bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 085: The Role of Afferent and Efferent Renal Nerves in T Lymphocyte Recruitment into the Kidneys and Development of Two Models of Salt-Dependent Hypertension
Authors: Foss Jason D , Fiege Jessica , Van Helden Dusty , Shimizu Yoji , Collister John P , Mayerhofer Tim , Wood Laurel , Osborn John W
Published: July 2021

Renal denervation (RDNX) attenuates hypertension (HT) in some animal models and human patients, yet the antihypertensive mechanisms involved are not known. Because it has been suggested that efferent renal nerves may cause recruitment of T lymphocytes (T cells) into the kidneys, and T cell-secreted cytokines can activate afferent neurons, we hypothesized that RDNX attenuates HT in the angiotensin II (AngII)-salt and deoxycorticosterone acetate (DOCA)-salt models by decreasing efferent renal nerve-dependent T cell trafficking to the kidneys and T cell-mediated activation of afferent renal nerves. Male Sprague-Dawley rats underwent SHAM surgery, RDNX or selective ablation of afferent renal nerves (renal-CAP) and were subjected to either AngII-salt or DOCA-salt HT. AngII-salt rats underwent SHAM, RDNX or renal-CAP (n = 8/group) and were instrumented with an IV catheter and radio-telemeter to measure mean arterial pressure (MAP). Rats were fed a 4% NaCl diet and, after a baseline period, AngII was infused IV (10 ng/kg/min) for 16 days. DOCA-salt rats were unilaterally nephrectomized and, after 2 weeks, instrumented with a radio-telemeter, subjected to SHAM (n = 5), RDNX (n = 7) or renal-CAP (n = 6) and given 0.9% saline to drink. After a baseline period, silicone pellets containing 100mg DOCA were implanted SC and rats were sacrificed 3 weeks later. Upon sacrifice, kidneys were harvested for flow cytometry to determine the number of CD4+ and CD8+ T cells in the kidneys. In AngII-salt rats, renal CD4 and CD8 T cell counts and the change in MAP from baseline (ΔMAP) were similar between groups (SHAM = 42.3 ± 6.4 mmHg, RDNX = 32.6 ± 5.9 mmHg, renal-CAP = 36.8 ± 5.0 mmHg). In DOCA-salt rats, RDNX and renal-CAP significantly attenuated ΔMAP (SHAM = 24.2 ± 3.1 mmHg, RDNX = 12.7 ± 2.4 mmHg, renal-CAP = 13.8 ± 1.7 mmHg). CD8 T cell counts in RDNX kidneys were significantly lower than SHAM (p < 0.01) and trended lower than renal-CAP. CD4 T cell counts in RDNX kidneys were significantly lower than renal-CAP (p < 0.05) and trended lower than SHAM. These data show that renal nerves do not contribute to AngII-salt HT, but suggest that afferent renal nerves contribute to DOCA-salt HT and that this may be T cell-mediated and dependent on efferent renal nerves. Funding: R01 HL116476-01.
Abstract 095: CD74 Deficient Mice are Resistant to Toll-Like Receptor-Induced Preeclampsia
Authors: Hatahet Mohamad , Gasheva Olga Y , Chiasson Valorie L , Chatterjee Piyali , Bounds Kelsey R , Tobin Richard P , Newell-Rogers M. K , Mitchell Brett M
Published: July 2021

Preeclampsia (PE) is a pregnancy-specific hypertensive disorder characterized by vascular endothelial dysfunction and excessive immunity and inflammation. Activation of the dsRNA receptor Toll-like receptor 3 (TLR3) or the ssRNA receptor TLR7 elicits a pregnancy-dependent PE-like syndrome in mice by inducing a pro-inflammatory immune response. CD74 (MHC Class II invariant chain) acts as a chaperone for MHC Class II surface expression on immune cells during antigen presentation and is cleaved into Class II-Associated Invariant Peptide (CLIP) following polyclonal activation of immune cell TLRs. The presence of CLIP in the groove of MHC Class II prevents T cell-dependent death leading to persistent immune cell activation. We hypothesized that genetic deletion of CD74 and subsequent depletion of CLIP on immune cells prevents TLR-induced immune responses and the development of PE in mice. Pregnant WT and CD74 KO mice were given i.p. injections of normal saline (P), poly I:C (TLR3 agonist; P-PIC), or R837 (TLR7 agonist; P-R837) on gestational days 13, 15, and 17 and euthanized on day 18. P-PIC and P-R837 WT mice had significantly increased splenic levels of pro-inflammatory CD3+/gd T cells and plasma levels of the gd T cell-derived cytokines IFNg, TNFa, and IL-17 compared to P WT mice whereas P-PIC and P-R837 CD74 KO mice had significantly increased anti-inflammatory CD3+/gd T cells and no significant increases in plasma IFNg, TNFa, and IL-17 levels. P-PIC and P-R837 CD74 KO mice did not develop the hypertension (gd17 SBP in mmHg: P WT=102±3, P CD74 KO=100±3, P-PIC WT=147±4*, P-PIC CD74 KO=95±3, P-R837 WT=133±2*, P-R837 CD74 KO=97±1; *p<0.05 vs. P WT), endothelial dysfunction, proteinuria, or placental necrosis seen in P-PIC and P-R837 WT mice. In conclusion, CD74 is crucial for the development of TLR-induced PE-like symptoms in mice and CD74/CLIP depletion may be a promising therapeutic target for women with PE.
Abstract 232: Urinary Proteomics in Patients with Type 2 Diabetes at High Cardiovascular Risk
Authors: Currie Gemma E , Friar Matthew , Brown Catriona E , Flynn Joanne , Mullen William , Mischak Harald , Delles Christian
Published: July 2021

Background:
We have previously demonstrated the value of a urinary proteomic classifier (CKD273) for diagnosis and prediction of diabetic nephropathy (DN). Whether CKD273 is only related to renal damage or also reflects generalised vascular damage in patients with diabetes remains unclear.

Methods:
We recruited 45 patients at high cardiovascular (CV) risk from secondary care: 15 normoalbuminuric; 15 with MA; and 15 with DN (urinary albumin:creatinine ratio: 1.1 (0-3.3), 7.7 (2.6-22.5) and 124.5 (0.8-412.6) mg/mmol; estimated glomerular filtration rate (eGFR): 74 (46-125), 69 (49-100) and 37 (6-60) mL/min/1.73m
2
). Participants underwent pulse wave analysis for assessment of heart rate-corrected central augmentation index (AIx@75) and ultrasound for assessment of carotid intima-media thickness (c-IMT). Spot urine samples were analysed using capillary electrophoresis coupled to mass spectrometry.

