bluuebunny/crossref_metadata_2025 huggingface dataset:
Superoxide Dismutase Mimetic Increases Neuronal Nitric Oxide Synthase Influence on Afferent Arteriolar Diameter in Spontaneously Hypertensive Rats
Authors: Ichihara Atsuhiro , Hayashi Matsuhiko , Hirota Nobuhisa , Saruta Takao
Published: July 2021

P108

This study was designed to determine the influence of increased superoxide anion production in neuronal nitric oxide synthase (nNOS)-dependent regulation of afferent arteriolar diameter in spontaneously hypertensive rats (SHR). Afferent arteriolar diameters of male Wister-Kyoto rats (WKY) and SHR (300-350 g) were assessed
in vitro
using the blood-perfused juxtamedullary nephron technique and averaged 21.6 ± 1.6 (n = 6) and 18.8 ± 1.2 μm (n = 7); respectively. Superfusion with the superoxide dismutase mimetic, tempol (1, 10, and 100 μmol/L), did not influence afferent arteriolar diameters of WKY, but significantly increased afferent arteriolar diameters of SHR by 20.6 ± 5.5%, 25.2 ± 5.4% and 23.3 ± 4.9%; respectively. In WKY (n = 6), superfusion with the nNOS inhibitor,
S
-methyl-L-thiocitrulline (L-SMTC; 10 μmol/L), and the NOS inhibitor,
N

w

-nitro-L-arginine (L-NNA; 100 μmol/L), significantly decreased afferent arteriolar diameters (19.6 ± 1.6 μm) by 11.9 ± 3.1% and 21.0 ± 3.9%; respectively. In SHR (n = 7), L-SMTC treatment did not influence afferent arteriolar diameters (21.0 ± 1.5 μm), but L-NNA treatment exerted a significant afferent arteriolar constriction (14.8 ± 3.2%) which was similar to that observed in WKY. Experiments were also performed in the presence of 100 μmol/L tempol. In WKY (n = 6), tempol treatment did not modulate basal afferent arteriolar diameters (21.5 ± 1.2 μm) or afferent arteriolar constrictor responses to L-SMTC (10.6 ± 2.1%) and L-NNA (19.3 ± 3.3%). In SHR (n = 8), tempol treatment significantly increased afferent arteriolar diameters by 22.5 ± 4.3% and enhanced afferent arteriolar constrictor responses to L-SMTC (18.4 ± 2.7%) and L-NNA (31.9 ± 2.6%). However, superfusion with the nitric oxide donor,
S
-nitroso-N-acetylpenicillamine (10 μmol/L), which exerted a similar afferent arteriolar vasodilation (19.7 ± 3.8%, n = 4), did not influence afferent arteriolar responses to L-SMTC (-2.1 ± 3.1%) or L-NNA (15.1 ± 3.3%). These results suggest that increased superoxide anion inhibits nNOS-dependent regulation of afferent arteriolar diameters in SHR.
5,6-Epoxyeicosatrienoic Acid Mediates the Enhanced Renal Vasodilator Response to Arachidonic Acid (AA) in the SHR.
Authors: Pomposiello Silvia I , Carroll Mairead A , McGiff John C
Published: July 2021

P109

The cytochrome P450-dependent renal vasodilator effect of arachidonic acid (AA) that is expressed when cyclooxygenase is inhibited and perfusion pressure (PP) is elevated, is enhanced in SHR. In kidneys perfused with Krebs’ solution and preconstricted with phenylephrine to increase PP to
ca
200 mmHg, 5 μg of AA reduced PP by 92 ± 10 mmHg in SHR compared to 44.5 ± 5 mmHg for WKY rats. In contrast, the dilator effects of acetylcholine (Ach; 0.1 μg) or sodium nitroprusside (SNP; 1 μg) were not different. As the response to AA was inhibited by ETYA (4 μM), an inhibitor of all AA oxygenase pathways, and also inhibited by the selective epoxygenase inhibitors, miconazole (0.3 μM) and MSPPOH (12 μM), we concluded that the vasodilator mediator(s) of the AA response was most likely an epoxyeicosatrienoic acid (EET). We therefore, assessed the renal vascular responses to EETs in the SHR and WKY. We found that 5,6-EET, 8,9-EET, and 11,12-EET were dilators in both SHR and WKY, whereas 14,15-EET produced constriction. 5,6-EET was the most potent dilator of the EET regioisomers in the SHR resulting in a decrease in PP of 56 ± 5 mmHg compared to 12 ± 4 mmHg for 8,9-EET and 15 ± 4 mmHg for 11,12-EET. The decrease in PP elicited by 5 μg of 5,6-EET was greater in SHR, 56 ± 5 mmHg compared to 29 ± 4 mmHg for WKY rats whereas the responses to the other EET regioisomers did not differ. We measured the release of EETs from SHR and WKY kidneys in response to 5 μg of AA, by GC-MS in negative ionization mode using selective ion monitoring. The release of 5,6-EET was increased in SHR (from a basal level of 0.5 ± 0.04 to 0.9 ± 0.1 ng/min; P<0.05) compared to WKY (from a basal level of 0.6 ± 0.2 to 0.7 ± 0.2 ng/min), whereas the release of the other EETs were not different. We suggest that 5,6-EET is the most likely mediator of the AA response and that the augmented vasodilator response to AA in the SHR kidney might be linked to increased production of and responsiveness to 5,6-EET.
Low Dietary Salt Regulates Renal Vascular 20-Hydroxyeicosatetraenoic Acid Levels.
Authors: Carroll Mairead A , Cheng Monica K , McGiff John C
Published: July 2021

