bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract P198: Differential Regulation of Endothelial Nitric Oxide Synthase Phosphorylation by Protease-Activated Receptors in Adult Human Endothelial Cells
Authors: Tillery Lakeisha C , Motley-Johnson Evangeline D
Published: July 2021

Protease-activated receptors (PARs) have been shown to regulate endothelial nitric oxide synthase (eNOS) through the activation of specific sites on the enzyme. It has been established that phosphorylation of eNOS-Ser-1177 leads to the production of the potent vasodilator nitric oxide (NO), and is associated with PAR-2 activation; while phosphorylation of eNOS-Thr-495 decreases NO production, and is coupled to PAR-1 activation. In this study, we demonstrate a differential regulation of the eNOS/NO pathway by the PARs using primary adult human coronary artery endothelial cells (HCAEC). Thrombin and the PAR-1 activating peptide, TFLLR, which are known to phosphorylate eNOS-Thr-495 in bovine and human umbilical vein endothelial cells, phosphorylated eNOS-Ser-1177 in HCAECs, and increased NO production. The PAR-1 responses were blocked using SCH-79797, a PAR-1 inhibitor, and L-NAME was used to inhibit NO production. A PAR-2 specific ligand, SLIGRL, which has been shown to phosphorylate eNOS-Ser-1177 in bovine and human umbilical vein endothelial cells, primarily regulated eNOS-Thr-495 phosphorylation and suppressed NO production in the HCAECs. PAR-3, known for its non-signaling potential, was activated by TFRGAP, a PAR-3 mimicking peptide, and only induced phosphorylation of eNOS-Thr-495 with no effect on NO production. In addition, we confirmed that PAR-mediated eNOS-Ser-1177 phosphorylation was calcium-dependent using the calcium chelator, BAPTA, and eNOS-Thr-495 phosphorylation was mediated via Rho kinase using the ROCK inhibitor, Y-27632. These data suggest a vascular bed specific differential coupling of PARs to the signaling pathways that regulate eNOS and NO production that may be responsible for the modulation of endothelial function associated with cardiovascular disease.
Abstract P204: Food Restriction Increases the Cardiovascular Response Evoked by Angiotensin-[1-10] and Angiotensin-[1-7] in Female Fischer Rats
Authors: Souza Aline M , Martins Talita A , Santo Laura G , Chianca Deoclecio A , Sandberg Kathryn , Menezes Rodrigo C
Published: July 2021

Anorexia is associated with cardiovascular dysfunction including hypotension, bradycardia and cardiac arrhythmias. In a rat model of food restriction (FR)-induced hypotension and bradycardia, we previously demonstrated that the activity of endothelial α1 adrenergic receptors and brain type 1 angiotensin receptors (AT1R) were increased. The aim of this study was to determine the role of the vasodilator, angiotensin (Ang)-[1-7] and the precursor Ang-[I-10] in the periphery and brain. Methods: Female Fischer rats (220g, 4 mo) were maintained for 14 days on a FR diet, that was 40% of the control group (C) food intake. Cannulas were implanted in the lateral ventricle (LV) on day 7. On day 14, the animals were catheterized with a polyethylene tube in the femoral artery for recording of mean arterial pressure (MAP) and heart rate (HR). Results: After 14 days, body weight (g) [C: 202±2, n=13 vs. FR: 176±2, n=14; p<0.05], MAP (mm Hg) [C: 105±1, n=35 vs. FR: 102±2, n=36; p<0.05] and HR (bpm) [C: 386±5, n=35 vs. FR: 353±8, n=36, p<0.001) were reduced in the FR compared to control rats. Injection of Ang-[1-7] caused a greater increase in MAP (Δ mm Hg) in the FR compared to C group when delivered either into the LV (25pmol) (C: Δ -2±1, n=8 vs. FR: Δ 7±1, n=10; p<0.05) or the femoral vein (1910pmol) [C: Δ 1±1, n=9 vs. FR: Δ 3±1, n=6; p<0.05]. Moreover, there was a greater increase in HR (Δ bpm) in the FR compared to the C group after LV injection (C: Δ 9±3, n=8 vs FR: Δ 29±9, n=10; p<0.05). Injection of Ang-[I-10] caused a greater increase in MAP (Δ mm Hg) in the FR compared to C group when delivered into the LV (25pmol) (C: 5±1, n=7 vs. FR: 13±2, n=7; p<0.005) or i.v (1910pmol) (C: Δ 1±1, n=7 vs. FR: Δ 15±2, n=8; p<0.05); however, there was no effect on HR. This Ang-[l-10]-induced increase in MAP was attenuated by administering captopril into the LV, in both the FR and C groups, suggesting this increase in MAP was mediated by angiotensin converting enzyme. Conclusion: These results suggest that increased Ang-[1-7] and Ang-[I-10] activity both in the periphery and brain play a role in the increased sympathetic activity observed in anorexia.
Abstract P214: Meta-analysis of the Effect of Mineralocorticoid Receptor Antagonists on Proteinuria and Progression of Chronic Kidney Disease
Authors: Currie Gemma E , Taylor Alison H , Jardine Alan G , Preiss David J , Mark Patrick B , Delles Christian
Published: July 2021

Background/Objective:
ACE-inhibitors (ACE-I) and angiotensin receptor blockers (ARB) are standard of care for patients with chronic kidney disease (CKD) and persistent proteinuria. Mineralocorticoid receptor (MR) activation has numerous “off target” effects on the kidney and vasculature and a number of studies have evaluated the use of MR antagonists (MRAs) in CKD. We conducted a meta-analysis of randomised controlled trials of MRA in addition to ACE-I and/or ARB in CKD to evaluate the potential reno-protective effects and risks of hyperkalaemia.

Methods:
MEDLINE (1966-2014) and EMBASE (1947-2014) were searched using a pre-specified strategy, and unpublished data were obtained from original study authors where possible (8 authors, 10 studies). Studies including dialysis or transplant patients and studies of less than 4 weeks duration were excluded from analysis. Results were pooled using random effects meta-analysis.

