bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 040: Comparative Genomic Analysis and Functional Insights for Blood Pressure-Associated Single Nucleotide Polymorphisms
Authors: Liu Yong , Mishra Manoj , Liang Eugene , Geurts Aron , Auer Paul W. , Liu Pengyuan , Rao Sridhar , Greene Andrew , Liang Mingyu
Published: September 2019

Genetic factors play an important role in the regulation of blood pressure (BP) and the development of hypertension. Recent genome-wide association studies (GWAS) in large cohorts have identified associations of hundreds of single nucleotide polymorphisms (SNPs) with human BP. Understanding the molecular function of these SNPs and linking them to BP regulation are among the most pressing challenges in BP research. Animal models, including mouse models, are essential tools for studying BP regulatory mechanisms. In this study, we performed a thorough analysis of the most comprehensive set of human BP-associated SNPs to date and experimental validation. We mapped 26,585 human BP-associated SNPs in 1,071 linkage disequilibrium (LD) regions to 9,447 syntenic regions in the mouse genome. 6,551 of these sites have the same nearest protein-coding genes in human and mouse. 87.0% (932) of the 1,071 LD regions do not contain any nonsynonymous SNP, including 21.8% (234) that are at least 10 kb from any protein-coding gene. Approximately 300 BP-associated SNPs are expression quantitative trait loci for a few dozen known BP physiology genes in tissues including specific kidney regions that are physiologically important for BP regulation. CCCTC-binding factors (CTCF) are essential for regulating long-distance actions of enhancers. Using a newly developed deep learning model, we identified 5 BP-associated SNPs that might influence the conservation of CTCF binding across 40 cell types, and two of them are predicted to have strong effect on CTCF binding. Using SpliceAI, a deep neural network model for predicting the effect of SNPs on mRNA splicing, we identified 19, 9, 22, and 13 SNPs that have a delta value of more than 0.2 for acceptor gain, acceptor loss, donor gain, and donor loss, respectively. Several SNPs are predicted to influence secondary structure of mRNAs and long noncoding RNAs. We experimentally validated that synonymous SNP rs9337951, predicted to influence the secondary structure of its host mRNA JCAD, influences JCAD protein expression. These findings provide novel insights into the potential functional significance of human BP-associated SNPs and form an extensive basis for developing experiments to test such significance in human cells and animal models.
Abstract 042: Carotid Body Ablation of TRPV1 Reduces Chemoreflex Sensitivity in Rats With Heart Failure
Authors: de Morais Sharon D , Zucker Irving H , Schultz Harold D
Published: September 2019

Resiniferatoxin (RTX), an ultrapotent analogue of capsaicin, activates and destroys TRPV1 expressing cells. RTX in the CB may reduce abnormal breathing patterns seen in rats with HF. The purpose of this study was to examine whether TRPV1 ablation in the CB of HF rats alters chemoreflex-evoked ventilation and cardiovascular function. We used the post-MI model of HF in male SD rats (200-250g). BP (radiotelemetry), cardiac function (echocardiography) and ventilation (plethysmography measured under hypoxic (10% and 15% of O2) and hypercapnic (5% and 7% of CO2) conditions) before and 4 weeks after application of vehicle or RTX (50ug/ml) to the CB. After 4 weeks of MI, the rats showed ejection fraction of 35.9±1.6% demonstrating HF condition. Immunofluorescence showed lower expression of TRPV1 in RTX treated comparing to vehicle-treated CBs. RTX treatment (n=4) reduced baseline MAP (Veh=93.3±2.3 vs RTX=87.1±1.4, p<0.001) and HR (Veh=335.8±10.5 vs RTX=297±10.2, p<0.001) and normalized the increased ventilatory response to hypoxia or hypercapnia in HF rats (Figure). Poincare analyses showed an increase in the breath-breath variability 4 weeks after treatment with veh (SD1=95.9±14.3 vs 198.2±31.9; SD2=135.3±20.2 vs 279.9±45.1), which was blunt by RTX (SD1=87.5±17.7 vs 97.2±23.6; SD2=123.4±25 vs 137.1±33.4; p<0.05). No changes in BP, HR and body temperature were observed during hypoxia and hypercapnia in CHF rats, treated or not with RTX. These data show that CB RTX treatment in HF rats reduces chemoreflex sensitivity. These data implicate an enhanced role for TRPV1 signaling in chemoreflex function in HF.
Abstract 043: Local Passive Heat in the Treatment of Nocturnal Supine Hypertension in Autonomic Failure
Authors: Okamoto Luis E , Celedonio Jorge E , Smith Emily C , Gamboa Alfredo , Shibao Cyndya A , Diedrich Andre , Paranjape Sachin , Black Bonnie K , Muldowney James A , Peltier Amanda C , Habermann Ralf , Crandall Craig G , Biaggioni Italo
Published: September 2019

Supine hypertension affects about half of patients with primary autonomic failure (AF), complicates the treatment of orthostatic hypotension (OH) and induces nocturnal polyuria, resulting in worsening of morning OH. We previously showed that short exposure to local passive heat (40-42°C applied for ≤2 hours over abdomen and pelvis with a heating pad) acutely decreased systolic blood pressure (BP) by -25±5 mmHg in AF patients with supine hypertension due to decreases in stroke volume (-29±5%) and cardiac output (-30±5%). In this study, we hypothesized that local passive heat therapy applied overnight would lower nocturnal BP and polyuria, and improve early morning OH in these patients. Ten AF patients with supine hypertension (age 76±2 years, 6 men, supine systolic BP 168±3 mmHg) received passive heat (38°C with a water-perfused heating pad placed under the torso) or placebo, in a 2-night crossover study. Supine BP was monitored every 2 hr from 8pm-8am. Heat therapy was applied from 10pm-6am. Morning orthostatic tolerance was assessed at 8 am. Systolic BP significantly decreased during overnight heat therapy compared to placebo (Figure; P<0.01 by mixed-effects model) with a maximal reduction of 30±6 mmHg at 4 hours of heat. Despite lowering nighttime BP, heat therapy did not decrease nocturnal diuresis or improve morning OH. In conclusion, low levels of local heat therapy applied overnight effectively lowers BP in AF patients with nocturnal supine hypertension. Thus, local heat therapy offers a novel non-pharmacologic approach to treat this condition, but future studies are needed to assess the long-term safety of this approach.
Abstract 046: Chronic Nicotine Inhalation Promotes the Development of Hypertension
Authors: Xu Jiaxi , Morris Tamara M , Oakes Joshua M , Gardner Jason D , Yue Xinping , Lazartigues Eric
Published: September 2019

