD) for 18 weeks. Vascular function was assessed in endothelium‐intact mesenteric arteries in the presence or absence of PVAT. Mice were treated with DNAse type 1 (10 mg/kg for 14 days, every other day). Molecular mechanisms were investigated. Experiments were approved by the Ethics Committee on Animal Research of the Ribeirao Preto Medical School, University of Sao Paulo (protocol nº 209/2016)ResultsValues are expressed relatively to the contraction triggered by 120 mM KCl [Emax (% KCl)]. In Balb/C mice, the PVAT decreased the contractile response in control arteries [Control PVAT (‐): 123.2±8.1 n=6; Control PVAT (+): 64.4±4.0 n=6]. However, HFD promoted partial loss of PVAT anti‐contractile effect [HFD PVAT (‐): 127.9±6.9 n=6; HFD PVAT (+): 98.3±7.4 n=6]. In PVAT of obese mice there was a greater amount of neutrophils, compared to PVAT from control mice [Control: 8.0% n=5; HFD: 18.9% n=5]. In PVAT of obese mice there was greater formation of NETs, and production of interleukin 6 (IL‐6), compared to PVAT from control mice [NETs in pg/ml – Control: 30.5±2.7 n=6; HFD: 71.4±3.2 n=6; IL‐6 in pg/100 mg of PVAT – Control: 45.6±1.9 n=6; HFD: 98.4±3.1 n=6]. DNAse treatment reduced the formation of NETs and IL‐6 in the PVAT of obese mice [NETs in pg/ml – HFD+DNAse: 18.3±1.2 n=6; IL‐6 in pg/100 mg of PVAT – HFD: 48.1±3.4 n=6]. DNAse treatment restored PVAT anticontractile function in obese mice [HFD+DNAse PVAT (+): 71.3±7.4 n=6]. Adipose tissue of obese humans exhibited greater formation of NETs and production of IL‐6 compared to adipose tissue of eutrophic humans [NETs in pg/ml – Eutrophic: 20.5±3.7 n=7; Obese: 79.2±1.3 n=7; IL‐6 in pg/100 mg of adipose tissue – Eutrophic: 9.7±2.9 n=7; Obese: 27.7±2.6 n=7].ConclusionThese results indicate that, in obesity, there is a greater neutrophils infiltration in the PVAT and, consequently, greater formation of NETs and inflammation, events that contribute to PVAT dysfunction.

Perivascular Adipose Tissue Anticontractile Function Is Mediated by Both Endothelial and Neuronal Nitric Oxide Synthase Isoforms.

The mechanism of the perivascular adipose tissue (PVAT) anticontractile effect is well characterized in rodent visceral vascular beds; however, little is known about the mechanism of PVAT anticontractile function in subcutaneous vessels. In addition, we have previously shown that PVAT anticontractile function is nitric oxide synthase (NOS) dependent but have not investigated the roles of NOS isoforms. Here, we examined PVAT anticontractile function in the mouse gracilis artery, a subcutaneous fat depot, in lean control and obese mice and investigated the mechanism in comparison to a visceral depot. Using the wire myograph, we generated responses to noradrenaline and electrical field stimulation in the presence of pharmacological tools targeting components of the known PVAT anticontractile mechanism. In addition, we performed ex vivo "fat transplants" in the organ bath. The mechanism of PVAT anticontractile function is similar between subcutaneous and visceral PVAT depots. Both endothelial and neuronal NOS isoforms mediated the PVAT anticontractile effect. Loss of PVAT anticontractile function in obesity is independent of impaired vasoreactivity, and function can be restored in visceral PVAT by NOS activation. Targeting NOS isoforms may be useful in restoring PVAT anticontractile function in obesity, ameliorating increased vascular tone, and disease.

NO, ROS, RAS, and PVAT: More Than a Soup of Letters.