Results:
Eighteen trials (1438 patients) were included. Addition of MRA reduced blood pressure (-5.9, [95% CI 9.6, -2.2mmHg] for systolic; -2.3, [95% CI -3.6, -1.0mmHg] for diastolic) and weighted mean protein/albumin excretion (- 40.6%) at the cost of a small reduction in glomerular filtration rate (GFR) (-3.4, [95% CI -5.7, -1.1 ml/min/1.73m
2
]). There was a 3-fold higher relative risk of hyperkalaemia above predefined trial limits (RR 3.21, [95% CI 1.19, 8.71]) equating to number needed to harm over one year of 23 [95% CI 7-267]. Average potassium increase was 0.21mmol/L [95% CI 0.08-0.33]. No studies reported life threatening hyperkalaemia events or hospitalisations as a result of hyperkalaemia. Baseline creatinine or diabetes status had no effect on hyperkalaemia risk in our analysis (
p
=0.15 for creatinine;
p
=0.38 for diabetes status).

Conclusion:
Addition of MRA is a promising therapeutic strategy for reducing blood pressure and proteinuria in patients with CKD, with a quantifiable risk of hyperkalaemia. This reduction in proteinuria could translate into reduced risk of progressive renal disease and cardiovascular events but appropriately designed larger studies with long term follow up are required.
Abstract P221: Physicians are More Prone to Causing White Coat Hypertension than Nurses or Cardiovascular Technicians: An Observational Study
Authors: Pandey Arjun K
Published: July 2021

Introduction:
Accurate assessments of blood pressure (BP) are critical for the effective diagnosis and treatment of hypertension. A substantial portion of patients labelled hypertensive have been shown to instead have White Coat Hypertension (WCH), where their BP is elevated exclusively when assessed in a clinic. In this study, we assessed whether the type of healthcare provider measuring individuals’ blood pressure impacted the incidence of WCH.

Methodology:
Following collection of baseline demographics, 106 participants had their BP measured by a physician, a nurse and a cardiovascular technician. The order of measurements was randomized. All healthcare providers used the same BP cuff for measurements and were instructed to measure BP following a standardized method. Following BP readings taken by healthcare providers, a 24-hour Ambulatory Blood Pressure Monitor (ABPM) was applied to all participants. The average of the daytime readings of the ABPM served as the control for this study.

Results:
Patients whose BP were greater than 140/90 mm Hg when measured by a healthcare provider, but whose control readings by ABPM were less than 135/85 mm Hg were classified as having WCH. Physicians caused 33% of participants (35 of 106) to have WCH. Nurses caused 23.5% of participants (25 of 106) to have WCH. Cardiovascular technicians caused 5.6% of participants (6 of 106) to have WCH. (p<0.0001).

Similar trends were observed based on analysis examining the percentage of accurate readings and the average of readings compared to the control ABPM, with technicians having the most accurate readings, nurses having moderately accurate readings, and physicians having the least accurate readings.

Conclusions:
The results of this study suggest that the incidence of WCH and BP measurement inaccuracy occur more frequently when BP is assessed by certain types of healthcare providers, potentially because patients may feel more anxious or stressed around these individuals. It may therefore be advisable for BP to be assessed by cardiovascular technicians, instead of nurses or physicians, to reduce the risk of White Coat Hypertension and inaccurate BP readings.
Abstract P223: How Does Patient Engagement and Gamification Correlate With Hypertension Control? Results From a Large-Scale Nationwide Network of Ambulatory Blood Pressure Kiosks
Authors: Siddiqui Khan , Goglia Ross
Published: July 2021

Background:
Ambulatory blood pressure (ABP) is known to provide prognostic information about cardiovascular disease better than office BP. Not much is known about the correlation of ABP control and patient engagement with gamification.

Objective:
To examine the relationship between ambulatory blood pressure and patient engagement with a nationwide ABP kiosk network.

Methods:
De-identified historic data from a nation wide ABP kiosk network (higi SH, llc www.higi.com) was analyzed from September 2012 to May 2015. At time of this abstract submission 9,926 ABP kiosks were deployed within the network. Only patients with initial BP readings in the hypertensive range and those who opted-in to share data for research purposes were included in the study. Level of engagement was defined as the total number of achievements, or badges, earned on the gamification platform. BP changes were defined as the difference between patients first and last reading on the kiosk network. Patient demographics, level of engagement with gamification platform and their ABP trends were analyzed and correlation measured.

Results:
A total of 153,092 patients qualified the inclusion criteria for the study. Mean age was 52 years with 56% (85,361) male and 44% (67,731) female. Almost half the patients were obese (49%, 74,587). The patients on the gamification platform earned a total of 898,130 achievements. There was a statistically significant difference in drop in systolic and diastolic blood pressure with number of achievements earned by patients (systolic BP: p-value < 0.0001, diastolic BP: p-value = 0.0033). Patients achieving greater than 20 achievements showed an average drop of systolic BP of 16.2 mmHg (p<0.01) and a drop of diastolic BP of 10.6 mmHg (p<0.01). Of the patients earning greater than 20 achievements 84.8% moved from hypertensive classification to normotensive classification.

Conclusion:
The results showed a statistically significant relationship between level of achievements earned on the gamification platform and lowering of blood pressure.
Abstract P228: Endothelial Dysfunction Markers and Coagulation Factors Are Elevated in Individuals With Hypertensive Emergency
Authors: Vilela-Martin Jose F , Andrade Days O , Cosenso-Martin Luciana N , Godoy Moacir F , Gregório Michele L , Souza Dorotéia R , Yugar-Toledo Juan C
Published: July 2021

Introduction:
Hypertensive crisis (HC) is characterized as sudden and symptomatic elevation of diastolic blood pressure (BP) > 120mmHg. HC can be classified in emergency (HE), condition that presents target organ damage (TOD) and hypertensive urgency (HU), situation without TOD. Evidences have shown that the coagulation markers and endothelial dysfunction (ED) play an important role in the pathogenesis of chronic elevation of BP. However, scarce studies have demonstrated the involvement of these markers in the complications of acute elevation of BP.