While smoking is a proven risk factor for stroke and heart attacks, whether it contributes to the development of hypertension is still unclear. Nicotine inhalation is thought to activate the sympathetic nervous system and could contribute to the development of dysautonomia and blood pressure (BP) dysregulation. The purpose of this study was to investigate a possible relationship between chronic nicotine inhalation and the development of hypertension. C57BL/6 mice (male, 8-12 weeks of age) were exposed to air or nicotine vapor (daily, 12 h on/off) for 8 weeks. BP was recorded weekly for 24 h (telemetry). Nicotine exposure was assessed by weekly measurement of serum cotinine levels, which showed an average of 599.0 ±54.3 ng/ml in nicotine-exposed mice. Nicotine exposure induced a time-dependent increase in systolic BP. Notably, 24-h BP recordings showed that by the end of the 7
th
week, systolic BP of nicotine-exposed mice was more often in the hypertensive zone (as defined by the area under the curve [AUC] above the 130 mmHg threshold within a 24-h recording) compared to their air-exposed counterparts (AUC: 555 ±299
vs.
112 ±30,
P=0.012
, n=12). This hypertensive response was validated in angiotensin-II (Ang-II)-infused mice (450 ng/kg/min, s.c., AUC: 1358 ±939
vs.
112 ±30,
P<0.001
, n=5). At the end of nicotine exposure, urinary norepinephrine (NE) showed significant increase in both nicotine (108.9 ±9.8
vs.
79.53 ±5.7 ng/1ng creatinine,
P=0.018
, n=9) and Ang-II groups (114.0 ±14
vs.
79.53 ±5.7 ng/1ng creatinine,
P=0.016
, n=9), suggesting increased sympathetic activity in those mice. In addition, mice with global deletion of angiotensin converting enzyme type 2 (ACE2) showed a dramatic elevation in urinary NE level after nicotine exposure (157.8 ±8.5
vs.
79.53 ±5.7 ng/1ng creatinine,
P<0.001
, n=9). These data suggest that ACE2 might be protective against nicotine inhalation-mediated rise in sympathetic activity. In summary, chronic nicotine inhalation promotes the development of hypertension, and this could be mediated through the elevated activity of sympathetic nervous system. Experiments are ongoing to confirm sympatho-excitation and a possible involvement of the renin-angiotensin system in chronic nicotine inhalation-induced hypertension.
Abstract 053: Gene Editing Yields a Novel Rat Model of Cardiometabolic Disease
Authors: Clark Karen C , Wagner Valerie A , Malek Jacob A , Arroyo Alyssa A , Grobe Justin L , Kwitek Anne E
Published: September 2019

Despite strong evidence that the Metabolic Syndrome (MetS) is highly heritable, the genetic etiology is complex and few causative genes are known. Using the Lyon Hypertensive (LH) rat we previously identified a novel candidate MetS gene (
C17h6orf52
) as a putative master regulator of gene expression with pleiotropic effects on body weight and blood pressure regulation.

In this study, CRISPR-Cas9 gene editing was used to generate mutations in
C17h6orf52
, and females from the resulting strains, designated M1 and M2, were characterized, with all measures presented as mean±SEM. Lean and fat body mass as a percentage of total body composition was measured via nuclear magnetic resonance (NMR), and resting metabolic rate (RMR) was measured via indirect calorimetry. Blood pressure (BP) measurements were obtained via radiotelemetry. Surprisingly, the M1 and M2 mutations in
C17h6orf52
result in different splicing variants that are shown to differentially modulate several cardiometabolic phenotypes.

The M1 mutation causes significant changes in BP, body composition and RMR, without affecting body weight. The M2 mutation significantly increased body weight and adiposity without changing BP or RMR. These data suggest that
C17h6orf52
is protective against cardiometabolic risk factors, given that the mutation confers a greater burden of MetS features on an already obese and hypertensive rat. The continued study of this rat model of MetS has the potential to functionally validate an uncharacterized regulatory gene and provide novel targets for pharmacological intervention in the treatment of cardiometabolic diseases.

*
p<0.05 by Students t-test

&

p<0.05 by ANCOVA

$

p<0.05 by 2-WAY ANOVA
Abstract 061: Systolic Blood Pressure is Maintained Despite Varying Degrees of Acute Caloric Restriction and Sodium-Glucose Cotransporter-2 Inhibition in Obese, Insulin Resistant Rats
Authors: Cornejo Manuel A , Nakano Daisuke , Nishiyama Akira , Ortiz Rudy M
Published: September 2019

A 5-10% reduction of body mass by caloric restriction (CR) has been effective in normalizing SBP in hypertensive patients. Moreover, SGLT2 inhibitors lowered SBP in spontaneous hypertensive rats and are considered a therapeutic alternative to patients with T2DM concomitant with hypertension. However, these potential benefits have not been fully addressed in a model of metabolic syndrome, which is characterized by multiple factors. Lean LETO and obese, insulin resistant OLETF rats from 2 independent studies were divided in the following groups (n=7): 1) ad lib LETO, 2) ad lib OLETF, 3) LETO 50% CR (10 d at 15 wks of age), 4) OLETF 50% CR, 5) LETO 30% CR (14 d at 12 wks of age), 6) OLETF 30% CR, 7) OLETF SGLT2i (Luseoglifozin 10 mg/kg/day x 4 wks at 10 wks of age), and 8) OLETF SGLT2i + 30% CR. SBP was measured weekly by tail-cuff plethysmography.SBP increased 19% (23 mmHg,
P
=0.002) consistently in OLETF compared to LETO (124 mmHg), but neither the CR-induced reduction in body mass nor the supplementation of SGLT2i modified the strain-associated increase in SBP (Fig. 1). Mean body mass in OLETF decreased by 18% with 50% CR (409 to 334 g) but decreased only 7% (34 g) with 30% CR and 8% (38 g) with SGLT2i and CR (454 g control). Serum Na
+
remained unaltered by 50% CR. Urinary Na+ excretion also was unchanged in 30% CR. While the CR treatments induced profound changes in body mass (and adiposity, not shown), they were not sufficient to alter Na+ balance and SBP. Furthermore, the addition of SGLT2i did not reduce SBP suggesting that in the OLETF model of metabolic syndrome factors beyond adiposity and impaired glucose regulation contribute to the maintenance of elevated arterial pressure.
Abstract 062: Elevated Cerebrospinal Fluid Sodium Concentration in Hypertensive Humans
Authors: Souza Lucas A , Trebak Fatima , Kumar Veena , Satou Ryousuke , Kehoe Patrick G , Yang Wei , Wharton Whitney , Feng Yumei
Published: September 2019