P110

20-hydroxyeicosatetraenoic (HETE), a prohypertensive cytochrome P450 (CYP)-derived arachidonic acid (AA) metabolite, is a principal eicosanoid of preglomerular microvessels. Vascular 20-HETE release is stimulated by angiotensin II (AII) and is subject to metabolism by cyclooxygenase (COX)-2. As dietary salt alters AII levels and COX-2 expression, we studied 20-HETE levels from microdissected arcuate and interlobular arteries and interlobar arteries obtained from male Sprague-Dawley rats fed a control (0.4% NaCl) or low salt diet (LS; 0.05% NaCl). In controls (n=6), metabolism of
14
C-AA (7μM) in the presence of NADPH (1mM) and indomethacin (INDO; 10μM) to 20-HETE was higher (P<0.05) in arcuate and interlobular arteries compared to interlobar arteries (25.2 ± 3.6 ng vs. 15.3 ± 2.3 ng/mg protein/30 min, respectively) based on reverse-phase HPLC retention times and on-line radiodetection. No regional differences in epoxide formation were evident between arcuate and interlobular arteries compared to interlobar arteries (21.9 ± 3.6 ng vs. 19.6 ± 3.7 ng/mg protein/30 min, respectively). LS treatment, for 7 days, selectively increased 20-HETE levels in interlobar arteries (24.6 ± 3.9 ng/mg protein/30 min) and increased medullary CYP-4A expression. However, in the absence of INDO, 20-HETE levels in arcuate and interlobular arteries, but not interlobar arteries, were diminished by 90% with LS treatment; changes that corresponded with induced cortical COX-2 expression. Thus, 20-HETE levels vary segmentally within the renal microvasculature. LS intake induces medullary CYP-4A expression and interlobar artery 20-HETE formation. Presumably, the diminished 20-HETE levels in arcuate and interlobular arteries, in the absence of COX inhibition, were a result of increased cortical COX-2 expression serving as a metabolic pathway for 20-HETE; the vasoconstrictor 20-HETE being metabolized by COX-2 to vasodilator prostaglandin analogs. Increased 20-HETE levels may account for the adverse effects of COX inhibitors with salt depletion.
Pulse Pressure Inversely Correlates with Telomere Length in French Men
Authors: Benetos Athanase , Okuda Koji , Lajemi Malika , Skurnick Joan , Labat Carlos , Aviv Abraham
Published: July 2021

P126

Cardiovascular risk increases with chronological age. Pulse pressure (PP) also increases with chronological age and is an independent indicator of cardiovascular risk. It is therefore possible that after age-adjustment, the biological age of persons with wide PP is more advanced than their chronological age would indicate. We explored whether an index of biological age, namely telomere length (measured by the mean length of the terminal restriction fragment, TRF), provides a better account than chronological age of about variation in PP. We studied 193 French men and women (age 56 ± 11 years) on no antihypertensive medications. Univariate analysis showed that in both genders, TRF length (in WBCs) was correlated inversely with age (p< 0.01). The rate of telomere attrition was 0.038 and 0.036 kb/year for men and women, respectively. Only in men, after adjustment for mean arterial pressure, TRF length accounted for 12% variability in PP (p<0.001), while chronological age accounted for 10% variability in PP (p<0.001). Age-adjusted TRF length was longer in women than men (8.67 ± 0.09 vs 8.38 ± 0.07 kb, p = 0.016). We conclude that TRF length adds additional information about variability in PP among men, such that men with shorter telomere length are more likely to exhibit a wider PP. The longer telomere length in women suggests that for a given chronological age, the biological age of men is more advanced than that of women. The results also underscore the sexual dimorphism in aging and blood pressure regulation. We propose that large-scale studies should be undertaken to explore the relation between telomere length and cardiovascular risk.
Transcriptional Activation of the Bradykinin Type 2 Receptor by the Tumor Suppressor P53.
Authors: El-Dahr Samir S , Saifudeen Zubaida , Du Hong
Published: July 2021

P145

The bradykinin type 2 receptor (BK2) is a developmentally regulated G protein-coupled receptor that mediates diverse actions such as vascular reactivity, salt and water excretion, inflammatory responses and cell growth. However, little is known regarding regulation of the BK2 gene. We report here that the rat BK2 receptor is transcriptionally regulated by the tumor suppressor protein p53. The 5’-flanking region of the rat BK2 gene contains two p53-like binding sites: a sequence at -70 bp (P1 site) that is conserved in the murine and human BK2 genes; and a sequence at -707 (P2) that is not. The P1 and P2 motifs bind specifically to p53, as assessed by gel mobility shift assays. Transient transfection into HeLa cells of a CAT reporter construct driven by 1.2-kb of the BK2 gene 5’-flanking region demonstrated that the BK2 promoter is dose-dependently activated by co-expression of wild-type p53. Co-expression of a dominant negative mutant p53 suppresses the activation of BK2 by wild-type p53. Promoter truncation localized the p53-responsive element to the region between -38 and -94 bp encompassing the p53-binding P1 sequence. Site-directed mutagenesis of the P1 site abolishes p53-mediated activation of BK2. Furthermore, p53-mediated activation of the BK2 promoter is augmented by the transcriptional co-activators, CBP/p300. Interestingly, removal of the P2 site by truncation or site-directed deletion amplifies p53-mediated activation of the BK2 promoter. These results demonstrate that the rat BK2 promoter is a target for p53-mediated activation and suggest a new physiological role for p53 in the regulation of G protein-coupled receptor gene expression.
Phospholipase A
2
Metabolites Mediate Endothelin Stimulation of the Human Brain Natriuretic Peptide Promoter
Authors: He Quan
Published: July 2021