Results:
There was no difference in age (61±8, 64±6 and 59±7 years;
P
=0.130), body mass index (34.4±6.2, 35.1±8.1 and 34.4±6.7 kg/m
2
;
P
=0.955) and blood pressure (144±15/83±7, 149±20/83±10 and 148±16/82±12 mmHg;
P
=0.765/0.910) between groups. In keeping with inclusion criteria participants were at high CV risk (Framingham score: 30±11, 38±12 and 32±12;
P
=0.141; ASSIGN Score: 36±15, 43±15 and 39±17;
P
=0.415) and had subclinical vascular damage (AIx@75: 22 (7-38), 23 (13-21) and 25 (4-35)%;
P
=0.993; c-IMT: 0.723 (0.563-1.276), 0.760 (0.614-1.082) and 0.704 (0.581-0.986) mm;
P
=0.305) independent of eGFR (
r
=0.259,
P
=0.086 for c-IMT;
r
=-0.082,
P
=0.598 for AIx@75). Despite similarities in CV risk and vascular phenotypes the CKD 273 classifier was significantly different between the groups (-0.169±0.373, 0.421±0.467 and 0.765±0.434;
P
=0.002) but not related to c-IMT (
r
=-0.075,
P
=0.747) or AIx@75 (
r
=-0.299,
P
=0.200). The previously defined CKD273 cut-off of 0.343 had 71% sensitivity for detecting MA or DN with specificity of 100%.

Conclusions:
CKD273 reliably distinguished normoalbuminuria from MA or DN in this high risk cohort independent of the vascular phenotype. Neither traditional renal markers nor a novel proteomic classifier appear to fully explain the vascular damage in our patients.
Abstract 238: Genetic Risk Score as a Predictor of Incident Hypertension and Blood Pressure Change Over Time
Authors: Niiranen Teemu , Mäki Juhani , Havulinna Aki S , Salomaa Veikko , Jula Antti
Published: July 2021

Little data exist regarding single nucleotide polymorphisms (SNPs) predicting blood pressure (BP) change over time. We genotyped 32 common SNPs in a nationwide cohort aged ≥30 years examined in years 2000 (n=5402) and 2011 (n=3373). We investigated whether genetic risk scores (GRSs) constructed of these variants would predict incident hypertension and BP change over time. We created GRSs for systolic and diastolic BP by multiplying the risk allele count of each SNP by the effect size estimated in published genome-wide association studies. In linear and logistic regression models adjusted for traditional risk factors, 1-unit increases in GRSs were associated with higher systolic and diastolic BP values both at baseline (β±SE, 1.04±0.14 mmHg and 1.11±0.13 mmHg; P<0.0001 for both) and at reinvestigation (β±SE, 0.86±0.18 mmHg and 0.74±0.16 mm Hg; P<0.0001 for both) and with increased hypertension prevalence (odds ratio [95% CI], 1.12 [1.08-1.16] and 1.18 [1.11-1.26]; P<0.0001 for both). Among all participants who were normotensive at baseline and participated in the re-examination (n=2045), the GRSs were not independently associated with BP change over time (β±SE, 0.016±0.18 mmHg and 0.019±0.18 mmHg; P≥0.27 for both). However, in the top tertile of the GRS, the predicted increase in BP compared with the bottom tertile was 1.59±0.78 mmHg greater for systolic BP (P=0.04) and 0.79±0.49 mmHg greater for diastolic BP (P=0.11) and the odds ratio for incident hypertension was 30% higher (P=0.04). Our data show that GRSs are strongly associated with BP and prevalence of hypertension, but only weakly associated with BP increase and incidence of hypertension in a general adult population.
Abstract 243: Echocardiographic Changes of Left Ventricular Geometry and Function Associated with Reverse Epidemiology of Blood Pressure in End Stage Kidney Disease.
Authors: Afshinnia Farsad , Segal Jonathan H
Published: July 2021

Background:
We hypothesize that reverse epidemiology of blood pressure (BP) defined as higher mortality associated with lower BP in ESKD is partially mediated by deteriorating left ventricular (LV) function and configuration.

Methods:
In a prospective observation 2136 patient-visits from 88 consecutive eligible prevalent ESKD patients on peritoneal dialysis from a single center between 2007 and 2012 were assessed. Monthly clinic BP was recorded. Routinely performed 285 interpretable serial echocardiograms were obtained. Patients were followed to primary end-point of all-cause mortality. Generalized linear mixed models using BP from last observation moving backward to assess trends, and time to event analysis using time varying Cox-survival models to estimate mortality hazard ratio of different BP categories were applied.

Results:
There were 47 males (53.4%). Mean age was 50 years (SD=15). Median follow up was 35 months. During follow up 22 patients (25%) died. As compared to BP of 140-159 mmHg, BP<120 mmHg was associated with 8.6-fold (95% CI: 1.8-40, P=0.006) higher mortality. As compared to survivors who did not show any change, in deceased patients systolic BP (SBP) declined to a statistically lower level 9 months prior to death (117 vs 141 mmHg, P<0.05), left ventricular mass index (LVMI) increased by 40% from 83 to 128 g/m
2
, EF<50% increased by 30%, and concentric left ventricular hypertrophy (LVH) decreased by 20% at the expense of a 20% reciprocal increase in eccentric LVH. After adjusting for LVMI, risk of mortality at SBP<120 mmHg decreased to 4.4 (95% CI: 0.8 to 24, P=0.09).

