scholarly journals Transglutaminases Are Active in Perivascular Adipose Tissue

2021 ◽  
Vol 22 (5) ◽  
pp. 2649
Author(s):  
Alexis N. Orr ◽  
Janice M. Thompson ◽  
Janae M. Lyttle ◽  
Stephanie W. Watts
Keyword(s):  
Adipose Tissue ◽  
Vascular Remodeling ◽  
Coagulation Factor ◽  
Sprague Dawley ◽  
Rt Pcr ◽  
Ideal Model ◽  
Perivascular Adipose Tissue ◽  
Tissue Sections ◽  
Insight Into ◽  
Immunohistochemical Analyses

Transglutaminases (TGs) are crosslinking enzymes best known for their vascular remodeling in hypertension. They require calcium to form an isopeptide bond, connecting a glutamine to a protein bound lysine residue or a free amine donor such as norepinephrine (NE) or serotonin (5-HT). We discovered that perivascular adipose tissue (PVAT) contains significant amounts of these amines, making PVAT an ideal model to test interactions of amines and TGs. We hypothesized that transglutaminases are active in PVAT. Real time RT-PCR determined that Sprague Dawley rat aortic, superior mesenteric artery (SMA), and mesenteric resistance vessel (MR) PVATs express TG2 and blood coagulation Factor-XIII (FXIII) mRNA. Consistent with this, immunohistochemical analyses support that these PVATs all express TG2 and FXIII protein. The activity of TG2 and FXIII was investigated in tissue sections using substrate peptides that label active TGs when in a catalyzing calcium solution. Both TG2 and FXIII were active in rat aortic PVAT, SMAPVAT, and MRPVAT. Western blot analysis determined that the known TG inhibitor cystamine reduced incorporation of experimentally added amine donor 5-(biotinamido)pentylamine (BAP) into MRPVAT. Finally, experimentally added NE competitively inhibited incorporation of BAP into MRPVAT adipocytes. Further studies to determine the identity of amidated proteins will give insight into how these enzymes contribute to functions of PVAT and, ultimately, blood pressure.

Abstract P071: Transglutaminases Are Active In Perivascular Adipose Tissue

Hypertension ◽  
2020 ◽  
Vol 76 (Suppl_1) ◽  
Author(s):  
Alexis Orr ◽  
Janice Thompson ◽  
Janae Lyttle ◽  
Stephanie W Watts
Keyword(s):  
Adipose Tissue ◽  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Coagulation Factor ◽  
Sprague Dawley ◽  
Perivascular Adipose Tissue ◽  
Tissue Sections ◽  
Mesenteric Resistance Artery ◽  
Immunohistochemical Analyses

Transglutaminases (TGs) are crosslinking enzymes best known for their vascular remodeling in hypertension. They require calcium to form an isopeptide bond, connecting a glutamine to a protein bound lysine residue or a free amine donor such as norepinephrine (NE) or serotonin (5-HT). We discovered that perivascular adipose tissue (PVAT) contains significant amounts of these amines, making PVAT an ideal model in which to test interactions of amines and TGs. We hypothesized that TG2 and FXIII are active in PVAT. Sprague-Dawley rat aortic, superior mesenteric (SMA), and mesenteric resistance artery (MR) PVAT express TG2 and blood coagulation factor XIII (FXIII) mRNA (Figure 1A). Consistent with this, immunohistochemical analyses support that PVATs all express TG2 and FXIII protein. The activity of TG2 and FXIII was investigated in tissue sections using substrate peptides that label active TGs and a catalyzing calcium solution, visualized with TRITC fluorescence (Figure 1B,C). Both TG2 and FXIII are active in rat aortic PVAT, SMAPVAT, and MRPVAT. Western blot analysis determined that the known TG inhibitor cystamine reduced incorporation of experimentally added amine donor 5-(biotinamido)pentylamine (BAP) into MRPVAT by 6.14% of total normalized signal (p<0.0001, N=7). Further Western blot analysis proved that experimentally added 5-HT competitively inhibits incorporation of experimentally added BAP into MRPVAT adipocytes, reducing total normalized signal by 10.75% (p=0.001, N=4). Further studies to determine what proteins TGs are amidating will give insight into how these enzymes contribute to the development of hypertension.

