bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 625: Angiotensin-(1-7) Protects From Central Nervous System Damage Induced By Shiga Toxin 2-producing Enterohemorrhagic Escheria Coli
Authors: Goldstein Jorge , Hocht Christian , Taira Carlos , Gironacci Mariela M
Published: July 2021

Several evidences showed a cerebroprotective action for angiotensin (Ang) (1-7) but neither of them demonstrated its cellular target for this protective effect. Our aim was to investigate the cellular type protected by Ang-(1-7) by transmission electron microscopy in the model of brain damage induced by Shiga toxin 2 (Stx2)-producing enterohemorrhagic Escherichia Coli. Adult male Wistar rats were injected with saline solution or Stx2 or Stx2 plus Ang-(1-7) or Stx2 plus Ang-(1-7) plus A779 into the anterior hypothalamic area (AHA). Rats received a single injection of Stx2 at the beginning while Ang-(1-7), A779 or saline was given daily as a single injection during 8 days. Ultrastructural changes were analyzed by transmission electron microscopy. Stx2 induced neurodegeneration, axon demyelination, alterations in synapse and oligodendrocyte and astrocyte damage, accompanied with edema. Ang-(1-7) partially prevented neuronal damage: 55.6±9.5 % of the neurons were protected from the damage triggered by the toxin. In addition, Ang-(1-7) hampered the Stx2-induced demyelination in 92±4% of the axons. Oligodendrocyte damage caused by Stx2 was prevented by Ang-(1-7) but atrocytes were partially protected by the peptide (38±5 % of astrocytes were preserved). The Stx2-induced synapse dysfunction was reverted by Ang-(1-7). The number of activated microglial cells induced by Stx2 was reduced by 50% by Ang-(1-7) treatment (P<0.05 by Student′s t test). All these beneficial effects elicited by Ang-(1-7) were blocked by the Mas receptor antagonist. We conclude that Ang-(1-7) protects mainly neurons and oligodendrocytes and partially astrocytes in the central nervous system through Mas receptor stimulation.
Abstract 634: Apelin as a Novel Therapeutic Target in Preeclampsia
Authors: Yamaleyeva Liliya M , Chappell Mark C , Brosnihan K. Bridget , Anton Lauren , Pirro Nancy , McGee Carolynne , Merrill David C , Mertz Heather L
Published: July 2021

Preeclampsia is a major cause for perinatal fetal and maternal morbidities and mortalities. The endogenous apelin system is an emerging target for the regulation of cardiovascular homeostasis; however, its role in pregnancy is not well understood. Apelin and its receptor are expressed in placental trophoblasts and fetal endothelial cells. We have reported that total immunoreactive apelin was lower in the placenta of preeclamptic women (PRE) compared to normotensive women (NT). The processing of the apelin precursor is complex and may yield a number of bioactive forms of the peptide. Presently, it is not known whether differences exist in the expression of the various molecular forms of apelin in the human placenta and whether the distribution pattern of apelin is altered in PRE. To address this issue, placental samples were pooled from NT (n=4) or PRE (n=4) at 37-38 weeks of gestation and the apelin forms resolved by HPLC coupled to an apelin radioimmunoassay (RIA). Total apelin content was lower in the placenta of PRE compared to NT (4.2 vs. 6.1 pg/mg wet weight). The HPLC analysis revealed negligible levels of apelin-36 from the placentas of NT (0.09 pg/mg wet weight) and PRE (0.11 pg/mg wet weight). A peak of apelin-17 was detected in both NT and PRE samples, but the shorter forms of apelin were more abundant. Therefore, separation conditions were tailored to achieve greater resolution of the lower molecular weight forms of apelin. This analysis revealed that the predominant apelin in both PRE and NT placentas was pyroglutamylated apelin-13 (Pyr-apelin-13; NT: 7.1 and PRE: 6.6 pg/mg wet weight); however, apelin-12 and apelin-13 were lower in PRE vs. NT placentas (1.8 and 22 fold, respectively). Lower expression of apelin-13 may reflect an inability to effectively influence blood pressure in PRE. Indeed, we further show that systemic infusion of apelin-13 (2 mg/kg/day) from day 13 to 20 of gestation reduced mean blood pressure (144±1 vs. 163±5 mmHg, n=4-6, p<0.01) in preeclamptic transgenic rats. In conclusion, we demonstrate that Pyr-apelin-13 is the predominant endogenous isoform of apelin in both NT and PRE placentas, while apelin-13 and apelin-12 are downregulated in PRE placentas. As a novel therapeutic agent, apelin may prevent the development of preeclampsia.
Abstract 639: Angiotensin 1-7 Contributes To Cardioprotective Effects Of Cardiac Overexpression Of Angiotensin II Type 2 Receptor In Mice Post-MI
Authors: Xu Jiang , Carretero Oscar A , Zhu Liping , Harding Pamela , Rhaleb Nour-Eddine , Shesely Edward G , Peterson Edward , Yang Xiao-Ping
Published: July 2021

