bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 031: Endoplasmic Reticulum (ER) Stress in the Subfornical Organ (SFO) Induces Peripheral Inflammation by Altering Autonomic Output
Authors: Lob Heinrich E , Song Jiunn , Chung Alvin , Butler Scott D , Mark Allyn L , Davisson Robin L
Published: March 2022

Endoplasmic reticulum (ER) stress is a contributing factor in a variety of chronic diseases. We previously showed that ER stress in the SFO is involved in the activation of the adaptive immune system, which is causally linked with hypertension. However, the exact mechanism by which ER stress in the SFO activates peripheral T cells remains unclear. Recently it has been shown that the SFO is involved in modulating autonomic output to stimulate T cells. Here we tested if ER stress in the SFO increases autonomic drive to stimulate peripheral T cell proliferation. Five days of intracerebroventricular (ICV) delivery of the ER stress inducer thapsigargin (Tg, 1 μg/day) resulted in significant increases in peripheral T cells in aorta and blood as measured by flow cytometry (Aorta: Veh, 2.1 ± 0.7 vs. Tg, 10.1 ± 2.8 x 10
3
cells/aorta; Blood: Veh, 17.8 ± 1.3 vs. Tg, 34.8 ± 4.7 % of CD45
+
cells, n=8-13, p<0.05). ER stress in the brain resulted in a marked increase in plasma norepinephrine (Veh: 285 ± 28 vs. Tg: 406 ± 45 pg/mL, n=4-5, p<0.05). We also found that the effect of ICV Tg to increase peripheral T cells was blocked by hexamethonium (Hex, i.p., 30 μg/g) (Aorta: Tg, 10.1 ± 2.8 vs. Tg+Hex, 3.1 ± 0.4 x 10
3
cells/aorta; Blood: Tg, 34.8 ± 4.7 vs. Tg+Hex: 21.4 ± 1.9 % of CD45
+
cells, n=6, p<0.05). Consistent with earlier findings, ICV infusion of Tg increased ER stress markers in the SFO (
CHOP
: Veh, 1.15 ± 0.106 vs. Tg, 3.285 ±1.04;
grp78
: Veh, 1.032 ± 0.13 vs. Tg, 3.065 ± 1.102;
p58IPK
: Veh, 0.85 ± 0.213 vs. Tg, 7.065 ± 2.588, fold change, normalized to 18S, n=4-6, p<0.05), but did not alter the expression of these markers in aortas. This suggests that T cell activation is centrally mediated and not caused by peripheral vascular ER stress or inflammation. In conclusion, our data suggest that ER stress in the SFO modulates autonomic nervous system output and activates the peripheral immune system, a hallmark of the development of hypertension.
Abstract 036: Family History of Hypertension and Sympathetic Neural Reactivity to Mental Stress in Humans
Authors: Fonkoue Ida T , Wang Min , Carter Jason R
Published: March 2022

A number of recent studies have highlighted large inter-individual variability of muscle sympathetic nerve activity (MSNA) responsiveness to mental stress in humans. It remains unclear if family history of hypertension (FHH+) influences this variability. The purpose of this study was to examine blood pressure (BP) and MSNA responsiveness to mental stress in a large and generalizable cohort of young adults, and control for a variety of baseline factors that can influence MSNA reactivity. We hypothesized that subjects with FHH+ would demonstrate greater sympathoexcitation to mental stress than subjects without a family history of hypertension (FHH-). A total of 85 subjects from recently published (within 3 yrs; n=37) and ongoing (n=48) studies were examined. Data are presented on 45 subjects with complete MSNA recordings (15 FHH+ subjects and 30 FHH- subjects; 31 men and 14 women; age, 18 to 33 years). Heart rate (HR), BP, and MSNA were recorded during five min of supine rest and five min of mental stress (via mental arithmetic). Age, sex, body mass index, baseline mean arterial pressure (MAP), and baseline HR were identified as covariates. Baseline MSNA and HR were not statistically different between FHH+ and FHH- groups (p>0.05), whereas MAP was higher in FHH+ (84±2 and 79±2 mmHg; p<0.05). Mental stress increased MSNA in FHH+ subjects ([[Unable to Display Character: ∆]]4.4±1.5 bursts/min), but not FHH- subjects ([[Unable to Display Character: ∆]]1.0±0.2 bursts/min; time х group, p<0.01). Differences between FHH+ and FHH- groups remained when MSNA was normalized to heart rate ([[Unable to Display Character: ∆]]2.8±1.0 bursts/100 heart beats vs. [[Unable to Display Character: ∆]]-3.1±0.5 bursts/100 heart beats; time х group, p<0.05). Mental stress increased MAP ([[Unable to Display Character: ∆]]11±1 mmHg and [[Unable to Display Character: ∆]]10±1 mmHg; p<0.001) and HR ([[Unable to Display Character: ∆]]18±4 beats/min and [[Unable to Display Character: ∆]]17±2 beats/min; p<0.001) in FHH+ and FHH- subjects, but these increases were not different between groups (time х group, p≥0.05). In conclusion, our findings demonstrate that FHH+ is associated with greater MSNA reactivity to acute mental stress when compared to FHH- individuals.
Abstract 039: Normalization of Uterine Interleukin (IL)-15 in the BPH/5 Preeclamptic Mouse Improves Decidual Natural Killer Cell Activation at the Maternal-Fetal Interface
Authors: Sones Jenny L , Heyward Christa , Lessard Dominique , Butler Scott , Lob Heinrich , Davisson Robin L
Published: March 2022

