bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 59: Loss of Cullin-3 in Smooth Muscle Increases RhoA, Vascular Contraction and Arterial Pressure
Authors: Pelham Christopher , Ketsawatsomkron Pimonrat , Grobe Justin , Faraci Frank , Sigmund Curt
Published: July 2021

Mutations in Cullin-3 result in early-onset hypertension in humans through an undefined mechanism. Cullin-3 RING E3 ubiquitin ligase complex mediates proteasomal degradation of RhoA, a key mediator of vascular smooth muscle tone. We reported that smooth muscle-specific expression of dominant negative PPARγ (S-P467L) in transgenic mice causes hypertension and aortic dysfunction via increased RhoA/Rho-kinase signaling. Dominant negative PPARγ caused decreased Cullin-3 protein (0.31±0.04-fold of Non-Transgenic (NT); P<0.05) and decreased ratio of active Nedd8-Cullin-3 / Cullin-3 (0.06±0.04 vs 0.51±0.18 NT; P<0.05), which correlated with increased protein levels of Cullin-3 substrates RhoA (2.6±0.2-fold of NT; P<0.05) and Cyclin E (2.0±0.1-fold of NT; P<0.05) in S-P467L medial aorta. We tested the hypothesis that Cullin-3 controls arterial pressure by regulating vascular function. siRNA-mediated knockdown of Cullin-3 (72 hr), confirmed by Western blot, increased RhoA (2.5±0.1-fold of NC1 negative control siRNA; P<0.05) and Cyclin E (1.9±0.2-fold of NC1; P<0.05) protein in primary rat aortic smooth muscle cells. Inhibition of cullin-RING ligase activity using the Nedd8-activating enzyme inhibitor, MLN4924 (1μM, 16 hr), also increased RhoA (4.3±1.0-fold of DMSO vehicle). Treatment of aortic rings from control mice with MLN4924 (1μM in DMEM/F12, 16 hr) resulted in enhanced agonist-mediated contraction in response to endothelin 1 (30nM: 0.35±0.05g vs 0.19±0.02g DMSO vehicle; P<0.05), serotonin (3μM: 1.2±0.07g vs 0.9±0.08g; P<0.05), and phenylephrine (3μM: 1.1±0.05g vs 0.7±0.05g, P<0.05) that was Rho-kinase-dependent, despite decreased contraction to KCl (100mM: 0.7±0.03g vs 1.0±0.03g; P<0.05). Finally, administration of MLN4924 (30mg/kg TID, 2 days) to control mice
in vivo
increased mean arterial pressure during the light phase (121±4 vs. 108±3mmHg baseline, P<0.05; dark phase: 122±4 vs. 123±4mmHg baseline, NS) as measured by radiotelemetry. Our results demonstrate that interference with PPARγ in smooth muscle causes vascular dysfunction via impaired Cullin-3-mediated regulation of RhoA/Rho-kinase signaling and provide a mechanistic link between mutations in Cullin-3 and hypertension.
Abstract 600: Neonatal Transient Hyperoxic Exposure Leads to Persistent Vascular DNA Damage and Increased Senescence Proteins in Rats: Potential Role in Developmental Programming of Hypertension
Authors: Huyard Fanny , Bertagnolli Mariane , Fallaha Catherine , Nuyt Anne Monique
Published: July 2021

Background:
Preterm infants have reduced antioxidant defenses vs. babies born at term and are exposed to high levels of oxygen (O2) both in intensive care and as compared to the intrauterine environment. We have shown that neonatal hyperoxic stress leads in adult rats to hypertension, endothelial dysfunction and arterial rigidity, all characteristic of vascular aging. Indeed, acutely, markers of vascular cells senescence are increased.

Aim:
To examine whether indices of vascular aging persists beyond neonatal hyperoxic exposure. For this purpose, vessels (aorta) are studied at 4 weeks (wks, pre-hypertension) and 16 wks (young adults-elevated blood pressure).

Methods:
Sprague-Dawley pups were kept at 80% O2 or in room air from postnatal days 3 to 10. Aortas were sampled at 4 and 16 wks (n=5-7 animals/group) to examine tumour suppressor p53 binding protein 1 (53BP1, indices of DNA damage) and senescence proteins (p53, p16 and p21) by immunofluorescence. Bax and bcl2 proteins (for apoptosis) were assessed by western blot at 4 wks.

Results:
At 4 wks, aortas of O2 exposed rats showed increased nuclear foci of 53BP1 (19.3±0.8 vs. 10.5±1.1 fluorescence intensity, p<0.0001), and increased expression of p53 (19.3±0.4 vs. 16.3±0.9, p<0.05) compared to controls. Expressions of Bax (pro-apoptotic protein) and the ratio Bax/Bcl2 were not different between groups (2±0.2 vs. 1.6±0.1, and 1.4±0.1 vs. 1.2±0.1, respectively). Bcl2 (anti-apoptotic protein) was decreased in O2 exposed rats compared to controls (1.6±0.03 vs. 1.3±0.07, p<0.01). At 16 wks, O2 exposed rats showed also increased nuclear foci of 53BP1 (23.9±3.8 vs. 14.2±0.9, p<0.05), and increased expression of p53 (21.6±1.6 vs. 13.8±0.5, p<0.01), and p16 (16.9±1.6 vs. 7.7±0.8, p<0.01), p21 was not changed (11.5±0.9 vs. 14±1.9, non significant).

Conclusions:
Neonatal exposure to O2 leads at 4 wks and 16 wks to DNA damage and expression of senescence associated proteins without evidence of apoptosis (at
4wks
). These changes are present before the adult consequences of neonatal exposure (i.e. hypertension and vascular dysfunction) and may contribute to their development.
Abstract 601: Total Plaque Area Measurement Is A Reliable Indicator Of Vascular Risk Factor Treatment Effect
Authors: Garcia Nestor H , Perez Hernan A , Spence J. David , Armando Luis J
Published: July 2021

Background:
Ultrasound measurement of carotid plaque area (TPA) is a noninvasive method for identifying patients at higher risk of cardiovascular events. Previous studies demonstrated that intensive medical therapy reduces TPA. In this study we examined effects of less intensive therapy on TPA, and the usefulness of TPA in risk stratification.