Objectives:
To characterize biochemical profile of HU and HE, and to evaluate the participation of C-reactive protein (CRP), intercellular adhesion molecule (ICAM-1) and coagulation factors (fibrinogen and PAI-1) in subjects with HC.

Methods:
It was a cross-sectional study with 74 normotensive (NT), 74 controlled hypertensive (CH), 50 with HU and 78 individuals with HE. It was used MULTIPLEX technique for evaluating the clotting factors. Analysis of variance was used for comparative study among the groups, with significant p-value<0.05.

Results:
The diastolic BP and heart rate were higher in the HC group (120mmHg and 85bpm, respectively) compared to the CH group (75mmHg and 68 bpm, respectively). Individuals with HE were older. Glycaemia was significantly higher in the HE group (113mg/dL) in comparison to NT and CH (91mg/dL and 98mg/dL, respectively; p<0.05). It was also higher in the HU group (109mg/dL) compared with NT. Potassium was significantly lower in HE group (4,2mEq/l) compared to NT, CH and HU groups (4.5, 4.4 and 4.4mEq/L, respectively). CRP, fibrinogen, and PAI-1 were significantly higher in patients with HE (0.75, 0.04 and 2.46, respectively) in comparison to NT (0.19, 0.01 and 2.1, respectively) and CH (0.14, 0.01 and 2.25, respectively) groups, except ICAM-1. Logistic regression showed that CRP and fibrinogen were markers for HC development with odds ratios of 2.6 (1.24 to 5.50) and 8.72 (4.07 to 18.68), respectively.

Conclusions:
Individuals with HC present biochemical changes. Markers of ED and coagulation factors are higher in EH group compared to controls. This suggests the role of ED markers and coagulation factors in the pathogenesis of acute hypertensive event.
Abstract P246: Salt-sensitivity of Angiogenesis Inhibition-induced Blood Pressure (BP) Rise: Role of Interstitial Sodium Accumulation?
Authors: Lankhorst Stephanie , Severs David , Marko Lajos , Rakova Natalia , Giménez-Rivera Vladimir A , Titze Jens , Müller Dominik N , Danser A H , van den Meiracker Anton
Published: July 2021

Objective:
Angiogenesis inhibition with the VEGF-inhibitor sunitinib, an established anti-cancer therapy, induces hypertension and proteinuria. Exposed to osmotic stress, the mononuclear-phagocyte-system (MPS) cells produces VEGF-C and exert homeostatic regulatory activity by promoting lymphatic Na+ drainage; interference with this process resulted in salt-sensitive hypertension. Therefore, we hypothesized that sunitinib, via blockade of the VEGF pathway, leads to Na+ accumulation in the skin and salt-sensitive hypertension.

Design and Methods:
In male WKY rats, mean arterial pressure (MAP) was monitored telemetrically during oral treatment with sunitinib (7 mg/kg.day, n=7-8) or vehicle (n=7-8) after a normal salt diet (NSD: 0.5-1.0% NaCl and tap water) or a high salt diet (HSD: 8% NaCl and saline water) for 2 weeks. After 8 days of sunitinib or vehicle administration, 24-h urine was collected. After sacrificing, blood was collected for biochemical measurements, skin for Na+ concentration ([Na+]) using dry-ashing, and ears for staining of MPS cells (CD68).

Results:
MAP during NSD was 101±0.9 mmHg. HSD increased MAP by 27±3 mmHg (P<0.05 vs. NSD). Sunitinib increased MAP by 15±1 mmHg during NSD (P<0.05 vs. NSD alone) and by 23±4 mmHg during HSD (P<0.05 vs. HSD alone). Although body weight, plasma [Na+] and plasma [cystatin-C] did not change in response to HSD and/or sunitinib, skin [Na+] increased from 90±1 (NSD) to 100±4 (HSD), and 108±4 mmol/L (HSD+sunitinib), respectively (P<0.01 for linear trend). Skin [Na+] correlated with MAP (r=0.57, P<0.01). Compared to NSD, proteinuria increased during HSD, rising further (P<0.05) with sunitinib. CD68 positive area increased during HSD from 0.29% to 0.43% (P<0.05), and increased even further with sunitinib (0.63%, P<0.05).

Conclusions:
Angiogenesis inhibition-induced hypertension is salt-sensitive. The parallel increases in BP and skin [Na+], in the face of unaltered plasma [Na+] and bodyweight, support the existence of a Na+-buffering compartment in the skin that may contribute to the salt-dependent volume and BP homeostasis during VEGF inhibition.
Abstract P254: Hypertension: A proposed Mechanistic Pathway for Arterial System Remodeling
Authors: Randall Otelio S , Lee Dexter , Retta Tamrat , Haddad Georges , Ketete Muluemebet , Kwagyan John , Xu Shichen
Published: July 2021

Arterial properties change during hypertension (HTN) development. The time course of the changes can be used as a marker of vascular remodeling in the development of HTN.

Study Aim:
To determine whether pressure induced changes in vascular properties result in alterations in pressure-flow patterns that impact the load for ventricular function: input & characteristic impedance,
A
rterial
C
ompliance (AC)= SV/PP (stroke volume)/(pulse pressure), AC=Tau/peripheral resistance (PR), and PP were investigated.

Methods:
Pressure and flow relations were investigated in dogs during the development of renal-HTN over 4-weeks (Wks). The sensitivity of hemodynamic markers of vascular remodeling was evaluated by changes in AC= SV/PP, and by AC= Tau/PR and PP.