High salt intake has been linked to the pathogenesis of hypertension and increased sympathetic activity, considered a hallmark of human hypertension. Recent studies showed that high-salt diets may lead to sequestration of sodium into interstitial tissues. Increased CSF sodium contributes to elevated sympathetic activity and high blood pressure (BP) in rodent models of hypertension. However, it is not known if there is an enhanced sequestration of sodium into cerebrospinal fluid (CSF) of chronic hypertensive humans. To address this question, we obtained CSF and plasma samples from 43 middle-aged, normotensive (n=26) and hypertensive (n=17) individuals. The sodium concentration ([Na
+
]) in plasma and CSF were measured using flame photometer. Ambulatory BP was monitored every 20 minutes while the participant was awake and once per hour while the participant was asleep over 24-hour period. Participants were deidentified and data were analyzed in conjunction with a retrospective analysis of patient history and diagnosis. Participants with uncontrolled hypertension had higher BP levels compared with the normotensive participants (systolic BP: 135.5±2.7 mmHg vs. 114.3±1.5 mmHg; diastolic BP: 81.9±2.1 mmHg vs. 71.6±1.2 mmHg,
P
<0.0001). Importantly, we found that CSF [Na+] was significantly higher in participants with uncontrolled hypertension (149.3±3.5 mM) compared with those of normotensive (139.5±2.12 mM,
P
=0.0156); while there was no difference in plasma [Na+] between groups (136±0.76 mM vs. 135.5±1 mM,
P
=0.73). Further multiple linear regressions analysis controlling for age, sex, race and body mass index revealed a significant positive correlation between CSF [Na
+
] and blood pressure levels (sBP: R
2
=0.4189,
P=
0.0007; dBP: R
2
=0.205,
P
=0.032), but showed no correlation between plasma [Na
+
] and BP. Furthermore, we found that plasma and CSF [Na
+
] were not correlated with each other (R
2
=0.0002174,
P
=0.9252), suggesting that increases in CSF [Na
+
] in hypertensive humans do not reflect simple passive diffusion from the circulation. In sum, CSF [Na
+
] was higher in chronic hypertensive humans and CSF sodium may play a key role in the pathogenesis of human hypertension.
Abstract 066: Beta-Hydroxybutyrate Synthesis Contributes to the Anti-Vascular Aging Effect of Autophagy
Authors: Mccarthy Cameron G , Chakraborty Saroj , Schreckenberger Zachary , Wenceslau Camilla F , Joe Bina
Published: September 2019

Diminished autophagy leads to the accumulation of debris, dysfunctional organelles, and consequently, cellular aging. Premature vascular aging occurs in hypertension. In fact, many factors that contribute to the deterioration of vascular function as we age are accelerated in hypertension. Previously, we have reported decreased autophagic activity in arteries from hypertensive rats and reconstituting autophagy ameliorated several vascular aging phenotypes. However, the mechanisms by which autophagy exerts anti-vascular aging effects are still unknown. Evolutionarily, autophagy functions to mobilize micronutrients in times of starvation. As a result, autophagy has also been recognized as a mediator of fatty acid oxidation and ketogenesis, predominantly from the liver. Therefore, we hypothesized that autophagy induction by fasting would increase the expression of ketone body beta-hydroxybutyrate (βOHB) in the livers of Wistar-kyoto rats and this would be prevented by systemic inhibition autophagy (3-methyadenine). As hypothesized, fasting increased βOHB, and this was prevented by 3-methyadenine (mmol/L, non-fasting: 0.43±0.04 vs. fasting: 0.58±0.04 vs. fasting+3-methyadenine: 0.35±0.01, p<0.05). Next we questioned whether βOHB could contribute to the anti-vascular aging effect of autophagy and hypothesized that βOHB administered directly to isolated arteries would cause endothelium-dependent relaxation. To test this hypothesis, we performed concentration-response curves to βOHB in mesenteric resistance arteries from Dahl salt-sensitive rats. We observed that βOHB was a potent endothelium-dependent vasodilator (%Relaxation, βOHB+endothelium-intact: 53±6 vs. βOHB+endothelium-denuded: 24±5, p<0.05). Inhibition of endothelium-derived vasodilators revealed that this relaxation was mediated by potassium channels (%Relaxation, βOHB: 53±6 vs. βOHB+tetraethylammonium: 19±9, p<0.05), but not nitric oxide (%Relaxation, βOHB: 53±6 vs. βOHB+L-NAME: 44±15, p>0.05). Overall, these data reveal βOHB as a novel endothelium-dependent vasodilator synthesized by the liver due to induction of autophagy. We conclude that βOHB synthesis is a mechanism by which autophagy exerts anti-vascular aging effects in hypertension.
Abstract 071: Endothelin-1 Contributes to Vascular Dysfunction in Peri-Menopausal Women
Authors: Wenner Megan M , Moreau Kerrie L
Published: September 2019

Declines in vascular function occur early in the menopausal transition, as previously demonstrated by a reduction in endothelial function in perimenopausal women (PERI). However, the mechanisms contributing to this decline remain to be elucidated. We have previously shown that vascular impairments in postmenopausal women (POST) are due in part to endothelin-1 (ET-1). Thus, the purpose of this study is to determine if ET-1 contributes to reductions in vascular function across the stages of the menopausal transition. We hypothesized plasma ET-1 levels would be elevated across menopausal stages, and negatively associated with endothelial function. We further hypothesized that ETB receptors would contribute to impaired vascular function in PERI.