Perivascular adipose tissue (PVAT) has recently entered in the realm of cardiovascular diseases as a putative target for intervention. Notwithstanding its relevance, there is still a long way before the role of PVAT in physiology and pathology is fully understood. The general idea that PVAT anti-contractile effect is beneficial and its pro-contractile effect is harmful is being questioned by several reports. The role of some PVAT important products or systems such as nitric oxide (NO), reactive oxygen species (ROS), and RAS may vary depending on the context, disease, place of production, etc., which adds doubts on how mediators of PVAT anti- and pro-contractile effects are called to action and their final result. This short review will address some points regarding NO, ROS, and RAS in the beneficial and harmful roles of PVAT.

Dysfunction of perivascular adipose tissue in mesenteric artery is restored by aerobic exercise in high‐fat diet induced obesity

This study investigated the function of perivascular adipose tissue (PVAT) on vascular contractility within resistant arteries in high-fat diet induced obese rats after long-term aerobic exercise. Male Sprague-Dawley rats were subjected to normal diet control group (N-CTRL), normal diet exercise group (N-EX), high-fat diet control group (H-CTRL), and high-fat diet exercise group (H-EX) (n=8 in each group). After intervention, adipose tissues morphology was observed. Vasomotor function of mesenteric arteries with or without PVAT were assessed; mesenteric PVAT isolated from each group were transferred to chambers bath with untreated vessels (without PVAT) to evaluate the independent effect. Isolated PVAT was further pre-treated with inhibitor of cystathionine-γ-lyase (CSE), a key hydrogen sulphide (H2 S) enzyme. Results showed that the size of lipid droplet around mesenteric arteries from H-EX was significantly reduced (P<.05); uncoupling protein1 (UCP1) in PVAT from H-EX was enhanced. In N-CTRL, N-EX, and H-EX, vessels without PVAT showed higher sensitivity to serotonin (5-HT) than that with intact PVAT. Vascular tension by 5-HT was significantly reduced in H-EX than H-CTRL (P<.05) in vessels with PVAT. Transferred PVAT from H-EX compared with H-CTRL significantly reduced vascular sensitivity to 5-HT (P<.05), and this effect was eliminated through inhibiting CSE. In summary, the anti-contractile effect of PVAT on resistance artery was impaired in obesity but restored by long-term aerobic exercise. The function of PVAT modified by obesity or by exercise has an independent influence on vascular reactivity, and PVAT derived H2 S may participate in this process.

Anticontractile Effect of Perivascular Adipose Tissue But Not of Endothelium Is Enhanced by Hydrogen Sulfide Stimulation in Hypertensive Pregnant Rat Aortae.

Perivascular adipose tissue (PVAT) modulates the vascular tone. Hydrogen sulfide (H2S) is synthetized by cystathionine gamma-lyase (CSE) in brown PVAT. Modulation of vascular contractility by H2S is, in part, adenosine triphosphate (ATP)-sensitive potassium channels dependent. However, the role of PVAT-derived H2S in hypertensive pregnancy (HTN-Preg) is unclear. Therefore, we aimed to examine the involvement of H2S in the anticontractile effect of PVAT in aortae from normotensive and hypertensive pregnant rats. To this end, phenylephrine-induced contractions in the presence and absence of PVAT and endothelium in aortae from normotensive pregnant (Norm-Preg) and HTN-Preg rats were investigated. Maternal blood pressure, fetal-placental parameters, angiogenesis-related biomarkers, and H2S levels were also assessed. We found that circulating H2S is elevated in hypertensive pregnancy associated with angiogenic imbalance, fetal and placental growth restrictions, which revealed that there is H2S pathway activation. Moreover, under stimulated H2S formation PVAT, but not endothelium, reduced phenylephrine-induced contractions in aortae from HTN-Preg rats. Also, H2S synthesis inhibitor abolished anticontractile effects of PVAT and endothelium. Furthermore, anticontractile effect of PVAT, but not of endothelium, was eliminated by ATP-sensitive potassium channels blocker. In accordance, increases in H2S levels in PVAT and placenta, but not in aortae without PVAT, were also observed. In conclusion, anticontractile effect of PVAT is lost, at least in part, in HTN-Preg aortae and PVAT effect is ATP-sensitive potassium channels dependent in normotensive and hypertensive pregnant rat aortae. PVAT but not endothelium is responsive to the H2S stimulation in hypertensive pregnant rat aortae, implying a key role for PVAT-derived H2S under endothelial dysfunction.

