bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 644: The Endothelin B Receptor Attenuates Endothelin Contraction Via An Endothelium Independent Mechanism In Mesenteric Arteries Of (mRen2)27 Rats
Authors: Pulgar Victor M , Jeffers Anne B , Chappell Mark C , Aileru Azeez A
Published: July 2021

Endothelin-1 (ET-1) and Angiotensin II (Ang II) are important modulators of vascular tone. Contraction to ET-1 is dependent on the activation of ETA and ETB receptors. The contractile receptor ETA is localized in the smooth muscle cells whereas ETB receptors are predominantly found in the endothelium. We investigated the role of ETB receptors in ET-1 contraction in a model of Ang II-dependent hypertension, the (mRen2)27 rat. Third branch mesenteric arteries were isolated from 12 weeks old male Sprague-Dawley (SD) control and (mRen2)27 hypertensive rats and mounted on a wire Multi Myograph (Model 620, DMT) for determination of isometric force. Dose response curves to ET-1 (10
-11
-10
-7
M) were performed in arterial segments that were either intact, endothelium denuded or preincubated with the ETB specific antagonist BQ788 10
-6
M. Endothelial denudation was performed by passing a human hair through the arterial lumen and functional loss of the endothelium was confirmed by the absence of vasodilatation to acetylcholine. Maximal responses were expressed as percent of maximal response to 75mM KCl (%K
MAX
) and the sensitivity as pD
2
(pD
2
= -Log[EC
50
]). In SD control rats, addition of BQ788 significantly increased ET-1 sensitivity in intact arteries (8.9±0.1 vs 9.3±0.1, p<0.05) and the increase was abolished by endothelial denudation with no change in maximal response. However, the ETB blocker BQ788 increased the maximal tension in both intact (127±2 vs 154±13 %K
MAX
, p<0.05); and denuded (156±9
vs
188±12 %K
MAX
, p<0.05) arteries from (mRen2)27 rats with no change in sensitivity. In summary, the present results demonstrate that endothelial ETB receptors influence vasodilatory function in control SD rats, but apparently attenuate the contraction to ET-1 in the (mRen2)27 rats. We conclude that the expression of ETB receptors in the endothelium and smooth muscle of resistance vessels in the (mRen2)27 rats may function as a compensatory mechanism to the increased blood pressure or activated angiotensin system found in this model of hypertension.
Abstract 70: Eplerenone Increases Dilation and the Diameter of Cerebral Penetrating Arterioles in Rats with Sustained Hypertension
Authors: Pires Paulo W , Jackson William F , Dorrance Anne M
Published: July 2021

Hypertension is linked to dementia in humans, as well as remodeling and dysfunction in large cerebral arteries. The effects of hypertension on cerebral microvessels, such as penetrating arterioles (PenA), are still unknown. These arterioles are the bottlenecks bridging the pial circulation to the deep parenchymal microcirculation, and they are vital for neurovascular coupling and functional hyperemia. Mineralocorticoid receptor (MR) antagonism reverses hypertension-induced changes in large cerebral arteries. Thus, we hypothesized that MR antagonism will improve PenA dilation and structure in adult rats with sustained hypertension. Twelve-week-old male stroke-prone spontaneously hypertensive rats (SHRSP) were treated with the MR antagonist eplerenone (EPL, 100mg/kg/day) or placebo for 6 weeks. PenA function and structure were studied by pressure myography. Data are means±SEM at 60mmHg intralumenal pressure, placebo vs EPL. Myogenic tone generation was not different between groups (%myogenic tone: 40.3±2.3 vs 41.3±2.0). PenA diameter after tone was higher in EPL (42.47±1.5 vs 49.21±2.4μm, p=0.02, t-test). Dilation of PenA to nifedipine (calcium channel blocker) was improved after EPL treatment, as evidenced by an increase in diameter change (10μM nifedipine: 13.44±1.94 vs 18.44±2.39μm, p<0.01, Two-way ANOVA) and PenA diameter (10μM nifedipine: 58.01±2.28 vs 68.58±3.74μm, p<0.01, Two-way ANOVA). PenA passive structure was improved in SHRSP+EPL, as evidenced by an increase in PenA diameter (65.67±1.17 vs 81.00±5.11, p<0.01, Two-way ANOVA) and lumen diameter (55.17±1.8 vs 73.00±5.00, p<0.01, Two-way ANOVA), and a reduction in the wall-to-lumen ratio (0.10±0.01 vs 0.07±0.01, p<0.01, Two-way ANOVA). PenA stiffness was increased after EPL (β-coefficient: 10.91±0.93 vs 14.84±1.83, p=0.03, t-test), but distensibility was unchanged (24.45±3.20 vs 21.88±2.68). These data suggest that MR antagonism improves the structure of cerebral arterioles and their responsiveness to calcium channel blockade in adult rats with sustained hypertension. These improvements have the potential to reduce the risk of developing vascular cognitive impairments and dementia in chronically hypertensive subjects.
Abstract 73: Nanoformulated Copper/Zinc Superoxide Dismutase: Alternative Therapeutic Strategy for Angiotensin II-dependent Neurogenic Hypertension
Authors: Savalia Krupa K , Manickam Devika S , Rosenbaugh Erin G , Tian Jun , Ahmad Iman , Kabanov Alexander V , Zimmerman Matthew C
Published: July 2021