Methods.
Baseline Framingham Risk Score (FRS), TPA, blood pressure (BP), body mass index and the post test probability using TPA (PtP TPA) were determined from patients at the primary care unit. TPA measurements were reported to primary physicians to encourage them to follow preventive guidelines. After 10±1 months patients were reevaluated to investigate changes from baseline.

Results:
among 140 patients enrolled between July 2009, and July 2011, 52% were male, BP was 142±1/81±1 mmHg, BMI 29±1 kg/m2, FRS 24±1, PtP TPA 50±2, and TPA 97±7 mm2. After 10±1 months of treatment, three groups were identified. Group 1 had a reduction of TPA (127±13 to 99±12 mm2, p<0.002), BP (142±3/80±2 to 137±2/75±2mmHg, p<0.002) and PtP TPA score (55±3 to 47±3, p<0.007). Group 2 patients had progression of TPA (from 90±8 to 138±12mm2, p<0.001) and reached a poorer PtP TPA score (51±2 to 58±2, p<0.001). Group 3 patients with stable TPA only decreased the BMI (from 28.5±0.9 to 27.8±1.0 kg/m2, p<0.05). TPA identified high-risk patients; 27% of these patients achieved regression of TPA, associated with a decrease of BP and PtP TPA. The rest of the patients only decreased the BMI, and the TPA remained stable. FRS did not change in any group in spite of changes on blood pressure or body weight. These data suggest that reduction in BP as well as reduction of BMI is required to prevent TPA progression.

Conclusions:
Plaque measurement may be useful for targeting preventive therapy and evaluating new treatments and response to therapy, and may improve cost-effectiveness of secondary preventive treatment.
Abstract 602: Determinants of Elevated Blood pressure with Water Immersion
Authors: Kawano Hiroshi , Mineta Mayuko , Gando Yuko , Asaka Meiko , Higuchi Mitsuru
Published: July 2021

Introduction
; Although exercise in water is appreciated in rehabilitation for patients with renal disease and obesity individuals with knee joint pain, elevated blood pressure (BP) is also induced by water immersion
via
water pressure. This elevation of BP in water may lead to incidence of acute heart diseases in rehabilitation exercise. It is important for prevention of such acute event to understand determinants of elevated BP with water immersion.

Hypothesis
; We assessed the hypothesis that elevated BP with water immersion is affected by venous volume or baroreflex sensitivity (BRS) as well as age or arterial stiffening, because blood pressure is associated with venous return (contributing cardiac output) or vascular functions.

Methods
; Thirty-eight young (21.2 ± 1.7 yrs) and 20 older (65.1 ± 3.2 yrs) men participated in this study. In all subjects, mean BP (oscillometric method), pulse wave velocity adjusted by BP (cardio-ankle vascular index: CAVI), venous volume (by using MRI and plethysmography), and BRS by using valsalva maneuver were measured at resting supine position on land. Furthermore, BP and heart rate at standing position on land and following in water (located surface of water at epigastrium) were determined.

Results
; On land, mean BP and CAVI were greater, and BRS was smaller in older men compared with young men. There was no different venous volume between 2 age groups. Change in increased systolic BP with water immersion was greater in older men (127 ± 12 mmHg → 145 ± 18 mmHg; P<0.05) than in young men (118 ± 129 mmHg → 129 ± 10 mmHg; P<0.05) (Interaction; P<0.05). Multiple-regression analysis revealed that the change in increased systolic BP with water immersion were independently associated with CAVI (beta = 0.406), when entering BRS, heart rate, venous volume, and CAVI.

Conclusion
; In conclusion, these results of the present study suggests that arterial stiffening may contribute to elevated BP with water immersion, but not BRS or venous volume.
Abstract 603: Silencing Of PKG1 Gene Mimics Effect Of Aging And Sensitizes Vascular Smooth Muscle Cells To Pro-fibrotic Effect Of Cardiotonic Steroids
Authors: Fedorova Olga V , Shilova Victoria , Lakatta Edward G , Bagrov Alexei Y
Published: July 2021

Marinobufagenin (MBG), an endogenous Na/K-ATPase inhibitor and a vasoconstrictor, induces vascular fibrosis via PKC-dependent inhibition of Fli-1, a nuclear transcription factor and a negative regulator of collagen synthesis. In vascular smooth muscle cells (VSMC), ANP, via cGMP/PKG1 dependent mechanism, reduces sensitivity of Na/K-ATPase to MBG. Because vascular fibrosis is a hallmark of cardiovascular aging, we hypothesized that VSMC from aged rats, in which cGMP/PKG-dependent signaling is down-regulated, would exhibit heightened sensitivity to the pro-fibrotic effect of MBG.

In primary culture of VSMC from thoracic aorta from young (3 month old) rats, 1 nmol/L MBG inhibited Na/K-ATPase by 15%, and induced a 60% down-regulation of Fli-1 and a concomitant 50% increase in the levels of collagen-1. Alpha-ANP (1 nmol/L) blocked these effects.

In VSMC from aged (24 months old) rats levels of PKG1 and Fli-1 in VSMC were markedly reduced, while PDE-5 was markedly upregulated. MBG (1 nmol/L) inhibited Na/K-ATPase by 26%, suppressed Fli-1 two-fold, and increased level of collagen-1 more than two-fold (P<0.01). Unlike that in young rats, alpha-ANP failed to oppose these effects of MBG in aged animals.

Silencing of the PKG1 gene in VSMC from young rats produced a marked increase in the sensitivity of VSMC to the pro-fibrotic effect of MBG; 1 nmol/L MBG increased levels of collagen-1 2.5-fold (P<0.01).