Conclusion:
Higher mortality with lower BP was likely mediated by worsening heart function.
Abstract 261: Chloroquine, an Inhibitor of Endosomal Toll-like Receptors, Improves Blood Pressure and Endothelial Function in Spontaneously Hypertensive Rats
Authors: McCarthy Cameron G , Wenceslau Camilla F , Goulopoulou Styliani , Ogbi Safia , Matsumoto Takayuki , Webb R. Clinton
Published: July 2021

HBPR 2014

Circulating mitochondrial DNA (mtDNA) is elevated in spontaneously hypertensive rats (SHR) and the enzyme responsible for its degradation, DNase II, has decreased activity in SHR tissues. Moreover, a synthetic mtDNA mimetic, specific for TLR 9, causes endothelial dysfunction and elevated blood pressure. Therefore, we sought to inhibit the contribution of mtDNA on innate immune system activation, via inhibition of Toll-like receptor (TLR)9, in SHR. We treated 12-15 week old SHR and Wistar-Kyoto rats (WKY) with chloroquine (CQ; 40mg/kg/day) or vehicle (veh; saline) for 21 days via intraperitoneal injection (
i.p.
). We hypothesized that CQ treatment would improve endothelial function and decrease blood pressure in SHR. Blood pressure was measured pre- and post-treatment via tail cuff and endothelial function was measured via mesenteric resistance artery (MRA) relaxation to acetylcholine (ACh; 10
-9
-10
-5
M) using a wire myograph. Treatment with CQ lowered post-systolic blood pressure in SHR (mmHg, Veh: 201±2 vs. CQ: 185±5 mmHg; p<0.05). No effects of treatment were observed on blood pressure in WKY (p>0.05). CQ abolished the contractile effect to high concentrations of ACh [AUC (%NE pre-contraction), Veh: 160±25 vs. CQ: 277±20; p<0.05] in MRA from SHR. However, no differences were observed in endothelium-independent relaxation to NO-donor sodium nitroprusside [E
max
(%NE pre-contraction), Veh: 97±12 vs. CQ: 102±26; p<0.05] in MRAs from SHR. Again, no effects of treatment were observed for endothelium-dependent or -independent relaxation in WKY (p>0.05). These data demonstrate the inhibition of endosomal TLRs, including TLR9, improves blood pressure and endothelial function in SHR. Inhibition of TLR9 abrogates the potentially deleterious contribution of increased mtDNA and impaired DNase II activity on innate immune system activation in hypertension.
Abstract 286: Immune Dysfunction in a Vasopressin-Induced Mouse Model of Preeclampsia
Authors: Scroggins Sabrina M , Santillan Donna A , Min James Y , Sangren Jeremy A , Pearson Nicole A , Gibson-Corley Katherine N , Grobe Justin L , Santillan Mark K
Published: July 2021

Preeclampsia is a disease of pregnancy and the postpartum period, affecting 5-7% of all U.S. pregnancies. We recently determined that elevated secretion of arginine vasopressin (AVP) in humans is detected as early as the 6th week of pregnancy in the women that subsequently develop preeclampsia. Further, we have determined that in C57BL/6 mice, chronic AVP infusion serves as a clinically-relevant and pregnancy-specific model of preeclampsia. Although the pathogenesis of preeclampsia is still poorly understood, an inflammatory immune response is clearly involved. AVP secretion is stimulated by proinflammatory cytokines and AVP in turn can act upon receptors present on lymphocytes. We hypothesize that hypersecretion of AVP results in maternal-fetal immune alterations in preeclamptics. Immune responses were evaluated in wild-type C57BL/6 female mice chronically infused with AVP (24 ng/hr or saline s.c.) throughout pregnancy. We observed an increase in the frequency of IL-12 (saline N=7: 1.1±0.2, vs AVP N=9: 2.2±0.4, P<0.05), IL-17 (saline N=7: 1.2±0.17, vs AVP N=9: 2.6±0.4, P<0.05) and TNFa (saline N=8: 0.9±0.1, vs AVP N=9: 1.6±0.1, P<0.005) expressing lymphocytes isolated from AVP mice. Additionally, the frequency of IFNg expressing lymphocytes was increased in AVP mice, approaching statistical significance (saline N=8: 2.2±0.9, vs AVP N=9: 3.0±0.4, P=0.059). The frequency of IL-6, IL-10, and LAP expressing splenic lymphocytes were similar between saline vs. AVP mice. Interestingly, maternal plasma revealed a significant reduction in IL-17 (saline N=4: 1.0e+7±3.8e+6, vs AVP N=5: 3.1e=5±1.3e+5 ng/g, P<0.05) in AVP mice. Neither IL-4 nor IFNg were detectable in maternal plasma of either group. Additionally, IL-17 trended higher in the amnionic fluid and lower in the placenta of AVP mice. IL-4 production trended lower in both the placenta and amnionic fluid of AVP-infused mice, while IFNg was similar between groups. These data support our hypothesis that AVP hypersecretion results in the maternal-fetal immune alterations associated with the development of preeclampsia. Ongoing experiments are aimed at identifying the lymphocytes and cytokines involved as well as local vs. systemic immune dysfunction associated with AVP-induced preeclampsia.
Abstract 298: Protective Role of Endogenous 20-HETE in Renal Ischemia-Reperfusion Injury
Authors: Muroya Yoshikazu , Fan Fan , Jacob Howard , Geurts Aron , Roman Richard
Published: July 2021

The present study compared renal ischemia-reperfusion (IR) injury in Dahl salt-sensitive (SS) rats that have a deficiency in the renal formation of 20-HETE versus CYP4A1 transgenic SS (SS.4A1) rats in which the renal production of 20-HETE is restored. The concentrations of free 20-HETE in the renal cortex and outer medulla were significantly greater in SS.4A1 than in SS rats. Renal 20-HETE levels rose to a greater extent in SS.4A1 than in SS rats following renal IR. Plasma creatinine level rose to 3.7 ± 0.1 in SS versus 1.8 ± 0.3 mg/dl in SS.4A1 rats (respectively, n=6) following 30 min of ischemia and 24 h reperfusion. The % of necrotic tubules and apoptotic cells were 4-fold higher in SS than in SS.4A1 rats. Administration of the 20-HETE synthesis inhibitor (HET0016, 10 mg/kg) abolished the resistance of SS.4A1 rats to renal IR injury and plasma creatinine level rose to 3.8 ± 0.1 mg/dl (n=6). Cortical blood flow in SS, SS.4A1 and HET0016 treated SS.4A1 rats immediately returned to control following IR. However, medullary blood flow in SS and HET0016 treated SS.4A1 rats fell to 30 % of control 3 h after IR (n=5), and it remained depressed for 24 h. In contrast, medullary blood flow did not decline following IR in SS.4A1 rats. Proximal intratubular pressure rose from 13 to approximately 40 mmHg, 2 h after IR in both SS and SS.4A1 rats. Proximal intratubular pressure remained much higher in SS than in SS.4A1 rats 24 h after IR (32 vs 19 mmHg). These data indicate that normalization of renal CYP4A activity and 20-HETE production opposes renal IR injury by preventing secondary fall in medullary blood flow and the prolonged renal medullary ischemia.
Abstract 311: Early Losartan Treatment Prevents the Developmental Programming of Cardiac Dysfunction Caused by Neonatal High Oxygen Exposure in Rats
Authors: Bertagnolli Mariane , Béland-Bonenfant Sarah , Dios Anne , Dartora Daniela R , Lukaszewski Marie-Amélie , Cloutier Anik , Sutherland Megan , DeBlois Denis , Paradis Pierre , Schiffrin Ernesto L , Nuyt Anne Monique
Published: July 2021