Excessive production of superoxide (O2•-) in the central nervous system has been widely implicated in the pathogenesis of angiotensin II (AngII)-dependent neurogenic hypertension (HTN). Our group has tried to overcome the failed therapeutic potential of currently available antioxidants by utilizing nanoformulated copper/zinc superoxide dismutase (SOD1), so-called SOD1 nanozymes, that specifically scavenges intracellular O2•-. These nanozymes consist of SOD1 protein wrapped with cationic block copolymers followed by covalent cross-linking of the polycation template (cl-nano). We hypothesize that cl-nano delivers active SOD1 protein to neurons and can effectively decrease blood pressure in a mouse model of AngII-dependent neurogenic HTN. As determined by electron paramagnetic resonance (EPR) spectroscopy, cl-nano retains SOD1 activity and scavenges O2•- to levels comparable with native SOD1 protein in a cell-free environment (EPR arbitrary units: vehicle 1.12e6 ± 1.79e5; native SOD1 protein 4.45e4 ± 3.00e3; cl-nano 6.78e4 ± 1.74e3, p<0.05 vs. vehicle). Experiments to examine neuronal uptake of cl-nano, analyzed by western blot and SOD1 activity assays, reveal that cl-nano delivers active SOD1 to central neurons in culture (CATH.a neurons) more efficiently than native SOD1 protein following 1 hour treatment (SOD1 activity in units/mg protein: vehicle 336; native SOD1 protein 313; cl-nano 718). Furthermore, in vivo studies demonstrate that HTN established by chronic subcutaneous infusion of AngII (400 ng/kg/min) is significantly attenuated following a single intracerebroventricular (ICV) injection of cl-nano for up to 7 days (mean arterial pressure (MAP) in mmHg: pre-AngII 87 ± 3; 9 days post-AngII 138 ± 6; 7 days post-ICV injection of cl-nano 112 ± 4, p<0.05 vs. pre-ICV injection). These data provide evidence for the efficacy of nanoformulated SOD1 in counteracting excessive O2•- and decreasing MAP in AngII-dependent hypertensive mice when injected directly into the brain. Although further experiments must be performed with more clinically relevant routes of cl-nano administration, such as intravenous injection, this study supports the further development of cl-nano with SOD1 as an alternative therapeutic option for HTN.
Abstract 74: The Nox4 Inhibitor GKT137831, Prevents Aldosterone Production by Adipocytes and Protects Against Adipose Tissue Inflammation and Fibrosis in Obese Mice
Authors: Even Sarah , Nguyen Dinh Cat Aurelie , Rios Francisco J , Tayze Antunes T , He Ying , Callera Glaucia E , Carswell Andy , Jenkins Carol , Jandeleit-Dahm Karin , Cooper Mark , Szyndralewiez Cedric , Page Patrick , Montezano Augusto C , Touyz Rhian M
Published: July 2021

Aldosterone (aldo) plays an important role in obesity-associated cardiovascular risk. We demonstrated that aldo is produced by adipocytes, an effect associated with increased generation of reactive oxygen species (ROS). These processes are exaggerated in obesity. The relationship between adipocyte aldosterone and ROS is unclear. We postulated that Nox4-derived ROS is important for aldo production in adipocytes and leads to a pro-inflammatory phenotype in obesity. Studies were performed in db/m (lean) and db/db (obese) mice, treated with low (20mg/kg/day) or high dose (60mg/kg/day) GKT137831 (GKT, Nox4 inhibitor, 16 weeks). Epididymal (EVAT) and perivascular (PVAT) fat were collected. Plasma and adipocyte aldo were measured by ELISA. Adipose tissue fibrosis was evaluated by picro Sirius red staining and inflammatory mediators by immunostudies. Body weight was increased in db/db mice (61.8g vs control 33.5g), with no effect of GKT. Epididymal adiposity was increased in db/db mice (0.098g vs. 0.067g, p<0.05). Plasma aldo levels in db/db (pg/mL: 518 vs. 272g) and aldo levels in culture media from db/db adipocytes were increased (pg/mL/μg RNA: 1964 vs. 388), p<0.05. All effects decreased by high dose GKT. In PVAT, CYP11B2 gene expression was increased in db/db (2.6±0.8 vs control 1.1±0.1, p<0.05), an effect blocked by Nox4 inhibition. Gene expression of adipocyte differentiation marker, AP2, was increased (3.5±1.1 vs control 1.4±0.4) while anti-inflammatory marker adiponectin was decreased (0.7±0.1 vs control 1.3±0.2, p<0.05)) in obese mice. GKT decreased AP2 levels. Adipocyte-derived TNFα was increased in db/db (4.9±1.8 vs control 1.6±0.6, p<0.05), an effect blocked by GKT. Pro-collagen I, marker of fibrosis, was increased in db/db mice (132±11 vs control 87±4, p<0.05). Sirius red staining was exaggerated in EVAT from db/db mice, and decreased by Nox4 inhibition. In conclusion, Nox4 plays a role in regulating adipocyte-derived aldosterone and promotes a pro-inflammatory and profibrotic adipose phenotype in obese db/db mice. These findings suggest that adipocyte Nox4 links hyperaldosteronism and inflammation/fibrosis in adiposity and as such may be a putative therapeutic target for obesity-associated cardiovascular damage.
Abstract 75: Nrf2 Mediates the Antioxidant Effect of D2r Via Dj-1 in the Kidney
Authors: Cuevas Santiago , Yang Yu , Assico Laureano , Feranil Jun , Jones John , Armando Ines , Jose Pedro A
Published: July 2021

Renal dopamine D2 receptor (D2R) dysfunction is associated with increased oxidative stress and high blood pressure. We have reported that DJ-1 is regulated by D2R in the kidney. Nrf2 is a transcription factor that regulates the expression of several antioxidant genes. We hypothesize that Nrf2 is involved in the DJ-1-mediated regulation of reactive oxygen species (ROS) production in the kidney. DJ-1 and Nrf2 co-inmunoprecipitate in mouse kidney. Selective renal silencing of D2R in mice by the renal subcapsular infusion of D2R siRNA (~50%) decreases Nrf2 expression (63.4%±3.4, n=7, P<0.05), and silencing of D2R expression in mouse proximal tubule cells (mRPTCs) decrease Nrf2 promoter activity (53±2%, n=5, P<0.05). Silencing DJ-1 expression in mRPTCs, via siRNA, decreases the expression of DJ-1 (75±4.5%, n=4, P<0.05), Nrf2 (63.2±6%, n=3, P<0.05), NQO1 (71±1%, n=3, P<0.05), and GST Yc-2 (42±1%, n=4, P<0.05) and Nrf2 promoter activity (58±1%, n=5, P<0.05). Selective renal silencing of DJ-1 in mice by renal subcapsular infusion of DJ-1 siRNA decreases expression of DJ-1 (31±2.6%, n=3, P<0.05), Nrf2 (57±7.9%,n=3, P<0.05), NQO1 (46±9.3%, n=3, P<0.05) and GST Yc2 (28±2.2%, n=3, P<0.05) and increases blood pressure (BP). DJ-1
-/-
mice also have increased systolic BP (30.7±1%, P<0.01, n=5 under anesthesia) and renal expression of nitro-tyrosine (76.8±13.5%, P<0.05, n=5), and decreases the expression of Nrf2 (46.8±6.8%, n=5, P<0.05) and NQO1 (19.4±2.7%, n=5, P<0.05). Silencing Nrf2 expression mRPTCs via siRNA decreases the expression of Nrf2 (34±1.8%, n=4, P<0.05), NQO1 (29.1±0.5%, n=4, P<0.05), and increases Nox2 (137±23.7%, n=4, P<0.05), but the expression of Nox4 and DJ-1 are not affected. Quinpirole a D2R agonist, does not modify the expression of Nrf2, NQO1, GST Yc-2, and Nrf2 promoter activity in MPCTS. However, in the presence of H
2
O
2
, quinpirole increases Nrf2 promoter activity (220±58.4%, n=5, P<0.05) and an effect that is blocked by pretreatment with D2R antagonist L741,626. Therefore, the inhibitory effect of DJ-1 on renal ROS production is, in part, mediated by positive regulation of Nrf2. Moreover, D2R and DJ-1 are necessary for normal Nrf2 activity to keep a normal redox balance in the kidney.
Abstract 94: High Sucrose Intake-induced Endoplasmic Reticulum Stress, Inflammation and Fibrosis in hte Pancreas: Role of Trpv1
Authors: Wang Donna H , Zhong Beihua
Published: July 2021