Methods:
We measured vascular endothelial function (flow-mediated dilation, FMD) and plasma ET-1 in 180 women classified as premenopausal (PRE, n=33, 33±7 years), early PERI (n=20, 50±3 years), late PERI (n=25, 50±4 years), early POST (n=36, 55±3 years), and late POST (n=58, 60±4 years) based on the STRAW criteria. In a separate, secondary study, we measured cutaneous microvascular endothelial function via local heating during microdialysis perfusions of lactated Ringers (control) or ETB receptor antagonist (BQ-788, 300 nM) in 6 PRE (22±2 years) and 8 PERI (50±3 years).

Results:
Plasma ET-1 concentration increased in a step-wise fashion across menopausal stage (PRE 5.4±1.1, early PERI 5.8±1.0, late PERI 6.1±1.6, early POST 6.4±1.6, late POST 6.4±1.5 pg/mL; P<0.01). Furthermore, there was a significant inverse correlation between plasma ET-1 and brachial artery FMD (r=0.28; P<0.001). In our secondary study, cutaneous microvascular vasodilation was lower in PERI compared to PRE (82±7 vs 92±8 %CVCmax,
P
<0.05). In PRE, ETB receptor blockade mediated dilation (control: 92±8 vs ETB: 83±9 %CVCmax,
P
<0.05), however this effect was lost in PERI (82±7 vs 92±4 %CVCmax,
P
<0.05).

Conclusions:
These data suggest that elevated ET-1 contributes to impairments in vascular endothelial function across the menopausal transition. Furthermore, the decline in endothelial function is evident in the macro- and micro- circulation during the perimenopausal transition, due in part to a loss of ETB mediated dilation.
Abstract 073: The Association Between Plasma Soluble Prorenin Receptor and Renin Activity Points Toward Renin Angiotensin System Activation and Cardiovascular Complications in Women With Type-2 Diabetes
Authors: Visniauskas Bruna , Arita Danielle Y , Rosales Carla B , Feroz Mohammed A , Lefante John , Thethi Tina K , Fonseca Vivian A , Mauvais-Jarvis Frank , Prieto Minolfa C
Published: September 2019

Activation of the renin-angiotensin system (RAS) increases the risk of hypertension and chronic kidney disease (CKD) in patients with type 2 diabetes mellitus (T2DM). The prorenin receptor (PRR), a component of the RAS, activates prorenin via a non-catalytic mechanism. High levels of the soluble form of PRR (sPRR) have been reported in patients with cardiovascular (CV) diseases, including essential hypertension, CKD, preeclampsia, and gestational diabetes. Increased systemic prorenin in T2DM subjects raises the concern whether CV complications are due to augmented levels of plasma sPRR. To test the hypothesis that increased plasma sPRR is associated with systemic RAS activation, CV complications, and decline in renal function, we measured plasma sPRR, plasma renin activity (PRA), and biomarkers of renal dysfunction in 269 patients (mean age, 48 ± 1 years; 42% men, 58% women), including 173 controls (CT) and 96 patients with T2DM. Plasma sPRR levels measured by ELISA were higher in T2DM patients (19.2 ± 7.6 ng/mL) compared with CT (16.5 ± 0.4 ng/mL; p<0.001). Plasma levels of sPRR contrasted between T2DM and CT patients of same sex (Women: 20 ± 1.11 vs. 15.3 ± 0.3 ng/mL; p<0.001; Men: 17 ± 1 vs. 18 ± 0.5 ng/mL; p=0.18). PRA was augmented in T2DM patients, including Men: 9.8 ± 3,6 vs. Ang I/mL/hr; p<0.001, and even greater in Women: 13.5 ± 1.1 vs. 3.1 ± 0.3 Ang I/mL/hr; p<0.001. Increases in sPRR and PRA paralleled with hypertension, declined eGFR, and albumin/creatinine ratio in T2DM patients. Interestingly, PRA was positively correlated with plasma sPRR in women (r=0.49; p<0.001) but not in men (r=0.15 1; p=0.207). Multiple regression analysis, adjusted by age, BMI, and groups supports the association between plasma sPRR and T2DM status in women (p<0.001) but not in men. In conclusion: 1) Plasma sPRR is associated with PRA in T2DM women but not in men; and 2) The association between sPRR and PRA predisposes women with T2DM to systemic RAS activation and CV complications, as hypertension and CKD. Plasma sPRR reflects systemic RAS activation and has the potential to be a biomarker of T2DM and CV complications, particularly in women.
Abstract 078: Nephron-Specific Deletion of Intraflagellar Transport 88 Alters Renal Electrolyte Excretion in the Absence of Cystic Kidney Disease in a Sex-Specific Manner
Authors: Hu Chunyan , Lakshmipathi Jayalakshmi , Stuart Deborah , Kohan Donald
Published: September 2019

Defects in primary cilia in the nephron can cause cystic kidney disease, however, the physiological role of nephron primary cilia is largely unknown. To address this, nephron primary cilia were disrupted by targeting the intraflagellar transport 88 (
IFT88
) gene. IFT88
f/f
mice containing loxP-flanked exons 4-6 of the
IFT88
gene were bred with doxycycline-inducible Pax8-rtTA/tetO7-CMV*-Cre knockin mice. Inactivation of IFT88 (KO) was induced with 2 mg/ml doxycycline in drinking water supplemented with 2% sucrose for 12 days at 2 months of age; controls were the same genotype but without doxycycline treatment; mice were studied 2-4 months after doxycycline treatment. Renal
IFT88
gene recombination was observed in KO mice associated with an 88±5% decrease in whole kidney IFT88 mRNA. Metabolic cage studies were conducted under a normal-Na
+
(0.3% Na
+
, 5 days), a low-Na
+
(0.03% Na
+
, 7 days), and a high-Na
+
(3.2% Na
+
, 7 days) diet (N=10 males and 10 females per group). No significant differences were found in food intake, water intake, urine volume (UV), urinary Na
+
excretion (UNaV), or urinary K
+
excretion (UKV) between KO and control mice under either a normal-Na
+
or a low-Na
+
diet. In contrast, water intake, UV, UNaV, and UKV were higher on days 2-7 of high-Na
+
diet in male, but not female, KO compared with control mice (p<0.05); no differences in food intake were noted. Plasma renin concentration was similar between high-Na
+
diet-treated KO and control mice. Both urinary nitrite/nitrate excretion (UNOxV) and urinary prostaglandin E2 excretion (UPGE2V) were increased in high-Na
+
diet-treated male, but not female, KO mice (p<0.05). No cysts or tubule dilation were evident in KO mice at the end of experiments. In summary, nephron-specific deletion of IFT88 increases urinary fluid and electrolyte excretion during high salt intake associated with increases in UNOxV and UPGE2V; these differences are evident in male, but not female, mice. These data suggest that primary cilia regulate nephron function independent of cystogenesis and in a sex-specific manner.
Abstract 082: Histone Deacetylase 1 Activates Renal Inner-Medullary Nitric Oxide Production
Authors: Sedaka Randee , Mironova Elena , Stockand James D , Hyndman Kelly A , Pollock Jennifer S
Published: September 2019