P155

Brain natriuretic peptide (BNP) gene expression accompanies cardiac hypertrophy and heart failure. The vasoconstrictor endothelin-1 (ET)may be involved in the development of these diseases. ET has also been shown to activate phospholipase A
2
(PLA
2
). Thus we studied whether ET and PLA
2
metabolites regulate BNP gene expression. The hBNP promoter (-1818 to + 100) coupled to a luciferase reporter gene was transferred into neonatal ventricular myocytes (NVM),and luciferase activity was measured as an index of promoter activity. ET (10
-7
M)induced BNP mRNA in NVM as assessed by Northern blot. It also stimulated the hBNP promoter 4-fold vs control, an effect completely inhibited by actinomycin D. To test the involvement of different PLA
2
isoforms, transfected cells were treated with the Ca
++
-independent PLA
2
(iPLA
2
)inhibitor bromoenol lactone (BEL), the cytosolic PLA
2
inhibitor methyl arachidonyl fluorophosphonate, or the secretory PLA
2
inhibitor ONO-RS-082 prior to stimulation with ET. Only the iPLA
2
inhibitor BEL prevented ET-stimulated hBNP promoter activity. The PLA
2
metabolite lysophosphatidic acid (LPA) also activated the hBNP promoter (2.2-fold; n = 3), but lysophosphatidylcholine did not. To test whether arachidonic acid metabolites are involved in ET’s effect, cells were pretreated with either a lipoxygenase (LO), cyclooxygenase, or p450 monooxygenase inhibitor. Only the LO inhibitor baicalein prevented ET stimulation of the hBNP promoter. Finally, we studied the involvement of
cis
elements in ET-stimulated hBNP promoter activity. Deletion of BNP promoter sequences from -1818 to -408 and from -408 to -40 reduced ET’s effect by 54% and 78%, respectively. Moreover, ET-stimulated luciferase activity was reduced by 53% when the GATA element (at position -85 relative to the start site of transcription) was mutated. These data suggest that: 1) ET activates the hBNP promoter through a transcriptional mechanism; 2) LPA, perhaps generated by a BEL-sensitive iPLA
2
, is involved in ET’s effect; 3) a LO pathway may also mediate ET signaling; and 4) ET regulation of the hBNP promoter targets both distal and proximal
cis
elements, including GATA.
Interactions Between Insulin and Angiotensin II on Calcium Mobilization in Skin Fibroblasts from Insulin Resistant Patients with Essential Hypertension
Authors: Giulio Ceolotto , Elisabetta Baritono , Roberto Valente , Alessandra Monari , Stefania Reato , Michelangelo Sartori , Roberto Trevisan , Andrea Semplicini
Published: July 2021

P181

Objectives: Insulin attenuates angiotensin II (Ang II)-induced Cai2+ mobilization in insulin sensitive (IS) control subjects. Therefore we compared the interactions between Ang II and insulin on Cai2+ mobilization in skin fibroblasts from insulin resistant (IR) and insulin sensitive (IS) patients with essential hypertension (HT). Methods: Skin fibroblasts from 9 normotensives (NT) and 18 HT were cultured and used after 4 passages. Ca2+ was measured in monolayers of 24h serum-deprived cells using Fura-2 AM. mRNA expression of Gαi2 subunit was determined by RT-PCR using a 5 and 3 specific primers. Results: Resting and Ang II-stimulated (100nM) Cai2+ peak were higher in fibroblasts from HT than NT (76±3 vs 62±3 nM, p<0.01 and 225±8 vs 164±12 nM, p<0.01). In the absence of extracellular Ca2+, the Cai2+ response to Ang II was still significantly enhanced in fibroblasts from HT (212±8 vs 158±9 nM, p<0.01). Thapsigargin (500 nM), a Ca2+ -ATPase inhibitor, induced a Cai2+ rise that was similar in HT and NT (from 42±7 to 93±15 vs 52±5 to 106±8 nM). Pertussis toxin (20ng/ml, 6h), inhibitor of Gαi2 subunit, reduced Ang II stimulated Cai2+ peak both in NT (from 153±7 to 98±8nM, p<0.01) and HT (from 189±16 to 138±12, p<0.01). In IS HT, insulin (100 nM, 20 min) blunted the Ang II stimulated Cai2+ response (from 208±9 to 167±9 nM, p<0.01), while in IR HT it did not (from 205±16 to 208±13 nM, ns). Pertussis toxin attenuated AngII stimulated Cai2+ peak in IS HT (from 208±4 to 112±2 p<0.01), but not in IR HT (from 207±10 to 165±9). mRNA expression of Gαi2 subunit was increased in IR HT compared to IS HT. Conclusion: In human skin fibroblasts the inhibitory effect of insulin on Ang II - induced Cai2+ mobilization is blunted in IR HT by a pertussis toxin sensitive mechanism, probably involving a Gαi2subunit.
Renin Gene Transfer Restores Angiogenesis and VEGF Expression in Dahl S Rats
Authors: Amaral Sandra L , Roman Richard J , Greene Andrew S
Published: July 2021

P204

To evaluate the importance of the renin angiotensin system (RAS) in VEGF expression and angiogenesis in skeletal muscle, we compared the angiogenic response to electrical stimulation in congenic strains of SS/Jr/Hsd rats using a complementation test design. We have previously demonstrated that both increases in VEGF expression and angiogenesis induced by electrical stimulation of skeletal muscle were absent in inbred Dahl S rats having a wildtype renin allele (S/ren
ss
). In contrast, the congenic S/ren
rr
in which a 10 cM segment of chromosome 13 containing the normally functioning salt resistant renin allele was transferred onto the Dahl S background, exhibit the expected changes in renin. In the present study we investigate the effects of electrical stimulation on VEGF expression and angiogenesis in these rats. Congenic S/ren
rr
and S/ren
ss
rats, fed a 0.4% salt diet were surgically prepared by chronic implantation of an electrical stimulator. Another group of S/ren
rr
rats was treated with lisinopril, 2 days before the surgery and throughout the stimulation protocol. Rats without any drug treatment were used as control. The right tibialis anterior (TA) and extensor digitorum longus (EDL) were stimulated (10 Hz, 0.3 ms duration) for 8 hours per day for 7 days. The contralateral muscles served as controls. Western blot analysis was performed to identify VEGF protein expression in these muscles. Seven days of electrical stimulation of the skeletal muscles produced no change in vessel density of S/ren
ss
(Δ=5.50 ± 3.8 % and 8.14 ± 2.0 % for EDL and TA respectively). Transfer of the resistant renin allele (S/ren
rr
) restored the angiogenic response (Δ=16% and 30% for EDL and TA, respectively) despite a significantly higher blood pressure (113.5 ± 2.25 mmHg and 148.67 ± 1.12 mmHg for S/ren
ss
and S/ren
rr
, respectively). Blockade of the RAS in S/ren
rr
restored the phenotype observed in the S/ren
ss
(Δ=1.46% and 1.9% to EDL and TA, respectively, p<0.05). In addition, increases in VEGF expression to electrical stimulation were observed only in S/ren
rr
. These results demonstrate that RAS plays an important role in the regulation of VEGF expression and angiogenesis in skeletal muscle.
Abstract 1: Loss of Nuclear Receptor 4a1 on a Hypertensive Genetic Background Promotes Immune-Mediated Renal Injury
Authors: Westbrook Lindsey , Regner Kevin R , Johnson Ashley C , Lee Jonathan , Kyle Patrick B , Mattson David L , Garrett Michael R
Published: July 2021