Excessive consumption of sugar may serve as a primary and independent factor for increased morbidity of diabetes. We test the hypothesis that the transient receptor potential vanilloid type 1 (TRPV1) channels play a protective role against sugar-induced pancreatic endoplasmic reticulum (ER) stress, inflammatory cell infiltration, and tissue fibrosis. Wild-type (WT, C57BL/6) or gene-targeted TRPV1-null mutant (TRPV1
-/-
) mice were fed a normal (CON) or high sucrose diet for 22-23 weeks. TRPV1 ablation impaired glucose tolerance and suppressed glucose-induced insulin release when fed a CON diet (p<0.05). Sucrose intake further impaired glucose tolerance and insulin release in TRPV1
-/-
compared to WT mice (p<0.05). Sucrose intake increased pancreatic ER stress, showing elevated levels of chop protein content in both strains and GRP78, XBP-1, and CREB-2 in TRPV1
-/-
mice only (p<0.05). TRPV1 ablation increased levels of phosphorylated JNK (WT, 0.12±0.14; TRPV1-/-, 0.31±0.13) and decreased glut2 contents (WT, 1.38±0.24; TRPV1-/-, 0.83±0.09) in the pancreas when fed a CON diet (p<0.05). Sucrose intake further increased levels of pancreatic phosphorylated JNK (WT, 0.89±0.19; TRPV1-/-, 1.05±0.19) and decreased glut2 contents (WT, 0.39±0.04; TRPV1-/-, 0.26±0.16) in both strains (p<0.05). Sucrose intake increased pancreatic TGF-β levels in both strains with exacerbation in TRPV1-/- mice (p<0.05), and increased pancreatic phosphorylated Smad2/3 and collagen levels in TRPV1
-/-
mice only (p<0.05). Quantitative immunostaining revealed that sucrose intake enhanced pancreatic infiltration of CD3 positive cells in both strains and macrophage in TRPV1
-/-
mice only (p<0.05). Thus, TRPV1 ablation impairs insulin release and glucose tolerance at baseline possibly via increased pancreatic phosphorylated JNK signaling and decreased glut2 expression. Sucrose intake aggravates impairment in insulin release and glucose tolerance when TRPV1 is ablation, possibly via enhancing ER stress, macrophage infiltration, and activation of the TGF-β/Smad pathway leading to pancreatic fibrosis, suggesting preserving TRPV1 expression and function may attenuate pancreatic injury via alleviating inflammatory responses.
Abstract 95: Glucagon Like Peptide-1 (GLP-1) Does Not Cause Vasodilation Even When Dipeptidyl Peptidase 4 (DPP4) is Inhibited
Authors: Devin Jessica K , Pretorius Mias , Billings Frederic T , Nian Hui , Brown Nancy J
Published: July 2021

Glucagon-like peptide 1 (GLP-1) causes direct vasodilation in animal models. Dipeptidyl peptidase 4 (DPP4) inhibitors improve glycemic control in diabetic patients by preventing the degradation of GLP-1. The direct effect of GLP-1 in the human vasculature, and how it is altered by DPP4 inhibition, has not been reported. This study tested the hypothesis that intra-arterial infusion of GLP-1 causes dose-dependent vasodilation, and that DPP4 inhibition potentiates the forearm blood flow (FBF) response to GLP-1 by decreasing its degradation.

Eight healthy, non-obese (BMI<30 kg/m
2
) subjects, age 28-54 years old (3 female) participated in this double-blind, placebo-controlled crossover study. On study days separated by at least one week subjects received DPP4 inhibitor (sitagliptin 200 mg p.o.) or placebo, followed by infusion of GLP-1 in the brachial artery at graded doses (0.45-3.60 pmol/min) for 5 minutes per dose.

Sitagliptin significantly decreased plasma DPP4 activity (p<0.001 vs. placebo). Sitagliptin did not significantly affect baseline heart rate or baseline FBF. Baseline mean arterial pressure was significantly higher during sitagliptin than during placebo [87.13 ± 2.10 mmHg versus 84.75 ± 3.28 mmHg, p=0.037]. GLP-1 concentrations were significantly higher after sitagliptin (Left Figure; N=5). There was no effect of GLP-1 on FBF either in the presence or absence of sitagliptin. Moreover, there was no interactive effect of GLP-1 and sitagliptin on FBF (Right Figure).