Neonatal oxidative stress is a major postnatal deleterious factor predisposing preterm born infants to classical complications of prematurity (retinopathy, bronchopulmonary dysplasia) which are characterized by impaired vascular development. Our group has previously shown that rats transiently exposed to high O2 as newborns (mimicking human preterms oxidative stress conditions) develop high blood pressure (BP), cardiac remodeling and dysfunction later in life, in part mediated by the renin angiotensin system (RAS). Cardiac RAS activation is characterized by AT1/AT2 receptors imbalance, with increased AT1R at adult age. In order to study the role of RAS at early stages of the developmental programming of cardiac dysfunction caused by high O2 exposure, we assessed whether an early and short-term treatment with AT1R blocker Losartan, prevents cardiac alterations at young male 4 wks-old rats (prior to the elevation of BP in this model). Sprague-Dawley newborns rats were kept with their mother in 80% O2 (O2 group, n=9) or room air (Ctrl, n=9) from days 3-10 of life (P3-P10). Losartan (LOS, n=10, 20 mg/Kg) or water was administered by gavage in O2 rats from P8-P10 (last 2 days of O2 to avoid impact on nephrogenesis). At 4 wks, echocardiography reveals that O2 rats have decreased fraction of shortening compared to Ctrl (FS: 37±2 vs 42±2 %), suggesting impaired systolic function in O2. Cardiac hypertrophy evaluated by heart/body weight and cardiomyocyte surface area (CSA) is also increased in O2 vs Ctrl (141±13 vs 118±4 μm2). LOS treatment prevented the impairment of systolic function in O2 by ameliorating FS (43±2 %) and reducing CSA (121±11 μm2). LOS treatment also modulated RAS genes expression (RT-PCR): LOS restored AT1/AT2 balance in O2 hearts by decreasing AT1b subunit (0.8±0.2 O2 vs 1.3±0.3 O2+LOS vs 0.9±0.2 Ctrl) as well as increasing ACE2 (1.5±0.4 O2 vs 0.8±0.1 O2+LOS vs 1.1±0.4 Ctrl) expressions. In conclusion, a short-term treatment with LOS during neonatal O2 exposure prevents the impairment of cardiac systolic function and hypertrophy at young age. This data reinforces the key role of RAS in the developmental programming of cardiac dysfunction and reveals LOS as an effective strategy to prevent early cardiac alterations caused by neonatal high O2 exposure.
Abstract 312: Jagged1 and Morphological Integrity of the Kidney
Authors: Belyea Brian C , Yu Fang , Sequeira-Lopez Maria Luisa S , Gomez R. Ariel
Published: July 2021

Renal vascular development is dependent on the participation of renin precursor cells, which localize to areas of new vessel formation and differentiate into smooth muscle cells (SMCs), mesangial and juxtaglomerular (JG) cells. However, the mechanisms that enforce this particular fate of renin precursor cells remain unclear. During embryogenesis, renin cells express a significant number of angiogenic factors including the Notch signaling ligand Jagged1. We therefore hypothesized that Jagged1 is necessary for renin precursors to differentiate into the various mural cells of the renal arterioles and that Jagged1 plays a critical role in renal vascular development. To generate conditional Jagged1 deletion in cells of the renin lineage, we crossed

Jagged1
fl/fl

mice with

Ren1d
cre/+

mice. We investigated kidney weight, morphology, and vascular architecture as well as the identity of cells composing the kidney arterioles including renin cells and SMCs. We performed quantitative RT-PCR on kidney cortex mRNA for
Jagged1
and
Renin
expression levels and immunohistochemistry for renin. The expression of
Jagged1
mRNA in conditional knockout (cKO) animals was reduced at 2 weeks (68.3% reduction) and 4 weeks (66.5% reduction) compared to control mice. We found no significant differences between control and cKO animals in their kidney weights,
renin
mRNA expression, and renin staining. The number of renin-positive juxtaglomerular apparatuses (JGA) was 49.25 +/- 3.301 (n=4) and 38.00 +/- 4.830 (n=4) for control and cKO mice respectively under physiologic conditions. Staining for α-smooth muscle actin (α-SMA) demonstrated an overall normal vascular anatomy in cKO kidneys, however there were focal areas containing activated pericytes and injured mesangial cells expressing α-SMA indicating an active fibrotic process. Mason’s trichrome staining confirmed areas of glomerular and interstitial fibrosis in cKO kidneys. This work suggests that while Jagged1 is dispensable for renin expression, its loss in renin cell descendants affects perivascular-interstitial integrity and glomerular structure. The findings indicate an important role of Jagged1 in the maintenance of the morphologic integrity of the kidney.
Abstract 328: High Fructose but not High Glucose Intake Induces Salt-sensitive Hypertension and Enhances NKCC2 Phosphorylation in Normal Rats.
Authors: Henson Emily , Ares Gustavo , Haque Mohammed , Ortiz Pablo
Published: July 2021