MR (Mineralocorticoid Receptor) Induces Adipose Tissue Senescence and Mitochondrial Dysfunction Leading to Vascular Dysfunction in Obesity.

Adipose tissue (AT) senescence and mitochondrial dysfunction are associated with obesity. Studies in obese patients and animals demonstrate that the MR (mineralocorticoid receptor) contributes to obesity-associated cardiovascular complications through its specific role in AT. However, underlying mechanisms remain unclear. This study aims to elucidate whether MR regulates mitochondrial function in obesity, resulting in AT premature aging and vascular dysfunction. Obese (db/db) and lean (db/+) mice were treated with an MR antagonist or a specific mitochondria-targeted antioxidant. Mitochondrial and vascular functions were determined by respirometry and myography, respectively. Molecular mechanisms were probed by Western immunoblotting and real-time polymerase chain reaction in visceral AT and arteries and focused on senescence markers and redox-sensitive pathways. db/db mice displayed AT senescence with activation of the p53-p21 pathway and decreased SIRT (sirtuin) levels, as well as mitochondrial dysfunction. Furthermore, the beneficial anticontractile effects of perivascular AT were lost in db/db via ROCK (Rho kinase) activation. MR blockade prevented these effects. Thus, MR activation in obesity induces mitochondrial dysfunction and AT senescence and dysfunction, which consequently increases vascular contractility. In conclusion, our study identifies novel mechanistic insights involving MR, adipose mitochondria, and vascular function that may be of importance to develop new therapeutic strategies to limit obesity-associated cardiovascular complications.

Effect of Peroxynitrite (ONOO&amp;lt;sup&amp;gt;-&amp;lt;/sup&amp;gt;) on the Function of Murine Perivascular Adipose Tissue

Perivascular adipose tissue (PVAT) surrounds the exterior of blood vessels and releases numerous substances such as adiponectin which positively modulate blood vessel tone. In some cardiovascular diseases such as diabetes and high blood pressure, the function of PVAT changes and we speculate that oxidant stress may play a role in this change. PVAT has the ability to generate both superoxide and nitric oxide and these can combine rapidly under physiological conditions to form peroxynitrite (ONOO-). In disease states, the production of ONOO- may be increased and so its effect on the function of PVAT is of great interest. Consequently, we studied the effects of acute addition of the oxidant species ONOO- on vascular tone and production of adiponectin by mouse thoracic aortic PVAT. Murine PVAT was immunostained for nitrotyrosine, indicating that ONOO- is formed in the PVAT. Exogenous ONOO- significantly increased the anticontractile effect of PVAT via increased adiponectin content but had no effect on eNOS expression or phosphorylation. These results suggest that generation of ONOO- within PVAT may be an important regulatory mechanism which influences the activity of PVAT. The effect of chronic exposure to raised levels of ONOO- on PVAT function remains to be determined.

Maternal separation enhances anticontractile perivascular adipose tissue function in male rats on a high-fat diet.

Clinical studies have shown that obesity negatively impacts large arteries' function. We reported that rats exposed to maternal separation (MatSep), a model of early life stress, display enhanced angiotensin II (ANG II)-induced vasoconstriction in aortic rings cleaned of perivascular adipose tissue (PVAT) under normal diet (ND) conditions. We hypothesized that exposure to MatSep promotes a greater loss of PVAT-mediated protective effects on vascular function and loss of blood pressure (BP) rhythm in rats fed a high-fat diet (HFD) when compared with controls. MatSep was performed in male Wistar-Kyoto rats from days 2 to 14 of life. Normally reared littermates served as controls. On ND, aortic rings from MatSep rats with PVAT removed showed increased ANG II-mediated vasoconstriction versus controls; however, rings from MatSep rats with intact PVAT displayed blunted constriction. This effect was exacerbated by an HFD in both groups; however, the anticontractile effect of PVAT was greater in MatSep rats. Acetylcholine-induced relaxation was similar in MatSep and control rats fed an ND, regardless of the presence of PVAT. HFD impaired aortic relaxation in rings without PVAT from MatSep rats, whereas the presence of PVAT improved relaxation in both groups. On an HFD, immunolocalization of vascular smooth muscle-derived ANG-(1-7) and PVAT-derived adiponectin abundances were increased in MatSep. In rats fed an HFD, 24-h BP and BP rhythms were similar between groups. In summary, MatSep enhanced the ability of PVAT to blunt the heightened ANG II-induced vasoconstriction and endothelial dysfunction in rats fed an HFD. This protective effect may be mediated via the upregulation of vasoprotective factors within the adipovascular axis.