GLP-1 does not cause vasodilation in healthy humans even when its degradation is inhibited. These data have implications for the cardiovascular effects of DPP4 inhibitors and GLP-1 receptor agonists.
Abstract 009: CETP inhibitors Torcetrapib, Dalcetrapib, and Anacetrapib induce adipocyte-derived Aldosterone production through Nox and STAT3 activation
Authors: Rios Francisco J , Neves Karla B , Nguyen Dinh Cat Aurelie , Even Sarah , Palacios Roberto , Jenkins Carol , Carswell Andy , Montezano Augusto C , Touyz Rhian M
Published: July 2021

Large clinical trials indicated that CETP inhibitors increase HDL levels, but had unexpected side effects, such as hyperaldosteronism and hypertension. Some CETP inhibitors appear to accumulate in adipocytes, which are a source of aldosterone (Aldo). As such, we questioned whether CETP inhibitors influence Aldo production in adipocytes and assessed the role of reactive oxygen species (ROS) and STAT3 in this process. Human adipocytes (SW872), expressing CETP, were studied and compared to mouse adipocytes (3T3-L1), lacking CETP. Cells were treated with torcetrapib (TOR), dalcetrapib (DAL), or anacetrapib (ANA). To evaluate the role of ROS, cells were pre-treated with N-acetylcystein (NAC-ROS scavenger), ML171 and GKT136901 (Nox1/4 inhibitor, respectively), and Rotenone (Rot, mitochondrial ROS inhibitor). ROS were measured by lucigenin and amplex red. Aldo production, measured by ELISA, was induced by TOR (668pg/mL), DAL (348pg/mL) and ANA (434pg/mL) (p<0.05 vs control 196pg/mL); an effect blocked by NAC. TOR, DAL and ANA increased superoxide (2-fold) and H2O2 (1.5-fold) production, and STAT3 phosphorylation (p<0.05). TOR increased Nox1, 4 and 5 (2-fold); DAL increased Nox4 (2-fold) and, ANA increased Nox1 and 4 (3-fold). In TOR treated cells, Aldo production was inhibited by GKT (62%), ML171 (62%), Rot (45%), and S3I-201 (STAT3 inhibitor, 66%). DAL-induced Aldo production was blocked by ML171(62%) and S3I-201 (74%). In ANA treated cells, Aldo production was inhibited by GKT (94%), ML171 (83%), and Rot (66%). We also assessed effects of CETP inhibitors on chemerin production, a novel adipokine associated with hypertension. TOR, but not ANA or DAL stimulated chemerin production (331pg/mL vs control 154pg/mL p<0.05); an effect blocked by GKT (95%) and S3I-201 (100%). In mouse adipocytes, TOR, DAL, and ANA induced Aldo (453pg/mL, 375pg/mL, and 445pg/mL vs cont 253pg/mL, p<0.05) and ROS production (1.7-fold, p<0.05). In conclusion, CETP inbitors induced aldosterone production in human and mouse adipocytes through redox-related mechanisms involving STAT3, Noxs and mitochondria. These novel findings have important clinical significance and may explain, in part, the hyperaldosteronism and hypertension reported in CETP clinical trials.
Abstract 012: Mineralocorticoid Receptors Mediate Western Diet-Induced Macrophage Polarization and Aortic Stiffness
Authors: Jia Guanghong , Aroor Annayya R , DeMarco Vincent G , Sun Zhe , Bostick Brian P , Meininger Gerald A , Jaffe Iris , Sowers James R
Published: July 2021

Mineralocorticoid receptors (MRs) are expressed in vascular smooth muscle cells (VSMCs), macrophages, and endothelial cells (ECs) and signaling through these receptors may play a key role in development of aortic stiffness. Accordingly, we hypothesized that these vascular MRs are involved in the western diet (WD) - induced macrophage polarization and aortic stiffness. The stiffness of primary cultured VSMC and tissue ECs or VSMCs from mice was measured using cell nano-indentation with atomic force microscopy (AFM). Both Ang II (100 nM) and aldosterone (10 nM) increased VSMC stiffness in vitro. Also, B57 mice fed a WD for 16 weeks increased EC and VSMC stiffness as determined by in vivo pulse wave velocity and ex vivo AFM techniques. The elevated aortic stiffness was accompanied by increased mRNA expression of M1 markers MCP-1, CD 86, and CD11b. However, treatment with low dose spironolactone (Sp) (1.0 mg/kg/day), a dose not affecting blood pressure, significantly attenuated WD-induced increases in stiffness in murine ECs and VSMCs. Sp increased M2 marker IL10 and the expression ratio of M2/M1 marker genes in aorta by real time PCR. Interestingly, mice with EC MR knockout did not develop WD- induced aortic vasodilation dysfunction under both Ach and SNP (10-9-10-4 mol/L) stimulation. Furthermore, low dose Sp inhibited WD- caused up-regulation of ERK 1/2 and down-regulation Akt/eNOS signaling pathways in the aorta. These findings support the notion that increased vascular and macrophage MR signaling play a key role in macrophage polarization and associated aortic stiffness that result from consuming a WD high in fructose and fat.
Abstract 020: Angiotensin Ii-induced Hypertension Has Significant Effects On Proliferation, Differentiation And Engraftment Efficacy Of Hematopoietic Stem Cell.
Authors: Kim Seungbum , Cogle Christopher R , Zingler Michael , Scott Edward W , Raizada Mohan K
Published: July 2021

Cyclosporin and other immunosuppressive drugs are used in bone marrow (BM) transplantation to increase engraftment efficacy and reduce rejection. However, their chronic clinical use is closely associated with increase in blood pressure and development of hypertension (HTN). Despite these significant side effects, little is known about the influence of high blood pressure on hematopoietic stem cell (HSC) and BM activity. Thus, the objective of this study was to investigate if Ang II induced HTN exerts influence on HSC proliferation, differentiation and engraftment in the BM. Infusion of Ang II (1000ng/kg/min for 21 days) and establishment of HTN resulted in increased proliferation of HSCs as evidenced by 87% increase in Sca-1+, c-Kit+, Lin- (SKL) HSC and 254% increase in CD150+, CD48- SKL long-term HSC in the BM. Furthermore, this was associated with significant accumulation of monocytes in both BM (30% increase) and spleen (250% increase). These changes in HSC and inflammatory cells were blocked by co-infusion of Ang II and losartan (60mg/kg/day), In order to understand the effect of Ang II on HSC homing, GFP+ HSCs were injected into the lethally irradiated and saline or Ang II infused C57BL6 mice. FACS analysis of GFP+ donor derived cells showed that hypertensive animals has poor engraftment efficacy on both BM and peripheral blood (35-52% compared to saline controls). Time-lapse in vivo imaging of mouse tibia showed that HSC failed to engraft to the BM osteoblastic niche in hypertensive mice. HSCs pretreated with 100nM Ang II for 18 hours in vitro also showed significantly diminished ability (16% compared to control) to engraft in normal recipient mice. These observations demonstrate that 1) chronic Ang II induced HTN regulates HSC proliferation and impairs the homing ability and reconstitution potential of HSC in BM, 2) These effects are mediated by the AT1 receptor on HSC and 3) Ang II accelerates HSC differentiation leading the increase of inflammatory cells in BM and spleen. The results suggest that hypertensive status and BP control should be strictly taken into account in consideration for BM transplantation.
Abstract 029: Experimental Modest Weight Gain Increases 24-h Blood Pressure in Lean Healthy Subjects: Implications of Increased Visceral Fat
Authors: Covassin Naima , Singh Prachi , Sert-Kuniyoshi Fatima H , Romero-Corral Abel , Davison Diane E , Lopez-Jimenez Francisco , Jensen Michael D , Somers Virend K
Published: July 2021