Nuclear hormone receptors of the NR4A subgroup have been implicated in cancer, atherosclerosis, and most prominently in metabolic disease. However, little is known about the role of NR4A1 in kidney health or disease. Fawn-hooded hypertensive (FHH) Nr4a1
-/-
deficient animals were evaluated for blood pressure, proteinuria, renal function and metabolic parameters from 4-24 weeks of age. At week 24, Nr4a1
-/-
rats exhibited 4-fold higher proteinuria (81±11.5 mg/24hrs/100gBW) compared to FHH (21±1.0). No difference in telemetry measured blood pressure was observed at any time point, but glomerular filtration rate was significantly decreased in the Nr4a1
-/-
(542±78.9 ul/min/gKW)
vs
FHH (937±75.2). Moderate increases in glomerulosclerosis were seen, but were not different between strains. The severity of tubular atrophy, casts, and fibrosis was significantly increased in Nr4a1
-/-
compared to FHH (19±1.86%
vs
12±0.8%, respectively). Tubulointerstitial macrophage infiltration was also significantly increased in kidneys from Nr4a1
-/-
(245±62 foci per field) compared to FHH (50±13.4) at week 24. Microarray analysis of kidney from week 8-24 found significant up-regulation of important immune mediators, including TGF-β1-2, CCL2 (MCP-1), CCL9 (MIP-1γ), and other chemokines and interleukins. The specific injury (tubulointerstitial), immune cell infiltration, and immune gene expression profile led to the hypothesis that loss of Nr4a1 contributes to the observed injury by an immune-mediated mechanism. To test this hypothesis, bone marrow (bm) cross transplantation studies were performed in 3 groups of irradiated (irr) animals: (1) bmFHH into irrFHH (FHH control); (2) bmFHH into irrNr4a1
-/-
; and (3) bmNr4a1
-/-
into irrNr4a1
-/-
(Nr4a1 control). The bmFHH into irrNr4a1
-/-
animals showed a significant attenuation of proteinuria starting as early as week 12 and continuing through week 24 (35±4.6 mg/24hrs/100gBW) compared to the Nr4a1
-/-
control (60±7.5). These data strongly suggest that the mechanism of Nr4a1
-/-
renal injury is mediated via an immune mechanism, likely due to the predisposition of the FHH to develop renal injury. Further studies are now underway to better understand the precise immune mechanism that promotes injury in this model.
Abstract 103: ACE2 Overexpression Prevents DOCA-Salt Hypertension by Modulating Oxidative Stress and Nitric Oxide in the Central Nervous System
Authors: Sriramula Srinivas , Xia Huijing , Lazartigues Eric
Published: July 2021

Elevated reactive oxygen species (ROS) in the central nervous system (CNS) through NADPH oxidase and diminished Nitric oxide (NO) levels are involved in the pathogenesis of hypertension. We previously reported that central Angiotensin Converting Enzyme 2 (ACE2) overexpression prevents the development of hypertension induced by DOCA-salt in a transgenic mouse model (syn-hACE2; SA) with human ACE2 targeted selectively to neurons in the CNS. While baseline blood pressure (BP; telemetry) was not different among genotypes, DOCA-salt treatment (1mg/g body wt DOCA, 1% saline in drinking water for 3 weeks) resulted in significantly lower BP level in SA mice (122 ±3 mmHg, n=12) compared to non-transgenic (NT) littermates (138 ±3 mmHg, n=8). To elucidate the mechanisms involved in this response, we investigated the paraventricular nucleus (PVN) expression of Nox-2 (catalytic subunit of NADPH oxidase), 3-nitrotyrosine, and endothelial nitric oxide synthase (eNOS) and anti-oxidant enzymes superoxide dismutase (SOD) and catalase in the hypothalamus. DOCA-salt treatment resulted in decreased catalase (95.2 ±5.6 vs. 113.8 ±17.6 mmol/min/ml, p<0.05) and SOD (4.1 ±0.4 vs. 5.9 ±0.2 U/ml, p<0.01) activities in hypothalamic homogenates of NT mice, which was prevented by ACE2 overexpression (141.8 ±9.9 vs. 142.1 ±9.2 mmol/min/ml and 5.9 ±0.3 vs. 7.9 ±0.2 U/ml, respectively). NT mice treated with DOCA-salt showed increased oxidative stress as indicated by increased expression of Nox-2 (61 ±5 % increase, n=9, p<0.001 vs. NT) and 3-nitrotyrosine (89 ±32 % increase, n=9, p<0.01 vs. NT) in the PVN which was attenuated in SA mice. Furthermore, DOCA-salt hypertension resulted in decreased phosphorylation of eNOS-ser1177 in the PVN (33 ±5 % decrease, n=9, p<0.05 vs NT) and this decrease was prevented by ACE2 overexpression. Taken together, these data provide evidence that brain ACE2 regulates the balance between NO and ROS levels, thereby preventing the development of DOCA-salt hypertension.
Abstract 109: Leptin Acts in POMC Neurons to Increase Sympathetic Nerve Activity
Authors: Harlan Shannon M , Morgan Donald A , Rahmouni Kamal
Published: July 2021

