Perivascular Adipose Tissue Function Research Articles

Single-cell view and a novel protective macrophage subset in perivascular adipose tissue in T2DM

BackgroundVasculopathy underlies diabetic complications, with perivascular adipose tissue (PVAT) playing crucial roles in its development. However, the changes in the cellular composition and function of PVAT, including the specific cell subsets and mechanisms implicated in type 2 diabetes mellitus (T2DM) vasculopathy, remain unclear.MethodsTo address the above issues, we performed single-cell RNA sequencing on the stromal vascular fraction (SVF) of PVAT from normal and T2DM rats. Then, various bioinformatics tools and functional experiments were used to investigate the characteristic changes in the cellular profile of diabetic PVAT SVF, their implications, and the underlying mechanisms.ResultsOur study reveals the single-cell landscape of the SVF of PVAT, demonstrating its considerable heterogeneity and significant alterations in T2DM, including an enhanced inflammatory response and elevated proportions of macrophages and natural killer (NK) cells. Moreover, macrophages are critical hubs for cross-talk among various cell populations. Notably, we identified a decreased Pdpn+ macrophage subpopulation in the PVAT of T2DM rats and confirmed this in mice and humans. In vitro and in vivo studies demonstrated that Pdpn+ macrophages alleviated insulin resistance and modulated adipokine/cytokine expression in adipocytes via the Pla2g2d-DHA/EPA-GPR120 pathway. This subset also enhances the function of vascular endothelial and smooth muscle cells, inhibits vascular inflammation and oxidative stress, and improves vasodilatory function, thereby protecting blood vessels.ConclusionPdpn+ macrophages exhibit significant vascular protective effects by alleviating insulin resistance and modulating adipokine/cytokine expression in PVAT adipocytes. This macrophage subtype may therefore play pivotal roles in mitigating vascular complications in T2DM. Our findings also underscore the critical role of immune-metabolic cross-talk in maintaining tissue homeostasis.Graphical

Phenotypic and functional disparities in perivascular adipose tissue.

The adipose tissue surrounding blood vessels is known as perivascular adipose tissue (PVAT), which represents a distinct ectopic fat depot that adheres to the majority of the vasculature. In recent years, owing to its unique location and function, PVAT has been regarded as a new type of adipose tissue distinct from traditional visceral fat. It releases adipokines with vasoconstrictive functions, which regulate vascular function through paracrine and endocrine mechanisms. Interestingly, PVAT can be categorized as white, brown or a mixture of both depending on its anatomical location. Brown adipose tissue (BAT) is located adjacent to the thoracic aorta in rodents, while a mix of brown and white tissue surrounds the abdominal aorta. PVAT exhibits regional phenotypic differences in different parts of the vasculature bed, which may lead to heterogeneity in the secretion profiles and norepinephrine (NE) content in regional PVAT and subsequently affect the regulation of specific adipokine signaling pathways in regional PVAT, resulting in differences in the regulation of vascular function. The aim of this review was to explore the potential factors that influence the anticontractile function of regional PVAT in the vasculature, including the heterogeneity of regional PVAT, the anticontractile function mediated by endothelial nitric oxide synthase (eNOS) in regional PVAT, the activity of the adiponectin-eNOS pathway in regional PVAT adipocytes, and the concentration of the sympathetic neurotransmitter NE in regional PVAT.

Apelin from Perivascular Adipose Tissue Is Involved in the Regulation of Vasorelaxation and Renal Function in Metabolic Syndrome

The perivascular adipose tissue (PVAT) regulates the arterial tone by releasing vasoactive molecules. PVAT dysfunction favoring the vasorelaxation response could contribute to the development of kidney disease in metabolic syndrome (MetS). Previously, we demonstrated that overactivation of angiotensin II signaling in the PVAT deteriorates the compensatory PVAT effects in rats with MetS (SHRSP.Z-Leprfa/IzmDmcr (SPZF) and SHR/NDmcr-cp (CP) rats). Apelin is an endogenous regulator of angiotensin II. Therefore, we investigated whether changes in apelin levels in the PVAT alter PVAT function and impair kidney function in MetS. Twenty-three-week-old male and female SPZF and CP rats were used. In the female CP rats, apelin mRNA levels in renal arterial PVAT, enhancing effects of the PVAT on acetylcholine-induced relaxation in renal arteries, and estimated glomerular filtration rate (eGFR) were the highest, and urine protein levels and homeostasis model assessment of insulin resistance (HOMA-IR) were the lowest. Apelin mRNA levels were positively correlated with the enhancing effects of the PVAT on vasorelaxation and eGFR but negatively correlated with urine protein levels and HOMA-IR. Moreover, apelin levels positively correlated with mRNA levels of angiotensin-converting enzyme 2 and angiotensin II type 1 receptor-associated protein, which are negative regulators of angiotensin II. This study suggests that a decline in apelin levels in the PVAT, probably owing to angiotensin II, is associated with PVAT dysfunction on vascular tone, resulting in impaired kidney function in MetS.