Preeclampsia (PE) is a hypertensive disease that affects up to 10% of pregnancies worldwide. Although a leading cause of maternal and fetal morbidity/mortality, the cause is unknown. Inadequate remodeling of decidual vessels and reduced placental perfusion are hallmarks of PE. Decidual Natural Killer cells (dNKs) are critical in this process. Early in pregnancy, dNKs are activated by uterine IL-15 to promote angiogenesis at the maternal-fetal interface. Activated dNKs produce IFN-γ to facilitate vasodilation of decidual vessels. In the BPH/5 mouse model that spontaneously develops late gestation hypertension, proteinuria and placentopathies, we have demonstrated an early pregnancy reduction in dNK activation and IFN-γ mRNA at the maternal-fetal interface. We functionally linked uterine IL-15 overexpression to dNK loss in C57 mice. Here we tested the hypothesis that uterine IL-15 in BPH/5 mice contributes to dNK dysregulation at the maternal-fetal interface prior to placenta formation. BPH/5 e7.5 implantation sites show a 5-fold increase in IL-15 protein vs C57 (.7±.07vs.35±.17, n=4, p<.05). We have shown that Cox2 inhibition early in BPH/5 pregnancy improves fetoplacental development. Cox2-derived products influence expression of the pro-inflammatory cytokine, IL-15. To address if Cox2 inhibition alters IL-15 in BPH/5 implantation sites, celecoxib (a selective Cox2 inhibitor) was administered at e6.5 (10mg/kg orally). This normalized IL-15 in BPH/5 e7.5 implantation sites compared to C57 veh-treated (n=4, p<.05). Flow cytometry confirmed a 2-fold increase in DBA+/CD122+ (dNK) cell numbers in BPH/5 e7.5 implantation sites from celecoxib vs veh-treated mice (16.6 vs 7.95). Moreover, the dNKs from e7.5 BPH/5 celecoxib-treated implantation sites had a 2-fold increase in IFNγ mean fluorescent intensity vs BPH/5 veh-treated. This demonstrates that uterine IL-15 overexpression in BPH/5 contributes to dNK loss and that normalization improves dNK activation by increasing their expression of IFNγ at the maternal-fetal interface. This highlights the significance of uterine inflammation in dNK regulation and suggests that aberrations in these cellular processes in early pregnancy may contribute to the placentopathies seen in PE pregnancies.
Abstract 040: Complement Activation in Placental Ischemia-induced Hypertension is not Dependent on Endothelin A Receptor Activation
Authors: Regal Jean F , Strehlke Megan E , Elmquist Barbara J , Gilbert Jeffrey S
Published: March 2022

Early onset preeclampsia is characterized by hypertension, reduced placental perfusion, intrauterine growth restriction and increased activation of complement, part of innate immunity. We previously reported that inhibiting complement activation attenuates reduced uterine perfusion pressure (RUPP)-induced hypertension in pregnant rat and the concomitant increase in complement activation product C3a. Studies by other groups indicate endothelin A receptor (ET
A
) activation plays a significant role in RUPP-induced hypertension. We hypothesized that ET
A
was important in placental ischemia-induced complement activation leading to hypertension. Thus, the effect of ET
A
antagonist atrasentan on hypertension and complement activation following placental ischemia in rat was determined. Dams received drinking water with or without 5 mg/kg/day atrasentan on gestation day (GD)13-19. On GD14, rats underwent Sham or RUPP surgery with clip placement on abdominal aorta and uterine arteries to decrease placental perfusion. On GD19, mean arterial pressure (MAP) via arterial catheter and serum C3a as indicator of complement activation were measured. RUPP surgery significantly increased MAP (109±5 mm Hg; n=7) compared to Sham (94±4 mm Hg; n=10) as well as C3a (0.36±0.08 to 0.69±0.12 units/ul). MAP was significantly reduced by atrasentan in RUPP (96±3 mm Hg; n=12) and unchanged by atrasentan in Sham (88±2 mm Hg; n=12). RUPP did not change preproendothelin mRNA in kidney cortex versus Sham, but did decrease fetal weight. In Sham, atrasentan significantly increased fetal weight (2.52±0.04 g; n=12) compared to no atrasentan (2.38±0.05 g; n=12). Importantly, atrasentan treatment did not significantly change RUPP-induced increase in C3a with 0.57±0.11 units/ul in RUPP atrasentan and 0.27±0.05 units/ul in Sham atrasentan. In contrast to other ET
A
antagonists (A-127722, FR-139317), the present data indicate atrasentan may promote fetal growth. However, the ET
A
receptor does not mediate increased complement activation and its subsequent involvement in placental ischemia-induced hypertension. Thus, our data suggest complement activation may precede involvement of endothelin in development and maintenance of high blood pressure due to placental ischemia.
Abstract 045: Epigenetic Modification of Brain (pro)renin Receptor Contributes to the Development of Doca-salt Hypertension via Epithelial Sodium Channel
Authors: Li Wencheng , Feng Yumei
Published: March 2022

We reported that elevated brain (pro)renin receptor (PRR) expression levels contributes to the development of salt-sensitive hypertension (SSH); however, the underlining mechanism on how PRR is regulated remains unknown. High-salt intake is a major life style epigenetic factors for the SSH. To test our hypothesis that epigenetic mechanism is involved in the regulation of PRR expression in SSH, we used deoxycorticosterone acetate (DOCA)-salt induced hypertension as a model of SSH. C57Bl/6J mice (n=5/group) were implanted with SHAM or DOCA (50mg) pellet and supplied with saline for 3 weeks. Histone modifications on PRR promoter were assessed in the hypothalamus using Chromatin Immunoprecipitation assays at the end of protocol. We found an increase (fold change) in histone 3 lysine 4 trimethylation (H3K4me3) on the PRR promoter in DOCA-salt (2.45±0.36) compared with SHAM treated mice (1.00±0.16, P<0.05). H3K4 methyltransferases (HMT) activity (OD/hour/mg protein), an enzyme complex responsible for H3K4 methylation is increased after DOCA-salt treatment (45±2, P<0.05) compared with SHAM (28±2). To examine the mechanisms of elevated HMT activity during DOCA-salt hypertension, we treated culture neurons (n=3 independent experiments/group) with control (155mM sodium), high salt (170 mM sodium), aldosterone (100 nM), or high salt+aldosterone with or without epithelial sodium channel (ENaC) blocker (amiloride, 100 nM) for 2 hours and test the HMT activity and PRR expression levels. High salt or aldosterone alone has minimal effects on HMT activity and PRR levels. High salt+aldosterone significantly increased HMT activity (41±3 vs. 24±2, P<0.05) and PRR mRNA (1.6±0.1 vs. 1.0±0.1 fold change, P<0.05) levels compare to control. Amiloride attenuated HMT activity (25±4.1, P<0.05) and PRR mRNA levels (0.98±0.1, P<0.05) compared to high salt+aldosterone treatment. In summary, elevated PRR expression is associated with increased HMT activity and H3K4me3 modification on PRR promoter. ENaC blockade attenuates the elevation of HMT activity and PRR levels induced by high salt+aldosterone in culture neurons. We conclude that ENaC may be responsible for epigenetic up-regulation of brain PRR and thus the increase in BP during salt-sensitive hypertension.
Abstract 050: Loss of Argininosuccinate Lyase Leads to Nitric Oxide Deficiency, Endothelial Dysfunction, Impaired Angiogenesis, and Hypertension
Authors: Kho Jordan , Tian Xiao Yu , Wong Wing Tak , Bertin Terry , Jiang Ming Ming , Bissig Karl-Dimiter , Nagamani Sandesh C , Lee Brendan H
Published: March 2022