Angiotensin II (Ang II) acting on AT1 receptor plays a pivotal role in the pathophysiology of cardiovascular disease, whereas AT2 has been considered cardioprotective, although the mechanisms are not fully understood. Recently studies suggest AT2 interacts with ACE2, an enzyme known to release Ang 1-7 from Ang II. Thus we hypothesize that Ang 1-7 contributes to cardioprotective effects of AT2, possibly via AT2/ACE2/Ang 1-7 cascade. Transgenic mice with AT2 specifically overexpressed in the heart (Tg-AT2) and their wild-type littermates (WT) were subjected to myocardial infarction (MI) or sham MI and divided into 1) sham MI; 2) MI + vehicle; and 3) MI + Mas receptor antagonist ([D-Ala7-Ang 1-7], A779, 0.5 mg/kg/day via osmotic mini pump). Treatments were started on the same day of MI and continued for 8 weeks. Our data show that AT2 and ACE2 protein expression in the heart was significantly increased in Tg-AT2 mice, whereas AT1 protein remained unchanged. Systolic blood pressure (SBP) and cardiac phenotypes did not differ between strains under basal conditions. MI increased myocyte cross-sectional area (MCSA), interstitial collagen fraction (ICF), left ventricular diastolic dimension (LVDd) and capillary density, and decreased LV ejection fraction (EF) in both strains; however, these pathological responses were diminished in Tg-AT2. Blockade of Mas receptor with A779 attenuated the cardioprotective effects observed in Tg-AT2 mice (Table). Infarct size (IS) did not differ among groups. Our findings suggest that overexpression/activation of AT2 protects against cardiac remodeling and dysfunction post MI, which is mediated in part through Ang 1-7 acting on the Mas receptor.
Abstract 641: Renal Denervation does not Lower Blood Pressure in Spontaneously Hypertensive Rats Treated with a Beta-1 Adrenergic Receptor Antagonist.
Authors: Phelps Jeremiah , Fink Gregory
Published: July 2021

We have used the SHR as an animal model to understand how renal denervation (RDN) lowers blood pressure. In order to determine how concomitant antihypertensive drug therapy could affect the blood pressure response to RDN, we performed RDN both before and after chronic (several days) treatment via the drinking water with the selective alpha-1 adrenergic antagonist prazosin and the selective beta-1 adrenergic antagonist atenolol. In the presence of prazosin treatment, RDN (n=7) significantly (p<0.05) lowered mean arterial pressure (MAP) compared to sham operated (SO; n=6) SHR (Δ=–4.0±1.7mmHg in RDN-SHR vs +1.5±1.3 in SO-SHR). Subsequent addition of atenolol during prazosin treatment reduced MAP significantly in both groups, but MAP in SO-SHR fell to the same level as in RDN-SHR, i.e. the antihypertensive effect of RDN was lost. After discontinuation of all drug treatment, the change in MAP from pre-treatment baseline was –7.7±3.5mmHg in RDN-SHR and +3.0±4.0mmHg in SO-SHR, p=0.06). To test whether the BP response to RDN would be lost during blockade of beta-1 adrenergic receptors alone, we performed RDN in SHR (n=5) pre-treated with atenolol. Atenolol alone significantly reduced MAP (Δ=-20.7±2.5mmHg). Steady-state MAP was not further reduced by RDN (Δ=+2.1±0.9mmHg). When atenolol was withdrawn MAP rose but remained reduced from baseline (Δ= -9.5±0.9mmHg). This magnitude of BP reduction is comparable to what we have previously observed after RDN in untreated SHR. In summary, in SHR RDN does not lower BP in the presence of an effective antihypertensive dose of a beta-1 antagonist. This indicates that RDN is reducing BP in SHR by interrupting a mechanism also affected by beta-1 adrenergic receptor signaling. Although further investigation is necessary, the data suggest that possibility that patients on clinically effective antihypertensive doses of beta-blockers may not exhibit a good response to RDN.
Abstract 646: The Existence of a Strong Correlation Between Leptin, TNF
α
Levels and BMI in Morbidly Obese Appalachian Females
Authors: Thompson Ellen , O'Hanlon Kathleen , Getty Morghan , Sodhi Komal , Maher Eamonn , Khitan Zeid , Abraham Nader G , Shapiro Joseph
Published: July 2021