Impact of the local renin–angiotensin system in perivascular adipose tissue on vascular health and disease

Perivascular adipose tissue (PVAT) is found locally around blood vessels. It has the ability to release vasoactive chemicals, such as factors that relax and contract blood vessels. PVAT is now recognized as an endocrine organ with metabolic activity and as a “protagonist” for maintaining vascular homeostasis. Angiotensin II, a powerful vasoconstrictor of the renin–angiotensin system (RAS) that can increase blood pressure and vascular tone, is produced locally by PVAT. To mitigate the multiple vascular effects of angiotensin II, PVAT also generates molecules such as angiotensin (1–7), adiponectin, and nitric oxide. Reactive oxygen species and proinflammatory cytokines are produced in greater amounts when PVAT-mediated angiotensin II is present, resulting in endothelial dysfunction, inflammation, atherosclerosis, and other vascular disorders. The anticontractile and procontractile nature of PVAT is frequently disrupted in obese individuals, which increases the production of angiotensin II and decreases the production of anti-inflammatory and vasodilatory factors. These changes in turn exacerbate vascular inflammation, hypertension, and atherosclerosis. PVAT, which is crucial for maintaining vascular homeostasis, loses its anticontractile effect in obesity due to adipocyte hypertrophy, inflammation, and oxidative stress, exacerbating endothelial dysfunction. Overactive RAS in PVAT facilitates the migration and proliferation of vascular smooth muscle cells and atherosclerotic plaques, both of which accelerate the development of atherosclerosis. Targeting PVAT and the local RAS can offer therapeutic benefits in treating hypertension, atherosclerosis, and other vascular diseases. This review highlights the scientific underpinnings of the function of PVAT in regulating the autocrine and paracrine activities of vascular RAS constituents.

Evaluation of the effects of pioglitazone on perivascular adipose tissue function, properties, and structure in a rat model of type-2 diabetes.

Perivascular adipose tissue (PVAT) plays an important role in many physiological and pathological processes, such as regulation of vascular tone. The aim of this study is to evaluate the effects of pioglitazone on functional, structural, and biochemical properties of PVAT in an experimental model of type-2 diabetes (T2DM). T2DM was induced by high-fat-diet/low-dose-streptozotocin in rats, and pioglitazone (20mg/kg/p.o.) was administered for 6 weeks. Changes in biochemical parameters, PVAT-mass, vascular-reactivity in thoracic-aorta, as well as PVAT adipocytokine and PPARG-expression levels, and histopathology were evaluated. Pioglitazone administration improved blood glucose and lipid profiles in T2DM. Pioglitazone did not change the anticontractile effect of PVAT on aortic contractile reactivity and besides, had no influence on endothelium-dependent and -independent relaxation responses. Pioglitazone administration increased PVAT-mass and tumor necrotizing factor-α levels, while adiponectin, leptin, and interleukin-6 levels were unchanged. Also, a prominent increase was observed in PPARG-expression in T2DM-Pio group. Moreover, pioglitazone decreased liver steatosis, aortic wall thickening, and myocardial damage, whereas increased adipocyte size and adiposity in PVAT. Overall, pioglitazone treatment changed the mass and in part the inflammatory profile of PVAT but did not modify vasoreactivity in T2DM. This study provides novel findings in relationship with the adipogenic effect of pioglitazone and PVAT function.