Abstract 333: Existence of Functional Norepinephrine Transport in Perivascular Adipose Tissue

Hypertension ◽  
2014 ◽  
Vol 64 (suppl_1) ◽  
Author(s):  
Nadia Ayala-Lopez ◽  
Robert Burnett ◽  
Janice M Thompson ◽  
Stephanie W Watts
Keyword(s):  
Blood Vessel ◽  
Contractile Function ◽  
Organic Cation Transporter ◽  
Sprague Dawley ◽  
Rt Pcr ◽  
Perivascular Adipose Tissue ◽  
Healthy State ◽  
Health And Disease ◽  
Organic Cation Transporter 3 ◽  
Beta 2 Microglobulin

Perivascular Tissue (PVAT) is a recent focus of studies for its regulation of blood vessel tone. Contraction to norepinephrine (NE) is reduced by PVAT through mechanisms not entirely clear. Loss of this anti-contractile function of PVAT occurs in obesity-related hypertension. We hypothesized that PVAT can remove NE from the blood vessel environment through transporters and reduce NE induced contraction in health. RT-PCR of mesenteric PVAT was positive for norepinephrine transporter (NET; Ct=33.2±1.4) and serotonin transporter (SERT; Ct=31.9±0.9) relative to beta-2-microglobulin. To study NE uptake in the healthy state, mesenteric and aortic PVAT from the male Sprague Dawley rat was incubated with 10 μM NE after pre-incubation with an inhibitor of transport (10 μM desipramine to inhibit NET and SERT, 100 μM corticosterone to inhibit organic cation transporter 3) or vehicle and quantified by HPLC for NE. Desipramine plus corticosterone inhibited NE uptake in mesenteric PVAT (figure) but desipramine (410.5±80.8 ng/g vs. 414.4±67.0 ng/g NE only; n=8) or corticosterone (225.0±26.1 ng/g vs. 319.2±35.9 ng/g NE only; n=5) alone did not, indicating a robust uptake capacity. In aortic PVAT, NE uptake was not inhibited by desipramine plus corticosterone (figure), desipramine (1763.5±460.8 ng/g vs. 1702.7±298.4 ng/g NE only; n=6), or corticosterone (1085.8±205.3 ng/g vs. 2245.8±506.8 ng/g NE only; n=4). Protein staining revealed bands positive for NET in mesenteric PVAT by Western blot. Taken together, we found that PVAT is a reservoir of NE that can remove NE from the blood vessel environment, an important finding to understand vasculature-PVAT interactions in health and disease.

scholarly journals Perivascular adipose tissue‐derived stromal cells contribute to vascular remodeling during aging

Aging Cell ◽  
2019 ◽  
Vol 18 (4) ◽  
Author(s):  
Xiao‐Xi Pan ◽  
Cheng‐Chao Ruan ◽  
Xiu‐Ying Liu ◽  
Ling‐Ran Kong ◽  
Yu Ma ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Stromal Cells ◽  
Vascular Remodeling ◽  
Perivascular Adipose Tissue

The P2Y12 Receptor Antagonist Ticagrelor Ameliorates Pulmonary Hypertension

2021 ◽  
Author(s):  
nannan li ◽  
jie yin ◽  
yugen shi ◽  
li sun ◽  
qingshan zhang ◽  
...  
Keyword(s):  
Vascular Remodeling ◽  
Adenosine Diphosphate ◽  
P2y12 Receptor ◽  
Sprague Dawley ◽  
Rt Pcr ◽  
Sprague Dawley Rats ◽  
Haemodynamic Changes ◽  
P2y12 Receptor Antagonist ◽  
Pulmonary Arterial ◽  
Adp Receptor

Abstract Background: Pulmonary arterial hypertension (PAH) is a disease that the pulmonary artery is abnormally elevated. P2Y12 is an adenosine diphosphate (ADP) receptor and it act as the target of thienopyridine antiplatelet drugs by controlling vascular remodeling. Inhibition of P2Y12 receptor in the process of PAH was explored in this study.Methods: The PAH model was established in Sprague-Dawley rats by single subcutaneous injection of 60 mg/kg monocrotaline (MCT). The ticagrelor solution (a selective P2Y12R inhibitor) was intraperitoneally injected into rats at a dose of 14 mg/kg from the time of MCT injection to day 28.Results: In the lung tissues of PAH rats, the marked P2Y12R was detected. Treatment with ticagrelor greatly decreased P2Y12R level and efficiently abolished the upregulation of α-SMA as demonstrated by Western blot and RT-PCR. The wall thickness and occlusion score of the pulmonary arterioles showed that blockade of P2Y12R could relieve lung remodeling caused by PAH. The haemodynamic changes at 4 weeks determined that P2Y12R inhibition affected RV pressure and right heart hypertrophy.Conclusions: P2Y12R might be involved in the pathogenesis of PAH. Blockade of P2Y12R has potential in treating PAH.