Results:
All BP components increased during the development of HTN. Changes in Input Impedance, increased through Wks 2 to 4. Increases in the steady component of the cardiac cycle were reflected by peripheral PR, and changes in the pulsatile component were reflected by changes in AC=SV/PP, and AC=Tau /RP; and changes in characteristic impedance. PP and PAP increased by Week 2, reflection of the cardiac and arterial workloads and vascular Tension RP that progressed through Wk 4

Conclusion:
Reliable and accessible markers that indicate changes in arterial pressure, flow & volume, are: PP and AC (SV/PP), and AC (Tau /TPR). These variables detected increases in the heart-load and alterations in pressure-flow patterns that impair cardiovascular-function resulting in HTN-CVD; input and characteristic impedance, also provides remodeling information about the arterial system but are not readily available to clinicians.
Abstract P256: Structural Alterations of Subcutaneous Small Resistance Arteries in Resistant Hypertension
Authors: De Ciuceis Carolina , Rossini Claudia , Muiesan Maria Lorenza , Salvetti Massimo , Porteri Enzo , Gavazzi Alice , Paini Anna , Pileri Paola , Agabiti Rosei Enrico , Rizzoni Damiano
Published: July 2021

Background:
It is was suggested that, in resistant hypertension, the presence of particularly pronounced microvascular alterations may contribute to explain the relative lack of response to treatment

Patients and Methods:
We investigated a population of 94 treated essential hypertensive patients. Secondary forms of hypertension were excluded, and in all patients a 24-hour blood pressure monitoring was performed in order to exclude a white coat effect. In all patients, we evaluated small resistance arteries morphology by a wire micromyographic approach. Subcutaneous tissue was obtained by local biopsy or during election surgery and subcutaneous small resistance arteries were dissected and mounted on a myograph; the media to lumen ratio (M/L) was then measured. We subdived our patents according to the presence or not of resistant hypertension (clinic blood pressure values above 140/90 mm Hg despite administration of three antihypertensive agent including a diuretic and 24-hour blood pressure values >130/80 mm Hg). Sixteen patients had true resistant hypertension, and were compared with the remaining 78 patients with non-resistant hypertension.

Results:
are summarized in the Table, The two groups were also different in terms of mean age (57±12 vs. 67±7 years, p=0.016 and pulse pressure/stroke volume, a rough index of large artery distensibility: 0.63±0.31 vs. 0.90±0.33, p=0.02).

Conclusion:
Hypertensive patients with true resistant hypertension have greater microvascular structural alterations compared with non-resistant hypertensive patients. This could partly explain the resistance to treatment and the high cardiovascular risk observed in these patients.
Abstract P608: Role Of Nitric Oxide In The Pathophysiology Of Hypertension Associated With Low Nephron Endowment.
Authors: Singh Reetu R , Easton Lawerance K , Schlaich Markus P , Head Geoff A , Moritz Karen M , Denton Kate
Published: July 2021

A low nephron endowment is a strong predictor for future risk of hypertension, kidney and cardiovascular disease. Previously, we have shown that fetal uninephrectomy (uni-x) results in elevated arterial pressure, reduced renal function and low plasma renin activity in adulthood in sheep. However, the mechanisms via which a reduction in renal mass in early life contributes to the development of hypertension and renal disease are not well understood. Nitric oxide (NO) plays an important role in driving the fall in renal vascular resistance (RVR) and rise in glomerular filtration rate (GFR) that characterizes the normal postnatal maturation of renal function in the newborn. We hypothesized that the early onset of renal dysfunction in individuals with a congenital reduction in renal mass is due to a reduced renal contribution of NO. Fetal uni-x (n=9) or sham operation (n=7) was performed at 100d of gestation (term=150 days). In conscious female lambs at 6 months of age, mean arterial pressure (MAP) was measured via an indwelling carotid arterial catheter and GFR and renal blood flow (RBF) were measured via clearance methods before and after nitric oxide synthase inhibition (L-NAME; 20mg/kg/h iv). Basal MAP was elevated in uni-x as compared to the sham sheep (sham: 80 ± 2 mmHg, uni-x: 86 ± 1 mmHg; P < 0.01). In response to systemic administration of L-NAME MAP increased similarly in both groups (sham: 13 ± 3 mmHg, uni-x: 13 ± 4 mmHg). Basal RBF (sham: 933 ± 29 ml/kg/h; uni-x 757 ± 43 ml/kg/h; P<0.01) and GFR (sham: 144 ± 3 ml/kg/h; uni-x 107 ± 3 ml/kg/h; P<0.01) was lower in the uni-x as compared to the sham sheep. In response to L-NAME the decline in RBF (sham: 73 ± 3 %; uni-x: 47 ± 3%; P<0.001) and GFR (sham: 54 ± 4 %; uni-x: 43 ± 1%; P<0.03) was less in uni-x as compared to sham sheep. Filtration fraction decreased significantly in the sham (P<0.001) but not the uni-x sheep in response to L-NAME. Renal dysfunction and hypertension in young hypertensive sheep born with a low nephron endowment maybe underpinned by impaired nitric oxide bioavailability. Targeting the nitric oxide system may offer therapeutic potential in children born with a reduced renal mass, in terms of treatment interventions and as a biomarker of prognosis.
Abstract P616: Renal Sympathetic Outflow And Beta-adrenergic Signaling Promote Dendritic Cell Activation In Hypertension
Authors: Xiao Liang , Kirabo Annet , Loperena Roxana , Foss Jason D , Mernaugh Raymond , Roberts L. Jackson , Itani Hana A , Chen Wei , Harrison David G
Published: July 2021