Fluid flow along the renal nephron, particularly within the inner-medullary collecting duct (IMCD), increases in response to high salt intake. We previously showed in a series of reports that high salt intake increases nitric oxide synthase 1 (NOS1) activity in the CD mediating inhibition of the epithelial sodium channel (ENaC) to promote natriuresis. However, the mechanism of how high salt increases NOS1 activity in the CD is not known. Histone deacetylase 1 (HDAC1) is known to mediate post-translational deacetylation in response to flow in other cell types. Hence, we hypothesized that HDAC1 activates renal medullary NOS1-dependent NO production in response to high salt intake. Renal medullas were isolated from C57bl/6J mice after one week of normal (NS, 0.4% NaCl) or high salt (HS, 4% NaCl) diet. Inner-medullary HDAC1 expression (NS: 1.00±0.1, HS: 1.61±0.1 RDU; n=5/group, p=0.001), but not activity (NS: 0.07±0.02, HS: 0.05±0.01μM/μg, n=5/group, p>0.05), was significantly elevated in mice on HS compared to NS. HDAC1-selective inhibitor, MS-275 (300 nM), was utilized in
in vitro
and
ex vivo
experiments to determine the contribution of HDAC1 to NOS activity. Nitrite production, analyzed by HPLC, increased >5-fold (static: 231±31.1, flow: 1291±183 pmol/mg pr/hr; n=7/group, p<0.001) in mouse IMCD cells in response to acute 10 dynes/cm
2
fluid-flow. MS-275 significantly blunted flow-responsive NO production (MS-275: 342±44.0 pmol/mg pr/hr; n=9/group, p<0.001). Likewise, ENaC activity, which is negatively regulated by NOS1, was significantly increased in isolated collecting ducts post
ex vivo
MS-275 treatment from mice on HS (HS: 0.34±0.11 NP
o
, HS+MS-275: 0.97±0.16 NP
o
; n=22/group, p<0.05). Moreover, acute DETA-NONOate treatment, an NO donor, reversed this finding (HS+MS-275: 0.77±0.14 NP
o
, HS+MS-275+DETA: 0.46±0.14 NP
o
; n=10/group, p<0.05). Lastly, acute MS-275 treatment
in vitro
did not increase NOS1 acetylation status as determined by co-immunoprecipitation, suggesting that HDAC1 may not directly deacetylate NOS1. Taken together, these data indicate that HS intake increases renal medullary HDAC1 to positively regulate NOS1-derived NO production.
Abstract 083: Divergent Metabolic and Cardiovascular Effects of FGF21
Authors: Morgan Donald A , Potthoff Matthew J , Rahmouni Kamal
Published: September 2019

Fibroblast growth factor 21 (FGF21) is a pleiotropic hormone that regulates energy homeostasis. We previously demonstrated that FGF21 act in the brain to increase thermogenic sympathetic nerve activity (SNA), and that this effect required for the weight-reducing action of this hormone. However, the role of FGF21 in cardiovascular sympathetic control has not been investigated. To address this, we examined the effect of FGF21 on renal SNA recorded simultaneously with thermogenic brown adipose tissue (BAT) SNA in mice. In line with our previous findings, intravenous infusion of FGF21 (1mg/kg for 30 min) led to a significant increase in BAT SNA (182±33%, p<0.05), but not renal SNA (-35±16%). Intracerebroventricular administration of FGF21 (1 μg) that increased BAT SNA also failed to increase renal SNA (-14±28%) in mice. Next, we tested the effect of FGF21 on telemetry blood pressure and heart rate in mice. Chronic treatment with FGF21 (1mg/kg BID, IP) for 5 days led to a significant decrease in body weight (p=0.01), but caused no significant change in blood pressure (MAP: 3±3 mmHg vs 4±4 mmHg in vehicle control group, p=0.43) and heart rate (16±24 bpm vs 14±24 bpm in controls, p=0.5). Similarly, chronic treatment with the potent long-acting rhFGF21 analog (10mg/kg, SC) decreased body weight (p=0.002), but failed to alter blood pressure (p=0.14) and heart rate (p=0.25). Finally, we studied transgenic mice that overexpress FGF21 leading to ~5-10 fold higher circulating FGF21 than in the control mice. Interestingly, transgenic FGF21 mice displayed lower blood pressure (MAP: 104±1 vs 117±1 mmHg in controls, p<0.001) without change in heart rate (p=0.09). Consistent with the lower body weight (17.7± 0.8g vs 34.4± 1.3g in wild type controls), transgenic FGF21 mice displayed a 125% greater (p<0.001) baseline BAT SNA relative to controls whereas renal SNA was only 16% higher (p=0.02) in the transgenic mice. Together, our findings show contrasting effects of FGF21 on SNA controlling metabolism relative to blood pressure. The selective action of FGF21 may contribute to the differential sympathetic regulation of metabolism vs cardiovascular function.
Abstract 084: Gut Dysbiosis Impairs Serotonergic Gut-Brain Axis and Increases Blood Pressure
Authors: Yang Tao , Larkin Riley , Bueno Silva Carla , Donertas Basak , Colon-Perez Luis , Awadallah Nora , Pino Reyes Jose Antonio , Malphurs Wendi , Torres Gonzalo , Baekey David , Febo Marcelo
Published: September 2019

Introduction:
Fecal microbiota transfer (FMT) from hypertensive to normotensive rats elevates blood pressure (BP). As gut microbiota can affect synthesis of serotonin (5-HT), a major neurotransmitter predominantly synthesized in the gut, our objective was to investigate the effect of gut dysbiosis on gut-derived 5-HT in control of BP.