Abstract P455: Integrins play a role in stress relaxation of perivascular adipose tissue

Perivascular adipose tissue (PVAT) is well known for being anti-contractile (reducing potency, maximum) in healthy tissues. We discovered a new function of PVAT, the ability to stress relax in response to a stretch. This is of note because stress relaxation has been attributed largely to smooth muscle, of which PVAT has none that is organized in a functional layer. Here we test the hypothesis that stress relaxation depends on the interactions of the cell membrane delimited heterodimer integrins with collagen. Our model is the thoracic aorta of the male Dahl SS rat on a normal diet. The PVAT and aorta were physically separated for most assays. Results from snRNA seq in PVAT, histochemistry and isometric contractility were used. Collagen was evident in PVAT as stained by Masson Trichrome. snRNA seq of whole PVAT determined expression of many collagen isoforms and integrins, with mRNA for both the α1 and β1 integrin being most highly expressed. Isolated aortic rings and separated PVAT were challenged with 2 or 4 grams of passive tension. Pharmacological inhibition of integrin/collagen interaction was effected by the specific α1β1 distintegrin obtustatin (5 uM) or general inhibitor RGD peptide (1 mM). RGD peptide but not obtustatin reduced the ability of the isolated PVAT ring to maintain applied tone at both the 2 and 4 gram challenge (~15% gain in relaxation/loss of tone, figure ). By contrast, stress relaxation in the aorta itself was not reduced by either inhibitor. In addition to a1b1 and collagen interaction, cell-cell communication inference identified integrins av and a5, two major RGD motif containing isoforms, as potential partners of collagens with fibroblast-like cells most likely involved. Collectively, these findings validate that stress relaxation can occur in a non-smooth muscle tissue, doing so at least in part through integrin-collagen interactions that may not include α1β1 heterodimers. The importance of this lies in considering PVAT as a vascular layer that possesses mechanical functions.

Abstract P449: A Murine Model of Mid-Thoracic Aortic Coarctation

Vascular pressure can directly affect cardiovascular structure and function. Understanding the mechanisms, however, is complicated because conditions or interventions that chronically raise blood pressure (e.g. excess angiotensin II) can also have direct effects on the vasculature that occur independent of changes in blood pressure. One approach to mitigating this problem is to obstruct flow in a vascular segment sufficiently to reduce blood pressure in the segment downstream of the obstruction, while pressure upstream either stays the same or increases. As a result, all factors other than pressure are expected to be the same in upstream and downstream vascular segments. We have found evidence that differences in intravascular pressure can also affect closely adjacent tissues, specifically perivascular adipose tissue (PVAT). To test this idea in vivo , we developed a method to produce chronic, partial occlusion (or coarctation) of the mid-thoracic aorta in mice. This exposes upstream aortic PVAT depots to significantly higher aortic pressure than downstream aortic PVAT depots. The key to the method is producing a stable occlusion with a diameter small enough to produce a pressure gradient but not so small that the downstream flow is inadequate to support tissue oxygenation. To this end, a thoracotomy was performed to expose the thoracic aorta in adult mice anesthetized with isoflurane and artificially ventilated. A coarctation of the mid-thoracic aorta was produced by tying a ligature around the aorta and a blunted needle on the aorta, then removing the needle. In other mice, a silicone rubber or nitrile O-ring of varying dimensions was placed around the aorta. Mice were allowed to recover for varying periods of time (weeks), then the degree of coarctation was evaluated using both imaging methods (ultrasound) and direct measurement of the pressure gradient (catheters placed in a carotid and a femoral artery) under isoflurane anesthesia. In a group of 9 male mice with coarctation and 4 male mice with sham coarctation, we found an average mean arterial pressure gradient of 11.7 ± 5.0 mmHg (mean ± SEM) in coarct mice and 1.6 ± 2.0 mmHg in sham mice. Systolic pressure gradients averaged 14.0 ± 5.9 mmHg in coarct mice and 1.0 ± 3.2 mmHg in sham mice. These results suggest it is possible to produce a stable pressure gradient across the thoracic aorta of mice as a tool to explore the impact of arterial pressure on vascular and PVAT structure and function.