Nitric oxide (NO) is an important mediator of vascular homeostasis and its deficiency in murine models results in hypertension. However, there are few monogenic causes of NO deficiency in humans and the effects of such genetic forms of NO deficiency on vasculature are not well-studied. We have recently shown that argininosuccinate lyase (ASL), a urea cycle enzyme, is necessary for synthesis of NO. ASL deficiency results in decreased production of NO and hypertension in humans and mice. To investigate whether loss of Asl-mediated NO synthesis in the vascular endothelium alone can cause hypertension, we generated a mouse model with endothelial-specific deletion of Asl (VE-Cadherin Cre(tg/+); Asl(flox/flox), or cKO). Asl cKO mice developed hypertension and had higher mean arterial pressure compared to control littermates (102.2± 2.9 vs. 89.8±3.6 mmHg, p<0.05). This hypertension was secondary to endothelial-specific NO deficiency as demonstrated by abnormal relaxation of aortic rings and correction with treatment with an NOS-independent NO supplement (MAP in sodium nitrite treated Asl cKO 97.7±3.8 vs. 97.6±7.3 mmHg in untreated control mice). To evaluate the human relevance of these findings, we developed human cell-based models from patients with ASL deficiency. Human induced pluripotent stem cells (hiPSCs) were generated and differentiated into endothelial cells. Interestingly, ASL-deficient hiPSCs differentiated less efficiently into endothelial cells as compared to control hiPSCs (9.7±4.0 vs. 20.8±3.7 % of CD144+; CD31+ cells, p<0.01). Furthermore, ASL-deficient hiPSCs-derived endothelial cells had a significantly reduced capacity to form capillary-like structures on Matrigel. Our study using a novel mouse model and hiPSCs-derived endothelial cells from patients with a rare Mendelian form of hypertension supports the hypothesis that structural and functional abnormalities in endothelial cells contribute to pathogenesis of hypertension. Our study is the first to use hiPSC-derived endothelial cells as a model system to study hypertension and highlights the utility of this technology in exploring the pathogenesis of other vascular diseases.
Abstract 051: The Role of Immunological Memory in Hypertension
Authors: Itani Hana A , Xiao Liang , Saleh Mohamed A , Wu Jing , Dale Bethany L , Barbaro Nataliya R , Foss Jason D , Kirabo Annet , Norlander Allison E , Chen Wei , Suto Ryo , Navar L. Gabriel , Bernstein Kenneth E , Harrison David G
Published: March 2022

Immunological memory provides protection to repeated antigen challenges and is a cardinal feature of adaptive immunity. We have previously shown that adaptive immunity contributes to hypertension and have observed memory T cells in several models. We hypothesized that memory T cells contribute to long-term renal damage in response to repeated hypertensive challenges. To impose repeated episodes of hypertension, we treated C57BL/6 mice with L-NAME (0.5mg/ml) in drinking water for two weeks, allowed a two-week normotensive interval and then fed high salt (4% NaCl) for three weeks. L-NAME followed by high salt increased SBP to 151 ± 14 mmHg and caused a two-fold increase in CD4
+
and CD8
+
memory T cells in the kidney and bone marrow, as identified by the surface marker CD44hi. Intracellular staining showed that memory T cells were predominant sources of the inflammatory cytokines IL-17A and IFN-γ. Development and reactivation of memory T cells require the interaction of CD27 on T cells with CD70 on antigen presenting cells. Flow cytometry revealed that L-NAME/High salt increased expression of CD70 on splenic macrophages by 5-fold and dendritic cells by 3-fold. Because memory T cells are a major source of IFN-γ, we examined the hypertensive response to the L-NAME/high salt protocol in IFN-γ
-/-
mice. The hypertension caused by L-NAME was identical between WT, CD70
-/-
and IFN-γ
-/-
mice. In contrast, the hypertension induced by subsequent salt administration was markedly attenuated in CD70
-/-
mice (123 ± 1.3 mmHg, p<0.02). Likewise, mice lacking IFN-γ developed blunted hypertension during the salt-feeding phase (127.6 ± 5.5 mmHg, p<0.04). Interestingly, CD70
-/-
and IFN-γ
-/-
mice failed to develop memory T cell formation in the kidney. The L-NAME/high salt caused striking albuminuria and increased urinary N-gal in WT mice, and these were absent in CD70
-/-
and IFN-γ
-/-
mice. In contrast, L-NAME/high salt had no effect on renal angiotensinogen levels. Thus, repeated hypertensive stimuli lead to accumulation of long-lived effector memory T cells that are major sources of inflammatory cytokines, which in turn promote renal dysfunction, salt sensitivity and hypertension. These studies provide further insight into how the adaptive immune system promotes hypertension.
Abstract 061: Kidney-specific Klotho Gene Deficiency Impairs Sodium Excretion and Causes Hypertension
Authors: Ali Quaisar , Wang Xiuqing , Sun Zhongjie
Published: March 2022

Background & Purpose:
Klotho was originally discovered as an anti-senescence gene. Mutation of klotho gene accelerates aging and shortens lifespan while overexpression of klotho gene extends lifespan in mice. We recently demonstrated that global mutation of klotho gene (+/-) increased blood pressure (BP). However, the underlying mechanism is not fully understood. The purpose of this experiment is to investigate if renal klotho plays a role in hypertension. Specifically, we assessed if kidney-specific deletion of klotho gene affects BP and renal function in male mice.