Abstract P067: Piezo1 Activation Inhibits Adipogenesis And Lipogenesis In Perivascular Adipose Tissue Preadipocytes

Hypertension ◽  
2020 ◽  
Vol 76 (Suppl_1) ◽  
Author(s):  
Cristian Javier Rendon Mora ◽  
Emma D Flood ◽  
Stephanie W Watts ◽  
G.Andres Contreras ◽  
Janice Thompson
Keyword(s):  
Adipose Tissue ◽  
Expression Patterns ◽  
Stromal Vascular Fraction ◽  
Serial Passage ◽  
Cell System ◽  
Sprague Dawley ◽  
Perivascular Adipose Tissue ◽  
Triglyceride Accumulation ◽  
Sprague Dawley Rats ◽  
Triglyceride Content

In adipogenesis, perivascular adipose tissue (PVAT) preadipocytes turn into adipocytes. In non-PVAT preadipocytes, mechanical forces affect the commitment and lipogenic stages of adipogenesis. The mechanism may involve PIEZO1, a mechanosensor, that boosts differentiation of progenitor cells towards osteogenic and fibroblastic lineages. Since hypertension causes changes in the vascular forces that could affect adipogenesis in PVAT, our objective was to evaluate PIEZO1 expression patterns in PVAT and the effects of PIEZO1 activation on the adipogenic potential of preadipocytes. We hypothesize that PIEZO1 activation limits the adipogenic potential of PVAT preadipocytes. PVAT from the thoracic aorta (APVAT) was collected from male Sprague Dawley rats at 10 weeks of age (n=5). Preadipocytes were obtained by Liberase digestion followed by serial passage of adherent cells. Preadipocyte progenitors, CD34+PDGFRα+, were harvested by magnetic-activated sorting. PIEZO1 expression was assessed by RT-qPCR and immunofluorescence (IF). Preadipocytes were induced to differentiate for 14 d in standard media (CON) or in the presence of PIEZO1 agonist Yoda (10μM) and inhibitor Dooku (5μM) during days 0-2 (commitment), 3-14 (lipogenesis), and 0-14 (overall adipogenesis). Adipogenesis was evaluated with IncuCyte Live-Cell system using the triglyceride stain Bodipy 493503. Triglyceride content is reported as Bodipy Intensity/Adipocyte Count. Piezo1 RNA was expressed in adipocytes and the stromal vascular fraction of APVAT. PIEZO1 IF signal was detected in SVF and preadipocyte. Triglyceride was reduced by Yoda (62 ± 14.3) and Dooku (49.3 ± 14) during 0-2 d compared to CON (312.5 ± 165.6). Neither Yoda nor Dooku for 12 d affected triglyceride accumulation compared to CON. In contrast, the lipid content of Yoda (77.7 ± 21.3) and Dooku (48.9 ± 15.2) treated cells during 14 d was reduced vs CON (312.5 ± 165.6). The expression of PIEZO1 in all PVAT fractions suggests mechanosensitivity. PIEZO1 activation during adipogenesis commitment impaired adipocyte maturation. These data provide evidence for the capacity of mechanosensory expressed in PVAT preadipocytes to modulate adipogenesis, underpinning the deleterious impact of hypertension on PVAT function.

scholarly journals Organic cation transporter 3 contributes to norepinephrine uptake into perivascular adipose tissue

2015 ◽  
Vol 309 (11) ◽  
pp. H1904-H1914 ◽  
Author(s):  
Nadia Ayala-Lopez ◽  
William F. Jackson ◽  
Robert Burnett ◽  
James N. Wilson ◽  
Janice M. Thompson ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Organic Cation ◽  
Organic Cation Transporter ◽  
Confocal Imaging ◽  
Uptake System ◽  
Sprague Dawley ◽  
Uptake Mechanism ◽  
Fluorescent Substrate ◽  
Perivascular Adipose Tissue ◽  
Organic Cation Transporter 3