Consumption of fructose as a sweetener has increased in the past three decades. A high-fructose diet has been implicated in the epidemic of diabetes, obesity, and hypertension. A third of the US population consumes 20-40% of their caloric intake from added sugars, with half of those calories from fructose. Little is known about the role of high fructose intake in renal salt handling and blood pressure regulation during high salt intake. In genetic models of salt-sensitive hypertension, the Na/K/2Cl cotransporter NKCC2 plays an important role by reabsorbing NaCl in the thick ascending limb (TAL). We hypothesized that 20% fructose in drinking water stimulates NKCC2 and sensitizes normal rats to high salt induced hypertension. Adult Sprague-Dawley rats were given 20% fructose or 20% glucose in drinking water for 1 week after which a high salt diet (4% Na in chow) was started. Systolic blood pressure (SBP) was measured every other day by tail cuff after 2 weeks of training. After one week of fructose or glucose alone, SBP did not change. In rats fed fructose, adding a 4% NaCl diet increased SBP to 128±6 mmHg by day 2 (p<0.01 vs glucose) and continued to increase up to 144±18 mmHg after 2 weeks on high salt (p<0.01 vs baseline; p<0.01 vs glucose). In glucose-fed rats high salt did not increase SBP (from 122±6 to 116±9 mmHg). 20% fructose alone for 3 weeks, or high salt alone did not change SBP. NKCC2 phosphorylation at Thr96,101 is associated with enhanced TAL NaCl reabsorption. We found that NKCC2 phosphorylation at Thr96,101 (normalized to total NKCC2) was higher in TALs isolated from rats fed fructose plus salt for 2 weeks compared to high salt alone (high-salt: 100%; fructose + high-salt: 250±40%, p<0.05). We concluded that a high fructose but not high glucose diet induces salt-sensitive hypertension in Sprague Dawley rats. This effect occurs within 1 week of a high fructose diet. In addition, a high fructose diet may stimulate NKCC2 activity by enhancing its phosphorylation. These data suggest that high fructose intake may increase blood pressure by preventing appropriate renal NaCl excretion during high dietary salt intake.
Abstract 331: Mutation in the PPARG Ligand Binding Domain Impairs PPARG-Mediated Turnover of the p65 Subunit of NF-kB
Authors: Mukoda Masashi , Stump Madeliene , Ketsawatsomkron Pimonrat , Quelle Frederick W , Sigmund Curt D
Published: July 2021

Peroxisome proliferator-activated receptor gamma (PPARγ) is a ligand activated transcription factor regulating metabolic and vascular function. PPARγ exerts anti-inflammatory actions, and recent data suggest this may be mediated by promoting the degradation of the p65 subunit of nuclear factor kappa B (NFκB). Transgenic mice expressing dominant negative PPARγ specifically in vascular smooth muscle or endothelium exhibited exacerbated atherosclerosis but the mechanism remains unknown. We hypothesized that PPARγ mutants promote inflammation because PPARγ-mediated p65 degradation is impaired. We tested this by co-transfection of HEK293T cells with vectors encoding p65, wildtype (WT) PPARγ, or various PPARγ mutants. The level of p65 protein expression was decreased by co-expression with WT-PPARγ (0.53±0.09 vs control, n=8). Whereas, the P467L PPARγ exhibited impaired degradation of p65 (1.0±0.06, n=10), the V290M (0.36±0.1), S273A (0.37±0.06), or K268R/K293R (0.41±0.03) mutations in PPARγ preserved p65 degradation. WT PPARγ was co-precipitated with p65 in co-transfected cells suggesting the mechanism of PPARγ-mediated p65 degradation involves a direct interaction between them. Consistent with this, the interaction between p65 and P467L PPARγ was severely impaired. To assess functional interactions between PPARγ and NFκB, we employed a model of interleukin-1β (IL-1β) mediated dysfunction in aortic rings. IL-1β dose-dependently induced NFκB activity as measured by increased phospho-p65 and decreased IκBα in aorta cultured for 2 hours with IL-1β. IL-1β dose-dependently reduced acetylcholine (Ach)-induced endothelial-dependent relaxation of aortic rings (80±12 vs 39±16, 20 pg/mL vs 11±3, 100 pg/mL, %). IL-1β-mediated loss of Ach vasodilation was reduced by the PPARγ agonist rosiglitazone (1 μM, 25 hr, n=3, p<0.05), or by transgenic over-expression of WT-PPARγ specifically in endothelium (n=6, p<0.05). We conclude that 1) p65 turnover may be regulated by PPARγ and that its mutation can result in impaired p65 degradation, 2) PPARγ activity can protect against vascular dysfunction associated with NFκB activation, and 3) loss of PPARγ-mediated p65 degradation may contribute to inflammation in hypertension and atherosclerosis.
Abstract 333: Existence of Functional Norepinephrine Transport in Perivascular Adipose Tissue
Authors: Ayala-Lopez Nadia , Burnett Robert , Thompson Janice M , Watts Stephanie W
Published: July 2021

Perivascular Tissue (PVAT) is a recent focus of studies for its regulation of blood vessel tone. Contraction to norepinephrine (NE) is reduced by PVAT through mechanisms not entirely clear. Loss of this anti-contractile function of PVAT occurs in obesity-related hypertension. We hypothesized that PVAT can remove NE from the blood vessel environment through transporters and reduce NE induced contraction in health. RT-PCR of mesenteric PVAT was positive for norepinephrine transporter (NET; Ct=33.2±1.4) and serotonin transporter (SERT; Ct=31.9±0.9) relative to beta-2-microglobulin. To study NE uptake in the healthy state, mesenteric and aortic PVAT from the male Sprague Dawley rat was incubated with 10 μM NE after pre-incubation with an inhibitor of transport (10 μM desipramine to inhibit NET and SERT, 100 μM corticosterone to inhibit organic cation transporter 3) or vehicle and quantified by HPLC for NE. Desipramine plus corticosterone inhibited NE uptake in mesenteric PVAT (figure) but desipramine (410.5±80.8 ng/g vs. 414.4±67.0 ng/g NE only; n=8) or corticosterone (225.0±26.1 ng/g vs. 319.2±35.9 ng/g NE only; n=5) alone did not, indicating a robust uptake capacity. In aortic PVAT, NE uptake was not inhibited by desipramine plus corticosterone (figure), desipramine (1763.5±460.8 ng/g vs. 1702.7±298.4 ng/g NE only; n=6), or corticosterone (1085.8±205.3 ng/g vs. 2245.8±506.8 ng/g NE only; n=4). Protein staining revealed bands positive for NET in mesenteric PVAT by Western blot. Taken together, we found that PVAT is a reservoir of NE that can remove NE from the blood vessel environment, an important finding to understand vasculature-PVAT interactions in health and disease.
Abstract 346: Subcutaneous Resistance Artery Remodeling And Function In Chronic Kidney Disease Patients
Authors: Fraulob-Aquino Julio C , Briet Marie , Barhoumi Tlili , Savoia Carmine , Paradis Pierre , Schiffrin Ernesto L
Published: July 2021

Background:
Chronic kidney disease (CKD) is associated with cardiovascular (CV) complications. However, interventional trials targeting classical CV risks factors have been often unsuccessful in advanced stage CKD, which emphasizes the need to better understand CKD-associated vascular disorders. Resistance arteries are a key determinant of blood pressure (BP) and their changes in different CV conditions contribute to target organ damage. The aim of the present study was to characterize resistance artery remodeling and function in CKD patients, compared to vessels from hypertensive (HTN) subjects.