Methods & Results:
Briefly, kidney-specific Cre mice and klotho floxed mice were bred for generating kidney-specific klotho heterozygous (KspKL+/-) mice. Daily food and water intakes and urine output were monitored in wild type and KspKL+/- mice (16 weeks) using metabolic cages. BP was by direct cannulation of the carotid artery. Deletion of renal klotho significantly increased BP by 25 mmHg in mice without affecting the heart rate. Food and water intakes were not different between KspKL+/- and WT mice. Interestingly, 24-hr urine flow (UF) was significantly decreased in KspKL+/- mice (WT: 5.140± 0.3, KspKL+/-: 4.052±0.2,μL/min/100 gm body wt). The decrease in UF was also associated with a decrease in urinary sodium excretion (UNaV) in KspKL+/- mice compared to wild type (WT: 0.18± 0.01, KspKL+/-: 0.13±0.01, μmol/μL) without altering urinary potassium excretion. The cumulative sodium balance remained positive during the observation period with no significant difference between KspKL+/- and WT mice WT. Kidney-specific klotho deficiency did not affect the glomerular filtration rate (GFR).

Conclusions:
This study demonstrates for the first time that kidney klotho deficiency is sufficient to cause hypertension. Klotho deficiency-induced hypertension may be due to impaired Na excretion. Klotho may be a therapeutic target for kidney dysfunction and hypertension. An additional study is warranted to investigate if klotho regulates tubular ENaC and Na, K-ATP synthase.
Abstract 067: Endothelial S1P/S1P1 Signaling is Critical Regulator of Blood Flow and Pressure
Authors: Cantalupo Anna , Liu Catherine , Hla Timothy , Di Lorenzo Annarita , Dautaj Elona
Published: March 2022

Background and objectives:
Sphingosine-1-phosphate (S1P) is emerging as a pivotal signaling molecule within the vasculature. S1P has been known as a potent endothelial nitric oxide synthase activator through G-protein coupled receptor S1P1 and less abundant S1P3 on endothelial cells (EC). Although it has been recently shown that S1P/S1P1 signaling mediates EC alignment in response to flow, the role of S1P1 signaling in the pathogenesis of hypertension remains unknown. FTY720, an S1P1 functional antagonist, recently approved by the FDA to treat autoimmune conditions, induces a modest and transient decrease in heart rate in both animals and humans, suggesting that drugs targeting sphingolipid signaling affect cardiovascular functions in vivo. Thus, the aim of this study is to investigate whether S1P-S1P1 signaling pathway is crucial in blood flow and pressure regulation.

Methods:
The BP was evaluated in conscious mice with tail cuff system, whereas the vascular reactivity was examined with pressure myograph system in mesenteric arteries isolated from age matched WT and EC-S1P1-/- mice. Results. Here we report for the first time that S1P1-mediated signaling protects from hypertension. Administration of SEW 2871, an agonist of S1P1, to angiotensin-II-induced hypertensive mice lowered blood pressure to normotensive levels (110.7±3.1 vs. 160.7±2.4 mmHg, SEW 2871 vs. vehicle respectively). Pharmacological inhibition of S1P1 with W146 (100 nM and 1 μM) markedly reduced flow-induced vasodilation (Emax 25.2±3.8 and 2.5±2.3 vs. 48.6±4.9 μm, W146 100 nM and 1μM respectively vs. WT). These data were corroborated by using a genetic approach: mice EC-S1P1-/- were hypertensive (124.7±1.9 vs. 109±1.7 mmHg, EC-S1P1-/- vs. S1P1f/f mice) and showed a reduced response to flow-induced vasodilation (Emax 15.2±3.0 vs. 53.8±5.0 μm, EC-S1P1-/- vs. S1P1f/f mice mesenteric artery).

Conclusion:
Our study identifies S1P-S1P1 signaling as a new regulatory pathway of blood flow and pressure homeostasis, providing a novel therapeutic target for the treatment of hypertension.
Abstract 068: Role for the Rho-GAP Graf3 in the Pathogenesis of Human Hypertension
Authors: Bai Xue , Mangum Kevin , Mack Christopher , Taylor Joan M
Published: March 2022

Activation of RhoA in vascular smooth muscle cells (SMC) has been linked to vasoconstrictor-induced hypertension (HTN), but the relevance of this pathway to human disease was undetermined. We identified GRAF3 as a RhoA-GAP expressed specifically in SMC in mice and humans and reported that global GRAF3-deficient mice exhibited significant basal HTN (+ 25 mm Hg) that was fully reversed by treatment with a Rho-kinase inhibitor (Nature Comm. 2013;4:2910). Importantly, we recently created a tamoxifen inducible SMC-GRAF3 re-expression model which resulted in a near complete reversal of MAP (from 123 mmHg to 95 mm Hg), indicating that GRAF3's ability to limit RhoA activity in SMC is required for the maintenance of normal BP. Given that a BP-associated locus within the GRAF3 gene was identified by Genome Wide Association, we next sought to characterize the mechanisms that control GRAF3 expression. Through the use of a series of siRNA-dependent approaches in cultured SMC, we found that SMC-specific expression of GRAF3 is mediated by the RhoA/MRTF-A/SRF-signaling axis. Interestingly, RhoA is known to be activated by mechanical forces and we found that GRAF3 expression was significantly increased (8-fold) in vessels subjected to pathological stress. The finding that GRAF3 expression was significantly increased in two separate hypertensive animal models relative to their littermate controls provides further evidence that GRAF3 expression is regulated as part of an auto-regulatory negative feedback loop to inhibit RhoA activity and SMC tone. Interestingly, the BP associated locus maps to the 80Kb first intron of the GRAF3 gene and we found that the minor GRAF3 allele that decreases BP was associated with a significant increase in GRAF3 mRNA in human tibial artery samples (3-fold increase; N=123; p=1e-10 ). We have identified regulatory elements within the hypertensive locus that exhibit SMC-selective transcriptional activity and have shown that a minor allele variation within one of these elements significantly increased transcriptional activity. Our studies add significantly to our understanding of the development and pathophysiologic consequences of hypertension and may lead to novel and perhaps individualized approaches to its treatment.
Abstract 071: CX3CR1 Deficiency Aggravates Renal Damage in Angiotensin II Induced Hypertensive Kidney Injury
Authors: Wenzel Ulrich , Ahzdahan Erfan , Paust Hajo , Kurts Christian , Rosendahl Alva
Published: March 2022

A dense network of macrophages and dendritic cells (DCs) expressing the chemokine receptor CX3CR1 populates the kidney. We recently reported that CX3CR1 regulates the abundance of DCs selectively in the kidney and promotes renal inflammation in glomerulonephritis. Given that inflammation plays also an important role in hypertensive renal injury, we hypothesized that CX3CR1 deficiency should attenuate renal injury in hypertension by reducing the number of renal DCs.