Abstract 12: Deficiency of Progranulin Decreases Mitochondrial Function and Induces a Whitening Phenotype in Perivascular Adipose Tissue, Suppressing Its Anti-Contractile Effects

Perivascular adipose tissue (PVAT) is recognized as an endocrine organ that secretes a variety of adipokines, which negatively regulate vascular contraction. Progranulin (PGRN), a glycoprotein encoded by the GRN gene, is highly expressed in adipocytes, and is implicated in inflammation, cellular proliferation, and repair mechanisms. The relationship between PGRN and PVAT remains unclear. We hypothesized that the lack of PGRN negatively affects PVAT quality. We used male C57BL/6J wild-type (WT) and global PGRN knockout (KO) mice, aged 10-12 weeks. Energy expenditure was analyzed via the Comprehensive Lab Animal Monitoring System (CLAMS). Vascular function was studied in endothelium-intact aortae with PVAT [PVAT(+)] and without PVAT [PVAT(-)]. Additionally, 3T3-L1 cells were used to evaluate the molecular mechanisms. The KO mice showed a decreased respiratory exchange ratio (RER) and a whitening phenotype of PVAT, which was accompanied by a loss of anti-contractile effects [(mN) WT_PVAT(-): 5.2±0.3 vs PVAT(+): 3.4±0.5*, *P<0.05 and KO_PVAT(-): 6.1±0.7 vs KO_PVAT(+): 5.7±0.4]. Additionally, PVAT from KO displayed a significant reduction in nitric oxide production [(RFU) WT_PVAT: 223.1± 12.3 vs KO_PVAT: 67.3±9.1*, *P<0.05] and endothelial NO synthase phosphorylation. Additionally, the lack of PGRN attenuated the expression of mitochondrial complexes [(AU) WT_PVAT: 1.3± 0.03 vs KO_PVAT: 0.6±0.06*, *P<0.05] measured by the OXPHOS cocktail antibody. Via high-resolution respirometry, we found that isolated mitochondria from KO_PVAT showed impaired succinate-driven respiration or complex II activity. To understand if the whitening phenotype of PVAT was leading to its dysfunction, we induced a browning phenotype of PVAT by treating the mice with a β3-agonist for 7 days (CL 316243, 1mg/kg/day). This treatment increased RER restored the PVAT phenotype, and improved PVAT anti-contractile effect and mitochondrial respiration. Using 3T3-L1 cells, we engineered the overexpression and knockdown of PGRN via lentivirus (LV) infection and siRNA, respectively. Suppression of PGRN increased mRNA of whitening markers (1.5-fold increase vs scramble siRNA), while overexpression elevated mRNA of browning and mitochondrial markers (1.3-fold increase vs scramble LV). In summary, our findings reveal that PGRN is crucial for maintaining the phenotype and anti-contractile function of PVAT and highlight PGRN as a pharmacological target for vascular outcomes in cardiometabolic conditions.

Egg white hydrolysate modulates the renin-angiotensin system in mesenteric perivascular adipose tissue in doca-salt hypertensive rats and restores its anticontractile ability

Perivascular adipose tissue (PVAT) regulates vascular tonus with an anticontractile effect, which may be dysfunctional in hypertension. An Egg White Hydrolysate (EWH), a bioactive food, restores endothelium dysfunction and reduces blood pressure levels in malignant hypertension. We aimed to evaluate whether dietary supplementation with EWH interferes with the PVAT function in the mesenteric resistance arteries (MRA) of DOCA-salt hypertensive animals and the mechanisms involved. Male rats were divided into 4 groups and treated for 8 weeks. For 4 weeks, DOCA-salt or vehicle-treated rats (SHAM) were induced, and after that, some rats were co-treated with DOCA-salt or vehicle plus EWH (1 g/kg/day) for 4 weeks more. MRA and/or their PVAT (mPVAT) were used for functional, molecular, and biochemical analysis. The anticontractile effect of mPVAT in response to norepinephrine (NE) was observed only in MRA rings with PVAT of the SHAM group, while this effect was abolished in DOCA-salt rings. The EWH treatment did not change the anticontractile effect of mPVAT in SHAM but partially restored it in DOCA-salt rats. Acute aliskiren incubation reduced NE-induced contraction only in MRA with PVAT of DOCA-salt rats. EWH prevented the overactivation of the renin levels and ACE activity in mPVAT of DOCA-salt rats; while did not change AT1R expression, but increased AT2R expression. In mPVAT, EWH blocked the increase of MDA, ROS and pro-inflammatory cytokines observed in DOCA-salt rats. EWH improved PVAT dysfunction in MRA of severe hypertension. This beneficial effect could be mediated by an ameliorated redox balance, inflammatory state, and RAS axis.