Methods:
Gut microbiota was depleted in normotensive adult male Wistar Kyoto (WKY) rats (N=24) using a cocktail of Vancomycin/Meropenem/Omeprazole at 50mg/kg/day for 5 days, followed by FMT from either adult male spontaneously hypertensive rats (SHR) (SHR-WKY, N=12) or age-, sex-matched WKY (WKY-WKY, N=12). At 8 weeks post-FMT, the following parameters were measured: BP by tailcuff and radiotelemetry; autonomic variables by spectral analysis of waveform BP; activity of cardioregulatory brain regions by manganese-enhanced MRI; fecal and proximal colonic 5-HT levels by HPLC; and levels of 5-HT
3
receptors in the proximal colon, nodose ganglia and the nucleus of the solitary tract (NTS) by qPCR. Lastly, BP and heart rate (HR) responses to intra-mesenteric i.v. injection of 5-HT
3
receptor agonist 1-phenylbigunide (35μg) were determined in anesthetized rats.

Results:
Compared with WKY-WKY, SHR-WKY showed a significant increase in systolic BP (Δ=34mmHg, P<0.05). This was associated with increased low frequency (by 13%, P<0.05) and decreased high frequency of BP (by 14%, P<0.05), indicative of sympathetic and parasympathetic variables respectively. A suppression of neural activity in the NTS (by 2 fold, P=0.06), and a reduction in 5HT
3
receptors in the NTS (by 1.5 fold, P<0.01), nodose ganglia (by 2 fold, P<0.05) and proximal colon (by 1.5 fold, P=0.09) in SHR-WKY resembled those in the SHR. Lower level of 5-HT in the proximal colon (by 4 fold, P<0.01), but not in the feces of SHR-WKY were observed. Similar to SHR, WKY-SHR showed significantly blunted drop in BP (by 5 fold, P<0.05) and HR (by 2 fold, P<0.05) in response to intra-mesenteric i.v. injection of 1-phenylbigunide, compared with WKY-WKY.

Conclusions:
FMT from SHR to WKY is associated with imbalanced autonomic variables and perturbations in serotonergic gut-brain axis that may contribute to hypertension partly via a dampened afferent vagal feedback to the NTS.
Abstract 085: Loss of Microglia Enhances Neuronal Activity in the Sympathetic Paraventricular Nucleus
Authors: Bi Qianqian , Wang Chao , Wei Bo , Liu Xiaoli , Gu Yan , Shen Xiao Z , Shi Peng
Published: September 2019

Microglia, the primary immune cells in the CNS, are critical in the maintenance of brain homeostasis such as participating neurogenesis and pruning synapses. Our previous work indicates that microglia are activated in hypertension; depletion of microglia in different activation status have opposite effects on blood pressure regulation, which could be due to tuning sympathetic outflow. To further delineate the mechanism how microglia modulate sympathetic nerve activity, we investigated the effect of microglial depletion on sympathetic neuronal activity in the hypothalamic paraventricular nucleus (PVN). In transgenic CD11b-DTR mice, intracerebroventricular (ICV) infusion of diphtheria toxin resulted in over 90% loss of microglia in the brain. Using the transgenic mouse brain slices, we examined the excitability of RVLM-PVN neurons by electrophysiological technique. Patch-clamp recordings obtained from the RVLM-projecting PVN neurons showed significant increases in firing frequency (3.59±0.72 vs. 1.56±0.51; P<0.05 by t-test) and half-width of action potential (2.53 ± 0.11 vs. 1.65 ± 0.07; P<0.001 by t-test); and a dramatic reduction in the outward K currents (I
A
and I
k
) in the absence of microglia compared to control. Single-cell RNA sequencing analysis of RVLM- PVN neurons revealed a significant decreased expression of I
A
(Kcnd2 abd Kcnd3) and I
k
(kcna2, Kcna3 and Kcnc2) subunits in neurons without microglia vs. the controls. These results indicate that loss of microglia increases PVN sympathetic neuronal activity, which could be mediated, at least in part, by the inhibition of outward K currents. Collectively, this study suggests a critical role of microglia in the maintaining the homeostasis of sympathetic PVN neurons in normal condition.
Abstract 092: Fine-Tuning of G
I/o
Signaling by Rgs2 and 5 is Critical to Ensuring Normal Ventricular Rhythm
Authors: Dahlen Shelby
Published: September 2019