Renal injury was induced by unilateral nephrectomy, angiotensin II (Ang II) infusion (1.5 ng/min/g) and salt (0.9% drinking water).

Ang II induced heavy proteinuria (measured as albumin/creatinine ratio in mg/mg) compared to controls (0.1±0.0) and proteinuria was unexpectedly more severe in knockout mice than in wildtype (WT) mice (day 3 after of Ang II infusion: 19.2±5.4 vs. 2.5±0.6 p<0.01, day 14: 32.4±8.5 vs. 9.2±2.6 p<0.05 n=13 and 11 respectively). Histologic evaluation of kidney sections revealed also an increased number of proteinaceous casts and more glomerular injury in knockout mice (casts 1.1±0.3 vs. 0.1±0.1/high power field, glomerular injury score 0.95±0.06 vs. 0.57±0.06, p<0.01). Also plasma cholesterol levels were significantly higher in knockout mice than in WT mice (145±11 vs. 105±6 mg/dl, p<0.01). No difference was found for blood pressure and cardiac injury (hypertrophy, fibrosis, fetal gene expression) between both groups. Flow cytometric analysis revealed that Ang II infusion significantly increased the number of renal CD11c
+
/MHCII
+
DCs in WT (38.5±1.4%, gated on CD45
+
cells) compared to controls (31.8±3.6%, p<0.05). However, a significant decrease of renal DCs was found in hypertensive CX3CR1 deficient mice (15.2±1.5%, p<0.001). In contrast, the number of CD11b
intermediate
/F4/80
high
macrophages was 8 times higher in the kidney of CX3CR1 deficient mice compared to wildtype mice (5.5±1.5 vs.0.9±0.2%, p<0.01). These findings show that CX3CR1 deficiency reduces renal DC numbers and increases numbers of renal macrophages in hypertension. Surprisingly, despite reduced DC numbers, CX3CR1 deficiency increases proteinuria and glomerular injury, thus, CX3CR1 inhibition should be avoided in hypertension because it may promote proteinuria and renal injury.
Abstract 077: In Vivo Evidence of AT1a Receptor-Mediated Uptake of Angiotensin II by the Proximal Tubule Visualized by Intravital Multiphoton Imaging
Authors: Li Xiao C , Sandoval Ruben M , Molitoris Bruce A , Zhuo Jia L
Published: March 2022

The development of all forms of angiotensin II (ANG II)-dependent hypertension is associated with higher levels of intrarenal ANG II, which are greater than can be explained on the basis of circulating ANG II and suppressed cortical renin expression. In the present study, we used intravital multiphoton imaging to test the hypothesis that AT1 (AT1a) receptor-mediated uptake of ANG II by the proximal tubules of the kidney plays a major role in the underlying mechanisms. Adult male Munich-Wistar rats were anesthetized with Inactin and continuously infused with a pressor dose of Alexa 488-conjugated ANG II (50 ng/min, i.v.) for 2 hr. Time-dependent proximal tubular uptake responses of Alexa 488-ANG II were studied with mean arterial blood pressure maintained at ~150 mmHg throughout the experiment. After 30 min infusion, very low levels of Alexa 488-ANG II were visualized within proximal tubules and cortical collecting ducts (CCDs) (p<0.05). However, high levels of Alexa 488-ANG II were accumulated in the lumen of CCDs, but not that of proximal tubules. After 1 hr infusion, moderate levels of Alexa 488-ANG II were visualized in the proximal tubules, but not in the glomeruli and CCDs. The most striking uptake responses were visualized in all segments of proximal tubules after 2 hr infusion. Internalized Alexa 488-ANG II was predominantly localized to the basolateral side, where it was colocalized with tetramethyl rhodamine methyl ester (TMRM), a mitochondrial membrane potential-dependent dye. TMRM is a lipophilic cationic dye that is primarily accumulated in the mitochondria of proximal tubules. Some internalized Alexa 488-ANG II was visualized around and over the nuclei. Furthermore, the uptake of Alexa 488-ANG II by proximal tubules was significantly attenuated in caveolin 1-knockout mice (p<0.01), and blocked in AT1a receptor-knockout mice (p<0.01). Our results provide strong intravital multiphoton microscopic evidence that circulating ANG II is primarily taken up by the proximal tubules of the kidney via an AT1a receptor-mediated mechanism, and that internalized ANG II may be transported to the mitochondria and the nucleus, where it may alter mitochondrial and nuclear function in the proximal tubules of the kidney.
Abstract 081: Intracerebroventricular (ICV) Infusion of Angiotensin-(1-7) Lowers Blood Pressure and Improves Autonomic Function in Antenatal Betamethasone Exposed Sheep
Authors: Lawson Matthew J , Figueroa Jorge P , Chappell Mark C , Diz Debra I , Shaltout Hossam A
Published: March 2022