Hypertension is associated with increased sympathetic outflow and activation of adaptive immunity. In hypertensive states, proteins that are oxidatively modified by highly reactive γ-ketoaldehydes (isoketals) accumulate in dendritic cells (DCs). These isoketal-protein adducts are immunogenic and lead to subsequent activation of T cells. We hypothesized that renal sympathetic nerves link the central nervous system to immune activation in hypertension. To test this, we performed bilateral renal denervation (RDN) in C57BL/6 mice by applying phenol to the renal artery. One week later, mice received an subcutaneous infusion of angiotensin II (490 ng/kg/min) for 14 days. RDN lowered the hypertensive response to angiotensin II infusion (130±3 vs. 161±3 mmHg) as measured by telemetry. By flow cytometry, we found that RDN reduced accumulation of T cells in the kidney. Ang II infusion caused resulted in 15-25% increases in expression of the maturation markers CD80 and CD86, as well as 2-fold increase in isoketal adducts in the DCs of spleen. It also increased IL-1α, IL-1β, and IL-6 production by splenic DCs by 4 to 6-fold. These increases were attenuated by RDN. Having confirmed that DCs express almost every subtype of adrenergic receptor by real time PCR, we further determined if sympathetic neurotransmitters contribute to DC activation by treating bone marrow-derived DCs with either norepinephrine (NE) or neuropeptide Y (NPY) in vitro. As measured by flow cytometry, NE dose-dependently increased isoketal-protein adducts in DCs (vehicle: 19 ± 3 vs. 3 μmol/L: 39 ± 4%). This was prevented by pretreating cells with the β-adrenergic antagonist propranolol (1 μmol/L), but not by blocking α
1
or α
2
adrenoreceptors with prazosin and yohimbine. In contrast, NPY did not affect DC isoketal-protein content. Therefore, our data indicate that renal sympathetic nerves increase isoketal-adduct formation in DCs via β-adrenergic signaling and that this contributes to the activation of adaptive immunity in hypertension. These data suggest that beta blockade might have previously unappreciated anti-inflammatory effects in the treatment of hypertension.
Abstract P623: Nutrient Stress Response of Cardiovascular Cells to β-Blockers, ARB and AT2R Agonists
Authors: Mahmood Abuzar , Pulakat Lakshmi
Published: July 2021

To determine how cardioprotective drugs modulate cell response to nutrient starvation, we investigated the effects of β-blockers (Nebivolol [Neb], Carvedilol [Car], Metoprolol [Met] and Atenolol [Aten] (3μM each), Angiotensin II (Ang II) (300nM), AT1R blocker (ARB-Losartan (1μM)) and AT2R agonists (CGP42112A [CGP] and a novel peptide agonist NP-6A4 (300nM each)) on serum-starved HL-1 cardiomyocytes. The Xcelligence Real-Time Cell Analyzer (RTCA), which measures area covered by cells in microtiter plates and displays it as a unitless quantity called Cell Index (CI), was used to assess cellular changes in response to drug treatment. To find whether changes in CI are due to altered proliferation or cell size, we utilized the MTS Proliferation Assay, a colorimetric assay that measures formazan dye produced by viable cells, and fluorophore-conjugated Wheat Germ Agglutinin (WGA) labeling to measure cell size respectively. Difference in CI units is reported as % of CI units of cells treated with vehicle. CI units were suppressed by β-blockers (Aten≤15%; Met≤15%; Neb≤17%; Car≤8%), increased by Ang II (≥9.6%), CGP (≥14%) and NP-6A4 (≥25%), but not by losartan (n≥4 and p≤0.05 for all treatments). MTS assay revealed that only NP-6A4 increased (19%) formazan dye as measured at 490nm (n=3, p<0.05). Differences in cell size of WGA stained cells was calculated as % of size of cells treated with vehicle. Neb and Car decreased cell size (Neb≤14%, Car≤10%, p<0.05 and p<0.1 respectively), suggesting their mechanism for decrease in CI. Other treatments showed no significant change. Next, we determined changes in Myeloid cell leukemia 1 (MCL-1) levels, an essential protein for cardiomyocyte survival, in response to drug treatments by immunoblotting and immunofluorescence. β-Blockers suppressed MCL-1 expression in HL-1 cardiomyocytes (Neb≤26%, Car≤24%, Met≤24%, Aten≤16%) while AT2R agonists increased (CGP≥17%, NP-6A4≥28%, n=3; p<0.05 for all). Treatment of Female Human Coronary Artery Vascular Smooth Muscle Cells by these drugs also showed a similar expression pattern of expression for MCL-1 (CGP≥23%, NP-6A4≥43%). Therefore, AT2R agonist NP-6A4 is more effective in protecting serum starved cardiovascular cells.
Abstract P625: Non-coding Rna Regulation Of Gene Expression In Angiotensin Ii-induced Vascular Damage
Authors: Huo Kugeng , Barhoumi Tlili , Fraulob-Aquino Júlio C , Richer Chantal , Lajoie Mathieu , Sinnett Daniel , Paradis Pierre , Schiffrin Ernesto L.
Published: July 2021

Introduction:
Non-coding RNAs (ncRNAs), including long ncRNAs (lncRNAs) and microRNAs (miRs), account for ~98% of the transcribed RNAs. They have been shown to play a role in cardiovascular disease. Vascular damage is an early manifestation and a cause of end-organ damage in hypertension. However, it is unknown whether ncRNAs are involved in the development of vascular injury in hypertension. We hypothesize that ncRNA regulation participates in mechanisms of vascular remodeling and plays an important role in the pathophysiology of hypertension.

Methods and Results:
Ten-week old male C57BL/6 mice were infused or not with angiotensin (Ang) II for 14 days. Systolic blood pressure (BP) determined by telemetry was increased by Ang II infusion compared to control (146±8 vs 113±5 mmHg, P<0.001). Total RNA was extracted from mesenteric arteries for total and small RNA deep sequencing using Illumina HiSeq-2500. Sequences were aligned to the mm10 genome with STAR, annotated and counted using HTSeq-count or miRDeep2. Differential expression analysis was done in R. Differentially expressed (DE) mRNAs (550 up & 266 down), lncRNAs (7 up & 42 down), miRs (23 up & 12 down) were identified in the Ang II-treated group (1.5 fold change, q<0.05). Targetscan was used to predict interactions between DE miRs and the inversely correlated DE mRNAs or DE lncRNAs. MEME Suite was used to predict DE transcription factor binding sites in the promoter region of genes encoding DE mRNAs, lncRNAs and miRs. Cytoscape was used to construct molecular networks integrating the above interactions and the gene expression profile and to perform functional enrichment analysis, which revealed enrichment of extracellular matrix and developmental processes in DE miR-targeting DE mRNAs (q<1E-20). Ten DE miRNAs whose expression levels correlated (P<0.05) with BP were identified, 9 of which are located in a single miRNA cluster that is conserved in humans.