Cardiac arrhythmia is attributed partly to abnormal G protein signaling that may result from the loss of signaling regulation by regulators of G protein signaling (RGS) proteins. RGS proteins act as GTPase activating proteins (GAPs) to control the kinetics and amplitude of G protein signaling; thus a decrease in their expression and/or function could lead to loss of fine-tuning of G protein signaling. Because RGS2 and 5 proteins are prominently expressed in the myocardium, we hypothesized that the dual absence of RGS2 and 5 predisposes to decreased heart rate variability (HRV) leading to the development of cardiac dysfunction. We generated mice dually lacking RGS2 and 5 (
Rgs2/5
dbKO). We used radiotelemetry to acquire diurnal blood pressure (BP) and ECG data from conscious mice to compute HRV in both time and frequency domains. Surgery-induced stress caused a marked increase in BP in
Rgs2/5
dbKO mice (WT: 140 ± 6 vs. dbKO: 170 ± 2 mmHg;
p
<0.01) initially, and gradually declined but remained elevated several days later. Standard deviation of cardiac cycles (SDNN) and total power (TP), both measures of HRV in time and frequency domain, respectively, were lower in male
Rgs2/5
dbKO relative to WT mice. SDNN in female WT and
Rgs2/5
dbKO were not significantly different. However, TP was markedly increased in female
Rgs2/5
dbKO compared to WT mice. To further determine how the changes in HRV correlate with cardiac events, we assessed excitation-contraction coupling in freshly isolated adult cardiomyocytes. Cells from male
Rgs2/5
dbKO mice showed decreased fractional shortening (WT: 16.1 ± 4.3 vs. dbKO: 7.4 ± 1.1 %;
p
<0.01) but high Ca
2+
transients (WT: 117 ± 20 vs. dbKO: 198 ± 50 au;
p
=0.07) at baseline. Application of low concentration of the non-selective β-adrenergic agonist, isoproterenol (ISO), triggered premature Ca
2+
transients, tachyarrhythmia and death of cells from
Rgs2/5
dbKO mice. Interestingly, cells from mice harboring just one copy of
Rgs2
(
Rgs2+/-,Rgs5-/-
) but not
Rgs5
(
Rgs2-/-,Rgs5+/-
) were resistant to low-dose ISO-induced arrhythmia, and blockade of Gi/o with pertussis toxin markedly reduced arrhythmia in
Rgs2/5
dbKO. We conclude that RGS2 and 5 are critical for proper control of cardiomyocyte excitation-contraction coupling and normal cardiac rhythm.
Abstract 094: Adaptive Cardiac Remodeling to Chronic Pressure Overload Requires the Expression of Placental Growth Factor in the Spleen to Recruit Reparative Macrophages to the Left Ventricle
Authors: Carnevale Daniela , Perrotta Sara , Carnevale Raimondo , Carnevale Lorenzo , Perrotta Marialuisa , Pallante Fabio , Cifelli Giuseppe , Lembo Giuseppe
Published: September 2019

Cardiac remodeling is the process established by the left ventricle (LV) to respond to different challenging stimuli. Although immunity has been clearly shown to participate to LV remodeling in myocardial infarction, how immune response participates to hypertensive chronic stress is poorly understood. We found that Placental Growth Factor (PlGF), belonging to the VEGF family, is necessary for adaptive remodeling to pressure overload and modulates the recruitment of innate immune cells in the LV. Here we asked where PlGF does exert its effects: i) directly in the myocardium or ii) by modulating immune organs? As TAC induced PlGF both in cardiac tissue and in the spleen, we devised a strategy to assess the contribution of these two tissues. We generated chimeric mice by spleen transplantation between WT and KO mice, obtaining: 1) WT mice with PlGF KO spleen and 2) PlGF KO mice with WT spleen (EF% 78 ± 0,7), finding that only mice with PlGF expressed in the spleen (EF% 68 ± 1,5) established adaptive remodeling to pressure overload, as assessed by echocardiography (p<0.01). Since in the absence of PlGF we observed a consistent reduction of LV macrophages, we characterized the subsets of innate immune cells by flow cytometry. While WT mice subjected to TAC showed early LV recruitment of non resident CD11b
+
CD64
+
Timd4
-
CCR2
low
CX
3
CR1
hi
macrophages, PlGF KO mice failed to show this response. To investigate the potential relevance of the spleen as reservoir of myeloid cells with adaptive/reparative functions for LV, we induced TAC in splenectomized mice, finding that they didn’t develop adaptive remodeling, exhibiting early HFrEF. To investigate the role of reparative macrophages, that are mostly characterized by the expression of the CX
3
CR1 receptor, and of pro-inflammatory monocytes that express high levels and accumulate through CCR2, we also subjected CCR2 KO and CX
3
CR1 KO mice to TAC. While CCR2 KO behaved similarly to WT mice, CX
3
CR1 KO were unable to establish adaptive remodeling and early developed HFrEF similarly to mice deprived of the spleen and to PlGF KO mice. Overall our results indicate that pressure overload induces PlGF in the spleen as an immunomodulator capable to deploy reparative macrophages that sustain adaptive remodeling of the LV.
Abstract 095: Mutation in the Furin Cleavage Site of the Prorenin Receptor Attenuates Angiotensin-II Induced Hypertension and Albuminuria
Authors: Ramkumar Nirupama , Stuart Deborah , Wheatley Will , Kohan Donald E
Published: September 2019

Cleavage of the extra-cellular domain of the (pro)renin receptor (PRR) yields a soluble fragment (sPRR) which can promote angiotensin-II (Ang-II) formation. Although alterations in plasma sPRR levels have been reported in hypertension, the causal role of sPRR in hypertension is unknown. To investigate this, we mutated the furin cleavage site (R276A, R279A) of the PRR using CRISPR/Cas9. Because the gene encoding PRR is on the X-chromosome and male mutant mice are infertile, only male mice were studied. Mutant mice had markedly lower plasma sPRR levels (control: 16.2 ± 0.3 vs mutant 0.2 ± 0.03 ng/ml). Mutant mice had normal survival and development and no histological renal abnormalities up to 12 months of age. During normal salt intake, no differences in blood pressure, body weight, urinary water or Na
+
excretion, or acid-base status were observed between control and mutant mice. Compared to controls, mutant mice had an attenuated hypertensive response to 2 weeks of Ang-II infusion (400 ng/kg/min) (Figure 1). Mutant mice also had lower albuminuria (control: 327 ± 114 vs mutant: 58 ± 8 μg/day) at day 7 post Ang-II infusion. No differences in urinary Na
+
excretion were detected between control and mutant mice after 7 days of Ang-II infusion (control: 33.6 ± 3.1 vs mutant: 35 ± 2.7 μmol/day/gram body weight). These results suggest that sPRR plays a role in Ang-II induced hypertension and renal injury.
Abstract 096: Renal-Specific Deletion of (Pro)Renin Receptor in Mice Induces Hyperkalemia During High Potassium Intake
Authors: Xu Chuanming , Chen Yanting , Xie Shiying , Wang Fei , Luo Renfei , Yang Tianxin
Published: September 2019