We have previously shown that antenatal betamethasone exposure in sheep is associated with a deficiency in angiotensin (Ang)-(1-7) levels in the cerebrospinal fluid of these betamethasone exposed (BMX) sheep and impaired baroreflex sensitivity for control of heart rate (BRS) by 6 weeks of age followed by persistent suppression of the reflex and elevation in mean blood pressure (MAP) by 6 months of age. In this study we sought to determine if supplementation of Ang-(1-7) administered via continuous infusion into the lateral ventricle of 4 month old BMX lambs (1-7, n=4) at a dose of 1.25ug/kg/hr for 1 week would improve the BRS, heart rate variability (HRV) and lower MAP and blood pressure variability (BPV). The control group received ICV artificial cerebrospinal fluid (aCSF, n=4). MAP and heart rate were recorded continuously via femoral arterial catheters and digitized before and one week after the ICV infusion. Both blood pressure and heart rate were analyzed to obtain measures of spontaneous BRS, HRV and BPV in the frequency and time domain. There were no differences in any of the measures between the two groups at baseline and no changes in the aCSF group after 1 week of ICV aCSF administration. Meanwhile, Ang-(1-7) ICV treatment significantly improved all measures of BRS compared to baseline, (HFa, 5.3±0.9 vs. 2.4±0.5 ms/mm Hg, p=0.03) and sequence ALL (6.5±1.6 vs. 1.9±0.9 ms/mm Hg, p=0.01). Ang-(1-7) also improved HRV measured as rMSSD (11.6±1.6 vs. 7±1.2 ms at baseline, p=0.04). Ang-(1-7) administration lowered MAP by 11.5 mm Hg (p < 0.05) compared to a rise of 2.0 mm Hg in the control group with no significant changes in heart rate or BPV in either group. These data support the hypothesis that Ang-(1-7) deficiency in the CSF contributes to the impaired autonomic function and reduced parasympathetic tone in betamethasone exposed sheep and replacement of the peptide in the brain will help restore these measures to normal levels. Elevating Ang-(1-7) levels in humans exposed antenatally to betamethasone may have beneficial cardiovascular effects.
Abstract 083: Metabolic and Cardiovascular Effects of BBS1 Ablation From the POMC-containing Neurons
Authors: Guo Deng F , Morgan Donald A , Grobe Justin , Sheffield Val , Rahmouni Kamal
Published: March 2022

Bardet-Biedl syndrome (BBS) is a pleiotropic autosomal recessive disorder associated with several features including obesity and hypertension. Deletion of
Bbs
genes globally, in the nervous system or in the leptin receptor-expressing cells recapitulated many of the BBS phenotypes including obesity and hypertension. Here, we assessed the effect of ablating the
Bbs1
gene from the neurons expressing proopiomelanocortin (POMC) neurons. Breeding Bbs1
flox
mice with POMC
Cre
mice created mice deficient in
Bbs1
gene only in the POMC-positive neurons (visualized by tdTomato expression). Importantly, POMC
Cre
/Bbs1
fl/fl
mice display an obesity phenotype as indicated by the increased (P<0.05) body weight (48.6±1.7 vs 34.9±0.8 g in controls) and fat pads (3.3±0.3 vs. 0.7±1.1 g for inguinal fat, 2.1±0.2 vs. 0.4±0.05 g for perirenal fat, 3.3±0.3 vs. 0.8±0.3 g for reproduction fat and 0.4±0.02 vs. 0.2±0.02 g for brow fat) associated with increased (P<0.05) food intake (3.79.±0.14 vs. 2.97±0.15 g in controls) in 25 weeks old mice. POMC
Cre
/Bbs1
fl/fl
mice displayed decreased (P<0.05) O
2
consumption (2.6±0.03 vs. 3±0.04 mL O
2
/100g/min in controls) and heat production (8.0±0.09 vs. 9.2±0.13 kcal/kg/hr in controls). These results indicate that hyperphagia and decreased energy expenditure contribute to the development of obesity in POMC
Cre
/Bbs1
fl/fl
mice. Next, we assessed the consequence on arterial pressure (AP) and sympathetic nerve activity (SNA) of ablating the
Bbs1
gene from POMC neurons. Interestingly, deletion of the
Bbs1
gene in POMC neurons did not recapitulate the hypertension phenotype of BBS as indicated by the slight, but not significant increase in mean AP (116±4.7 vs 109±6.1 mmHg in controls). However, conscious renal SNA was significantly higher in POMC
Cre
/Bbs1
fl/fl
mice relative to controls (132.4±11 vs 74.3±4.2 spikes/sec, P<0.05). Finally, the depressor effect of ganglionic blockade (hexamethonium) was exaggerated in POMC
Cre
/Bbs1
fl/fl
mice (-68.6±4.4 vs -45.2±9.4 mmHg in control, P=0.017). These findings demonstrate that
Bbs1
gene in the POMC neurons is critical for energy homeostasis, but not for arterial pressure regulation.
Abstract 086: Insulin Receptor Signaling in the Subfornical Organ Protects Against the Development of Metabolic Syndrome
Authors: Horwath Julie A , Butler Scott D , Mark Allyn L , Davisson Robin L , Young Colin N
Published: March 2022

Metabolic syndrome encompasses a combination of conditions including obesity, diabetes, dyslipidemia and hypertension. Brain insulin resistance has emerged as a contributor to the development of metabolic syndrome, although the neural regions involved remain unclear. While most studies have focused on hypothalamic areas, recent evidence suggests that the subfornical organ (SFO), a circumventricular organ well-known for cardiovascular control, is also involved in metabolic regulation. We therefore hypothesized that the SFO insulin receptor protects against the development of metabolic syndrome. Male mice (6.5 wks) harboring a conditional allele of the insulin receptor gene (
INSR)
underwent SFO-targeted delivery of an adenovirus encoding a control vector (AdLacZ, n=7) or Cre-recombinase (AdCre, n=11) for selective removal of the SFO insulin receptor. Both groups remained on normal chow for 10 wks. Removal of the SFO insulin receptor did not influence food intake, but resulted in an ~40% greater increase in body weight (AdLacZ vs AdCre: Δ2.2±0.3 vs. 3.8±0.4 g; p<0.05). Consistent with the increased body weight, SFO insulin receptor deletion was associated with overall elevations in adiposity (e.g., abdominal fat, AdLacZ vs AdCre: 0.19±0.04 vs. 0.33±0.05 g; p<0.05). Analysis of the liver revealed substantial hepatic triglyceride accumulation in SFO-targeted AdCre mice (AdLacZ vs. AdCre: 62 ± 16 vs. 209 ± 29 mg/dl; p<0.05), with histological examinations (Oil Red O) revealing large lipid droplet accumulation following removal of the SFO insulin receptor. Parallel elevations in circulating triglycerides were also found (AdLacZ vs. AdCre: 1.4 ± 0.2 vs. 3.3 ± 0.6 mg/dl; p=0.05). These data demonstrate that ablation of SFO insulin receptors resulted in an overall deleterious metabolic state including increases in body weight, elevations in adiposity, hepatic steatosis and hypertriglyceridemia. These findings suggest that impairments in insulin signaling within the SFO contribute to the development of metabolic syndrome. Studies are ongoing to investigate the effect of SFO insulin receptor removal on blood pressure.
Abstract 088: Exercise Training Restores miRNA-1 and -29c in Obese Preventing Pathological Cardiac Hypertrophy via Targets in the Collagen and Calcium- Signaling Pathway
Authors: Silveira André C , Fernandes Tiago , Soci Ursula P , Negrao Carlos Eduardo , Oliveira Edilamar M
Published: March 2022

Introduction:
Overweight and obesity are risk factors in several cardiovascular diseases that lead to the pathological cardiac hypertrophy (CH) phenotype. We previously reported that aerobic exercise training (AET) counteracts CH in obesity. Here, we evaluate the role of microRNAs (miRs) and target genes involved in the AET-induced prevention and improvement of ventricular compliance in CH of obese Zucker rats.