BETA3-ADRENOCEPTOR MEDIATED ADIPONECTIN SECRETION IS ESSENTIAL IN MODULATION OF VASCULAR TONE BY PERIVASCULAR ADIPOSE TISSUE

Objective: Healthy perivascular adipose tissue (PVAT) exerts an anti-contractile effect which is vital in regulating blood pressure. Evidence suggests that sympathetic nervous stimulation of PVAT triggers activation of adipocyte beta3-adrenoceptors, and the subsequent release of the vasoactive adipokine adiponectin. In obesity plasma adiponectin is reduced, which may result in a loss of PVAT function and subsequent hypertension. Therefore we examined beta3-adrenoceptor and adiponectin function in the PVAT anti-contractile effect. Design and method: Body weight, blood pressure, and blood glucose were measured in control C57 mice, and adiponectin knockout (Adipo-/-) mice. Following sacrifice, electrical field stimulation (EFS) profiles of isolated C57 and Adipo-/- mesenteric and skeletal muscle resistance arteries (<250 μm, ± PVAT) were characterised using wire myography. Sympathetic denervation of PVAT using 6-hydroxydopamine (6-OHDA) was performed. Beta3-adrenoceptor function was investigated using the agonist CL-316,243 and antagonist SR59203A. The role of the adipokine adiponectin was examined using exogenous adiponectin and a blocking peptide for adiponectin receptor 1 (ABP). The concentration of adiponectin secretion upon EFS was measured using an ELISA kit. Results: Adipo-/- mice exhibit significant elevations in both blood pressure and blood glucose. During EFS of isolated ± PVAT arteries, mesenteric and skeletal muscle PVAT from C57 mice elicited an anti-contractile effect, which was absent in the Adipo-/-. Sympathetic denervation of both mesenteric and skeletal muscle PVAT abolished the anti-contractile effect. Inhibition of beta3-adrenoceptors in C57 mesenteric and skeletal muscle PVAT using SR59230A significantly reduced the anti-contractile effect, however activation of beta3-adrenoceptors in Adipo-/- PVAT using CL-316,243 did not restore function. Application of exogenous adiponectin to C57 mesenteric and skeletal muscle –PVAT arteries caused a significant vasodilation. In ± PVAT arteries, incubation with ABP significantly reduced the anti-contractile effect. Using an ELISA, adiponectin secretion from PVAT upon EFS was significantly reduced when incubated with SR59230A. Conclusions: These results demonstrate that upon sympathetic stimulation, PVAT from two different vascular beds releases adiponectin via activation of beta3-adrenoreceptors, exerting an anti-contractile effect on the blood vessels. In the absence of adiponectin PVAT anti-contractile function is lost, resulting in hypertension and hyperglycaemia, highlighting the importance of adiponectin in vascular function.

Palmitic Acid Methyl Ester and Its Relation to Control of Tone of Human Visceral Arteries and Rat Aortas by Perivascular Adipose Tissue.