Conclusions:
We have identified a conserved miRNA cluster that may play a pivotal role in the regulation of vascular damage in hypertension. A sub-network of genes that participates in the interaction between the miRNA cluster and other BP-correlated RNAs was selected for future investigation to identify therapeutic targets.
Abstract P632: The Blood Pressure in Dialysis (BID) Pilot Study
Authors: Zager Philip G , Miskulin Dana , Gassman Jennifer , Ploth David , Jhamb Manisha , Rahman Mahboob , Negrea Lavinia , Kendrick Cynthia
Published: July 2021

Background:
The optimal blood pressure (BP) target for hypertensive hemodialysis (HD) patients is unknown. Current KDOQI guidelines have been extrapolated from data in the general population. The BID pilot, funded by NIDDK and DCI, is the first trial to randomize hypertensive HD patients to intensive (110-140 mm Hg) vs. usual (155-165 mm Hg) control of systolic blood pressure (SBP). The study’s goal is to assess the feasibility of conducting a full-scale trial.

Methods:
BID consortium consists of 5 clinical centers, a cardiac MRI reading center and a data coordinating center. Standardized predialysis SBP, measured in the dialysis unit in accord with AHA recommendations (SDUSBP), guide therapy. To be eligible for randomization patients needed a 2-week running mean SDUSBP ≥ 155 mm Hg. Home BP measurements (HBPM) are obtained twice on the day after the midweek dialysis. Ambulatory Blood Pressure Monitoring (ABPM) during a 44h interdialytic period is obtained quarterly. We compared SDUSBP, HBPM and ABPM.

Results:
We enrolled 281 and randomized 126 participants. Major reasons for drop out during the baseline period were 2-week mean SDUSBP < 155 mm Hg (40.6%), no cardiac MRI (13.0%) and perception of protocol as burdensome (11.2%). Adherence with prescribed SDUSBP was satisfactory. The percent of patients with ≥ 4, ≥ 8 and ≥12 SBP per month were 96, 88 and 57% in month 1 and 78, 68 and 37% in month 12. In a constant cohort of participants followed ≥ 330 days 2-week mean SDUSBP were 144 ± 17.4 and 156 ± 15.2 mm Hg in the intensive and usual arms, respectively. Major reasons that participants in the intensive arm did not achieve target SBP included large interdialytic weight gain (27.2%), non-adherence with medications or dialysis prescription (40.9%), and intradialytic hypotension (31.9%). Differences between the SDUSBP and both HBPM and ABPM were often ≥ 10 mm Hg. Optimal control of BP requires measurements in and out of the dialysis unit. Rates of adverse events were similar to those in other NIDDK funded ESRD studies.

Conclusion:
The difference in BP between arms was achieved and maintained throughout the study. It is feasible to conduct a full-scale clinical trial of intensive vs. usual treatment of hypertension in HD patients.
Abstract P637: Pigment Epithelium Derived Factor (PEDF) Deficiency Increases Blood Pressure And Accentuates Glomerular Pathology In Diabetic Mice
Authors: Ye Minghao , Volpert Olga , Wysocki Jan , Batlle Daniel
Published: July 2021

Pigment Epithelium Derived Factor (PEDF) encoded by SERPINF1 gene has potent anti-angiogenic and cytoprotective activities. It has been reported that PEDF protein levels are reduced in the kidneys of rodents with experimentally induced diabetes. However the effect of PEDF on blood pressure has not been elucidated. Here, we used SERPINF1 KO mice to examine the impact of PEDF deficiency on kidney pathology and blood pressure in the STZ-induced mouse model of diabetes.

Twelve weeks after diabetes induction by STZ, SERPINF1 KO mice showed exacerbated glomerular damage with ~ 2-fold increase in mesangial matrix (2.8±0.14 vs. 1.2±0.14 arbitrary units, p<0.01) and ~ 20% decrease in podocyte counts (8.2±0.0.4 vs. 9.7±0.2 podocytes/glomerulus, p<0.01) compared to the wild type controls. Of note, STZ-treated SERPINF1-/- mice displayed elevated systolic blood pressure (SBP) (127±4 vs. 109±4 mmHg, p<0.01, n=11).

Our data indicate that global PEDF deficiency intensifies glomerular injury in STZ-induced mouse model of diabetes. An important drawback of most murine models of diabetic kidney disease is the lack of hypertension, a known key factor that accelerates progression to CKD in humans. In contrast, we found that STZ-treated SERPINF1-/- mice become hypertensive. Together, our findings point to SERPINF1-/- mice as an attractive model to study diabetic kidney disease and hypertension in mice and suggest an important causative role of PEDF downregulation in glomerular pathology in diabetes.
Abstract P645: Chronic Angiotensin-(1-7) Improves Whole-Body Insulin Sensitivity in High-Fat Fed Mice by Enhancing Skeletal Muscle Glucose Uptake
Authors: Williams Ian M , Wasserman David H , Biaggioni Italo , Arnold Amy C
Published: July 2021