Sex-Specific Effects of Cholesteryl Ester Transfer Protein (CETP) on the Perivascular Adipose Tissue.

Cholesteryl ester transfer protein (CETP) increases the atherosclerosis risk by lowering HDL-cholesterol levels. It also exhibits tissue-specific effects independent of HDL. However, sexual dimorphism of CETP effects remains largely unexplored. Here, we hypothesized that CETP impacts the perivascular adipose tissue (PVAT) phenotype and function in a sex-specific manner. PVAT function, gene and protein expression, and morphology were examined in male and female transgenic mice expressing human or simian CETP and their non-transgenic counterparts (NTg). PVAT exerted its anticontractile effect in aortas from NTg males, NTg females, and CETP females, but not in CETP males. CETP male PVAT had reduced NO levels, decreased eNOS and phospho-eNOS levels, oxidative stress, increased NOX1 and 2, and decreased SOD2 and 3 expressions. In contrast, CETP-expressing female PVAT displayed increased NO and phospho-eNOS levels with unchanged NOX expression. NOX inhibition and the antioxidant tempol restored PVAT anticontractile function in CETP males. Ex vivo estrogen treatment also restored PVAT function in CETP males. Moreover, CETP males, but not female PVAT, show increased inflammatory markers. PVAT lipid content increased in CETP males but decreased in CETP females, while PVAT cholesterol content increased in CETP females. CETP male PVAT exhibited elevated leptin and reduced Prdm16 (brown adipocyte marker) expression. These findings highlight CETP sex-specific impact on PVAT. In males, CETP impaired PVAT anticontractile function, accompanied by oxidative stress, inflammation, and whitening. Conversely, in females, CETP expression increased NO levels, induced an anti-inflammatory phenotype, and preserved the anticontractile function. This study reveals sex-specific vascular dysfunction mediated by CETP.

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.

Protective role of perivascular adipose tissue in the cardiovascular system.

This review provides an overview of the key role played by perivascular adipose tissue (PVAT) in the protection of cardiovascular health. PVAT is a specific type of adipose tissue that wraps around blood vessels and has recently emerged as a critical factor for maintenance of vascular health. Through a profound exploration of existing research, this review sheds light on the intricate structural composition and cellular origins of PVAT, with a particular emphasis on combining its regulatory functions for vascular tone, inflammation, oxidative stress, and endothelial function. The review then delves into the intricate mechanisms by which PVAT exerts its protective effects, including the secretion of diverse adipokines and manipulation of the renin-angiotensin complex. The review further examines the alterations in PVAT function and phenotype observed in several cardiovascular diseases, including atherosclerosis, hypertension, and heart failure. Recognizing the complex interactions of PVAT with the cardiovascular system is critical for pursuing breakthrough therapeutic strategies that can target cardiovascular disease. Therefore, this review aims to augment present understanding of the protective role of PVAT in cardiovascular health, with a special emphasis on elucidating potential mechanisms and paving the way for future research directions in this evolving field.

Homocysteine impairs the anticontractile/vasorelaxing activity of perivascular adipose tissue surrounding human internal mammary artery.

Perivascular adipose tissue (PVAT) surrounding human internal mammary artery (IMA) possesses anticontractile property. Its function under pathological conditions is barely studied. We previously reported that homocysteine impairs the vasodilator function of IMA through endothelium and smooth muscle-dependent mechanisms. This study investigated the effect of homocysteine on the function of PVAT and the associated mechanisms. Residual IMA tissues were collected from patients undergoing coronary artery bypass grafting. Vasoreactivity was studied using myograph. Adiponectin was measured by ELISA. Expressions of adiponectin receptors (AdipoRs), eNOS and p-eNOS were determined by RT-qPCR and Western blot. Exposure to homocysteine augmented the contractile responses of PVAT-intact IMA to U46619 and potassium chloride, regardless with or without endothelium. Such augmentation was also observed in skeletonized IMA with transferred, homocysteine-exposed PVAT. Homocysteine attenuated the relaxant response of PVAT-intact while endothelium-denuded vessels to acetylcholine. Homocysteine lowered adiponectin content in the PVAT, downregulated the expression of AdipoR1 and AdipoR2 as well as eNOS and p-eNOS in skeletonized IMA. The relaxant response of skeletonized IMA to AdipoR agonist AdipoRon was blunted by homocysteine or eNOS inhibitor, and homocysteine significantly attenuated the inhibitory effect of eNOS inhibitor on AdipoRon-induced relaxation. Homocysteine impairs the anticontractile/vasorelaxing activity of PVAT surrounding the IMA through inhibiting adiponectin/AdipoR/eNOS/nitric oxide signalling pathway.