Background: Perivascular adipose tissue (PVAT) exerts anti-contractile effects on visceral arteries by release of various perivascular relaxing factors (PVRFs) and opening voltage-gated K+ (Kv) channels in vascular smooth muscle cells (VSMCs). Palmitic acid methyl ester (PAME) has been proposed as transferable PVRF in rat aorta. Here, we studied PVAT regulation of arterial tone of human mesenteric arteries and clarified the contribution of Kv channels and PAME in the effects.Methods: Wire myography was used to measure vasocontractions of mesenteric artery rings from patients undergoing abdominal surgery. Isolated aortic rings from Sprague-Dawley rats were studied for comparison. PVAT was either left intact or removed from the arterial rings. Vasocontractions were induced by external high K+ (60 mM), serotonin (5-HT) or phenylephrine. PAME (10 nM−3 μM) was used as vasodilator. Kv channels were blocked by XE991, a Kv7 (KCNQ) channel inhibitor, or by 4-aminopyridine, a non-specific Kv channel inhibitor. PAME was measured in bathing solutions incubated with rat peri-aortic or human visceral adipose tissue.Results: We found that PVAT displayed anti-contractile effects in both human mesenteric arteries and rat aortas. The anti-contractile effects were inhibited by XE991 (30 μM). PAME (EC50 ~1.4 μM) was capable to produce relaxations of PVAT-removed rat aortas. These effects were abolished by XE991 (30 μM), but not 4-aminopyridine (2 mM) or NDGA (10 μM), a lipoxygenases inhibitor. The cytochrome P450 epoxygenase inhibitor 17-octadecynoic acid (ODYA 10 μM) and the soluble epoxide hydrolase inhibitor 12-(3-adamantan-1-ylureido)-dodecanoic acid (AUDA 10 μM) slightly decreased PAME relaxations. PAME up to 10 μM failed to induce relaxations of PVAT-removed human mesenteric arteries. 5-HT induced endogenous PAME release from rat peri-aortic adipose tissue, but not from human visceral adipose tissue.Conclusions: Our data also suggest that Kv7 channels are involved in the anti-contractile effects of PVAT on arterial tone in both rat aorta and human mesenteric arteries. PAME could contribute to PVAT relaxations by activating Kv7 channels in rat aorta, but not in human mesenteric arteries.

Modulation of Vascular Reactivity by Perivascular Adipose Tissue (PVAT).

In this review, we discuss the role of perivascular adipose tissue (PVAT) in the modulation of vascular contractility and arterial pressure, focusing on the role of the renin-angiotensin-aldosterone system and oxidative stress/inflammation. PVAT possesses a relevant endocrine-paracrine activity, which may be altered in several pathophysiological and clinical conditions. During the last two decades, it has been shown that PVAT may modulate vascular reactivity. It has also been previously demonstrated that inflammation in adipose tissue may be implicated in vascular dysfunction. In particular, adipocytes secrete a number of adipokines with various functions, as well as several vasoactive factors, together with components of the renin-angiotensin system which may act at local or at systemic level. It has been shown that the anti-contractile effect of PVAT is lost in obesity, probably as a consequence of the development of adipocyte hypertrophy, inflammation, and oxidative stress. Adipose tissue dysfunction is interrelated with inflammation and oxidative stress, thus contributing to endothelial dysfunction observed in several pathological and clinical conditions such as obesity and hypertension. Decreased local adiponectin level, macrophage recruitment and infiltration, and activation of renin-angiotensin-aldosterone system could play an important role in this regard.

P-386 - Contribution of perivascular adipose tissue to vascular dysfunction in type 2 diabetes

Perivascular adipose tissue (PVAT) surrounds most large blood vessels and plays an important role in vascular homeostasis. The present study was conducted to investigate the contribution of PVATto vascular dysfunction in a rat model of type 2 diabetes (GK) fed with a high-fat diet (HFD). Aortic vascular reactivity was performed in GK, GK fed with HFD and compared with Wistar rats in the absence (PVAT−) or in the presence (PVAT+) of thoracic PVAT. We also measured vascular oxidative stress Endothelium-dependent relaxation to acetylcholine, assessed by wire myography, was impaired in GK and GKHFD rats, not affected by NG-nitro-L-arginine methyl ester, inhibitor of endothelial nitric oxide (NO) synthase, and improved by the antioxidant tempol, suggesting reduced NO bioavailability and increased oxidative stress. Furthermore, vascular superoxide and peroxynitrite production was increased in the vascular wall of diabetic rats. The presence of PVAThad an anti-contractile effect in response to phenylephrine in Wistar rats. In GK rats, the anti-contractile effects of PVAT were lost. Our data suggest that this rat model of type 2 diabetes is associated with perivascular adipose dysfunction, oxidative stress and endothelial dysfunction.