Introduction:
Although BMI contributes to increased levels of adipokines, such as leptin, and tumor necrosis factor (TNFα) and to a decrease in adiponectin, few studies have examined the association between leptin, TNFα and body mass index (BMI) in Appalachian females.

Methods:
A total of 74 subjects from the Appalachian region were examined for the effect of BMI on the levels of leptin, TNF
α
, and adiponectin. Patients without overt cardiovascular disease were included. Linear regression analysis was used to analyze for changes in the relationship between BMI, leptin, TNFα and adiponectin. Nonlinear regression was used to determine the odds ratio and confidence intervals.

Results:
Serum TNFα and leptin levels were significantly increased in subjects with an increased BMI of 45-72 (9.9pg/ml ± 0.56 and 7.7pg/ml ± 0.6 respectively; p<0.05), compared to subjects with a BMI of 20-32. This was paralleled by increased levels of leptin, TNFα and IL-6. IL-6 was elevated in subjects with a BMI of 45-72 (4.83pg/ml ± 0.40; p<0.02). IL-6 was increased to a greater extent than leptin. In contrast, adiponectin levels were decreased (p<0.05) in subjects with a BMI of > 45-72 (6.63ng/ml ± 0.5; p<0.05), compared to subjects with a BMI of 20-32 (14.43ng/ml ± 0.32).

Conclusion:
Our study demonstrates for the first time the unique inflammatory biomarkers profile in morbidly obese females, even before diagnosis of coronary disease, highlighting the possible pathogenic potentials and opens the door for potential use of these biomarkers as a prognostic tool and a therapeutic target that can ameliorate obesity induced cardiovascular dysfunction.
Abstract 657: Increased Urinary Angiotensin Converting Enzyme 2 (ACE2) In Type 2 Diabetic Patients
Authors: Gutta Sridevi , Grobe Nadja , Osman Hassan , Saklayen Mohammad , Elased Khalid M
Published: July 2021

Diabetes and its associated chronic kidney disease (CKD) is a major health burden and there is an urgent need for new sensitive biomarkers to detect and monitor the progression of CKD. Albuminuria is still the gold standard for the evaluation of kidney function. However, its sensitivity and reliability have recently been questioned. ACE2 is highly expressed in renal tubules and has been shown to be shed in the urine of diabetic patients with CKD. The aim of the study was to investigate whether urinary ACE2 is increased in diabetic patients with CKD before the onset of microalbuminuria. Participants were recruited from Dayton VA Medical Center (Dayton, OH, USA). Baseline urinary albumin creatinine ratio (UACR) and estimated glomerular filtration rate (eGFR) were determined three months before initiation of the study in non-diabetic patients (UACR <30 mg/g, eGFR=97.40±16 ml/min/1.73 m
2
), and in diabetic patients with normoalbuminuria (UACR <30 mg/g, eGFR=83.08±17 ml/min/1.73 m
2
), microalbuminuria (UACR = 30-300 mg/g, eGFR=47.13±23 ml/min/1.73 m
2
), and macroalbuminuria (UACR >300 mg/g, eGFR=39.68±20 ml/min/1.73 m
2
). Using fluorogenic and mass spectrometry-based enzyme assays, we measured urinary and plasma ACE2 activity in patients. Urinary ACE2 activity was significantly increased in diabetic patients with normoalbuminuria (0.58±0.2 nmol/hr/mg creatinine), microalbuminuria (1.19 ±0.5 nmol/hr/mg creatinine), and macroalbuminuria (2.265±0.4 nmol/hr/mg creatinine) compared with non-diabetic controls (0.06 ± 0.02 nmols/hr/mg creatinine) (p<0.0001). These results were confirmed by detecting the ACE2 product Ang-(1-7) (
m/z
899) in incubations of urine samples with the natural substrate Ang II (
m/z
1046) using mass spectrometry-based enzyme assays. In addition, urinary ACE2 expression was significantly increased in diabetic patients as determined by western blot analysis (p<0.05). Plasma ACE2 activity was not detectable in control and diabetic patients. In conclusion, urinary ACE2 is increased in diabetic patients with CKD which suggests that urinary ACE2 could be used as an early, noninvasive biomarker for diabetic nephropathy before the onset of microalbuminuria.
Abstract 662: Association Analysis of Raptor Gene Variants with Overweight and Hypertension in American Men of Japanese Ancestry
Authors: Morris Brian J , Carnes Bruce A , Chen Randi , Donlon Timothy A , He Qimei , Grove John S , Masaki Kamal H , Elliott Ayako , Willcox D. C , Allsopp Richard , Willcox Bradley J , Willcox Bradley J
Published: July 2021