Angiotensin (Ang)-(1-7) is a vasodilatory peptide implicated in the pathophysiology of hypertension, in part by opposing deleterious Ang II cardiovascular actions. Recent studies show that Ang-(1-7) restoration lowers blood pressure and improves glycemic control in animal models of cardiometabolic syndrome. The tissue-specific sites of action and blood pressure dependence for these metabolic effects, however, remain unclear. We hypothesized that Ang-(1-7) improves insulin sensitivity by enhancing peripheral glucose delivery. To test this hypothesis, adult male C57BL/6 mice were placed on standard chow or 60% high-fat diet for 11 weeks, with Ang-(1-7) [400 ng/kg/min] or saline given during the last 3 weeks of diet by subcutaneous osmotic mini-pump. Hyperinsulinemic (4 mU/kg/min) euglycemic clamps were performed in conscious, unrestrained mice at the end of the treatment period. High-fat fed mice exhibited modest hypertension (systolic blood pressure: 137±3 high-fat vs. 123±5 mmHg chow; p=0.043), which was not altered by Ang-(1-7) infusion (141±4 mmHg; p=0.516). Body weight, body composition, and fasting plasma glucose and insulin levels were not significantly different following Ang-(1-7) treatment in chow or high-fat fed mice. Ang-(1-7) increased the glucose infusion rate (GIR) needed to maintain euglycemia in high-fat fed mice (steady-state GIR: 31±5 Ang-(1-7) vs. 16±1 mg/kg/min vehicle; p=0.017) indicating enhanced whole-body insulin sensitivity, with no significant effect in chow fed mice. The improvement in insulin sensitivity in high-fat fed mice was due to an enhanced rate of whole-body glucose disappearance (R
d
: 34±5 Ang-(1-7) vs. 20±2 mg/kg/min vehicle; p=0.049), with increased rates of glucose uptake in gastrocnemius, vastus, and soleus muscle. There was no effect of Ang-(1-7) on insulin-mediated suppression of hepatic glucose production. Our data shows that Ang-(1-7) has direct insulin-sensitizing effects on skeletal muscle, which are independent of changes in body weight or systemic blood pressure. These overall findings provide new insight into mechanisms by which Ang-(1-7) improves insulin action, and provide further support to targeting this peptide for treatment of cardiometabolic disease.
Abstract 001: Sympathetic Innervation Promotes Bone Marrow Homing of Hypertension-specific CD8
+
Effector Memory T Cells
Authors: Xiao Liang , Itani Hana A , Foss Jason D , Harrison David G
Published: July 2021

We have recently identified a critical role of hypertension-specific effector memory T lymphocytes (T
EM
cells) in the blood pressure elevation and renal dysfunction caused by repeated hypertensive stimuli. Formed during an initial immune challenge, T
EM
cells reside in the bone marrow (BM) in a quiescent state for prolonged periods, and can be reactivated upon re-exposure to the hypertensive stimulus. Hypertension is associated with increased sympathetic outflow. We therefore hypothesized that sympathetic nerves regulate accumulation and reactivation of hypertension-specific T
EM
cells in the BM. We performed unilateral superior cervical ganglionectomy (SCGx) in wild-type C57BL/6 mice, causing selectively sympathectomy of the forelimb on the surgical side. After recovery, mice received Ang II infusion for two weeks. To determine the changes of T cells in the BM that were specific to hypertension, 5х10
6
BM cells were isolated from either the SCGx or control limbs, loaded with proliferation marker CFSE, and co-cultured with 0.5х10
6
splenic dendritic cells isolated from another Ang II-infused mouse. We found 30% less CD8
+
T cell proliferation in the SCGx BM compared to control side (1.8±0.1 vs. 2.6±0.3х10
4
), but no difference in CD4
+
T cells. To further study the effect of sympathetic nerves on BM T cell homing, 1х10
7
pan T cells were isolated from CD45.2
+
wild-type mice after Ang II infusion and adoptively transferred to CD45.1 mice that had previously undergone unilateral SCGx. Flow cytometry indicated that 7 days after transfer, 25% fewer CD8
+
T
EM
cells from the hypertensive donors homed to the SCGx BM compared to the innervated BM (27.8±2.6 vs. 20.8±2.5 per 10
3
total BM cells). This effect was specific for hypertension as homing of OT-I T
EM
cells, which are responsive to ovalbumin, was not influenced by sympathectomy. Further adoptive transfer studies using mice lacking beta 2 adrenergic receptors (β2AR) indicate that β2AR in the bone marrow niche, rather than T cell β2AR is critical for T
EM
cell homing. These data define a novel role of sympathetic nerves in regulation of memory T cell trafficking, and this likely contributes to the predisposition to hypertension and end-organ damage for prolonged periods following an initial episode of hypertension.
Abstract 006: Hypertension is Accompanied With Over Activity of the Adaptive Immune System
Authors: Zhao Tuantuan , Shen Xiao Z , Bernstein Kenneth E
Published: July 2021

Adaptive immune response plays an important role in the pathogenesis of hypertension, but how does hypertension influence on the immune system is still poorly understood. To understand this, we studied antigen presenting cells (APCs) isolated from normotensive and experimental hypertensive mice induced by angiotensin II (490 ng/kg/min, 2 weeks). The APCs were loaded with ovalbumin(OVA) or its MHC class I epitope SIINFEKL (SKL) for 3h. Then, splenocytes from OT-I mice, containing OVA-specific, CD8
+
T cells (OT-I cells), were added for 4h. Significantly more activated T cells expressing surface marker CD69 and cytokine IFN-γ were detected by flow cytometry when stimulated with APCs from hypertensive mice than equivalent cells from normotensive mice (P<0.05). Thus,
in vitro,
APCs derived from hypertensive mice more effectively present antigen to T cells. To study antigen presentation
in vivo
, we immunized AngII and sham treated mice with OVA and adjuvant. Consistently more OVA-specific CD8
+
T cells were induced in the blood (2.7 ± 0.23% vs. 1.2 ± 0.47%, P<0.05) and the spleen (1.51 ± 0.17% vs. 2.62 ± 0.31%, P<0.05) of AngII mice vs sham when measured by flow using an H-2k
b
-SKL tetramer. We also evaluated the APC and T cell activation signals before and after hypertension. Flow analysis showed that splenic DCs divided from AngII treated mice resulted in a substantially increased surface expression of co-stimulatory factor CD86 compare with DCs from normotensive controls (MFI: 131±7.8 vs. 97±5.3). RIP-mOVA mice were used to study if hypertension affects APC cross-presentation of self-antigens. This transgenic mouse line expresses membrane-bound OVA in pancreatic islet β cells. When OT-I cells (5x10
6
) are injected into RIP-mOVA mice, they activate by cross-presentation of OVA and cause insulitis and diabetes. RIP-mOVA mice were made hypertensive with either AngII or L-NAME (1.5 mg/ml, 4 weeks). When OT-I cells were
i.v.
infused into RIP-mOVA mice, the hypertensive mice had more OT-I cells infiltration into the pancreas (AngII 75%, L-NAME 73%, control 13%) and developed severer diabetes (Blood glucose: AngII 331, L-NAME 315, control 168 mg/dl). In conclusion, high blood pressure itself is associated with over activity of the adaptive immune system.
Abstract 016: Combined AT
2
Receptor Stimulation/ AT
1
Receptor Blockade Does Not Cause Renal Dysfunction
Authors: Baraldi Dhaniel , Hilliard Lucinda M , Gaspari Tracey A , Denton Kate M , Widdop Robert E
Published: July 2021