Abstract 13047: The Modulatory Effects of Perivascular Adipose Tissue on the Renal Arterial Vasorelaxation and Its Relationship With Kidney Function in Metabolic Syndrome

Metabolic syndrome (MetS) is a cluster condition that increases the risk of cardiovascular diseases and type 2 diabetes. Perivascular adipose tissue (PVAT) regulates the arterial tone. We have demonstrated that the renal arterial PVAT enhances the vasorelaxation response in MetS model strains, SHRSP.Z- Lepr fa /IzmDmcr rats (SPZF) and SHR/NDmcr-cp rats (CP). Furthermore, we have found that overactivation of angiotensin II type 1 receptor (AT1R) signaling and a decrease in apelin levels in PVAT leads to a decline in compensatory PVAT effects. MetS is associated with chronic kidney disease, and the dysfunction of the PVAT favoring the vasorelaxation response could be a reason for kidney disease development. Therefore, we investigated the relationship between PVAT function and kidney function using MetS model rats.Renal arteries were isolated from the male and female SPZF and CP at 23 weeks of age. Vasorelaxation and mRNA transcript levels in PVAT were examined using organ bath methods and quantitative real-time polymerase chain reaction, respectively. Systolic blood pressure (sBP) was measured using a tail-cuff method. Insulin, glucose, and creatinine in the serum and protein in the urine were measured using commercial kits and eGFR, and the HOMA-IR were calculated.sBP was higher in SPZF than CP, but lower in females than males. HOMA-IR and urine protein levels were the lowest, while eGFR and enhancing effects of PVAT on acetylcholine-induced relaxations were the highest in female CP among the groups. eGFR was negatively correlated with sBP, HOMA-IR, and urinary protein levels. The enhancements of relaxations by PVAT were negatively correlated with sBP, HOMA-IR, and AT1R activity in PVAT.This study suggests that the insulin resistance and high blood pressure are associated with PVAT dysfunction, resulting from AT1 activation in PVAT, while it does not support the hypothesis that the disappearance of the effects of PVAT directly leads to kidney dysfunction.

Nicotine Impairs the Anti-Contractile Function of Perivascular Adipose Tissue by Inhibiting the PPARγ-Adiponectin-AdipoR1 Axis.

Nicotine is an addictive compound found in cigarette smoke that leads to vascular dysfunction and cardiovascular diseases. Perivascular adipose tissue (PVAT) exerts an anti-contractile effect on the underlying vasculature through the production of adipokines, such as adiponectin, which acts on adiponectin receptors 1 (adipoR1) to cause vasorelaxation. Peroxisome proliferator-activated receptor gamma (PPARγ) is a transcription factor that regulates adiponectin gene expression and PVAT development. This study aimed to determine the effect of nicotine on the anti-contractile function of PVAT via the PPARγ-adiponectin-adipoR1 axis. Male Sprague Dawley rats were divided into a control group (given normal saline), a nicotine group (given 0.8 mg/kg of nicotine), and a nicotine + PPARγ agonist group (given nicotine and 5 mg/kg of telmisartan). Thoracic aorta PVAT was harvested after 21 days of treatment. The results showed that nicotine reduced the anti-contractile effect of PVAT on the underlying thoracic aorta. Nicotine also decreased the gene and protein expression of PPARγ, adiponectin, and adipoR1 in PVAT. Treatment with telmisartan restored the anti-contractile effect of PVAT and increased the gene and protein expression of PPARγ, adiponectin, and adipoR1 in PVAT. In conclusion, nicotine attenuates the anti-contractile function of PVAT through inhibition of the PPARγ-adiponectin-adipoR1 axis.