Leptin acts in the brain to decrease food intake and promote energy expenditure by increasing sympathetic nerve activity (SNA) to thermogenic brown adipose tissue (BAT). Leptin also increases SNA to other beds including kidney with implications for obesity-associated hypertension. We previously demonstrated the importance the arcuate nucleus (Arc) of the hypothalamus in mediating leptin-induced increases in regional SNA, but the specific neuronal population within the Arc that mediates these responses is unknown. We hypothesized that proopiomelanocortin (POMC) neurons of the Arc are critical for the increases in SNA in response to leptin. To test this, we generated mice lacking the leptin receptor (ObR) specifically in POMC neurons (ObR
flox/flox
/ POMC
Cre
). We used multifiber sympathetic nerve recording to assess the SNA effects of leptin. ObR
flox/flox
/POMC
Cre
mice exhibited a significantly (P<0.01) blunted renal SNA response to intravenous administration of 120 μg of leptin (4±18%) as compared to wild type controls (161±35%). Intracerebroventricular (ICV) injection of leptin (2 μg) also increased renal SNA in wild type mice (220±77%), but not in ObR
flox/flox
/POMC
Cre
mice (-1±13%). In contrast, the renal SNA response to ICV insulin (100 μU) was not different (P=0.27) in ObR
flox/flox
/POMC
Cre
mice (150±24%) as compared to littermate controls (195±68%) demonstrating that the loss of renal SNA response to leptin in mice lacking ObR in POMC neurons is a specific effect. Next, we investigated the requirement of ObR in POMC neurons for the sympathoexcitatory effects of leptin in other beds. Strikingly, ObR
flox/flox
/POMC
Cre
mice had a significantly (P<0.05) blunted lumbar SNA response to ICV leptin (14±22%) as compared to controls (226±38%). In addition, the BAT SNA response to ICV leptin was attenuated (P<0.01) in ObR
flox/flox
/POMC
Cre
mice (126±28%) relative to controls (309±80%). These results demonstrate the leptin receptor signaling in POMC neurons in the control of regional sympathetic nerve activity.
Abstract 113: Cytokines Expression in Hypothalamic Paraventricular Nucleus Account for Cardiovascular Deficits and Improved Functions after Training in Spontaneously Hypertensive Rats.
Authors: Masson Gustavo S , Costa Tassia S , Michelini Lisete C
Published: July 2021

Aim:
To analyze the sequence of adaptive responses induced by training (T) on both hemodynamic profile and cytokines gene expression within the paraventricular nucleus (PVN).

Methods:
SHR submitted to T (treadmill, 55% maximal capacity, 1h/day, 5 d/week) or kept sedentary (S), were cannulated for measurement of pressure (BP), heart rate (HR) and baroreflex sensitivity (BrS) at rest on weeks 0, 1, 2, 4 and 8 of protocols. HR and BP variability (power spectral analysis) were also evaluated. After euthanasia, brains were removed for isolation of PVN. Total mRNA was extracted for quantification of TNFα and IL-6 expression by real time PCR (HPRT as the endogenous gene). WKY were used as time controls. One-way ANOVA, with Fisher post-hoc test and Pearson correlation were employed.

Results:
At the beginning of the protocols, SHR exhibited high mean BP and HR, decreased BrS, increased BP variability and decreased HR variability (170±3 mmHg, 356±7 b/min; 2.3±0.2 b/min/mmHg, 14±2 mmHg
2
and 14±2 ms
2
) when compared to WKY (119±2 mmHg, 307±6 b/min; 3.6±0.3 b/min/mmHg, 6±1 mmHg
2
and 22±2 ms
2
, respectively). Pro-inflammatory cytokines expression in PVN are also elevated in SHR
vs
WKY (TNF-α= 1.9±0.3
vs
1.2±0.1, IL-6= 4.9±0.8
vs
1.1±0.3 UA). In the SHR, T caused significant increases on BrS (4.0±0.2 b/min/mmHg, T
2
-T
8
), HR variability (21±1 ms
2
, with a 60% increase of HF component from T
4
-T
8
) and prevented the increase on BP variability in the SHR (13±2
vs
20±3 mmHg
2
, T
8
vs
S
8
, with 29% reduction of LF component at T
8
). T caused a prompt normalization of TNFα and IL-6 mRNA expression within the PVN (1.3±0.2 and 0.9±2 UA at T
2
), accompanied by HR reduction (338±4 b/min, T
4
-T
8
) and mean BP fall (159±6 mmHg, T
8
), with significant correlations between TNFα x HR variability (Y=-3.6x+24.3, r= -0.29; P=0.04), TNFα x BrS (Y=-0.7x+4.3, r=-0.35; P=0.01), TNFα x BP (Y=9x+140, r= 0.26; P=0.04) and TNFα x HR (Y=18x+309, r= 0.29; P=0.03). PVN IL-6 content was also correlated with HR variability (Y=-1.8x+21.5, r=-0.36; P=0.01), BrS (Y= -0.3x+4.2, r=-0.44; P<0.001) and HR (Y=7x+324, r=0.30; P=0.02).

Conclusions:
Data suggested the involvement of PVN pro-inflammatory cytokines in the mediation of cardiovascular deficits in the SHR and the efficacy of T to prompt normalize these deleterious effects.
Abstract 118: Fetal Betamethasone Exposure Enhances the Peptide Ratio of Angiotensin II to Angiotensin-(1-7) in the Dorsal Medulla of Adult Sheep
Authors: Marshall Allyson C , Shaltout Hossam A , Nautiyal Manisha , Chappell Mark C , Diz Debra I
Published: July 2021

Betamethasone (BMS) is administered to women entering into early preterm labor to facilitate fetal lung development and decrease infant mortality; however, fetal steroid exposure may lead to deleterious consequences long term. In a sheep model of BMS exposure in utero, we show elevated mean arterial pressure (MAP) and decreased baroreflex sensitivity (BRS) for control of heart rate at 6- and 20-months (mo) of age. The changes in MAP and BRS are associated with alterations in components of both the circulating and intra-renal renin-angiotensin systems (RAS) that may ultimately shift the balance to the pro-hypertensive peptide, angiotensin (Ang) II. In the brain solitary tract nucleus, Ang II actions through the AT
1
receptor oppose the beneficial actions of Ang-(1-7) at the
mas
receptor for BRS regulation. We found decreased expression of the
mas
protein in the dorsal medulla of BMS-exposed animals at 6- and 20- months of age with no changes in the AT
1
receptor expression. To determine whether tissue Ang peptides may contribute to altered BRS, the present study utilized three selective radioimmunoassay (RIAs) to quantify Ang I, Ang II and Ang-(1-7) in the dorsal medulla of 6-mo old male sheep. Comparisons of tissue peptide concentrations were not significantly different between groups; however, BMS-exposed offspring have a significant increase in the ratio of Ang II to Ang-(1-7) (Control: 0.61 ± 0.11, BMS: 2.3 ± 0.62; p<0.05). There was a significant increase in the ratio of Ang II to Ang I (Control: 0.66 ± 0.20, BMS: 2.35 ± 0.46), with no change in the ratio of Ang-(1-7) to Ang I. The altered peptide ratios potentially reflect a decrease in the ACE2 enzyme, which converts Ang II to Ang-(1-7), and an increase in ACE activity which concerts Ang I to Ang II. Future studies are required to determine enzymatic activity in dorsal medullary tissue, but data are consistent with previous observations on altered ACE and ACE2 in the circulation and kidney of BMS-exposed animals. We conclude that BMS-depedent fetal programming may influence the central RAS through alterations in the balance of Ang II and Ang-(1-7) that contribute to the elevated MAP, lower BRS, and altered renal function in this model. Support: HD0474584
Abstract 15: Increased Intrinsic Stiffness of Aortic Vascular Smooth Muscle Cells as a Mechanism for Increased Aortic Stiffness in Spontaneously Hypertensive Rats
Authors: Sehgel Nancy L , Zhu Yi , Gao Shumin , Sun Zhe , Trzeciakowski Jerome P , Hong Zhongkui , Hunter William C , Vatner Dorothy E , Meininger Gerald A , Vatner Stephen F , Qiu Hongyu
Published: July 2021