These results demonstrate that age-associated reduction in vascular PKG1 levels and resultant decline in cGMP signaling sensitize VSMC to pro-fibrotic effect of MBG. Silencing of PKG1 in VSMC from young rats mimics these effects of aging.
Abstract 608: Unilateral Renal Denervation Enhances Baroreflex Function in Conscious Rabbits
Authors: Schiller Alicia M , Curry Pamela L , Zucker Irving H
Published: July 2021

Renal nerve denervation (DNx) is currently being assessed as a therapy for drug-resistant hypertension and other disorders characterized by sympatho-excitation, such as chronic heart failure (CHF). We hypothesized that renal DNx enhances arterial baroreflex (BR) control of heart rate (HR) in normal and CHF rabbits. All animals were instrumented with ventricular pacing leads and an arterial pressure (AP) radiotelemetry transducer. After recovery the left kidney was denervated by stripping the renal artery of all visible fibers. Intact animals were subjected to a sham operation. Two weeks later, CHF animals were subjected to rapid ventricular pacing. BR changes in HR and AP were induced by iv sodium nitroprusside and phenylephrine (PE) infusions alone and after injection of atropine or metoprolol. BR function was determined by fitting mean AP and HR data to a 4 parameter logistic equation. Renal DNx increased HR range from 156 ± 34 bpm in intact rabbits compared to 298 ± 39 bpm in DNx rabbits (p = 0.022). This was primarily due to a decrease in the minimum HR following PE infusion. During blockage with metoprolol HR range was increased from 140 ± 39.35 bpm in Sham to 262.75 ± 9.97 bpm in DNx rabbits (p = 0.016) suggesting an improvement in the vagal components of BR function. In intact rabbits with CHF, HR range was 110 ± 16 bpm compared to 294 ± 15 bpm in CHF renal DNx animals (p = .002). These data suggest renal DNx enhances the autonomic components of BR control of HR in the normal and CHF states.
Abstract 61: Adipose Tissue Specific HO-1 Induction Regulates the Crosstalk between Wnt and β-catenin Pathways and Leads to Restoration of Vascular Endothelial Function in Mice Fed a High Fat Diet
Authors: Cao Jian , Peterson Stephen J , Schwartzman Michal L , Sodhi Komal , Rezzani Rita , Rodella Luigi F , Abraham Nader G
Published: July 2021

The principal goal of the present study is to examine the contributions of adipocyte specific over expression of the cytoprotective gene heme oxygenase-1 (HO-1) in modulating vascular function in an animal model of diet induced obesity. A lentiviral construct of the human HO-1 was synthesized under the control of an aP2-activated promoter so as to ensure HO-1 targeting to murine adipocytes. Lentiviruses (50 μl, 2x109 TU/ml in saline) were injected into the C57BL/6 mice by a single intracardiac injection. Mice (6-7 wks old) were divided into 4 groups: Control, high fat diet (HF) mice receiving the control Lenti-aP2-GFP (HF-GFP) and high fat diet mice receiving the lenti-aP2-HO-1 (HF-HO-1) with and without treatment with SnMP. Transduction of lenti-aP2-HO-1 increased human HO-1 expression in adipose tissues without affecting endogenous mice HO-1 (p<0.01). In mice fed a HF diet, lenti-aP2-HO-1 transduction attenuated the increases in body weight (from 52.0±1.0 g to 35.6±2.1g; p<0.01), blood glucose (from 255±3.5 g to 140±2.9mg/dl), blood pressure (from 135±2.8mmHg to 101±2.2mmHg; p<0.01) and inflammatory cytokines as well as the content of both visceral (from 5.5±0.15g to 2.9±0.22g; p<0.05) and subcutaneous fat (p<0.05). Transduction of lenti-aP2-HO-1 increased the numbers of adipocytes of small cell size (p<0.05), insulin sensitivity (p<0.05), adiponectin levels (from 32.4±1.9μg/ml to 19.9±2.1μg/ml; p<0.05) as well as vascular relaxation to acetylcholine ( p<0.05) compared to HF mice administered the lenti-aP2- GFP. Adipocytes of mice fed a HF diet expressed high levels of PPARγ, aP2, C/EBP and Wnt5b proteins and displayed marked increases in Peg1/Mest (p<0.03). Lenti-aP2-HO-1 transduction lowered the elevated levels of these proteins and increased Shh, Wnt10b and β-catenin (p<0.05). Inhibition of HO activity by administration of tin mesoporphyrin (SnMP) to HF-fed mice transduced with the lenti-aP2-HO-1 reversed the decrease in Peg 1/Mest, TNFα and MCP-1 levels. This novel study demonstrate that adipocyte-specific overexpression of HO-1 attenuates HF-mediated adiposity and vascular dysfunction, increases insulin sensitivity and improves adipocyte function by increasing adiponectin, Shh and WNT10b and decreasing inflammatory cytokines.
Abstract 614: Mechanism Involved In Fructose Induced Cardiovascular Risk: Evidence In An Experimental Model Of Hypertension And Menopause
Authors: Conti Filipe F , Brito Janaina d , Bernardes Nathalia , Dias Danielle d , Sanches Iris C , Irigoyen Maria-Cláudia , De Angelis Kátia
Published: July 2021

The metabolic syndrome is a bunch of ordinary pathologies: abdominal obesity associated to an excess of visceral fat, insulin resistance, dyslipidemia and hypertension. Moreover, the prevalence of hypertension and metabolic dysfunctions sharply increases after the menopause. However, the mechanisms involved in these changes were not well understood. Thus, the aim of this study was to evaluate the effects of ovarian hormone deprivation (9 wks) in hypertensive (or not) female rats submitted (or not) to fructose overload on cardiovascular autonomic control and cardiac oxidative stress. Female Wistar and SHR rats were divided into (n=8/group): control (C), ovariectomized (O), hypertensive ovariectomized (HO) and hypertensive ovariectomized submitted to fructose overload (100g/L in drinking water) (FHO). The arterial pressure (AP) signals were directly recorded. The cardiovascular autonomic modulation was evaluated by pharmacological blockade and by spectral analysis. Oxidative stress profile was evaluated by chemiluminescence (CL) and redox balance of glutathione (GSH/GSSG) determinations. The AP showed increased in the O and HO groups when compared to C group. The FHO group showed increased blood glucose and tryglicerides, insulin resistance; an additional increase in mean AP (174±4 vs. C:108±1; O:121±2; HO:164±5 mmHg), resting tachycardia, reduced vagal tonus (5±3 vs. C:55±5; O:37±6; HO:35±7bpm), increased sympathetic tonus (91±18 vs. 39±10bpm in C), decreased high frequency band of the pulse interval (vagal modulation) and exacerbated increased systolic AP variance and decreased low frequency band of the pulse interval (sympathetic modulation) in relation to other groups. The CL was higher in O and HO groups than C group. FHO group showed additional increase in CL (15043±1333 vs. C: 2661±358; O: 5134±277; HO: 6514±547 cps/mgprot) when compared to other groups. The cardiac GSH/GSSG was lower in FOH group (8.94±0.80 umol/g) in relation to HO group (13.0±1.4 umol/g), indicating impaired redox balance. In conclusion, the fructose overload induced metabolic dysfunctions associated with additional impairment in cardiovascular autonomic control and oxidative stress profile in hypertensive rats submitted to ovarian hormones deprivation.
Abstract 623: Robust Strategy to Identify Metabolomic Markers of Preeclampsia
Authors: Delles Christian , Sungatullina Liliya , Burgess Karl E , Tsiropoulou Sophia , Carty David M , McBride Martin W
Published: July 2021

Background.
Despite considerable progress in recent years the pathogenesis of preeclampsia remains incompletely understood. Metabolomic studies have the potential to discover novel disease pathways through an unbiased approach but are prone to exaggerating small but statistically significant differences in metabolite levels.