Introduction:
Obesity is linked with heightened vulnerability to cardiovascular diseases including hypertension. Nonetheless, mechanistic studies addressing the effects of experimental weight gain on blood pressure are lacking. We sought to examine whether experimental weight gain raises ambulatory blood pressure in healthy individuals and identify any relationship between changes in blood pressure and changes in regional fat distribution.

Methods:
Twenty-six normal weight subjects were randomized to a 8-week period of weight gain through overfeeding (N=16; age 30.4±6.6 years, BMI 23.5±3.5 kg/m
2
) or weight maintenance (N=10; age 27.1±7.7 years, BMI 23.6±2.7 kg/m
2
). Measurements of body composition by dual-energy X-ray absorptiometry and abdominal computed tomographic scans and 24-h ambulatory blood pressure monitoring were obtained at baseline and at follow-up.

Results:
Overfeeding resulted in an increase in body weight of 3.7±1.5 kg (
p
<0.001) in weight gainers, with increments seen in total (21994±8247.4 to 25180.7±8563.1 gr,
p
<0.001), visceral (61.6±32.7 to 75.5±30.9 cm
2
,
p
=0.002) and subcutaneous fat (135.5±77.4 to 167.9±82.9 cm
2
,
p
<0.001). No changes occurred in the maintenance group. Weight gainers exhibited an increase in 24-h systolic blood pressure at follow-up (113.7±8 to 117.7±7.9 mmHg,
p
=0.009) and mean blood pressure (MAP) (85.1±4.9 to 86.8±5.1 mmHg,
p
=0.02), while blood pressure was unchanged in controls. Changes in MAP were positively correlated only with changes in visceral fat (rho=0.452,
p
=0.02) but not with changes in weight or any other body composition measure.

Conclusion:
Modest weight gain leads to marked elevation in 24-h blood pressure in lean healthy subjects. The association between increased MAP and abdominal visceral fat accumulation suggests that visceral deposition of adipose tissue may contribute specifically and mechanistically to the enhanced risk of hypertension associated with weight gain.
Abstract 034: Transient Receptor Potential Vanilloid-1 (TRPV1) Channels Mediate the Sympathoexcitatory and Pressor Responses to Central NaCl Stimulation
Authors: Stocker Sean D , Simmonds Sarah S , Browning Kirsteen N
Published: July 2021

Accumulating evidence indicates a high salt diet increases cerebrospinal fluid sodium concentration to elevate sympathetic nerve activity (SNA) and arterial blood pressure (ABP) in salt-sensitive hypertension. Recent evidence suggests TRPV1 channels underlie the ability of the brain to sense changes in osmotic pressure. The present study investigated whether the sympathoexcitatory response to intracerebroventricular (ICV) infusion of hypertonic NaCl was mediated by central TRPV1 channels. In Inactin-anesthetized Sprague-Dawley rats, ICV infusion of 1M NaCl (n=8) significantly increased lumbar SNA (140±12%), heart rate (20±7bpm), and mean ABP (9±1mmHg). ICV pretreatment with the broad-spectrum TRPV channel blocker ruthenium red (5mM, 2uL, n=8) eliminated the sympathoexcitatory response to ICV 1M NaCl: lumbar SNA (106±5%), heart rate (-6±7bpm), and mean ABP (1±2mmHg). Similarly, ICV pretreatment with the selective TRPV1 channel antagonist SB366791 (3mM, 2uL, n=8) prevented any change in lumbar SNA (106±3%), heart rate (4±6), and mean ABP (1±1mmHg). ICV injection of ruthenium red or SB366791 alone did not alter any variable. Surprisingly, ICV infusion of 1M NaCl (1μL) produced similar pressor responses in wild-type versus TRPV1-/- mice (12±2 vs 11±2 mmHg, n>4). Pretreatment with SB366791 attenuated the pressor response in wild-type but not TRPV1-/- mice. In vitro patch clamp recordings of neurons in the ventral lamina terminalis reveal that increases in bath osmolality produced dose-dependent membrane depolarization (5 mOsm: 5.0±0.63 mV, 10 mOsm: 6.1±1.24 mV, 15 mOsm: 12.4±2.1; n=5, P<0.05) and increase discharge frequency (5 mOsm: 208±32%, 10 mOsm: 246±55%, 15 mOsm: 274±101%, 20 mOsm: 496±53%; n=5, P<0.05). These responses were attenuated by bath application of the broad spectrum TRPV blocker ruthenium red. Collectively, these observations suggest that central TRPV channels mediate changes in neuronal excitability and cardiovascular responses to central osmotic stimulation.
Abstract 035: Cholinergic Agonists Protect from the Development of Hypertension during Chronic Inflammatory Disease
Authors: Mathis Keisa W
Published: July 2021