Method:
Twenty-two stage 4 CKD patients (aged 63.6±3.1 years) and 16 HTN subjects (45.6±16.1 years) were included in the present study. They all underwent a subcutaneous biopsy under local anaesthesia. Small artery remodeling and function were studied on a pressurized myograph, and subcutaneous fat CD3 infiltration and media fibronectin expression by immunostaining. Vascular smooth muscle cells (VSMCs) were counted after hematoxilin-eosin staining.

Results:
CKD systolic BP was similar to HTN (133±18 vs. 143±10 mmHg, respectively). Vasodilatory responses to acetylcholine were lower in CKD compared to HTN (maximal relaxation (%), 74.3±3.4 vs. 87.5±2.7, P<0.05). Media/lumen at 60 mmHg was lower in CKD than in HTN (6.7±0.5 vs 8.8±0.7, P<0.05). Resistance artery stiffness was lower in CKD compared to HTN (strain at 120 mmHg, 0.845±0.126 vs 0.585±0.099, P<0.05). Fibronectin staining in resistance arteries was lower in CKD than HTN (8.2±0.8 vs 23.3±1.7 RFU/μm2, P<0.001). Less VSMCs were present in the arterial wall of CKD compared to HTN (5.4±0.4 vs 7.2±0.5 cells/μm2, P<0.05). Subcutaneous fat presented fewer CD3+ cells in CKD than HTN (12.8±4.1 vs 23.7±12.8 cells/mm2, P<0.05).

Conclusion:
Despite higher levels of BP, resistance arteries isolated from CKD patients exhibited no vascular remodeling and lower arterial stiffness compared with HTN patients. These results are in line with the maladaptive hypotrophic remodeling observed in large vessels in CKD, suggesting a generalized vascular defect in mechanotransduction in CKD.
Abstract 355: Calculating Pulse Wave Velocity from a Single Pressure Waveform Using the Intrinsic Frequency Method
Authors: Pahlevan Niema M , Petrasek Danny , Rinderknecht Derek G , Tavallali Peyman , Gharib Morteza
Published: July 2021

INTRODUCTION:
Increased aortic stiffness is correlated with many clinically adverse cardiovascular outcomes. The “gold standard” quantitative index for arterial stiffness is the pulse wave velocity (PWV). We have developed a new method called the
Intrinsic Frequency
(
IF
), which views the arterial pressure waveform as a piecewise combination of two coupled systems, the heart and arterial system which are decoupled upon closure of the aortic valve. Each of these dynamical systems has an inherent frequency of operation (ω
1
and ω
2
) which gives information about LV function (ω
1
) as well as arterial dynamics (ω
2
).

METHODS:
IF
methodology is based on Sparse Time-Frequency Representation method. It uses an effective L
2
-minimization to extract the second intrinsic frequency (ω
2
) from an aortic pressure waveform. To examine the clinical relevance of this method,
IF
was applied to aortic pressure waveforms taken from published works. These aortic waveforms were selected from a healthy population free of any cardiovascular diseases (CVD).

RESULTS:
Our results show that ω
2
represents information about the arterial system and that these measurements are highly correlated with PWV (
r=0.9
).

CONCLUSION:
These results show ω
2
can potentially be used to evaluate aortic rigidity and calculate aortic PWV from one pressure waveform. Increased aortic rigidity is a common feature in normal aging and is accelerated in many CVDs including diabetes. One unique advantage of the method is that only a single measurement of the pressure waveform is required to extract the result. Therefore, ω
2
may be employed as a clinically effective noninvasive assessment of cardiovascular health.
Abstract 400: Blood Pressure Effect Of Aldosterone Increased With Age
Authors: Tu Wanzhu , Eckert George , Hannon Tamara , Pratt John Howard
Published: July 2021

Aldosterone often plays a central role in the regulation of blood pressure (BP) by acting on the distal nephron to promote sodium and water retention. As people age, levels of aldosterone and renin tend to decrease. It is unclear how these changes affect BP. To investigate, we examined the temporal changes in plasma aldosterone concentration (PAC) and plasma renin activity (PRA) in a cohort of healthy blacks and whites (ages: 7 - 38 years). We examined the relationship between PAC and BP at different ages. Our data showed that while blacks on average had lower PRA levels than whites at any given age (p<0.0001), PRA decreased with age (p<0.0001) in both race groups. PAC also decreased with age (p<0.0001) but decline was faster whites. Aldosterone-renin ratio (ARR), on the other hand, increased with age in both race groups. PAC effects on SBP in blacks and whites were estimated and presented graphically (Figure 1). The figure showed that aldosterone effect on BP increased with age in blacks and whites (p<0.008); but at any given age, the BP effect of PAC was greater in blacks than in whites, and this racial difference became more pronounced with an increase in age. In conclusion, BP becomes more sensitive to aldosterone with age. An accumulated sodium load with volume expansion may sensitize BP to even small increment of aldosterone. Blacks appear to be especially vulnerable to such an increased level of sensitivity.
Abstract 528: Dose- And Salt-dependency Of Angiogenesis Inhibition-induced Blood Pressure Rise And Renal Toxicity
Authors: Lankhorst Stephanie , Kappers Mariëtte H , Sleijfer Stefan , Danser A H , van den Meiracker Anton H
Published: July 2021

Angiogenesis inhibition with the VEGF inhibitor sunitinib is an established anti-cancer therapy, inducing hypertension and nephrotoxicity. We explored the dose- and salt-dependency of these side effects. In male WKY rats, mean arterial pressure (MAP) was monitored telemetrically during oral treatment with a high (27.5 mg/kg.day, n=14), an intermediate (14 mg/kg.day, n=6) and low dose (7 mg/kg.day, n=6) of sunitinib or vehicle (n=8) after normal salt diet for 2 weeks. The low dose-model was also combined with a high salt diet (8% NaCl and saline water). Eight days after administration rats were sacrificed and blood and 24h urine samples collected for biochemical measurements.