An increase in vascular stiffness is a fundamental component of hypertension. Our hypothesis is that increased large artery stiffness in hypertension is due in part to intrinsic mechanical properties of vascular smooth muscle cells (VSMC). Adult spontaneously hypertensive rats (SHR) (16 weeks old) and age-matched Wistar-Kyoto normotensive (WKY) rats were studied. Aortic pressure, measured with a Millar catheter, was higher in SHR than WKY (MAP 123±4 versus 96±6 mmHg,
p
<0.05). Aortic stiffness, measured with pulse wave velocity, in vivo, was 2 to 2.5 fold higher in SHR rats (7.65 mm/ms) compared to normotensive rats(3.27 mm/ms,
p<0.01
). Aortic VSMCs were isolated from the SHR and WKY rats. Stiffness in single VSMC was continuously recorded in vitro using atomic force microscopy and temporal oscillations in VSMC elasticity, which represent the dynamic properties of elasticity, were observed and analyzed by Eigen decomposition. VSMC stiffness was consistently increased by 1.5 to 2 fold in SHR (24.0 kPa) versus WKY (11.8 kPa) (
p<0.05)
. Moreover, the oscillations in elasticity were significantly different between the two groups in terms of frequency and amplitude (
p<0.05)
. ML-7, a highly specific inhibitor of myosin light chain kinase (MLCK), produced a greater reduction in the VSMC elasticity from SHR aorta compared to WKY rats (
p<0.05
), and also eliminated the difference between the SHR and WKY. Furthermore, ML-7 altered the oscillation in VSMC elasticity in SHR, but not WKY. These observations indicate that there is an inherently altered VSMC stiffness in hypertension that is associated with unique dynamic cytoskeletal behavior. These dynamic elements may be involved mechanistically in increasing aortic stiffness in SHR rats. MLCK participates in the regulation of VSMC stiffness in hypertension via altering both basal VSMC elasticity and temporal oscillations.
Abstract 150: Disparities in Blood Pressure Screening in Adults: An Analysis of NAMCS-NHAMCS Data
Authors: Nair Madhuri , Thomas Joshua R , Chandrasekharan Gopal , Zhao Yan D , Fox Mark D
Published: July 2021

BACKGROUND:
Blood pressure (BP) screening in adults has been identified by the US Preventive Services Task Force as a Grade A recommendation.

OBJECTIVE:
To compare BP screening rates in adult patients among payer sources and provider types in two national probability samples of outpatient office visits.

DESIGN/METHODS:
An analysis of visit data for all patients 18 years and older in the National Ambulatory Medical Care Survey (NAMCS) and National Hospital Ambulatory Medical Care Survey-Outpatient Department (OPD) during the years 2005-2009 was performed. Data on 279,510 patient visits were analyzed, including adults seen in ambulatory or office-based settings (NAMCS, N=147,675) and hospital outpatient departments (OPD, N=131,835). BP screening rates were compared based on provider type (Family Practice, Internal Medicine, Obstetrics-Gynecology, and Cardiovascular Specialist) and payer source (Private Insurance, Medicare, Medicaid, and Self Pay).

Results:
Overall, BP screening rates were higher in the OPD sample (65.70% vs, 55.83%, p < .001), despite a higher mean age and higher proportion of privately insured or Medicare patients in the NAMCS sample. The odds ratio for BP screening was 1.538 for insured patients compared to self-pay in the NAMCS sample (95% CI: 1.473-1.606); however, the disparity was not observed in the OPD data. Cardiovascular specialists demonstrated the most adherence to BP screening, regardless of payer source. Overall BP screening rates are shown in Table 1.

Conclusions:
Analysis of two national probability samples reflected marked disparity in BP screening in adult patients based on payer source and provider type.
Abstract 153: Born with a Single Kidney versus Nephrectomy: Similar End Point, Different Mechanism of Injury
Authors: Wang Xuexiang , Johnson Ashley , Lee Jonathan , Solberg-Woods Leah , Garrett Michael R
Published: July 2021

A relatively common abnormality of the urogenital tract in humans is the development of only a single kidney (1:500 to 1:1000). Clinical studies suggest that patients born with a single kidney can develop proteinuria, hypertension, and even renal failure later in life. In contrast, studies in children who undergo nephrectomy or adults who serve as kidney donors appear to exhibit little difference in renal function compared to two-kidney subjects. Invasive techniques such as nephrectomy or renal ablation have been used to generate animal models to recapitulate this human congenital disorder. The progression of injury in these models is attributed to hyperfiltration which refers to changes in hemodynamics that cause glomerular damage leading to hypertension. Recently, our lab developed a new genetic animal model [heterogeneous stock derived model of unilateral renal agenesis, (HSRA)] that develops with a single kidney in 50-75% of offspring. The model is characterized by reduced nephron number, kidney hypertrophy, and renal injury that leads to a decline in renal function. Time course evaluation of blood pressure, renal hemodynamics, and renal injury was performed in 4 groups; (1) HSRA-S (1-kidney), (2) HSRA-C (2-kidney littermates), (3) HSRA-UNX3 (uninephrectomy-week 3) and (4) HSRA-UNX8 (uninephrectomy-week 8). Nephrectomized animals demonstrated hyperfiltration, whereas single kidney animals (HSRA-S) did not. This suggests a different pathophysiological mechanism of injury between congenital and nephrectomized rats. At later time points, proteinuria for HSRA-UNX3 (82±22.9 mg/24h) and HSRA-UNX8 (46±18.1) were significantly higher than HSRA-C (11±6.4), while HSRA-S (109±15.7) demonstrated the highest proteinuria. GFR was lowest in HSRA-S (656±123.9 ul/min/gKW), followed by HSRA-UNX3 (1151±112.4), HSRA-UNX8 (1309±98.3) and HSRA-C (1544±111.7). Microarray studies have identified several developmental genes (
Hox5b
,
Smoc2
and c-
Kit
) that may be linked to reduced nephron number and other structural changes that could predispose the HSRA-S to kidney injury later in life. These results demonstrate that rats born with a single kidney are more prone to renal injury than nephrectomized rats and the mechanism is likely different.
Abstract 160: Characterization of a Comt Knock-out Rat for Blood Pressure and Associated Phenotypes.
Authors: Casati Marc , Hoffman Mathew , Schilling Rebecca , Flister Michael , Geurts Aron M , Dwinell Melinda , Lazar Jozef , Jacob Howard J , Hernandez Isabel , Moreno Quinn Carol P
Published: July 2021