Increased O-GlcNAcylation of Endothelial Nitric Oxide Synthase Compromises the Anti-contractile Properties of Perivascular Adipose Tissue in Metabolic Syndrome.

Under physiological conditions, the perivascular adipose tissue (PVAT) negatively modulates vascular contractility. This property is lost in experimental and human obesity and in the metabolic syndrome, indicating that changes in PVAT function may contribute to vascular dysfunction associated with increased body weight and hyperglycemia. The O-linked β-N-acetylglucosamine (O-GlcNAc) modification of proteins (O-GlcNAcylation) is a unique posttranslational process that integrates glucose metabolism with intracellular protein activity. Increased flux of glucose through the hexosamine biosynthetic pathway and the consequent increase in tissue-specific O-GlcNAc modification of proteins have been linked to multiple facets of vascular dysfunction in diabetes and other pathological conditions. We hypothesized that chronic consumption of glucose, a condition that progresses to metabolic syndrome, leads to increased O-GlcNAc modification of proteins in the PVAT, decreasing its anti-contractile effects. Therefore, the current study was devised to determine whether a high-sugar diet increases O-GlcNAcylation in the PVAT and how increased O-GlcNAc interferes with PVAT vasorelaxant function. To assess molecular mechanisms by which O-GlcNAc contributes to PVAT dysfunction, thoracic aortas surrounded by PVAT were isolated from Wistar rats fed either a control or high sugar diet, for 10 and 12 weeks. Rats chronically fed a high sugar diet exhibited metabolic syndrome features, increased O-GlcNAcylated-proteins in the PVAT and loss of PVAT anti-contractile effect. PVAT from high sugar diet-fed rats for 12 weeks exhibited decreased NO formation, reduced expression of endothelial nitric oxide synthase (eNOS) and increased O-GlcNAcylation of eNOS. High sugar diet also decreased OGA activity and increased superoxide anion generation in the PVAT. Visceral adipose tissue samples from hyperglycemic patients showed increased levels of O-GlcNAc-modified proteins, increased ROS generation and decreased OGA activity. These data indicate that O-GlcNAcylation contributes to metabolic syndrome-induced PVAT dysfunction and that O-GlcNAcylation of eNOS may be targeted in the development of novel therapies for vascular dysfunction in conditions associated with hyperglycemia.

Increased mitochondrial ROS generation mediates the loss of the anti-contractile effects of perivascular adipose tissue in high-fat diet obese mice.

Obesity is associated with structural and functional changes in perivascular adipose tissue (PVAT), favouring release of reactive oxygen species (ROS), vasoconstrictor and proinflammatory factors. The cytokine TNF-α induces vascular dysfunction and is produced by PVAT. We tested the hypothesis that obesity-associated PVAT dysfunction was mediated by augmented mitochondrial ROS (mROS) generation due to increased TNF-α production in this tissue. C57Bl/6J and TNF-α receptor-deficient mice received control or high fat diet (HFD) for 18 weeks. We used pharmacological tools to determine the participation of mROS in PVAT dysfunction. Superoxide anion (O2.- ) and H2 O2 were assayed in PVAT and aortic rings were used to assess vascular function. Aortae from HFD-fed obese mice displayed increased contractions to phenylephrine and loss of PVAT anti-contractile effect. Inactivation of O2.- , dismutation of mitochondria-derived H2 O2 , uncoupling of oxidative phosphorylation and Rho kinase inhibition, decreased phenylephrine-induced contractions in aortae with PVAT from HFD-fed mice. O2.- and H2 O2 were increased in PVAT from HFD-fed mice. Mitochondrial respiration analysis revealed decreased O2 consumption rates in PVAT from HFD-fed mice. TNF-α inhibition reduced H2 O2 levels in PVAT from HFD-fed mice. PVAT dysfunction, i.e. increased contraction to phenylephrine in PVAT-intact aortae, was not observed in HFD-obese mice lacking TNF-α receptors. Generation of H2 O2 was prevented in PVAT from TNF-α receptor deficient obese mice. TNF-α-induced mitochondrial oxidative stress is a key and novel mechanism involved in obesity-associated PVAT dysfunction. These findings elucidate molecular mechanisms whereby oxidative stress in PVAT could affect vascular function. This article is part of a themed section on Molecular Mechanisms Regulating Perivascular Adipose Tissue - Potential Pharmacological Targets? To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.20/issuetoc.