Methods.
We selected 36 women who participated in the Proteomics in Preeclampsia (PIP) study from whom plasma samples at gestational weeks 16 and 28 were available. Of these, 18 subsequently developed preeclampsia (cases) and 18 had normotensive pregnancies (controls). Metabolomic studies were performed in plasma using hydrophilic interaction liquid chromatography coupled to Orbitrap high resolution mass spectrometry (HILIC-MS). Statistical analysis was performed using IDEOM version 13.0. More than 2-fold change and false discovery rate (FDR) < 0.05 were considered as statistically significant between the groups (i.e., cases and controls at each gestational age, and between different gestational ages within cases and controls separately).

Results.
We detected 133 metabolites which were differentially expressed between the groups. We identified the amino sulfonic acid taurine as differentially regulated between cases and controls with 1.47 and 2.09-fold lower levels in cases compared to controls at weeks 16 and 28, respectively, and with a 1.42-fold increase from week 16 to 28 in controls as opposed to no change in cases. Significant increases (2.28 to 2.92-fold) in progesterone and related metabolites (pregnanediol-3-glucuronide and [ST (2:0)] 5 alpha-cholesta-8,24-dien-3beta-ol) during pregnancy in all women were expected and add further confidence to our data.

Conclusion.
Our longitudinal study design and an analysis strategy based not only on statistical significance but also on fold changes detects taurine as marker of preeclampsia. Taurine has anti-inflammatory properties and reduced levels could provide a mechanistic explanation for development of preeclampsia in some women.
Abstract 625: Role Of Proximal Na+ Reabsorption In The Fructose-induced Hypertension
Authors: Queiroz-Leite Gabriella D , Neri Elida A , Rebouças Nancy A , Malnic Gerhard
Published: July 2021

Methods/Results:
Wistar rats were fed for 2 or 8 weeks with control diet (CTRL) or isocaloric 60% fructose diet (HF). Systolic blood pressure (SBP) was measured weekly by tail cuff plethysmography. There was no difference in the SBP (mmHg) after 2 weeks of HF intake (110 ± 1.7 CTRL x 121 ± 5.8 HF); however, there was an increase in SBP after 8 weeks (111.9 ± 1.3 CTRL x 143.3 ± 1.5 HF, P < 0.0001). Urine of 24h collected in metabolic cages showed a reduction in urine flow (mL/min) (2 weeks - 0.018 ± 0.002 CTRL x 0.009 ± 0.001 HF, P = 0.0007 and 8 weeks - 0.017 ± 0.002 CTRL x 0.007 ± 0.0004 HF, P = 0.001). Glomerular filtration rate (GFR) was not altered after 2 weeks (1.31 ± 0.3 CTRL x 1.22 ± 0.5 HF), but was decreased after 8 weeks (1.82 ± 0.4 CTRL x 0.37 ± 0.1 HF, P = 0.001). Na+ fractional excretion (Na-FE), in % of creatinine clearance, was decreased after 2 weeks (0.37 ± 0.05 CRTL x 0.13 ±0.03 HF), but was increased after 8 weeks (0.18 ± 0.06 CTRL x 0.54 ± 0.14 HF, P = 0.038). After 2 or 8 weeks, rats were subjected to in situ microperfusion experiments and had their proximal tubules (TP) perfused with an alkaline solution to investigate Na+ reabsorption by means of bicarbonate flux (JHCO3-, in nmol/cm2 x s). JHCO3- was increased after 2 weeks (1.78 ± 0.09 CTRL x 2.7 ± 0.14 HF), but was reduced after 8 weeks (2.00 ± 0.09 CRTL x 1.19 ± 0.04 HF). The perfusion of PT with S3226, a specific NHE3 inhibitor, showed that fructose acts stimulating NHE3 activity after 2 weeks (0.82 ± 0.09 CRTL + S3226 x 1.83 ± 0.14 HF + S3226, P < 0.0001) and inhibiting NHE3 activity after 8 weeks (0.85 ± 0.09 CRTL x 0.22 ± 0.04 HF, P < 0.0001).

Conclusions:
Reduced Na+-FE and urine flow, associated with increased NHE3 activity after 2 weeks of HF suggest that, initially, fructose leads to a state of Na+ overload, which may contribute to the development of hypertension observed after 8 weeks of HF intake. Kidney damage was increased after 8 weeks, since GFR was decreased, explaining the persistence of reduced urine flow, despite reduced NHE3 activity. These data, together with the increase in Na-FE and installation of hypertension, suggest that the mechanism of pressure-natriuresis was activated after 8 weeks, in order to compensate for volume expansion. Our data suggest a role of increased Na+ reabsorption in the development of fructose-induced hypertension.
Abstract 630: Lymphocytes Exert Homeostatic Immune Function For Blood Pressure Control Via Inhibition Of The MPS / VEGF-C Regulatory Axis.
Authors: Brenner Sven , Schröder Agnes , Rakova Natalia , Jantsch Jonathan , Titze Jens
Published: July 2021

Background:
We have shown previously that electrolyte homeostasis is characterized by local regulation of water-free Na
+
storage in the skin. Macrophages infiltrate the Na
+
overloaded skin interstitium and induce interstitial electrolyte clearance by TonEBP- / VEGF-C-driven lymph capillary network hyperplasia, therefore providing a buffering mechanism for hypertension. Whether such extrarenal control for volume and blood pressure homeostasis is exclusively achieved by innate immune cells, or whether components of the adaptive immune system modulate this macrophage-specific response, is unclear. We hypothesized that lymphocytes might influence macrophage/TonEBP/VEGF-C driven regulation of skin electrolyte homeostasis and thereby modulate blood pressure.