The mechanistic target of rapamycin (mTOR) pathway is pivotal for cell growth and has been implicated in aging, cardiovascular disease, obesity, diabetes and cancer. mTOR signaling is involved in cardiac leptin-mediated cardiac hypertrophy and fibrosis associated with obesity. mTOR is a key component of two multiprotein complexes, mTOR complex 1 and mTOR complex 2. The former is pro-growth and contains a unique protein, raptor. The present study tested for the first time whether genetic variation across the raptor gene (
RPTOR
) is associated with overweight/obesity, essential hypertension (EHT) and isolated systolic hypertension (ISH). We genotyped 61 common (allele frequency ≥ 0.1) tagging single nucleotide polymorphisms (SNPs) that captured most of the genetic variation across
RPTOR
in 374 subjects of normal lifespan and 439 subjects with a lifespan exceeding 95 years. Subjects were drawn from the Honolulu Heart Program, a homogeneous population of American men of Japanese ancestry, well characterized for phenotypes relevant to conditions of aging. Hypertension status was ascertained when subjects were 45–68 years old. Statistical evaluation was performed by contingency table analysis, logistic regression and recursive partitioning (RP), which is regarded as amongst the most powerful methods for statistical analysis of large complex sets of genetic information. After analysis of
RPTOR
genotypes by each statistical approach we found no significant association between genetic variation in
RPTOR
and either EHT or ISH. For EHT, RP revealed that even the most predictive SNPs (
rs4969322
and
rs4890052
) provided little contribution to correctly assigning individuals to EHT or NT (
P
=0.22 by Z test). In the case of ISH, RP revealed that only one SNP (
rs2589118
) made a noticeable contribution, and that this was no better than the contribution from the weakest laboratory/examination variable (overweight/obesity). In contrast, for overweight/obesity, the RP model revealed that
RPTOR
SNPs significantly enhanced the predictive capacity of the model (
P
=0.008 by one-tailed Z test). In conclusion, genetic variation across
RPTOR
is associated with overweight/obesity, but not EHT or ISH in the populations of normal lifespan and of long-lived subjects studied.
Abstract 041: Elevated Circulating Copeptin in Mid-Gestation is Associated with Increased Aortic Stiffness and Vascular Endothelial Dysfunction in Pregnant Women at High Risk for Preeclampsia
Authors: Pierce Gary L , Santillan Donna A , Fleener Diedre , Scroggins Sabrina M , Leslie Kimberlly K , Hunter Stephen K , Grobe Justin L , Sigmund Curt D , Haynes William G , Santillan Mark K
Published: July 2021