Systemic lupus erythematosus (SLE) is an autoimmune disorder with prevalent hypertension. Previous studies using a genetic mouse model of SLE (NZBWF1) suggest chronic inflammation is an important contributor to SLE hypertension. A novel neuroimmune pathway involving the α7 subunit of the nicotinic acetylcholine receptor (α7nAChR) suppresses splenic cytokine release and reduces systemic inflammation upon stimulation. To test whether activation of this ‘cholinergic anti-inflammatory pathway’ at the level of the α7nAChR attenuates the development of hypertension during SLE, female SLE and control (NZW) mice were infused with nicotine hydrogen tartrate salt (2 mg/kg/day, SC) or saline for 7 days. Nicotine-treated SLE mice had lower splenic protein expression of TNF-α and IL-6 (normalized to β-actin) relative to saline-treated SLE mice (1.09±0.06 vs. 1.37±0.06 and 0.36±0.04 vs. 0.55±0.10; all p<0.05), suggesting efficacy of the therapy. Mean arterial pressure (MAP; mmHg) was increased in SLE mice compared to controls (140±4 vs. 114±2; p<0.001). Nicotine prevented the rise in MAP in SLE mice (129±4; p=0.022), but not controls (121±3). This protection from hypertension coincided with a 46±5% lower renal cortical TNF-α in nicotine-treated SLE mice compared to saline-treated SLE mice (0.39±0.04 vs. 0.73±0.18), which is important because it has been previously shown that renal TNF-α plays a mechanistic role in the development of hypertension during SLE. Because nicotine acts on both ganglionic and peripheral cholinergic receptors, in a subsequent study mice were administered the selective α7nAChR agonist, PNU-282987 (0.38 mg/kg/day, IP), or vehicle for 28 days. PNU-282987-treated SLE mice had lower splenic protein expression of TNF-α and IL-6 relative to saline-treated SLE mice (0.33±0.01 vs. 0.54±0.03 and 0.40±0.08 vs. 0.86±0.05; all p<0.05). MAP was increased in SLE mice compared to controls (138±2 vs. 122±5). PNU-282987 prevented the rise in MAP in SLE mice (128±4), but not controls (125±5). These data suggest the anti-inflammatory effects of cholinergic agonists may protect from SLE hypertension and that the cholinergic anti-inflammatory pathway may be an important target in hypertensive patients with chronic inflammatory diseases.
Abstract 046: High Fructose Intake Increases Phosphorylation and Trafficking of the Na/K/2Cl Cotransporter (NKCC22) In Rat Thick Ascending Limbs
Authors: Ares Gustavo R , Haque Mohammed Z , Ortiz Pablo A
Published: July 2021

A third of the US population consumes 20-40% of their caloric intake from added sugars with half of those calories from fructose. Fructose consumption is linked to salt-sensitive hypertension in humans and rodents. We found that feeding Sprague-Dawley rats 20% fructose in their drinking water did not increase blood pressure unless a high salt diet was added. The thick ascending limb (TAL) reabsorbs 25% of filtered NaCl, primarily
via
NKCC2. NKCC2 activity is increased by enhanced expression at the apical membrane and phosphorylation at Thr
96/101
. We found enhanced NKCC2 activity and trafficking in genetic models of salt-sensitive hypertension. However, the effect of fructose on NKCC2 regulation is unknown. Thus, we hypothesized that a fructose enriched diet stimulates NKCC2 activity. Sprague-Dawley rats were fed control diet or 20% fructose in drinking water for 1 week. TALs were isolated and phosphorylated and total NKCC2 was measured by surface biotinylation followed by western blot. A fructose-enriched diet increased surface-to-intracellular NKCC2 ratio by 60 ± 23% (
p<
0.05) and increased NKCC2 phosphorylation at Thr
96/101
by 8.02 ± 2.67 fold (
p
<0.05). NKCC2 phosphorylation at the plasma membrane was also increased by 4.5 fold (
p
<0.05). Total NKCC2 expression was reduced by 40.1 ± 8.9% (
p<
0.05). Since phosphorylation of NKCC2 at Thr
96/101
is mediated by STE20- and SPS1-related proline and alanine-rich kinases (SPAK) and oxidative stress-responsive kinase 1 (OSR1), we studied whether fructose stimulates expression and/or activity of SPAK/OSR1. Total SPAK/OSR1 expression was not enhanced by one week of a fructose diet. However, phosphorylation at Ser373 was enhanced by 2.8 ± 0.3 fold (
p<
0.05). We concluded that a fructose enriched diet increases phosphorylation and trafficking of NKCC2, enhancing its accumulation at the apical membrane. Moreover, a fructose diet increased SPAK/OSR1 kinases phosphorylation, suggesting they may be responsible for the enhanced NKCC2 phosphorylation. Our data suggest that a fructose-enriched diet promotes salt-sensitive hypertension in part by stimulating NKCC2 and TAL-dependent NaCl reabsorption.
Abstract 052: Alpha1a-Adrenoceptor Genetic Variant-Triggered Hyperproliferation in Cardiovascular Cells is Mediated by Novel Interacting Protein Spinophilin
Authors: Babaeva Ekaterina , Gradinaru Irina , Schwinn Debra A , Oganesian Anush
Published: July 2021

Objectives:
Alpha1-Adrenergic Receptors (α1ARs), members of the G protein-coupled receptor (GPCR) superfamily, play a major role in regulating cardiovascular (CV) function. Recently, we discovered that naturally occurring human α1aAR-G247R (247R) genetic variant, identified in the 3rd intracellular loop (3iL) of the receptor in highly hypertensive patient, triggers constitutive hyperproliferation in fibroblasts, cardiomyoblasts and smooth muscle cells (SMC). Specific proteins mediating this signaling remain unknown. Spinophilin (SPL) is a ubiquitously expressed protein controlling GPCR signaling by binding its 3rd intracellular loop (3iL). We hypothesize that SPL mediates α1aAR signaling and examined whether SPL directly interacts with α1aAR-WT (WT) and 247R.

Methods:
Cells were co-transfected with HA-α1ARs and full length Myc-SPL or its fragments to determine SPL domains responsible for binding to α1ARs. Cell lysates were co-immunoprecipitated with HA-tag antibodies. SPL levels were analyzed by Western blotting. SPL knockdown experiments were performed by transiently transfecting cells with SPL or scrambled siRNA, cell proliferation was determined by cell counting.

Results:
We demonstrate a distinct interaction of SPL with WT and 247R, WT interaction being the strongest. Different domains of SPL differentially interact with WT or 247R. SPL 1-480aa fragment interacts stronger with WT indicating interaction with 3iL, while SPL 480-817 fragment interacts stronger with 247R. Endogenous SPL levels are increased in 247R cells compared to WT or control cells. Inhibition of SPL expression in SMC with siRNA reduces 247R-triggered hyperproliferation to near normal levels and has no effect in WT cells.