Methods:
Seven wt and six RAG
-/-
-mice, which have no B- or T-cells, were fed a low-salt diet for 2 consecutive weeks. In the end of the experiments, blood pressure was measured via an arterial line, skin electrolyte composition was analyzed, and the macrophage/TonEBP/VEGF-C regulatory axis was characterized.

Results:
RAG
-/-
-mice had lower skin Na
+
(0.11±0.01 vrs. 0.15±0.04 mmol/g; P<0.05) and water content (1.06±0.14 vrs. 1.42±0.35 ml/g; P<0.05) than wt mice, showed increased VEGF-C mRNA expression in the skin (1.78±0.41 vrs. 1.00±0.66 arbitrary units; P<0.05), and displayed hyperplastic subcutaneous lymph capillaries (7.0±1.8 vrs. 4.0±2.0 arbitrary units; P<0.05). In parallel with this extracellular volume loss, mean arterial blood pressure was reduced in RAG
-/-
mice (97±8 vrs. 108±7 mmHg; P<0.05).

Conclusion:
We conclude that the absence of B- and T-cells disinhibits VEGF-C expression from subcutaneous macrophages, resulting in inadequate hyperplasia of subcutaneous vessels and extracellular volume loss from skin, which leads to hypotension in RAG
-/-
-mice. The findings suggest that B- or T-cells are part of a regulatory network in which macrophages modulate interstitial electrolyte homeostasis and blood pressure.

Perspectives:
Additional experiments in mice with selective B- or T-cell depletion, and adoptive transfer of B- or T-lymphocyte subsets into RAG
-/-
-mice will allow identification of specific lymphocyte subpopulations involved in inhibition of the TonEBP/VEGF-C regulatory axis in macrophages. Further experiments will also focus on molecular mechanisms of lymphocyte / macrophage interaction. We aim to demonstrate that cells of the innate and adaptive immune system form a network of "homeostatic immune function" for the control of electrolyte and blood pressure homeostasis.
Abstract 64: Microvascular Endothelial Dysfunction, Oxidative Stress and Reduced Arginine: Adma in Premenopausal Women With HIV
Authors: Wang Dan , Aslam Shakil , Hu Haihong , Melancon Joseph K , Verbesey Jennifer , Lee Dexter L , Young Mary , Wilcox Christopher S
Published: July 2021

Cardiovascular disease (CVD) is common in persons infected with the human immunodeficiency virus (HIV) who are receiving highly active antiretroviral therapy (HAART). Since CVD generally begins with microvascular dysfunction, we tested the hypothesis that women with HIV have microvascular endothelial dysfunction. We related this to reactive oxygen species (ROS), NOS substrate: inhibitor ratio (arginine: ADMA) and inflammation. Subcutaneous microvascular arterioles (SMAs) were isolated from gluteal skin biopsies from 11 premenopausal, virally suppressed African American HIV positive women receiving HAART but without CVD risk factors and 10 healthy matched controls in the Women’s Interagency HIV Study (WHIS). Preconstricted SMAs were mounted in a Mulvany-Halpern myograph and relaxed with graded concentrations of acetylcholine (ACh). Endothelium derived relaxation (EDR), endothelium-derived relaxation factor (EDRF; NOS-dependent relaxation), endothelium derived hyperpolarizing factor (EDHF; K
+
-channel dependent relaxation) and endothelium derived contracting factor (EDCF; ACh-induced contraction with relaxation pathways inhibited) were related to plasma cytokines (array) and plasma arginine: ADMA ratio (measured by capillary zone electrophoresis).Compared to controls, HIV infected subjects had reduced EDRs (50±6 vs 78±3%; P< 0.01), EDRF (27±4 vs 38±3%, P< 0.05), EDHF(18±4 vs 39±4%, P< 0.05) and decreased NO activity (0.18±0.05vs 0.69±0.17ΔUnits, P< 0.05 from DAF-FM fluorescence).They had an ACh-induced EDCF (21±2vs7±3%, P< 0.01) and increased microvascular ROS (0.14±0.04 vs 0.04±0.04 Δunits; P< 0.01 from tempo-9ACfluorescence). Plasma cytokines in HIV subjects showed enhanced inflammation and plasma arginine: ADMA concentrations were reduced (178±12 vs 203±13 μmol/ μmol; P< 0.005). In conclusion, pre-menopausal women receiving HAART with virally suppressed HIV who were free of CVD risk factors developed defective microvascular EDRF and EDHF with reduced NO and NOS substrate: inhibitor ratio. They had inflammation and their microvessels developed a new ACh-induced EDCF and enhanced ROS. These profound microvascular disturbances in apparently healthy young women may herald the development of premature CVD.
Abstract 647: Toll-like Receptor 4 (TLR4) Activation Impairs the Nitrergic Relaxation Response in Rat Corpus Cavernosum
Authors: Stallmann-Jorgensen Inger S , Davis Jason E , Carrillo-Sepulveda Maria Alicia , Webb R. Clinton
Published: July 2021