Methods and Results:
Zucker rats were assigned into four groups: 1) lean group (LZR), 2) obese (OZR), 3) trained lean group (LZR+TR) and 4) trained obese group (OZR+TR). The AET consisted of swimming training with 10 weeks duration sessions of 60 min, 1x/day, 5x/week with overload by 4% of body weight. Our results showed that there were no differences in the cardiomyocytes diameter, however; heart weight was higher in OZR (29% - 0,031 to 0,024mg/cm) compared to LZR group, due to increased cardiac intramuscular fat and collagen. AET prevented pathological CH in OZR+TR group normalizing heart weight. Cardiac miR-29c expression was reduced (47% - 43 to 100% of LZR) in OZR compared to LZR group paralleled by an increase of collagen fraction. AET restored miR-29c expression in OZR-TR along with the expression of its target gene collagen. In addition, miR-1 targets NCX1 gene. MiR-1 was upregulated (63% - 163 to 100%) while their target gene NCX1 was reduced (51% - 49 to 100%) in OZR compared to LZR group. Corroborate with NCX1, pPLB
Ser16
was reduced in OZR group, indicating damage in calcium handling by obesity. Interestingly, AET normalized cardiac miR-1 and calcium signaling protein levels in OZR-TR group.

Conclusion:
Our findings show the AET as a non-pharmacological therapy for the prevention and even reversal of pathological CH and dysfunction in obesity. Together, the data suggests that miR-1 and -29c play key role in cardiac remodeling which could be used as potential therapeutic target for cardiac disorders.
Abstract 092: Low Birth Weight per se Is Not a Determinant of Vascular Dysfunction and Presumably Future Cardiovascular Risk in Humans
Authors: Rimoldi Stefano , Rexhaj Emrush , Brenner Roman , Sartori Claudio , Raio Luigi , Allemann Yves , Scherrer Urs
Published: March 2022

Background:
Epidemiological studies suggest that low birth weight (BW) is associated with systemic vascular dysfunction and premature cardiovascular morbidity and mortality later in life. However, this association could be due to confounding factors associated with low BW, such as genetic factors, pathological events during intrauterine life, and maternal or early-life environmental insults. Studies of monozygotic twins born with significantly different BW provide a unique opportunity to control for these potential confounding factors, thereby allowing to directly study the effect of low BW per se on systemic vascular function later in life.

Methods:
We, therefore, measured flow-mediated dilation (FMD) of the brachial artery, carotid-femoral pulse wave velocity (PWV) and carotid intima-media thickness (IMT) in 13 monozygotic, monochorionic healthy twins pairs (mean±SD age, 13.5±3.4 years) who were born with significantly different BW (defined as >20% BW difference within pairs, 2310±600 vs. 1800±520 g, P<0.0001) and in 26 healthy age- and sex-matched control subjects born at term with normal BW (3460±360 g). None of the participants had suffered from intrauterine or perinatal complications.

Results:
The major new findings were two fold; 1) systemic vascular function was comparable in low and high BW twins (FMD, 9.0±1.8 vs. 8.7±2.0%; PWV, 6.5±1.3 vs. 6.6±0.9 m/s; IMT, 390±30 vs. 390±20μm, all P values >0.2, low vs. high BW) and 2) systemic vascular function in twins was similar to the one observed in control singletons (FMD, 8.8±1.3 vs.8.9±1.8%, P=0.95; PWV: 6.5±1.1 vs. 6.6±1.2 m/s, P=0.80; IMT: 390±25 vs. 385±20μm, P=0.33, singletons vs. twins). Finally, there existed no relationship between BW and vascular function in the whole study population (FMD: r=0.001, P=0.99).

Conclusions:
This study provides the first direct evidence in humans that low BW per se is not a determinant of systemic vascular dysfunction (and presumably future cardiovascular risk). Moreover, it indicates that independent of BW, vascular function in twins is normal. We suggest that the association between low BW and increased cardiovascular risk reported in earlier epidemiological studies was related to confounding factors associated with low BW.
Abstract 093: Could Spironolactone Be More Effective Than Sympathetic Renal Denervation to Treat True Resistant Hypertension? Preliminary Results From the DENERVHTA Study.
Authors: Oliveras Anna , Armario Pedro , Sans Atxer Laia , Clarà Albert , Vázquez Susana , Vera Maria , Ho Tai Mooi , Faura Anna , Pascual Julio , de la Sierra Alejandro
Published: March 2022

Aim:
To compare the efficacy and safety between two therapeutic strategies to reduce 24h-SBP in patients with resistant hypertension: renal denervation or the addition of spironolactone.

Methods:
Twenty-one patients with office-SBP ≥150 mmHg and 24h-SBP≥140 mmHg despite receiving ≥3 full-dose antihipertensive drugs, one a diuretic, but without aldosterone antagonists, were randomized to renal denervation (by Simplicity®) or to spironolactone (25-50 mg), as add-on therapy. Changes in both office- (3 averaged readings) and 24h- BP (by Spacelabs®-90207) were evaluated at 6 months. Comparisons between treatment groups were performed using analysis of variance adjusted by age, gender and baseline values.