The catecholamine system plays an important role in the control of blood pressure and sodium excretion. One of the inactivation pathways of catecholamines is the enzymatic metabolism by catechol-O-methyltransferase (Comt). There are conflicting data regarding the role of Comt on the development of salt-sensitive hypertension, so the goal of our study was to evaluate the importance of
Comt
in the context of a susceptible background
in vivo.
Comt
was mutated in the Dahl SS rat by zinc finger nucleases (ZFNs) injections targeting the sequence CTGTTCCAGGTCACCATCctcaatGGGGCATCCCAGGATCTT into SS/JrHsdMcwi (Dahl S) rat embryos. The resulting mutation was a 14-bp frameshift deletion in exon 4. Conscious blood pressure was measured by telemetry on male and female Comt knockout and wild type (WT) rats on low salt diet (0.4% NaCl) and during three weeks of high salt diet (8%NaCl). Disruption of Comt caused the protein not to express in the kidneys of the Dahl S rat. There were no differences in mean arterial pressure (MAP) between the
Comt
-/- and the
Comt
+/+ male rats at any time point during the day-night cycle at low or high salt diet. Body and organ weights, and protein and electrolyte excretion was also unchanged by the
Comt
mutation. On the other hand, female
Comt
-/- rats evidenced a higher MAP, which was only significantly higher at night during low salt diet (112±2 mmHg in
Comt
+/+ vs 125±2 mmHg in the
Comt
-/-, n>6) and both during day and night after 21 days of high-salt diet (∼ 30 mmHg difference between
Comt
+/+ and
Comt
-/- strains at both day and night). Systolic blood pressure differences were mostly responsible for the observed blood pressure diferences in females KO of the Comt gene, despite blood pressure effect, was not followed by a parallel difference in urine flow, electrolyte excretion or renal damage (protein and albumin excretion). In conclusion, we are the first ones to show that disruption of
Comt
enhances salt-sensitive hypertension in a gender-dependent manner.
Abstract 165: Angiotensinogen M235T Modifies the Relationship between PAI-1 and Renal Blood Flow in Caucasian Hypertensive Individuals
Authors: Shafiq Qaiser , Sun Bei , Williams Jonathan S
Published: July 2021

Studies have reported an association of angiotensinogen (AGT) gene polymorphisms with hypertension and renal blood flow (RBF). The T allele of rs699 (AGT 235T) is associated with increased levels of angiotensinogen and reduced RBF. Plasminogen activator inhibitor-1 (PAI-1) is an inhibitor of the fibrinolytic pathway. Elevated PAI-1 is associated with increased renin-angiotensin system (RAS) activity and cardiovascular damage. We hypothesized that hypertensive individuals with the TT genotype of AGT 235 would display higher levels of PAI-1. We further hypothesized an inverse relationship would be observed between PAI-1 and RBF, and that the strength of this correlation would be dependent on genotype status. A total of 159 Caucasian hypertensive participants (mean age 49.3y, BMI 27.6) had data available for AGT235, PAI-1 and RBF from the HyperPATH study. RBF was measured by para-aminohippurate clearance method while on a liberal salt diet (>200 mmol Na/day for 1 week). Genotype frequencies were in Hardy-Weinberg Equilibrium (MM 41/MT 94/TT 24). Multivariate regression demonstrated the TT genotype was associated with higher PAI-1 levels compared with MT + MM (41.0±8.1 v. 26.9±7.1ng/ml, p=0.005). Genotype status modified the relationship between PAI-1 and RBF whereby only the TT genotype showed a significant correlation after adjusting for age, BMI and BP (TT pr2=0.22, Beta= -1.66 p=0.001 v. MT+MM pr2=0.05, Beta= -0.82, p=0.21). AGT 235T is associated with elevated PAI-1 and influences the relationship between PAI-1 and RBF in Caucasian hypertensives. This provides mechanistic clues to explain the genetic underpinnings involving AGT polymorphisms and hypertension.
Abstract 177: Obesity Alters Circulating T Cell Profile in Female African American Teenagers
Authors: De Miguel Carmen , Wang Xiaoling , Ownby Dennis , Pollock Jennifer
Published: July 2021