Conclusions:
We identified SPL as a novel interacting protein involved in mediating intracellular signaling of α1aAR and its genetic variant in CV cells. Different domains of SPL differentially bind to WT or 247R indicating that SPL has a distinct role in regulation of their signaling pathways. Our findings also reveal that SPL is critical for 247R-triggered EGFR transactivation pathways. Thus SPL could be considered as a potentially novel target in α1aAR-mediated cardiovascular disorders.
Abstract 054: Mutations In Pde3a Explain Mendelian Hypertension With Brachydactyly
Authors: Maass Philipp , Aydin Atankan , Luft Friedrich C , Toka Hakan , Hollfinger Irene , Wefeld-Neuenfeld Yvette , Bartels-Klein Eireen , Mühl Astrid , Klussmann Enno , Toka Okan , Bähring Sylvia
Published: July 2021

We identified Mendelian type E brachydactyly (BDE) with hypertension in six different families from Turkey, America, France, Canada and South Africa. The two phenotypes, hypertension and BDE, invariably coincide. The blood pressure in affected individuals increases with increasing age, the mean arterial blood pressure at 50 years exceeds 150 mm Hg, resulting in stroke. Earlier we suggested that a chromosomal rearrangement on 12p could be responsible. We now used whole-genome sequencing and discovered six different so far unknown missense mutations in the gene encoding PDE3A. The mutations are not identical, but adjacent to each other. Each is responsible for an amino acid substitution in a conserved portion of the protein. The mutations were not present in any non-affected family members, in more than 200 additional controls, and not found in the “1000 genome” project. We performed in vitro cell transfections and found that mutated PDE3A showed gain-of-function with increased cAMP hydrolysis. Mesenchymal stromal cell-derived vascular smooth muscle cells and chondrocytes gave insight into the molecular pathogenesis; PDE3A and vasodilator-stimulated phosphoprotein (VASP) were differently phosphorylated and parathyroid hormone-related peptide (PTHrP) was dysregulated. Our preliminary results reveal that the mutated PDE3A enzymes show gain-of-function with increased cAMP hydrolysis and sensitivity to cGMP inhibition. We suggest that the mutations are responsible for both phenotypes. This Mendelian hypertension is the first that is not attributable to increased sodium reabsorption in the distal nephron.
Abstract 059: Additive effect of the GSTM1(0) Null and APOL1 High Risk Alleles in the African American Study of Kidney Disease (AASK) Trial
Authors: Bodonyi-Kovacs Gabor , Ma Jennie Z , Lipkowitz Michael S , Kopp Jeffrey B , Winkler Cheryl A , Le Thu H
Published: July 2021

Background:
Apolipoprotein L-1 (
APOL1
) G1 and G2 alleles are strongly associated with end stage renal disease (ESRD) in non-diabetic African American (AA) patients. Glutathione-S-transferase Mu 1 (
GSTM1
) null allele,
GSTM1(0)
, is also associated with accelerated progression of chronic kidney disease (CKD) in hypertensive AA patients. We hypothesized that the
GSTM1(0)
and
APOL1
high risk alleles interact to influence the clinical outcomes in the AASK Trial participants.

Methods:
A total of 682 patients were classified into the combination of homozygous null for
GSTM1
(0/0)
, heterozygous
(1/0)
, and homozygous active
(1/1)
and
APOL1
high (
High
) and low (
Low
) risk genotypes. The survival differences among the six groups were assessed by log-rank test and in the Cox regression for time to the event of glomerular filtration rate (GFR, 50% or 25ml/min/1.73m
2
decline), dialysis, or death, or their composite events. Regression analyses were adjusted for age, gender, proteinuria, baseline GFR, MAP, CVD, drug class, and population admixture.

Results:
The 6 groups differed significantly in the time to composite events of GFR, or dialysis or death (log-rank p=0.0033) (Fig. 1). Compared to
(1/1)_Low
group, the hazard ratios (HR) for the composite events were 2.6 (95% CI 1.27 – 5.32, p=0.009) and 2.4 (95% CI 1.15 – 4.99, p= 0.019) for the
(0/0)_
Low and the
(1/0)_High
groups, respectively. The HRs for the other groups did not reach statistical significance [
(1/1)_High
(HR 1.2, p = 0.73),
(1/0)_Low
(HR 1.78, p = 0.079), and
(0/0)_High
(HR 1.91, p = 0.097)].

Conclusion:
GSTM1(0)
and
APOL1
high risk alleles have independent and additive deleterious effects on CKD progression among hypertensive AAs.
Abstract 060: Adipose Derived Stem Cell (ADSC) Therapy Prevents The Progression Of Hypertension And Improves Cardiac Functions
Authors: Qi Yanfei , Rathinasabapathy Anandharajan , Zhang Juan , Zingler Michael , Wong Fong , Petersen John W , Katovich Michael J , Raizada Mohan K , Pepine Carl J
Published: July 2021

Despite major advances in pharmacological and interventional therapies, 30% of hypertensive patients do not achieve their blood pressure (BP) target and have apparent treatment resistant hypertension (TRH). Thus, innovative strategies are urgently needed for the control of TRH considering its prevalence and deleterious complications. The objective of the current study was to evaluate effects of ADSC on BP in spontaneously hypertensive rats (SHR). The rational for using ADSC is that they have multipotent differentiation capacities and potent anti-inflammatory effects. Moreover, autologous ADSC are an attractive therapeutic option for clinical trials based on availability, less invasive isolation procedure, and safety in several hundred patient studies. ADSC were isolated from inguinal adipose tissue of a normotensive WKY rat, characterized using flow cytometry (CD44
+
,CD90
+
,CD34
-
and CD45
-
), and their differentiation ability to adipocyte was confirmed by Oil Red “O” staining. Basal systolic BP for SHR at 13 week age was 191.6 ± 5.6 mmHg measured by a tail-cuff method. SHR animals were then randomly assigned to control and ADSC-treated groups. ADSC (5х10
6
cells/animal) was intravenously administered through jugular vein into treated SHR. Three weeks after treatment, left ventricular function was assessed using echocardiography and direct BP was measured using a Millar catheter. The SHR control exhibited 82% increase in systolic BP (198.4 ± 25.9 vs WKY: 109.0 ± 1.6, mmHg) and 14% elevation in ventricular mass (3.3 ± 0.2 vs WKY: 2.9 ± 0.1, g/kg). Furthermore, the SHR control demonstrated 42% increase in deceleration time of mitral valve (58.1 ± 6.0 vs WKY: 40.8 ± 2.3; ms) and 21% decrease in ejection fraction (69.9 ± 4.2 vs WKY: 88.2 ± 1.3, %). Single injection of ADSC resulted in a 28% decrease in systolic BP (142.5 ± 25.8mmHg), 18% reduction in ventricular mass (2.7 ± 0.3g/kg), 36% decrease in deceleration time of mitral valve (36.9 ± 16.3ms), and 17% increase in ejection fraction (80.0 ± 3.4%). Collectively, our data demonstrate that ADSC arrested the progression of hypertension and associated left ventricular dysfunction in the hypertensive animals. It warrants additional study on ADSC intervention as a potential alternative treatment for hypertension.
Abstract 061: Prevention Of Oxidative Stress In The Paraventricular Nucleus (PVN) Does Not Attenuate Hypertension During High Fat Diet (HFD), Despite Blunting Weight Gain.
Authors: Lob Heinrich E , Song Jiunn , Butler Scott D , Mark Allyn L , Davisson Robin L
Published: July 2021