Perivascular adipose tissue (PVAT) reduces vasoconstriction to norepinephrine (NE). A mechanism by which PVAT could function to reduce vascular contraction is by decreasing the amount of NE to which the vessel is exposed. PVATs from male Sprague-Dawley rats were used to test the hypothesis that PVAT has a NE uptake mechanism. NE was detected by HPLC in mesenteric PVAT and isolated adipocytes. Uptake of NE (10 μM) in mesenteric PVAT was reduced by the NE transporter (NET) inhibitor nisoxetine (1 μM, 73.68 ± 7.62%, all values reported as percentages of vehicle), the 5-hydroxytryptamine transporter (SERT) inhibitor citalopram (100 nM) with the organic cation transporter 3 (OCT3) inhibitor corticosterone (100 μM, 56.18 ± 5.21%), and the NET inhibitor desipramine (10 μM) with corticosterone (100 μM, 61.18 ± 6.82%). Aortic PVAT NE uptake was reduced by corticosterone (100 μM, 53.01 ± 10.96%). Confocal imaging of mesenteric PVAT stained with 4-[4-(dimethylamino)-styrl]- N-methylpyridinium iodide (ASP+), a fluorescent substrate of cationic transporters, detected ASP+ uptake into adipocytes. ASP+ (2 μM) uptake was reduced by citalopram (100 nM, 66.68 ± 6.43%), corticosterone (100 μM, 43.49 ± 10.17%), nisoxetine (100 nM, 84.12 ± 4.24%), citalopram with corticosterone (100 nM and 100 μM, respectively, 35.75 ± 4.21%), and desipramine with corticosterone (10 and 100 μM, respectively, 50.47 ± 5.78%). NET protein was not detected in mesenteric PVAT adipocytes. Expression of Slc22a3 (OCT3 gene) mRNA and protein in PVAT adipocytes was detected by RT-PCR and immunocytochemistry, respectively. These end points support the presence of a transporter-mediated NE uptake system within PVAT with a potential mediator being OCT3.

scholarly journals Effect of Resistance Training on NADPH Oxidase and Adiponectin in PVAT of OVX Rats

2021 ◽  
Vol 30 (2) ◽  
pp. 205-212
Author(s):  
Erling Guo ◽  
Jin-Hwan Yoon ◽  
Wooyeon Jo ◽  
Jaeho Jin ◽  
Sang Ki Lee
Keyword(s):  
Adipose Tissue ◽  
Resistance Training ◽  
Nadph Oxidase ◽  
Abdominal Aorta ◽  
Sham Group ◽  
Ovariectomized Rats ◽  
Sprague Dawley ◽  
Perivascular Adipose Tissue ◽  
Ovx Rats ◽  
Sprague Dawley Rats

PURPOSE: Perivascular adipose tissue (PVAT) is a type of adipose tissue that surrounds vessels to provide anti-contractile effects. This study aimed to investigate the effect of resistance training on NADPH oxidase, adiponectin, and endothelial NOS (eNOS) expression in the abdominal aorta and PVAT of ovariectomized rats.METHODS: Sprague-Dawley rats at 20 weeks of age were divided into three groups: sham control (Sham, n=10), OVX-control (OVX_ Con, n=10), and OVX-resistance exercise (OVX_Rex, n=10). Resistance training was performed by climbing a ladder for 12 weeks. Western blotting was used to analyze target protein expression in the rat abdominal aorta and PVAT.RESULTS: NADPH oxidase (p67phox) expression was significantly higher in the OVX_Con group than in the sham group, but it was significantly decreased in the OVX_Rex group. The expression of adiponectin, AKT, and eNOS in both abdominal aorta and PVAT was significantly reduced in the OVX_Con group than in the Sham group, but it was improved in the OVX_Rex group.CONCLUSIONS:The results suggest that regular resistance training inhibits p67phox and increases adiponectin expression and phosphorylation of AKT and eNOS in abdominal aortic PVAT of ovariectomized rats.

scholarly journals Loss of anti-contractile effect of perivascular adipose tissue in offspring of obese rats

2016 ◽  
Vol 40 (8) ◽  
pp. 1205-1214 ◽  
Author(s):  
K E Zaborska ◽  
M Wareing ◽  
G Edwards ◽  
C Austin
Keyword(s):  
Adipose Tissue ◽  
Maternal Obesity ◽  
Later Life ◽  
Independent Effect ◽  
Sprague Dawley ◽  
Response Curves ◽  
Perivascular Adipose Tissue ◽  
Contractile Effect ◽  
Sprague Dawley Rats ◽  
Pregnancy And Lactation