Extensive evidence supports a role of the immune system in the development of cardiovascular disease. Moreover, at-risk cardiovascular patients display pro-inflammatory mediators years before disease is openly declared. Given the greater prevalence of obesity among African Americans, we hypothesized that obesity leads to alterations in the circulating immune cell profile in a population of African American teenagers. Lean (BMI < 50th percentile) and obese (BMI ≥ 95th percentile) African American teenagers (14-20 years-old; n=13-22/group) of both genders were recruited from public schools of Augusta, GA. Cell differentials and circulating specific immune cell populations were analyzed by fluorescence-activated cell sorting or FACS flow cytometry. Whole blood cell counting with five differentials (including neutrophils, lymphocytes, monocytes, eosinophils and basophils) were conducted in all the subjects. We did not observe any significant difference in the proportion of these five major cell types between obese cases and lean controls either in the overall samples or in the sex-stratified analysis. Obese African American females presented significantly elevated percentages of circulating T lymphocytes (CD3+ cells: (34.6 ± 1.8% vs. 28.1 ± 2.2%; p<0.05) and T helper cells (CD4+ cells: 29.37 ± 1.5% vs. 26.4 ± 1.5%; p<0.05) when compared to lean females. African American obese females also tended to have greater percentages of activated CD3+ cells (CD3+/CD69+ cells) than lean females, although significance was not reached (0.67 ± 0.2% vs. 0.39 ± 0.1%). No significant differences were found between African American male obese and lean subjects in any of the circulating immune cell populations. In conclusion, obese female African American teenagers present higher circulating percentages of CD3+ and CD4+ cells than their lean counterparts without changes in the percentages of total lymphocytes. These results suggest that certain subsets of T cells may be key players in obesity-related immune disorders. The identification of the obesity-related T-cell activation may provide new etiologies and biomarkers for obesity-induced cardiovascular disease.
Abstract 180: Interleukin (IL) 16 Acts as a Regulator of Skin Electrolyte Metabolism and Blood Pressure
Authors: Jantsch Jonathan , Schröder Agnes , Brenner Sven , Rakova Natalia , Goss Jennifer , Cruikshank William W. , Titze Jens
Published: July 2021

We showed that Na
+
is stored in the skin without commensurate water retention. In response to interstitial electrolyte load, mononuclear phagocyte system (MPS) cells secrete VEGF C, as regulated by tonicity enhancer binding protein (TonEBP) attaching to the VEGF C promoter. VEGF C induces hyperplasia of skin lymphcapillaries. As IL16 harbors a TonEBP binding site, we tested the hypothesis that IL16 participates in this process. We cultured bone marrow-derived macrophages in isotonic or hypertonic cell culture media and measured IL16 expression. We fed male FVB mice a low salt diet (LSD; < 0.1% NaCl + tap) or a high salt diet (HSD; 4% NaCl + 0.9% saline) for two weeks. We treated the groups with either IL16 specific antibodies or no antibody. After two weeks we measured mean arterial blood pressure (MAP), skin Na
+
, Cl
-
, and water content (rSKNa, rSKCl, rSKW), quantified lymphcapillary density, VEGF C and TonEBP expression. IL16 was specifically secreted from macrophages in response to hypertonicity
in vitro
. HSD increased skin electrolytes, activated TonEBP/VEGF C, and resulted in lymphcapillary hyperplasia. Anti IL16 treatment under LSD increased skin electrolyte, water content and MAP. The IL16 antibody blunted the VEGF C driven lymphcapillary response despite salt load. No differences in skin electrolytes and MAP were found between IL16 antibody treated and untreated mice after HSD. We conclude that IL16 is a regulator of electrolyte and water metabolism and MAP, even in the absence of HSD. The finding that the IL16 promoter contains a TonEBP binding site suggests that IL16 could act as an osmoprotective gene that is involved in regulation of electrolyte homeostasis and MAP control.
Abstract 19: Chronic Infusion of Angiotensin-(1-7) into the Lateral Ventricle of the Brain Improves Cardiovascular and Metabolic Parameters in Fructose-Fed Rats
Authors: Guimaraes Priscila S , Oliveira Mariana F , Braga Janaina F , Santos Robson A , Campagnole-Santos Maria J
Published: July 2021

We evaluated whether chronic intracerebroventricular (ICV) infusion of angiotensin (Ang)-(1-7) improves cardiovascular and metabolic parameters in fructose-fed rats. Six-weeks-old Sprague-Dawley rats were fed with either normal rat chow (CTL, n=11-13) or the same diet plus 10% fructose in drinking water (FRUT, n=7-10) for 10 wks. On the 6
th
wk of fructose diet, a set of rats were subjected to ICV infusion of Ang-(1-7) (FRUT-A7; 200 ng/ hour; n=7-9) for 28 days. Baseline mean arterial pressure (MAP), cardiac baroreflex sensitivity (BRS, phenylephrine bolus
iv
), and the cardiac autonomic tone (pharmacological blockade of β-adrenergic and muscarinic receptors) were evaluated. Another group of rats were subjected to glucose tolerance test (2 g/kg rat, 50% solution
ip
) and extraction of blood and tissues after 6-8 h fasting. Compared to CTL, FRUT had reduced BRS (0.53 ± 0.05 vs 0.96 ± 0.08 ms/mmHg), with no changes in MAP, and cardiac autonomic tone. Moreover, FRUT displayed hyperglycemia (142 ± 2 vs 128 ± 3 mg/ dL), hyperinsulinemia (1.8 ± 0.1 vs 1.3 ± 0.1 ng/ ml), hyperleptinemia (5 ± 0.2 vs 3 ± 0.5 ng/ ml), and hypertriglyceridemia (106 ± 13 vs 79 ± 5 mg/ dL). These changes were followed by impaired response to tolerance glucose test (258 ± 19 vs 177 ± 39 mg/ dL), sustained for 60 min, and liver hypertrophy (33 ± 1 vs 29 ± 0.5 mg/ g body weight) with lower hepatic glycogen levels (9 ± 2 vs 15 ± 1 mg/ g liver). In contrast, FRUT-A7 rats had normalized BRS (0.96 ± 0.07 ms/ mmHg), reduced cardiac sympathetic tone (20% change vs CTL and FRUT), with no change in parasympathetic tone or MAP. Similar to FRUT, FRUT-A7 showed hypertriglyceridemia (115 ± 9 mg/ dL), and hyperleptinemia (5 ± 0.4 ng/ ml). However, serum insulin (1.4 ± 0.1 ng/ ml) and glucose (129 ± 4 mg/ dl), response to tolerance glucose test (178 ± 41 mg/ dL), liver relative weight (30 ± 0.7 mg/ g) and glycogen deposition (14 ± 1.8 mg/ g) were normalized in these rats. Total cholestherol (∼10%) and high density lipoprotein (∼20%) were increased in FRUT-A7 (p<0.05). There was no change in glucagonemia, body weight gain, or relative weight of left ventricle, and adipose tissues among all groups. These data showed that chronic ICV infusion of Ang-(1-7) produced important benefits on cardiovascular and metabolic parameters in fructose-fed rats.