Cardiovascular disease etiology of hypertension and atherosclerosis has been shown to involve pattern-recognition receptor activation, including toll-like receptor 4 (TLR4). Indeed, our lab showed that TLR4 activation was implicated in blood pressure regulation, and its blockade lowered blood pressure in spontaneously hypertensive rats. However, whether TLR4 activation plays a role in erectile dysfunction is not known. Therefore, we hypothesized that TLR4 activation using its ligand, lipopolysaccharide (LPS), would increase reactive oxygen species (ROS) and impair corpus cavernosum (CC) relaxation. To test our hypothesis, rat CC smooth muscle cells were incubated for 15 min with LPS (100 ng/mL) and ROS detected with a fluorescent probe. Treatment with LPS resulted in an increase in ROS, compared to baseline (Fig.1). To investigate vascular reactivity to TLR4 activation, we performed myograph studies on CC isolated from 13-15 week old Sprague-Dawley rats, then incubated 3 hours in DMEM with LPS (+LPS) or vehicle (VEH). Concentration response curves to phenyleprine (PE, 10
-9
-10
-4
M) and to non-adrenergic, non-cholinergic (NANC) stimulation were used to investigate CC reactivity. Contractility to PE did not differ between groups, but +LPS-induced relaxation responses decreased significantly to 20-volt electrical field stimulation compared to VEH (8Hz: 16.3 vs 24.9, p< 0.01 and 16 Hz: 21.2 vs 35.1, p< 0.001). In conclusion, treatment with the TLR4 agonist, LPS, decreased NANC relaxation in the CC. Increased ROS generation may result in scavenging and reducing availability of nitric oxide that could impair relaxation in the CC.
Abstract 649: Heme-oxygenase-1 Induction Improves Type-1 Cardiorenal Syndrome Only In Mice With Impaired AngII-induced Lymphocyte Activation (SCID Mice)
Authors: Pesce Paola , Sacerdoti David , Monu Sumit R , Sodhi Komal , Boldrin Massimo , Abraham Nader G
Published: July 2021

Rational:
The absence of lymphocyte activity protects SCID mice from AngII-induced hypertension facilitating blood pressure-induced sodium excretion, possibly via the stimulation of eNOS- and COX-2-dependent pathways. Type-1 cardiorenal syndrome (CRS-1), characterized by acute kidney dysfunction secondary to deterioration in cardiac function, is caused by renal arteriolar vasoconstriction, mediated by the activation of renin-angiotensin and sympathetic nervous systems (RAAS, SNS). Heme Oxygenase-1 (HO-1) induction improves renal function, but not renal vasoconstriction, in AngII-induced hypertension, and causes the desensitization of vascular smooth muscle to phenylephrine.

Objectives:
We evaluated whether AngII resistant SCID mice develop CRS-1 as occurs in control mice, and the differential effects of HO1 induction on renal hemodynamics in CRS-1.

Methods:
Post ischemic heart failure was induced in C57 and SCID mice by left anterior coronary artery ligation. Mice were divided in 4 groups: sham, myocardial infarction (MI), MI treated with cobalt protoporphyrin (CoPP), an inducer of HO-1, in the presence and absence of HO activity inhibitor, stannous mesoporphyrin (SnMP). All mice underwent echocardiography (ejection fraction, EF) and renal echoDoppler (arterial pulsatility index, K-PI) examination 30 days after surgery.

Results:
EF was significantly reduced both in control and SCID mice after MI (C57: sham 0.60±0.07, MI 0.45±0.04, p<0.05; SCID: sham 0.60±0.06, MI: 0.46±0.1, p<0.01). K-PI was significantly increased in MI groups compared to sham groups (C57: sham 0.98±0.05, MI 1.12±0.11, p<0.05; SCID: sham 0.72±0.08, MI 1.37±0.37, p<0.05). HO1 induction improved renal vasoconstriction only in SCID mice (SCID K-PI: MI+CoPP 0.9±0.19 p<0.5çC57 K-PI: MI+CoPP 1.05±0.15, n.s.). In SCID mice SnMP reversed the effect of CoPP on renal vasoconstriction.

Conclusion:
CRS-1 is similar in SCID and control mice and is associated with increased renal arterial resistance. HO-1 induction improves renal vasoconstriction only in SCID (AngII resistant) mice with CRS-1, suggesting that increased HO-1 activity cannot protect the kidney from AngII-induced lymphocyte activation, but only from SNS-induced vasoconstriction.
Abstract 657: Inhibition Of MMPs In Hypertensive Cerebropathy Protects Blood-Brain Barrier In Dahl-Salt Sensitive Rats
Authors: Vacek Joanthan C , Sathnur Pushpakumar , Metreveli Naira , Kundu Sourav , Tyagi Suresh C , Tyagi Neetu
Published: July 2021

Hypertensive cerebropathy is a pathological condition associated with cerebral edema and breakdown of the blood-brain barrier (BBB). Previous studies have shown that increased activity of matrix metalloproteinases (MMPs) leads to alteration in the extracellular matrix (ECM) which is associated with impairment of the neurovasculature, resulting in destruction of the BBB. However, the molecular pathways leading to this condition remains obscure. In this study, we hypothesize that pharmacological inhibition of MMPs modulates BBB disruption by decreasing MMP activity and thus ameliorating hypertensive cerebropathy.

Methods:
We used Dahl-salt sensitive (Dahl-SS) and Lewis rats which were fed a high salt diet. The groups were 1) Dahl-SS, 2) Dahl-SS+GM6001 (non-specific MMP inhibitor), 3) Lewis, and 4) Lewis+GM6001. GM6001 was given at 0.5mg/mL by intra-peritoneal injection on alternate days for 3 weeks. Blood pressure was measured by tail-cuff. The brain tissues were analyzed for oxidative stress, matrix metalloproteinase-9 (MMP-9) activity and tight junction proteins by Western blot and Zymography. Immunohistochemistry was used to determine the collogen/elastion ratio.

Results:
Mean arterial blood pressure was 169.10 ± 0.57 mm Hg in hypertensive Dahl-SS rats compared to 134.12 ± 1.2 mm Hg in Dahl-SS+GM6001 rats. The mean arterial pressures in Lewis and Lewis+GM6001 groups were 96.66 ± 0.54 and 87.69 ± 2.9 mm Hg respectively. There was an up-regulation in p47 (oxidative stress) and nitrotyrosine protein expression but down-regulation in super oxide dismutase-1(SOD-1) and tight junction proteins levels (zonula-occluden-1 and occludin) in hypertensive Dahl-SS and Lewis rats compared to GM6001 treated rats. MMP-9 protein expression as well activity was increased in hypertensive Dahl-SS and Lewis rats compared to GM6001 treated rats. Brain section stained for collagen and elastin showed an increase in collagen level in hypertensive Dahl-SS and Lewis rats compared to GM6001 treated rats, whereas the elastin level decreased in hypertensive Dahl-SS and Lewis rats compared to GM6001 treated rats.