Abstract 018: Overexpression Of ERα In Adipose Tissue Contributes To The Anti-contractile Effect Of Mesenteric PVAT By NOX4-induced H 2 O 2

Perivascular adipose tissue (PVAT) induces a significant anti-contractile effect on vasculature, secreting relaxing and constricting factors. It is well established that the PVAT anti-contractile function is lost in cardiometabolic diseases, including obesity. Estrogen receptor alpha (ERα) is known to play a crucial role in the function of adipose tissues and its activation has been associated with adipogenesis by redox balance. In addition, NADPH oxidase 4 (NOX4) is an enzyme that contribute to adipose tissue differentiation and intracellular H 2 O 2 generation. While ERα are expressed in adipose tissue, but their potential role in PVAT function is unknown. To test the hypothesis, adipose tissue specific ERα overexpressing female mice at 30 weeks old were used. We aimed to determine whether the overexpression of ERα would enhance the anti-contractile effect of PVAT by NOX4-induced H 2 O 2 . The high-fat diet (40% fat) was administered for 20 weeks and mesenteric resistance arteries, with and without PVAT, were isolated and used for measurements of vascular function, protein levels (Western blot) and redox signaling (Amplex Red). The higher-fat diet increases the weigh in ~ 80% in Wild Type (WT) and ERα (p>0.05). The anti-contractile effect of PVAT was exacerbated in arteries of ERα mice than WT [Area under the curve (AUC): WT 369±2 vs. ERα 230±1 p<0.05). NOX1/4 inhibitor (GSK136901) increased the vasoconstriction only in arteries from ERα mice (WT 348 ±4 vs. ERα 580±5 p<0.05). NOX4 (0.19±0.01 vs 2.3±0.6) and H 2 O 2 (> 70%) levels were higher in PVAT from ERα mice than WT. In addition, 6 uM of H 2 O 2 , concentration corresponding to the observed in PVAT from ERα mice, induced vasodilatation in arteries from WT. On the other hand, a high-fat diet abrogates the anti-contractile effect of PVAT (WT with PVAT 233±1 vs. without PVAT 241±4) and has partially restored in the ERα, analyzing LogEC 50 (with PVAT 5±0.7 and without PVAT 6±0.8 M). In addition, H 2 O 2 levels (>50%) were higher in PVAT from the high fat diet fed ERα mice, but NOX4 levels did not change. In summary, overexpression of ERα increases H 2 O 2 levels derived from NOX4, contributing to the anti-contractile effect of PVAT in healthy animals. This effect is partially restored in mice on a high-fat diet with overexpressed ERα.

Isolation, Culture, and Adipogenic Induction of Stromal Vascular Fraction-derived Preadipocytes from Mouse Periaortic Adipose Tissue.

Perivascular adipose tissue (PVAT) is an adipose tissue depot that surrounds blood vessels and exhibits the phenotypes of white, beige, and brown adipocytes. Recent discoveries have shed light on the central role of PVAT in regulating vascular homeostasis and participating in the pathogenesis of cardiovascular diseases. A comprehensive understanding of PVAT properties and regulation is of great importance for the development of future therapies. Primary cultures of periaortic adipocytes are valuable for studying PVAT function and the crosstalk between periaortic adipocytes and vascular cells. This paper presents an economical and feasible protocol for the isolation, culture, and adipogenic induction of stromal vascular fraction-derived preadipocytes from mouse periaortic adipose tissue, which can be useful for modeling adipogenesis or lipogenesis in vitro. The protocol outlines tissue processing and cell differentiation for culturing periaortic adipocytes from young mice. This protocol will provide the technological cornerstone at the bench side for the investigation of PVAT function.

TUDCA treatment restores anticontractile function of perivascular adipose tissue (PVAT) in post-weaning protein-restricted mice