Abstract Rationale: Maternal obesity pre-programmes offspring to develop obesity and associated cardiovascular disease. Perivascular adipose tissue (PVAT) exerts an anti-contractile effect on the vasculature, which is reduced in hypertension and obesity. Objective: The objective of this study was to determine whether maternal obesity pre-programmes offspring to develop PVAT dysfunction in later life. Methods: Female Sprague–Dawley rats were fed a diet containing 10% (control) or 45% fat (high fat diet, HFD) for 12 weeks prior to mating and during pregnancy and lactation. Male offspring were killed at 12 or 24 weeks of age and tension in PVAT-intact or -denuded mesenteric artery segments was measured isometrically. Concentration–response curves were constructed to U46619 and norepinephrine. Results: Only 24-week-old HFD offspring were hypertensive (P<0.0001), although the anti-contractile effect of PVAT was lost in vessels from HFD offspring of each age. Inhibition of nitric oxide (NO) synthase with 100 μM l-NMMA attenuated the anti-contractile effect of PVAT and increased contractility of PVAT-denuded arteries (P<0.05, P<0.0001). The increase in contraction was smaller in PVAT-intact than PVAT-denuded vessels from 12-week-old HFD offspring, suggesting decreased PVAT-derived NO and release of a contractile factor (P<0.07). An additional, NO-independent effect of PVAT was evident only in norepinephrine-contracted vessels. Activation of AMP-activated kinase (with 10 μM A769662) was anti-contractile in PVAT-denuded (P<0.0001) and -intact (P<0.01) vessels and was due solely to NO in controls; the AMPK effect was similar in HFD offspring vessels (P<0.001 and P<0.01, respectively) but was partially NO-independent. Conclusions: The diminished anti-contractile effects of PVAT in offspring of HFD dams are primarily due to release of a PVAT-derived contractile factor and reduced NO bioavailability.

Abstract P291: High Fat Diet Promotes Hypertension and Vascular Remodeling but Not Vascular Fibrosis or Altered Contractility in Dahl Salt Sensitive Rats

Hypertension ◽  
2016 ◽  
Vol 68 (suppl_1) ◽  
Author(s):  
Roxanne Fernandes ◽  
Patricia A Perez Bonilla ◽  
Hannah Garver ◽  
James J Galligan ◽  
Gregory D Fink ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
High Fat Diet ◽  
Vascular Remodeling ◽  
Vascular Reactivity ◽  
Vascular Dysfunction ◽  
Mesenteric Arteries ◽  
High Fat ◽  
Perivascular Adipose Tissue ◽  
Morphological Studies

Obesity associated hypertension in rodent models is commonly associated with altered vascular reactivity to sympathetic neurotransmitters and inflammation-induced vascular remodeling/fibrosis. Dahl salt-sensitive (SS) rats exhibit elevated sympathetic activity and vascular remodeling. We hypothesized that diet-induced obesity in Dahl SS rats would promote hypertension, vascular dysfunction and remodeling/fibrosis. Male Dahl SS rats were placed on high fat diet (HFD, 60% kcal from fat with final concentrations of 0.33% NaCl and 1% K + , n=5) or normal-fat diet (NFD; 10% kcal from fat, 0.24% NaCl, 0.36% K + , n=5) for 24-26 weeks after weaning (3 weeks of age). Compared with NFD rats, HFD rats displayed severe hypertension (MAP, 165±4 mmHg vs 133±6 mmHg, P<0.05), higher body-weight (470±6g vs 433±7g, P<0.05), and hyperlipidemia (cholesterol, 211±22 mg/dl vs 138±23 mg/dl, P=0.05). HFD rats did not show significant changes in plasma levels of fasting glucose (85±5 mg/dl vs 75±5 mg/dl), insulin (2.6±0.8 ng/ml vs 2.2±1.1 ng/ml), leptin (0.77±0.18 ng/ml vs 0.44±0.06 ng/ml), or aldosterone (249±3 pg/ml vs 234±3 pg/ml) (all P>0.05). HFD did not affect pressurized mesenteric arterial (~300 μm inner diameter, 60 mmHg) reactivity to norepinephrine or ATP in vitro . Pressurized mesenteric arteries from HFD rats displayed thicker walls (Ca 2+ free buffer, 40±1 μm vs 36±1 μm, P<0.05), but showed slightly increased distensibility. Morphological studies did not reveal greater fibrosis in adventitia of mesenteric, intrarenal and coronary arteries from HFD rats. However, HFD induced inflammation in mesenteric perivascular adipose tissue, as shown by increased CD3 positive cell infiltration and histological evidence of fibrosis and angiogenesis. Our studies indicate that HFD in male Dahl SS rats promotes hypertension, perivascular adipose tissue inflammation and vascular remodeling, but not vascular fibrosis. Alteration of vascular contractility to sympathetic neurotransmitters, however, is not required for obesity associated hypertension in Dahl SS rats.

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