Conclusion:
Our results suggest that in hypertensive Dahl-SS rats, pharmacological inhibition of MMPs attenuates high blood pressure and prevents disruption of blood brain barrier.
Abstract 66: Increased Circulating Inflammatory Endothelial Cells (IEC) in African-American vs. Caucasians with Essential Hypertension
Authors: Eirin Alfonso , Herrmann Sandra M , Gloviczki Monika L , Zhu Xiangyang , Tang Hui , Jordan Kyra L , Juncos Luis A , Calhoun David A , Lerman Amir , Textor Stephen C , Lerman Lilach O
Published: July 2021

Introduction:
Morbidity and mortality attributable to hypertension are higher in African American (AAEH) compared to Caucasian essential hypertensive (EH) patients, possibly related to a differential effect on vascular injury and repair. While circulating endothelial progenitor cells (EPC) preserve endothelial integrity IEC detach from sites of injury and represent markers of vascular damage. We hypothesized that plasma levels of IEC and inflammatory markers would be higher in AAEH compared to EH patients.

METHODS:
Inferior vena cava levels of CD34+/KDR+ (EPC) and VAP-1+ (IEC) cells were measured by FACS in EH and AAEH under fixed sodium intake (150 mEq/d) and blockade of the renin-angiotensin-system, and compared to systemic levels in normotensive control subjects (n=19 each). Systemic levels of inflammatory cytokines and EPC homing factors were measured by Luminex.

Results:
Blood pressure, serum creatinine, lipids, antihypertensive medications, and EPC levels did not differ between EH and AAEH patients. Circulating IEC were higher in AAEH, and inversely correlated with EPC levels (Figure). Systemic levels of inflammatory cytokines and EPC homing factors were higher in AAEH compared to EH patients (Table), and correlated directly with IEC.

Conclusion:
Despite preserved kidney function and controlled BP, circulating inflammatory markers were elevated in AAEH and correlated with increased IEC and decreased EPC levels. Increased release of cytokines and IEC in AAEH may impair EPC reparative capacity and predispose to hypertensive vascular injury. This process may aggravate vascular damage and accelerate hypertension-related morbidity/mortality rates in AAEH.
Abstract 663: Angiotensin-(1-7)-MAS Receptor Gene Knockout Impairs Post-ischemic Hindlimb Neovascularization
Authors: Pereira Jousie M , Santos Robson A , Schirmer Brigida G , Barcelos Lucía S
Published: July 2021

Introduction:
Angiotensin-(1-7) is known by its cardiovascular protective effects by activating MAS receptor. It has been shown that Angiotensin-(1-7) stimulates proliferation of endothelial progenitor cells in vitro, but exerts antiangiogenic effects both in inflammatory and tumor environments in vivo. The role of Angiotensin-(1-7)-MAS axis on ischemic conditions was never reported. Hypothesis: We assessed the hypothesis that MAS receptor signaling plays a role in reparative neovascularization after hindlimb ischemia in mice.

METHODS:
C57BL/6 wild-type (WT) and MAS receptor knockout (MAS-KO) mice (8-10 weeks) were subjected to unilateral permanent left femoral artery occlusion (FAO). Hindlimb blood flow was measured before and immediately after FAO, and 7 and 14 days after FAO by laser Doppler perfusion imaging (LDPI).To further assess the vascularization in the ischemic limb, 28 days after FAO, we used the technique of microbubles contrast enhanced ultrasound perfusion imaging that enables a more sensitive discrimination of blood flow in deep microvessels. Capillary and arteriole density were evaluated by histological analysis at day 14 after FAO.

Results:
Vascular density was similar in normoperfused muscles from MAS-KO and WT mice. However, the neovascularization response to hindlimb ischemia was significantly reduced in MAS-KO muscles at capillary (p<0.05; n=4/group) and arteriole (p<0.05; n=4/group) level, which led to lesser perfusion recovery of the ischemic limb of MAS-KO mice as evaluated by LDPI (Day 7: 0.67±0.04 vs 0.85±0.04 ischemic/contralateral ratio, p <0.01; Day 14: 0.62±0.15 vs 0.91±0.03, p<0.001; n=8-12/group). In accordance, we observed that MAS-KO mice require shorter time to peak enhancement of microbubles contrast in ischemic adductor muscles compared with WT mice (p<0.01; n = 5-6/group).

Conclusion:
In conclusion, our data suggests that post-ischemic hindlimb neovascularization and blood flow recovery are impaired in Angiotensin-(1-7)-MAS receptor knockout mice.
Abstract 665: Endothelial Nitric Oxide Negatively Modulates Phenylephrine-induced Contraction In Matrix Metalloproteinase 2 (mmp-2)-filled Rabbit Aortic Rings
Authors: Prado Alejandro F , Pernomian Laena , Bendhack Lusiane M , Gerlach Raquel F
Published: July 2021

Endothelial nitric oxide negatively modulates Phenylephrine-induced contraction in Matrix metalloproteinase 2 (MMP-2)-filled rabbit aortic rings

The present study aimed to evaluate the vascular function of aortic rings after intraluminal injection of matrix metalloproteinase 2 (MMP-2) and contribution of endothelial nitric oxide (NO) in this response.

METHODS:
thoracic aorta isolated from
New Zealand
rabbits (3.0 Kg) was isolated and were performed in situ gelatinolytic assay (intraluminal exposure with Krebs solution or MMP-2 1.2 μg/mL, at 37° C for 30 minutes, with measure of fluorescence intensity - FI) or vascular reactivity. Cumulative concentration-effect curves for phenylephrine (PE, 0.1 nM - 10 μM/L) were performed in endothelium-intact (E+) or denuded (E-) aortic rings, in the absence or presence of L-NAME (non-selective inhibitor of NOS, 100 μM) or Hydroxycobalamine (scavenger of intracellular NO), 30 min before the PE curves on E+ rings. The vascular contraction was normalized by grams of tension by dry tissue (g/g) and were analyzed the agonist maximum effect (Emax) and potency (pD2). Data represent mean ± S.E.M and the results were statistically analyzed using Student t test, one-way or two-way ANOVA with Bonferroni pos-hoc (p<0,05).