With the high dose of sunitinib, MAP increased from 94.7±0.9 mmHg to 125.8±1.5 mmHg (Δ31.1±0.9 mmHg, p<0.001). The intermediate and low doses induced MAP rises of 24.3±2.7 mmHg (p<0.001) and 13.4±3.3 mmHg (p<0.001), respectively. The low dose of sunitinib with high salt, induced a MAP rise of 43.5±2.2 mmHg (p<0.001 compared to normal salt). With the high dose, circulating ET-1 increased from 0.6±0.1 pg/ml to 1.6±0.2 pg/ml (p<0.01) and serum cystatine-C from 4.5±0.1 mg/L to 6.6±0.3 mg/L (p<0.001). Comparable increases in circulating ET-1 were seen with the intermediate and low doses, whereas serum cystatine-C did increase with the intermediate dose (to 6.3±0.1 mg/L, p0.05). Serum cystatine-C levels with low and high salt were identical. With the high dose of sunitinib, proteinuria increased from 7.5±1.3 to 33.3±4.8 mg/day (p<0.05). The rise in proteinuria was attenuated with the intermediate (16.2±2.1 mg/day, p<0.01) and low dose (19.9±3.3 mg/day, p<0.01), but increased to 40.4±30.1 mg/day (p>0.05) with high salt.

Angiogenesis inhibition-induced hypertension and nephrotoxicity are dose-dependent with a lower threshold for the rise in BP than for renal toxicity. The BP rise observed with the low dose of sunitinib observed in normotensive rats is comparable to the sunitinib-induced BP rise observed in patients and clearly is salt-sensitive. Since cystatine-C levels during normal and high salt diet were comparable, the BP rise during high salt seems independent of renal dysfunction.
Abstract 542: Blockade of Angiotensin-(1-7)-mediated Vascular Remodeling by a CB2 Cannabinoid Receptor Antagonist
Authors: Tallant E. Ann , Howlett Allyn C , Gallagher Patricia E
Published: July 2021

Angiotensin-(1-7) [Ang-(1-7)] inhibits vascular smooth muscle cell (VSMC) growth and reduces cardiac hypertrophy and vascular remodeling in Ang II-infused hypertensive rats. We previously showed that Ang-(1-7) increased the endocannabinoid 2-AG in VSMCs and that inhibition of VSMC growth by Ang-(1-7) was prevented by blockade of CB2 cannabinoid receptors. VSMCs isolated from mice with ablated CB2 receptors were treated with platelet-derived growth factor to stimulate growth; neither Ang-(1-7) nor the CB2 receptor agonist HU308 reduced VSMC growth (98.7±9.9% and 105.4±13.0% of stimulated growth, respectively, n = 3-6), suggesting that growth inhibition by Ang-(1-7) is mediated through activation of the CB2 receptor. Ang II-infused Sprague-Dawley rats were treated for 28 days with Ang-(1-7), in the presence or absence of the CB2 receptor antagonist SR144528, to determine whether CB2 receptor blockade prevents the Ang-(1-7)-mediated reductions in Ang II-stimulated cardiac hypertrophy and vascular remodeling. Systolic blood pressure was similar in both groups, before or after treatment (115±4.6 mmHg with SR144528 and 115.7±10.2 mmHg with Ang-(1-7)/SR144528 before treatment and 186.7±6.2 mmHg and 182.0±2.1 mmHg after Ang II treatment, respectively, n=4). Ang II increased the mean cross-sectional area of cardiac myocytes which was reduced 55% by Ang-(1-7); CB2 receptor blockade prevented the Ang-(1-7)-mediated reduction in myocyte size. Fibrosis (interstitial and perivascular) and vessel size (M/L) were unchanged by Ang-(1-7) in the presence of SR144528 compared to a 79%, 85% and 90% reduction of the Ang II-mediated increase in interstitial fibrosis, perivascular fibrosis and M/L, respectively, by Ang-(1-7) alone. Brain natriuretic peptide and atrial natriuretic peptide also were unchanged by Ang-(1-7) in the hearts of rats treated with Ang II and SR144528 compared to a 91% and 99% reduction by Ang-(1-7) in Ang II-treated rats. Thus, treatment with a CB2R antagonist blunts the anti-hypertrophic and anti-fibrotic responses to Ang-(1-7) in hypertensive Ang II-treated rats, independent of blood pressure, suggesting that the growth inhibitory properties of Ang-(1-7) involve a novel pathway which includes activation of the CB2 receptor.
Abstract 560: Inflammation Converts Glucose Into A Deleterious Agent In Human Aortic Smooth Muscle Cells
Authors: Sanchez-Ferrer Carlos F , Peiró Concepción , Romacho Tania , Azcutia Verónica , Villalobos Laura , Fernández Emilio , Bolaños Juan P , Moncada Salvador
Published: July 2021