Combined ACE inhibition and angiotensin type 1 receptor (AT
1
R) blockade (dual RAS) blockade) causes renal failure which limits their combined use. Angiotensin type 2 receptor (AT
2
R) stimulation evokes renoprotective effects but potential adverse renal effects in drug combination are not known. We examined the effects of the AT
2
R agonist compound 21 (C21) combined with either candesartan cilexetil or perindopril on renal function in a preclinical model that is predictive of clinical outcome. Adult (~25 week-old) male SHR were placed on a normal salt (0.35%, n=8) or a low salt (LS; 0.05%) diet for 17 days with rats on a LS diet randomised to receive one of the following treatments for the final 10 days: untreated (n=8); C21 (0.3mg/kg/d s.c., n=5); candesartan (2 mg/kg/d, n=6); perindopril (0.5mg/kg/d, n=5); C21+candesartan (n=8); C21+perindopril (n=5); or candesartan+perindopril (dual RAS blockade, n=7). Systolic arterial pressure (SBP) was measured via tail cuff at days 0, 7 and 17. At the end of the treatment, renal function was assessed by measuring plasma creatinine, urea, K+ and glomerular filtration rate (GFR) in conscious rats via transdermal assessment of elimination half-life kinetics of FITC-sinistrin (3-5mg/100g i.v.). Candesartan (-43±10 mmHg) or perindopril (-49±12 mmHg) reduced SBP to a similar extent alone or combined with C21, whereas dual RAS blockade markedly reduced SBP (-115±14 mmHg; P<0.01 versus all groups). Plasma creatinine (424 ±65 μmol/L), urea (>50 mmol/L cut-off) and K+ (5.94 ±0.82 mmol/L) levels were all significantly elevated by dual RAS blockade compared with untreated SHR on normal diet (creatinine 39 ±2 μmol/L; urea 7.08±0.22 mmol/L; K+ 4.23±0.13 mmol/L; all P<0.01) or LS diet (creatinine 29 ±2 μmol/L; urea 5.2±0.57 mmol/L; K+ 4.29±0.17 mmol/L; all P<0.01), whereas plasma measurements for C21+candesartan and C21+perindopril were similar to control groups. Estimated GFR in LS group (1.18±0.06 ml/min/100g BW) was similar to other groups except during dual RAS blockade where GFR was markedly impaired (0.32±0.10 ml/min/100g BW; P<0.01 versus LS). Collectively, these data suggest that, unlike dual RAS blockade, an AT
2
R agonist combined with either ACE inhibition or AT
1
R blockade is not likely to cause renal dysfunction.
Abstract 020: Heterozygote Knockout of TMEM16B in Intestinal Vagal Afferents Causes Cholecystokinin Insensitivity and Obesity in Male but Not Female Mice
Authors: Wang Runping , Lu Yongjun , Cicha Michael Z , Benson Christopher J , Chapleau Mark W , Abboud Francois M
Published: July 2021

Cholecystokinin (CCK) is a well-known satiety peptide that inhibits food intake. The CCK induced satiety signal in intestinal vagal afferents is attenuated in mice on a High Fat Diet (HFD). We have shown that down regulation of a Ca
2+
-activated Cl
-
channel (CaCC) downstream of CCK receptors is responsible for the neuronal insensitivity to CCK. We also found that the CaCC subunit Ano2/TMEM16B is essential for the CCK-induced current in nodose neurons. In this study we tested the hypothesis that reduction of this subunit
in vivo
contributes to weight gain. One allele of the Ano2/TMEM16B was knocked out in sensory neurons by crossing the ano2
fl/
fl
mice with Nav1.8Cre mice to generate the Nav1.8Cre;ano2
fl
/wt
mice. The Cre negative littermates were used as control. We found that CCK-induced suppression of food intake is eliminated in male Nav1.8Cre;ano2
fl
/wt
mice. Food intake measured over 4 hours was 1.21±0.11g (n=5) in male wild type (wt) mice injected with saline and was reduced to 0.77±0.18 g (n=7, p<0.05) in mice injected with CCK-8 (3μg/kg). Those values were 0.63±0.15 g (n=6) in saline injected and 0.91±0.13 g (n=7, p>0.05) in CCK injected male Nav1.8Cre;ano2
fl
/wt
mice. However, the CCK injection did not affect food intake in either female wt or Nav1.8Cre;ano2
fl
/wt
mice. The male Nav1.8/ano2
fl/wt
mice were on the average 5.5g heavier than wt mice at 40 weeks of age (39.8±1.4 g, n=13 vs 34.3±1.4 g, n=14, p<0.01). Body weight of females was significantly lower than in males but was not different between wt and Nav1.8Cre;ano2
fl
/wt
mice (30.4±1.0 g, n=14 vs 28.5±1.0 g, n=11, p=0.20 respectively). Single cell mRNA level of Ano2 and CCK-induced TMEM16 currents in nodose neurons were reduced significantly in male Nav1.8Cre;ano2
fl/wt
mice compared to Cre negative controls, but such changes were not seen in female mice. We conclude that heterozygote knockout of Ano2/TMEM16B specifically in sensory neurons causes neuronal insensitivity to CCK and excessive weight gain in male but not female mice. Reduction of this subunit may contribute to the HFD induced obesity. The reason for the phenotype and allele expression variability between sexes is unclear.