Results:
mean age was 62.7 ± 7.6 yr; men: 61.9% (13 of 21); diabetes: 47,6% (10 of 21). Mean BMI: 32.3 ± 6.1 Kg/m
2
. Duration of hypertension: 13.4 ± 7.2 yr. Number of antihypertensive drugs: 4,1 ± 0,7. Mean office-BP: 167,5 ± 20,1 / 91,7 ± 12,4 mmHg. Mean 24h-BP: 151,8 ± 9,1 / 81,7 ± 8,4 mmHg. Baseline characteristics were not different between groups (p=NS for all). Comparison between groups of main changes over time is shown in Table.

Reduction of 24h-SBP was higher in the spironolactone group after adjusting by age, sex and baseline 24h-SBP (p=0.016).

Reduction of eGFR was higher in the spironolactone group after adjusting by baseline eGFR (p=0.033).

Conclusions:
1) As compared to renal denervation, spironolactone was more effective to reduce 24h-SBP after 6 months in patients with resistant hypertension. These differences were not significant as regards office-BP.

2) Spironolactone add-on treatment significantly decreased eGFR as compared to renal denervation treatment.
Abstract 100: Sorting Nexin 1: Molecular Mechanisms and Pharmacogenomics
Authors: Yang Jian , Asico Laureano , Beitelshees Amber , Wang Xiaoyan , Jones John , Armando Ines , Cuevas Santiago , Zeng Chunyu , Felder Robin , Weinman Edward , Jose Pedro , Villar Van Anthony M
Published: March 2022

Sorting nexin 1 (SNX1) plays a pivotal role for the normal activity of renal dopamine D5 receptor (D5R). Kidney-restricted, Snx1-siRNA depletion of SNX1 results in impaired natriuretic response to salt load and hypertension in mice. Genetic ablation of the Snx1 gene (Snx1-/-) resulted in increased oxidative stress, impaired sodium excretion, and elevated systolic blood pressure (SBP, 131.3±6.4 mm Hg, n=5) in mice. The D5R has antioxidant properties by negatively regulating the expression of the NADPH oxidase (NOX). We found that NOX1, NOX2, and p47phox, as well as the antioxidant PON2, conceivably as compensation, were increased in Snx1-/- mice compared with wild-type littermates. Snx1-/- mice had higher ROS (218.6±7.7%), NOX activity (43.9±3.3 AU/mg protein/min vs. 25.98±3.5), and other markers of oxidative stress, e.g., malonyldialdehyde (32.5±3.4 pmol/mg protein vs. 17.2±2.1) and 3-nitrotyrosine (128.3±4%), which were all normalized by 10-day renal infusion of apocynin, a drug that prevents NOX assembly. The SBP in Snx1-/- mice was also normalized by apocynin (131.3±4.8 mm Hg to 105.7±0.3). Compared with human renal proximal tubule cells obtained from normotensive Caucasian males (NT cells), those from hypertensive subjects (HT cells) had reduced expression of SNX1 (160±2.1%, n=4=5/group), increased ROS (182±10.5%), and blunted cAMP response (110.8±35.3%) and sodium transport inhibition (101.2±1.9%) in response to D1-like receptor stimulation. These observations were corroborated by results in siRNA-induced gene silencing in NT cells or “genetic rescue” in HT cells. We also evaluated 12 SNPs in the SNX1 gene as possible genetic predictors of BP response to monotherapy with HCTZ among hypertensive patients enrolled in the Pharmacogenomic Evaluation of Antihypertensive Responses study (n=768). Three of the 12 SNPs (rs12591947, rs11854249, and rs11635627) associated with poor BP response to thiazide ([[Unable to Display Character: ∆]] SBP of -1.8 mm Hg vs. 113.5) among blacks. An SNX1 SNP (rs1802376) was associated with essential hypertension in a Caucasian population (n=502). Our data demonstrate the novelty and relevance of SNX1 in human pathology and pharmacogenomics of essential hypertension.
Abstract 101: Interleukin 17A Upregulates Both Renal Proximal and Distal Tubule Sodium Transporters in Angiotensin II-dependent Hypertension
Authors: Norlander Allison E , Kamat Nikhil , Ko Benjamin , Kirabo Annet , Gnecco Juan , Saleh Mohamed A , Hoover Robert S , McDonough Alicia , Madhur Meena S
Published: March 2022

We have previously shown that angiotensin II (Ang II)-induced hypertension is associated with an increase in T cell production of interleukin 17A (IL17A), and that IL17A promotes hypertension and end-organ damage. However, the precise mechanism is unknown. Recently, we reported that Ang II infusion into C56Bl/6J wild type (WT) mice blunted the rate of natriuresis following an acute saline challenge, while the rate of salt and water excretion in IL17A-/- mice was unaffected by Ang II. Following 2 weeks of Ang II infusion (490 ng/kg/min), proximal tubule sodium hydrogen exchanger 3 (NHE3) abundance was depressed in IL17A-/- but not WT mice, suggesting enhanced pressure natriuresis in IL17A-/- mice. We then performed renal transporter profiling on mice deficient in IL17A, or the related isoform IL17F, after prolonged (4 weeks) of Ang II infusion (490 ng/kg/min), a time when the blood pressure reduction in IL17A-/- mice is most prominent. Interestingly, at this time, deficiency of IL17A, but not IL17F, blunted the activation of distal tubule transporters, specifically sodium-chloride cotransporter (NCC) and the epithelial sodium channel (ENaC). We hypothesized that IL17A directly modulates renal sodium transporters as a mechanism to regulate salt and water excretion and hypertension. To test this hypothesis, we treated cultured human renal proximal tubule cells and mouse distal convoluted tubule (mDCT15) cells with recombinant IL17A or IL17F. We found that IL17A, but not IL17F, increased NHE3 protein levels (1.4-fold, p=0.003) and SGK1 mRNA expression (3.9-fold, p=0.01). In mDCT15 cells, IL17A but not IL17F, increased NCC activity as measured by thiazide-inhibited sodium uptake (1.78 vs 1.62 μmol/mg/20min, p<0.001), and this increase was significantly blunted with an SGK1 inhibitor (GSK 650394) and in cells lacking Nedd4-2 (an E3 ubiquitin ligase downstream of SGK1). Moreover, in mDCT15 cells, acute IL17A treatment caused phosphorylation of SGK1 on Ser78. These studies are the first to describe a mechanistic link by which IL17A modulates renal sodium transporters and suggests that targeting IL17A may improve renal function and slow the progression to renal failure in hypertension and other autoimmune disorders.