Although hyperglycemia is an independent risk factor for vascular diseases, the links between glucose metabolism and atherosclerosis still require elucidation. We have previously shown that vascular cells, which regulates the glucose entry, are not damaged by high glucose concentrations unless they are primed with an inflammatory stimulus like interleukin (IL)1β. We now analyze the mechanisms accounting for the synergism between high glucose and IL1β. Under high glucose conditions (22 mmol/L), cultured human aortic smooth muscle cells (HASMC) exhibited excess glucose uptake and consumption (from 4.2±0.6 to 7.5±0.7 pmol per cell/24 h) associated to increased GLUT1 transporters expression only when co-stimulated with 10 ng/mL IL1β. However, the simple excess entry of glucose was not deleterious in these cells, as the inhibition of mitochondrial respiration with 0.5 mmol/L sodium azide increased glucose uptake and consumption (from 6.0±0.1 to 13.2±0.8 pmol per cell/24 h) without triggering inflammatory responses, measured by NF-κB activation and iNOS expression. We found that, besides allowing glucose entry, IL1β enhances glucose-6-phosphate dehydrogenase (G6PD) expression by 3.6±1.0 fold and activates the pentose phosphate pathway (PPP) from 9.6±0.7 to 17.4±1.5 nmol/h.mg prot in HASMC submitted to high glucose, thus permitting some of the excess glucose to be metabolized by this route. This provides additional substrate for enhancing the NADPH oxidase enzymatic activity by from 472±30 to 785±41 RLUS/μg prot/min, producing superoxide anions that are required for the activation of NF-κB and iNOS. The higher the concentration of glucose the more the PPP pathway is activated, giving rise to an increased inflammatory condition which cannot be counterbalanced by the simultaneous regeneration of reduced glutathione. We conclude that IL1β transforms excess glucose into a deleterious agent in HASMC by increasing glucose uptake, which is diverted into the PPP, promoting the pro-oxidant conditions required for the exaggeration of inflammatory pathways. Interestingly, all these pathways were blocked with the IL1 receptor antagonist anakinra (1 μmol/L), suggesting this anti-inflammatory drug can be effective for preventing diabetic vasculopathy.
Abstract 573: Uncovering the Molecular Mechanisms Underlying the Hypertension in
Snx1
Knockout Mice
Authors: Yang Jian , Asico Laureano D , Feranil Jun B , Jones John E , Armando Ines , Weinman Edward J , Jose Pedro A , Villar Van Anthony M
Published: July 2021

We have reported that sorting nexin 1 (SNX1) is crucial for renal dopamine D
5
receptor (D
5
R) trafficking, signal transduction, and function in human renal epithelial cells and in C57Bl/6J and BALB/cJ mice, as shown by the development of hypertension and impaired natriuretic response to agonist stimulation after an acute SNX1 depletion in the kidney. Thus, we elucidated the renal molecular mechanisms for these phenotypes in
Snx1
-/-
mice, which have congenital absence of SNX1. These mice have increased expression of glycosylated AT
1
R (123.8±2.1
vs
. 100±2.0% in wild-type littermates, P<0.05, Student’s
t
-test, n=5/group), a receptor with pro-oxidant and hypertensinogenic effects. We next determined the expression profiles of the components of the NADPH oxidase (NOX), an enzyme complex that is a major source of reactive oxygen species (ROS). We found an increased expression of renal NOX1 (153.4±12.2%
vs
. 100±4.1%, P<0.05), NOX2 or gp91
phox
(129.9±5.5%
vs
. 100±7.7%, P<0.05), and p47
phox
(118.2±2.7%
vs
. 100±5.0%, P<0.05), suggesting increased oxidative stress in these mice. Interestingly, the
Snx1
null mice have elevated renal D
5
R (142.9±4.7%
vs
. 100±6.8%, P<0.05) and D
3
R (134.3±5.3%
vs
. 100
±
1.6%, P<0.05), receptors with anti-oxidant activity, as well as the antioxidant paraoxonase 2, perhaps as compensatory mechanisms; the loss of SNX1 impairs the function of D
5
R. To corroborate our findings, we treated the
Snx1
-/-
mice and controls with a 10-day renal infusion of apocynin, a drug that blocks NOX assembly by preventing p47
phox
translocation to NOX2. Apocynin treatment resulted in the amelioration of systolic blood pressure (SBP) in
Snx1
-/-
mice (131.3±4.8 mm Hg to 105.7±1 mm Hg, P<0.05). There was no difference in the SBP with vehicle treatment in both strains, or with apocynin in control mice. Basal NOX activity was higher in
Snx1
-/-
mice (169±12.8 units/mg protein/min
vs
. 100±13.3 in controls, P<0.05), which was normalized by apocynin (99.4±16.5), while basal ROS levels were 2-fold higher in the
Snx1
-/-
mice (218.6±7.7 units/mg protein
vs
. 100±17.9, P<0.05), which was also normalized by apocynin (125.8±20.4). Our data indicate that the hypertension in
Snx1
-/-
mice is due to impaired D
5
R activity, higher NOX expression and activity, and increased AT
1
R.
Abstract 620: Development Of A New Mouse Model Exhibiting A Specific Deficiency Of Prorenin Renin Receptor In Adipocytes.
Authors: Yiannikouris Frederique B , Nguyen Genevieve
Published: July 2021

Objectives:
Recent studies demonstrated that the prorenin renin receptor (PRR) is present in adipose tissue. In adipose tissue stromal cells, PRR has the ability to bind renin and prorenin and contributes to the generation of angiotensinI (AngI). However, the contribution of adipocyte PRR to the generation of the vasoactive peptide, AngII and therefore to the regulation of blood pressure in physiological condition is unknown. The purpose of this study was to develop and characterize a new mouse model with adipocyte-specific PRR deficiency and define the role of adipose PRR in normal physiology.

Methods and results:
Female mice with 2 loxP sites flanking exon 2 of the PRR gene (floxed alleles, PRRfl/fl) were bred with aP2-Cre or with Adi-Cre male mice. Since PRR is located in the X chromosome, the male mice generated from the breeding were homozygotes for the deletion (PRRaP2 and PRRAdi). From the breeding, 5 PRRfl/fl, 2 PRRaP2 and 5 PRRAdi male mice were generated suggesting that the deletion of PRR in adipocyte was not lethal. Mice were fed on chow diet during 20 weeks. The body weight, the fat, lean mass and the blood pressure were quantified. Preliminary data suggest that the body weights (BW) were slightly decreased in PRRaP2 and PRRAdi compared to PRRfl/fl (PRRfl/fl: 29±1g; PRRaP2: 25±5g; PRRAdi: 28±1g). The slight reduction in BW was attributed to a reduction in fat mass (PRRfl/fl: 4.8±0.9g; PRRaP2: 3.8±1.8g; PRRAdi: 1.9±0.4g). Blood pressure was measured by plethysmography and by radiotelemetry. Preliminary data demonstrated that under physiological conditions, the SBP was not changed in PRRaP2 male mice compared to PRRfl/fl mice (plethysmography: PRRfl/fl: 108±1 mmHg; PRRaP2: 99±7 mmHg; radiotelemetry: PRRfl/fl: 129±2 mmHg; PRRaP2: 128±6 mmHg). The SBP of PRRAdi is currently under investigation.

Conclusions:
These results demonstrate the viability of mice with specific adipocyte deficiency of PRR. Future studies will define the effects of adipocyte PRR deficiency on obesity-induced hypertension.