Results:
In situ
gelatinolytic activity was increased in the vessels filled with MMP-2 (19.19+0.93U,n=5) compared to vehicle (10.75+0.45U,n=5). In the vascular reactivity, E- aortic rings filled with MMP-2 increased Emax of PE (1893,28+122,40g/g,n=9) versus vehicle E- (1485.73+76.82g/g,n=9), but no differences were observed on Emax of E+ aortic rings (vehicle: 1416.13+120.87g/g,n=7; MMP-2: 1893.28+122.40g/g,n=9). The presence of L-NAME induced an increase in Emax of E+ aortic rings filled with vehicle (1817.60+76.37g/g,n=7) or MMP-2 (2340.50+130.43g/g,n=5). Hydroxycobalamine also increased the Emax of E+ aortic rings with vehicle (1889,81+129,17,n=6) or MMP-2 (2480,12+192,57,n=5). No difference of pD2 value was observed in this study.

Conclusion:
Data suggest this protease cross the aorta layers from the lumen. An increased vasoconstriction in E- vessels suggests a vascular smooth muscle activity of MMP-2. In the presence of L-NAME and Hydroxycobalamine, vessels filled with MMP-2 shows a greater maximum effect suggesting that NO exerts a negative modulation of the PE-induced vasoconstriction on MMP-2 aortic rings.
Abstract 67: Fractalkine is Increased in Cardiomyocyte-Specific EP4 Knockout Hearts and Decreases Cardiomyocyte Contractility
Authors: Harding Pamela , Xu Jiang , Yang Xiao-Ping , Undrovinas Albertas , Taube David
Published: July 2021

Introduction:
Our laboratory has reported that approximately half of aged male mice with cardiomyocyte-selective knockout of the prostaglandin E2 EP4 receptor sub-type (EP4 KO) exhibit reduced cardiac function. Gene array on left ventricles (LV) of such mice showed elevated expression of fractalkine, a novel chemokine that is implicated in human heart failure, when compared with age-matched controls.

Hypothesis:
We therefore hypothesized that fractalkine protein is increased in EP4 KO hearts and contributes to depressed contractility.

Methods:
To test this hypothesis, echocardiography was performed on 28 week old male EP4 KO and wild type controls (WT), and fractalkine was measured in LV by ELISA. Additionally, the effect of fractalkine on single myocyte contractility and intracellular calcium was determined using the IonOptix myocyte contractility system. Myocytes (AVM) from male 16-20 week old C57Bl/6 mice were loaded with 1 μM Fura-2 AM and the response to 5 ng/ml fractalkine measured under basal conditions and after stimulation with 0.1 μM isoproterenol (Iso) with pacing at 3 Hz.

Results:
LV fractalkine was greater in EP4 KO than WT controls (0.34 vs 0.23 ng/mg protein, n=4, p <0.05). Under basal conditions, treatment of AVM for 10 min with 5 ng/ml fractalkine decreased both the speed of contraction and relaxation (from - 164.1 ± 22.8 to -100.4 ± 18.9 μm/sec, p < 0.05; and from 83.3 ± 16.8 to 38.2 ± 11.7 μm/sec respectively, p = 0.065, n = 22-33 cells). After stimulation with Iso, fractalkine also decreased both the speed of contraction and relaxation, from -568.8 ± 30.6 to -467.5 ± 40.5 μm/sec and from 407.0 ± 28.4 to 304.7 ± 37.5 μm/sec respectively, p < 0.05. In addition, fractalkine decreased the percent change in contraction after Iso stimulation from 10.3 ± 0.5 to 8.4 ± 0.8, p = 0.05. Surprisingly, fractalkine increased the percent change in the Fura-2 ratio after Iso stimulation from 115.8 ± 6.6 to 135.8 ± 6.2, p < 0.05 despite reducing contractility.

Conclusion:
Our results suggest that fractalkine directly depresses myocyte contractility by mechanisms that are downstream of changes in intracellular calcium. Furthermore, these effects may contribute to the impaired contractility observed in EP4 KO mouse hearts.
Abstract 75: Chronic Treatment With At2 Receptor Agonist Rescues High Fat Diet-induced Obesity in Female Mice.
Authors: Khan Mohammed A , Samuel Preethi , Nag Sourashish , Hussain Tahir
Published: July 2021

Obesity in itself is a disease condition and a major risk factor in the development of hypertension, dyslipidemia, and hyperglycemia. Therefore, successful strategies for improving obesity and related metabolic risk factors are needed. Role of renin-angiotensin system (RAS) has been implicated in obesity and metabolic dysfunction. Recently, we have shown that AT2R knock-out in female mice caused a greater body weight gain and hyperinsulimia in response to high fat diet (HFD). In the present study, we hypothesize that AT2R activation rescues diet-induced obesity in females. To test this hypothesis, we injected AT2R non-peptide agonist C21 (0.3mg/kg/day i.p) in C57BL6 female mice on HFD for 12 weeks. C21-treatment did not affect the HFD calorie intake (HFD: 937±18 Kcal; C21HFD: 886±37 Kcal) but caused lesser body weight gain compared to control (HFD: 4.4± 0.4g; C21HFD: 3.06± 0.4g). Similar to the body weight gain pattern, gonadal fat weight and adipocyte size were decreased significantly in C21-treated mice on HFD compared to control HFD group (HFD: 4.4± 0.4 g; C21 HFD: 3.06± 0.4g) and (HFD: 6404±161.6μm2 ; C21HFD: 3874±103.2μm2 ) respectively. Moreover, the C21-treated females on HFD had lower levels of plasma insulin, improved glucose tolerance, and decreased plasma free fatty acids and hepatic triglycerides. Western blot revealed that phospho-Ser79-acetyl CoA carboxylase (p-Ser79-ACC-1) was reduced, an index of increased lipogenic activity and decreased β-oxidation process, in both adipose (Adi) and hepatic (Hep) tissues of HFD fed groups (Adi: 86% and Hep: 73% of 100% controls); C21-treatment revered the decrease in p-ser79-ACC-1 in Adi (104% of control) and caused an increase in Hep (122% of control) respectively. The HFD feeding lowered the estradiol level (ND: 38.8±2.6 vs HFD:11.3±1.2ng), which was modestly reversed by C21 treatment (C21HFD:17.4± 1.5ng) in HFD mice. Our results strongly suggest that stimulation of AT2R in female mice positively contribute, predominantly independent of estrogen, to rescue body weight gain and adipocyte size increase in response to HFD. We propose reduced lipogenesis and enhanced lipid β-oxidation as potential mechanisms linked to AT2R action in reducing obesity and its related metabolic disorders in females.