We have previously shown that systemic administration of (pro)renin receptor (PRR) decoy inhibitor PRO20 impaired kaliuretic response to high K
+
(HK) intake (5% KCl in diet). The present study extends this observation by examining the phenotype of inducible renal tubule-specific PRR knockout (RT PRR KO) mice fed a HK diet. The null mice were generated by breeding Floxed PRR mice with mice transgenic for Pax8-rtTA and LC-1 transgene. To induce RT PRR KO, mice with homozygous for Floxed PRR gene and hemizygous for the Pax8-rtTA and the LC-1 transgene were given 2 mg/ml doxycycline in 5% sucrose drinking water for 14 days followed by 4-wk off doxycycline. Following 1-wk HK intake, RT
PRR
KO mice had decreased urinary K
+
(by 33.6%) excretion and elevated plasma K
+
level (KO+HK: 5.38 ± 0.21
vs.
Floxed+HK: 3.98 ± 0.03 mM,
P
<0.001), accompanied with a reduction of urinary prorenin/renin content (by 56.5%), urinary renin activity (by 69.6%), urinary free and total Aldo excretion (by 41.2% and 25.3%, respectively), and kidney cortical and medullary Aldo levels (by 62.0% and 64.2%, respectively), without affecting plasma Aldo or renin levels. HK upregulated renal protein expression of fPRR (cortical: 170.4%, medullary: 195.0%), sPRR (cortical: 138.0%, medullary: 180.1%), prorenin (cortical: 217.5%, medullary: 164.0%), renin (cortical: 183.2%, medullary: 153.1%), Aldo synthase CYP11B2 (cortical: 321.6%, medullary: 243.8%), ROMK (165.4%), α-BK (161.4%), α-Na
+
-K
+
-ATPase (160.3%), β-ENaC (130.2%), and cleaved-γ-ENaC (542.0%), and downregulated phosphorylated NCC-T53 (pNCC-T53) protein expression, all of which were significantly blunted in RT
PRR
KO mice (by 25-80%). In Flp-In T-REX 293 NCC cell line (stably expressing NCC), PRR siRNA significantly increased pNCC-T53 (by 72.3%) and partially reversed prorenin (100 nM)-induced inhibition of pNCC-T53 (by 38.2%). sPRR-His (10 nM) inhibited pNCC-T53 (by 48.9%), which were significantly reversed by losartan or angiotensin II receptor type 1 (AT1R) siRNA, without affecting total NCC expression. Together, these results suggest that PRR/sPRR contributes to the kaliuretic response through an impact on the intrarenal RAAS and the K
+
secretory machinery in the distal nephron.
Abstract 105: Blood Pressure Variation and Renal Functional Changes in Chronic Renal Insufficiency Patients With or Without Diabetes in a Large Cohort Study
Authors: Wang Ling , Pezeshkian Kevin , Wang Donna
Published: September 2019

Hypertension and diabetes mellitus (DM) are leading causes for chronic kidney disease (CKD). Regardless, interplays of blood pressure, diabetes status, and renal function are largely unknown. We hypothesize that systolic blood pressure (SBP) variation affects renal function disproportionally in CKD patients with or without DM. Data from the Chronic Renal Insufficiency Cohort Study (CRIC) that enrolled 3,939 participants with CKD aged 21-74 years and followed between 2003-2013 were used. SBP was categorized into 3 levels over time: ≤120 as low, 121-140 as medium, and >141 as high, and relationship of these SBP levels with four renal indices in DM versus non-DM patients was analyzed using Generalized Estimating Equation (GEE) and Curve Fitting methods. Regression results showed that after controlling for confounding factors, for each category of low, medium or high SBP, the predicted eGFR was 4.5, 3.4 and 3.1 units lower for DM comparing to non-DM patients (p<0.01), respectively. Similarly, albuminuria was 0.19, 0.54 and 0.76 units higher (p<0.01), serum creatinine was 0.43, 0.33 and 0.29 units higher (p<0.01), and urinary creatinine was 5.24, 6.27 and 7.37 units lower (p<0.01) with DM comparing to without DM for low to high SBP, respectively. Curve Fitting results showed that slopes of curves for eGFR at all SBP levels were similar for DM and non-DM patients. In contrast, the slopes for albuminuria curve in all SBP levels was much steeper for DM comparing to non-DM patients. Thus, while renal function is worse in CKD patients with DM comparing to non-DM at any given SBP, lowering blood pressure may improve renal function with bigger beneficial effects on albuminuria in CKD patients with DM.
Abstract 109: Changes in Plasma Renin Activity After Renal Denervation in the Spyral Htn-Off Med Trial
Authors: Mahfoud Felix , Kandzari David E , Kario Kazuomi , Pocock Stuart , Schmieder Roland E , Townsend Raymond R , Tsioufis Konstantinos , Weber Michael A , Lauder Lucas , Fahy Martin , Böhm Michael
Published: September 2019

Results from recent sham-controlled clinical trials demonstrate the efficacy of renal denervation (RDN) in blood pressure reduction, however the effects of RDN on the renin-angiotensin system are not clear. We sought to evaluate changes in plasma renin activity (PRA) and serum sodium levels after RDN in the absence of anti-hypertensive medications in the SPYRAL HTN-OFF MED trial.

SPYRAL HTN-OFF MED is a randomized, sham-controlled trial to evaluate the impact of RDN in the absence of anti-hypertensive medications. Patients were enrolled with an office systolic BP (SBP) ≥150 and <180 mmHg, office diastolic BP ≥90 mmHg, and a mean 24-hour SBP ≥140 and <170 mmHg. Patients were randomized 1:1 to RDN with the SYMPLICITY Spyral™ catheter (N=38) or sham procedure (N=42). PRA and serum sodium levels were measured at baseline and 3 months post-procedure. Changes were expressed using medians and interquartile ranges and compared between groups using the Mann-Whitney U-test. Change in PRA from baseline to 3 months was significantly greater for the RDN group (n=23) -0.15 ng/mL/h [-0.76, -0.02] compared to the sham control group (n=33) 0.02 ng/mL/h [-0.16, 0.16]; p=0.018 (
Figure
). Comparison of 3-month changes in PRA using a Wilcoxon test stratified by 3 equal groups of baseline PRA also found a significant difference between groups (p=0.033). Changes in sodium levels were similar between groups: 0 mmol/L [-2, 2] (RDN, N=37) vs. 0 mmol/L [-1, 1] (control, N=41), p=0.920).

In this sham-controlled feasibility trial, renal denervation was associated with a reduction in PRA but no difference in serum sodium levels was observed. The impact on sodium excretion will need to be assessed in future studies.