Healthy perivascular adipose tissue (PVAT) has anticontractile, anti-inflammatory and antioxidant actions that can be disrupted by cardiometabolic risk factors. In the present study we hypothesized that protein restriction in the early phases of development could result in PVAT dysfunction and tauroursodeoxycholic acid (TUDCA) could be beneficial by inhibiting endoplasmic reticulum stress (ERS). To test this hypothesis, post-weaning male and female mice fed a normoprotein (NP, 14% protein) or a low protein (LP, 6% protein, isocaloric) diet for 15 weeks. In the last 2 weeks, some NP and LP animals received TUDCA (300 mg/kg/day) or vehicle. At the end of treatment, thoracic aorta was isolated to assess vascular responses with or without adjacent PVAT, as well as PVAT gene expression and histology. As results, the presence of PVAT reduced the phenylephrine-induced contraction in aorta of female NP and LP and male NP but not in male LP, suggesting LP-induced PVAT dysfunction in males only. In males, aortic PVAT from LP group showed less lipid content and increased collagen deposition. Decreased expression of PRDM-16, PPAR-γ, PGC-1α and leptin was observed in PVAT of LP group. These functional and morphological PVAT alterations in males were reversed by TUDCA treatment. Because TUDCA inhibits ERS, next we evaluated gene expression of ERS markers. There was an increased expression of GRP78, ATF4/6, IRE1α, and CHOP in PVAT from LP group, which was normalized by TUDCA. Also, TUDCA treatment normalized the reduced PVAT expression of the TUDCA receptor FXR in LP group. In conclusion, our data suggest that post-weaning protein-restriction impairs PVAT function and structure in males. These changes were associated with decreased expression of adipogenesis and adipocyte differentiation factors, and increased ERS markers. TUDCA reversed protein-restriction-induced PVAT abnormalities. Therefore, TUDCA emerges as a potential therapy for vascular complications associated with early undernutrition. The authors have nothing to disclose. 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.

Perivascular adipose tissue: a driver of sex-specific differences in the vascular outcomes of obesity

Obesity is a potent predictor of cardiovascular disease (CVD) and associated risk factors, including hypertension. Perivascular adipose tissue (PVAT) surrounds most blood vessels and modulates vascular tone. Under physiological conditions, PVAT exerts anti-contractile effects and sex differences are apparent in PVAT function. Under obese conditions, PVAT becomes dysregulated which may contribute to vascular dysfunction; how sex impacts the remodelling of PVAT and altered vascular contractility during obesity is unclear. We hypothesise that sex specific PVAT dysfunction in the setting of obesity is an important driver of sex differences in vascular pathologies and CVD risk.Adult (8wk) male and female C57Bl/6J mice received either chow diet (CD; 2.5%kcal fat) or high fat diet (HFD; 58%kcal fat) for 16 weeks (n=8/sex and diet). At 15 weeks of diet, blood pressure was assessed via tail-cuff plethysmography. At 16 weeks of diet, mesenteric arteries were isolated, and PVAT left intact or removed. Wire myography was used to assess the dilatory response to endothelium-dependent acetylcholine (ACh) and endothelium independent sodium nitroprusside (SNP). LogEC50 and %Preconstriction were calculated for each concentration response curve. PVAT was dissected from mesenteric arteries (mPVAT) and flash frozen for bulk RNA sequencing or fixed for histology. Data are mean ± SD and were analysed by two-way ANOVA.Under CD, the removal of PVAT did not change vasodilatory response to ACh or SNP in either sex. Under HFD, male mice showed reduced sensitivity (logEC50 PVAT vs no PVAT, p=0.0349) and maximal dilatory response (%Preconstriction PVAT vs no PVAT, p=0.0119) to ACh. This effect was not observed in females. Moreover, PVAT exerted no effect on dilatory responses to SNP in HFD groups of either sex, indicating an endothelium-dependent effect. Blood pressure was not altered in either sex by HFD. Transcriptomic analysis using RNA sequencing revealed female mPVAT is clearly distinguished from male mPVAT under HFD, but not CD conditions, with significantly down-regulated extracellular matrix remodelling and pro-inflammatory pathways, and significantly upregulated oxidative phosphorylation pathways. Histological analysis revealed potentiated adipocyte hypertrophy and collagen staining in mPVAT from males compared to females under HFD (p=0.034 and p=0.018 respectively).Our study shows that female mice are more protected from adverse effects of HFD, including PVAT-mediated endothelial dysfunction. Our RNA sequencing data provides insights into how pathophysiologic remodelling of mPVAT may contribute to the vascular outcomes of obesity-associated hypertension and suggests that this may be modulated by sex hormones. Further studies will unpick the underlying mechanism to identify potential targets for therapeutic intervention in obesity-associated hypertension. Funded by British Heart Foundation grant FS/19/55/34890. 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.

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.