The role of perivascular adipose tissue in obesity-induced vascular dysfunction.

Under physiological conditions, perivascular adipose tissue (PVAT) attenuates agonist‐induced vasoconstriction by releasing vasoactive molecules including hydrogen peroxide, angiotensin 1–7, adiponectin, methyl palmitate, hydrogen sulfide, NO and leptin. This anticontractile effect of PVAT is lost under conditions of obesity. The central mechanism underlying this PVAT dysfunction in obesity is likely to be an ‘obesity triad’ (consisting of PVAT hypoxia, inflammation and oxidative stress) that leads to the impairment of PVAT‐derived vasoregulators. The production of hydrogen sulfide, NO and adiponectin by PVAT is reduced in obesity, whereas the vasodilator response to leptin is impaired (vascular leptin resistance). Strikingly, the vasodilator response to acetylcholine is reduced only in PVAT‐containing, but not in PVAT‐free thoracic aorta isolated from diet‐induced obese mice, indicating a unique role for PVAT in obesity‐induced vascular dysfunction. Furthermore, PVAT dysfunction has also been observed in small arteries isolated from the gluteal/visceral fat biopsy samples of obese individuals. Therefore, PVAT may represent a new therapeutic target for vascular complications in obesity. A number of approaches are currently being tested under experimental conditions. Potential therapeutic strategies improving PVAT function include body weight reduction, enhancing PVAT hydrogen sulfide release (e.g. rosiglitazone, atorvastatin and cannabinoid CB1 receptor agonists) and NO production (e.g. arginase inhibitors), inhibition of the renin–angiotensin–aldosterone system, inhibition of inflammation with melatonin or cytokine antagonists, activators of AMP‐activated kinase (e.g. metformin, resveratrol and diosgenin) and adiponectin releasers or expression enhancers.Linked ArticlesThis article is part of a themed section on Molecular Mechanisms Regulating Perivascular Adipose Tissue – Potential Pharmacological Targets? To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.20/issuetoc

Interaction of perivascular adipose tissue and sympathetic nerves in arteries from normotensive and hypertensive rats.

The inhibitory action of perivascular adipose tissue (PVAT) in modulation of arterial contraction has been recently recognized and contrasted with the prohypertensive effect of obesity in humans. In this study we demonstrated that PVAT might have opposing effect on sympatho-adrenergic contractions in different rat conduit arteries. In superior mesenteric artery isolated from normotensive Wistar-Kyoto rats (WKY), PVAT exhibited inhibitory influence on the contractions to exogenous noradrenaline as well as to endogenous noradrenaline released from arterial sympathetic nerves during transmural electrical stimulation or after application of tyramine. In contrast, the abdominal aorta with intact PVAT responded with larger contractions to transmural electrical stimulation and tyramine when compared to the aorta after removing PVAT; the responses to noradrenaline were similar in both. This indicates that PVAT may contain additional sources of endogenous noradrenaline which could be responsible for the main difference in the modulatory effect of PVAT on adrenergic contractions between abdominal aortas and superior mesenteric arteries. In spontaneously hypertensive rats (SHR), the anticontractile effect of PVAT in mesenteric arteries was reduced, and the removal of PVAT completely eliminated the difference in the dose-response curves to exogenous noradrenaline between SHR and WKY. These results suggest that in mesenteric artery isolated from SHR, the impaired anticontractile influence of PVAT might significantly contribute to its increased sensitivity to adrenergic stimuli.