We recently reported that PVN-specific deletion of the NADPH oxidase subunit p22
phox
attenuates oxidative stress in this brain region and prevents diet-induced obesity (DIO) in mice. In addition to its importance in the regulation of energy homeostasis, the PVN is a well-described cardiovascular regulatory center. Since increased oxidative stress in the central nervous system is causally linked to both hypertension and obesity, we hypothesized that attenuating oxidative stress in the PVN would reduce sympathetic nerve activity (SNA) and improve hypertension during HFD. In mice harboring loxP-sites flanking the

p22
phox

gene (p22
phox/flox
), we tested if p22
phox
deletion in the PVN reduces arterial pressure (AP) and heart rate (HR) during HFD. For these experiments, p22
phox/flox
mice underwent bilateral PVN-targeted injection of adenoviral vectors encoding Cre recombinase (AdCre) or control LacZ (AdLacZ). Following 1-week recovery, mice were started on a 42% HFD or normal chow (NC). After 7 weeks of HFD, radiotelemeters were implanted and 3 weeks later (after 10 weeks HFD) we recorded AP and HR. HFD caused a significant elevation in AP (AdLacZ NC: 104 ± 2.8 mmHg vs. AdLacZ HFD 113 ± 0.6 mmHg n = 3-5, P < 0.05) and HR (AdLacZ NC: 487 ± 7 bpm vs. AdLacZ HFD 537 ± 12 bpm; n = 3-5, P < 0.05). Importantly, despite a significant reduction in body weight after deletion of p22
phox
in the PVN (AdLacZ HFD: 32 ± 1.2 g, vs. AdCre HFD 27 ± 0.6 g, P < 0.05), removal of oxidative stress in the PVN did not blunt AP (AdLacZ HFD 113 ± 0.6 mmHg vs. AdCre HFD: 112 ± 2.0 mmHg, n = 5, P > 0.05) or HR (AdLacZ HFD 537 ± 12.3 bpm vs. AdCre HFD: 540 ± 6.3 bpm, n = 5, P > 0.05). We utilized power spectral analysis of AP variability to assess SNA in mice during NC and HFD. We found that HFD elevated SNA similarly in AdLacZ and AdCre mice (1.7-fold and 1.5-fold, respectively vs. NC mice, P > 0.05). Thus, while our data demonstrate that oxidative stress in the PVN is a major player in the regulation of metabolism and adiposity, these results suggest that PVN oxidative stress does not contribute to increased SNA, AP or HR during HFD. While the underlying mechanism(s) of this divergence is currently being investigated, these data emphasize that there is differential central neural regulation of cardiovascular and metabolic responses to HFD.
Abstract 067: Downregulation Of Vascular Factor-erythroid 2-related Factor-2 And Associated Antioxidant Enzymes In Hypertension
Authors: Lopes Rheure A , Neves Karla B , Montezano Augusto , Tostes Rita , Touyz Rhian
Published: July 2021

Oxidative stress plays an important role in vascular dysfunction in hypertension. While mechanisms regulating vascular pro-oxidants are emerging, there is a paucity of information on anti-oxidant systems. Factor-erythroid 2-related factor-2 (Nrf2) is a master regulator of antioxidants and its role in hypertension remains elusive. We assessed vascular Nrf2 in hypertension by studying mesenteric vessels and VSMCs from WKY and SHRSP rats. Cells were stimulated with Ang II (10-7M) in the absence/presence of Nrf2 activators (bardoxolone or L-sulforaphane). ROS generation was assessed by chemiluminescence and amplex red. mRNA expression of anti-oxidant enzymes was assessed by qPCR. Nrf2 activity was analyzed by ELISA. Nrf2 activity was decreased in arteries (18%) and VSMCs (48%) in SHRSP (p<0.05 vs WKY). mRNA levels of antioxidant enzymes were reduced in SHRSP (SOD 1 (64%), catalase (60%), peroxiredoxin 1 (75%) and glutathione peroxidase (54%) Ang II increased Nrf2 activity in VSMCs from WKY (197%, 4h) and SHRSP (44%, 4h) (p<0.05, vs. vehicle). This was associated with increased antioxidant mRNA expression in WKY rats (SOD1-32%, catalase-42%, thioredoxin-71%, peroxiredoxin 1-90%, quinone oxidoreductase-84%; p<0.05 vs. vehicle) but not in SHRSP. ROS production and glucose-6-phosphate dehydrogenase (source of NADPH) mRNA levels were increased in SHRSP. Ang II-induced ROS generation in VSMCs from WKY and SHRSP was blocked by Nrf2 activators. Vascular function assessment, by wire myography, demonstrated that increased contractility (Emax Phe: WKY 113.4±5,67 vs. SHRSP 159.0±8.29) and decreased endothelial-dependent relaxation (Emax ACh: WKY 88.7±3.13 vs. SHRSP 74.7±3.25, p<0.05) in SHRSP were corrected by bardoxolone and L-sulforaphane. In conclusion, vascular dysfunction in SHRSP is associated with oxidative stress, decreased Nrf2 activity and reduced Nrf2-regulated antioxidant enzymes. A similar molecular phenotype was observed in Ang II-stimulated VSMCs. Nrf-2 agonists ameliorated vascular dysfunction in SHRSP. Our findings suggest that Nrf-2 downregulation may contribute to redox-sensitive vascular dysfunction and could be a therapeutic target in hypertension. Financial Support: ScWB.