Circulating copeptin, a stable biomarker of vasopressin (AVP) secretion, is elevated throughout pregnancy in women who develop preeclampsia (PreE) and is a strong predictor of PreE as early as the 6th week gestation. Reduced vascular endothelial function and increased aortic stiffness occur in mid-gestation before clinical signs/symptoms of PreE manifest, suggesting that maternal vascular dysfunction may be an early event in the pathogenesis of PreE. However, it is unknown whether elevated copeptin/AVP in early/mid gestation contributes to vascular dysfunction in pregnant women who subsequently develop PreE. Therefore, we hypothesized that elevated copeptin would be associated with increased aortic stiffness and reduced vascular endothelial function in early/mid gestation of pregnant women at high risk for PreE. Pregnant women in the 1st trimester (n=72; age=30 ±1 yrs; BMI=34 ± 1 kg/m2) with at least 1 risk factor for PreE were enrolled. Aortic stiffness (carotid-femoral pulse wave velocity, CFPWV), vascular endothelial function (brachial artery flow-mediated dilation, FMD), blood pressure (BP) and plasma copeptin (ELISA) were assessed in both the 1st (11.7 ± 0.2 wks) and 2nd (18.8 ± 0.4 wks) trimesters. In the 1st trimester, CFPWV (7.3 ± 0.2 vs. 7.3 ± 0.5 m/sec, P=0.86), brachial artery FMD (12.9 ± 1.1 vs. 14.3 ± 2.0%, P=0.53), BP, BMI and age did not differ between women in the highest (1513 ± 221 pg/ml) vs. lowest (279 ± 12 pg/ml) quartile of copeptin (P<0.01). In contrast, 2nd trimester CFPWV was greater (7.2 ± 0.2 vs. 6.4 ± 0.2 m/sec, P<0.05) and brachial artery FMD was lower (10.2 ± 2.8 vs. 16.5 ± 1.3 %, P<0.05) among women in the highest (1714 ± 481 pg/ml) vs. the lowest (249 ± 13 pg/ml) quartile of copeptin (P<0.01), in the absence of differences in BP, BMI or age. For the entire cohort, (log)copeptin was significantly correlated with CFPWV (r=0.23, P=0.04) and tended to correlate with FMD (r=-0.23, P=0.06) in the 2nd but not in the 1st trimester. These data suggest that elevated copeptin in mid-gestation is associated with aortic stiffness and vascular endothelial dysfunction in pregnant women at high risk for PreE, but whether increased copeptin/AVP causes vascular dysfunction in pregnancies destined for PreE requires further studies using animal models.
Abstract 082: High Fat Diet Promotes Haplotype Dependent Differential Transcriptional Regulation of the Human Angiotensinogen Gene
Authors: Rana Anita , Jain Sudhir , Puri Nitin , Eren Deniz , Sirianni Natalie , Kumar Ashok
Published: July 2021

Angiotensinogen (AGT) is the only known precursor to angiotensin II. Systemic renin angiotensin aldosterone system (RAAS) is activated in human and experimental models of obesity. RAAS activation in obesity is linked to the development of cardiovascular pathophysiologies. We have identified polymorphisms in 2.5 Kb promoter of human angiotensinogen gene (hAGT) that forms two haplotype (Hap) blocks: -6A/G (-1670A/G, -1562C/T, -1561T/C) and -217A/G (-532T/C, -793A/G, -1074T/C,& -1178G/A). Hap -6A/-217A (Hap -6A) is associated with human hypertension whereas, Hap -6G/-217G (Hap -6G) reduces cardiovascular risk. Here, we examine high fat diet-mediated allele-specific transcriptional regulation of the hAGT gene in adipose tissue, in vivo, in transgenic (TG) mice engineered with either haplotype of the hAGT gene. Twelve-week-old male TG mice with Hap –6A or –6G were fed normal diet (10% kcal as fat) and high fat diet (60% kcal as fat) for 10 weeks. Using Q - RT PCR and western blot we show increased hAGT expression in adipose tissue of the Hap -6A-TG mice after high fat diet than control diet (Hap -6A-0.68±0.04 vs. Hap -6G- 0.33±0.03 A.U., p<0.05). ChIP assay shows greater chromatin binding of GR, MR, CEBPβ and STAT3 transcriptional factors (Hap -6A- 0.80±0.04 vs. Hap -6G- 0.26±0.06 A.U., p<0.05) to the hAGT transgenes in Hap -6A TG mice after high fat diet. No significant change was observed in the endogenous mouse AGT gene. In addition, after high fat diet, change ([[Unable to Display Character: ∆]]) in inflammatory and redox markers was significantly (p<0.05) greater in TG mice with Hap I including, IL1 (4.6±0.8 vs. 2.1±0.49 fold), IL6 (4.0±0.69 vs. 2.1±0.2 fold) and NOX1 (8.3±0.4 vs. 2.5±0.6 fold). This is accompanied by reduction in levels of antioxidant defenses (SOD1: 0.97±0.0 vs. 1.4±0.1 fold; HO1: 0.77±0.1 vs. 1.3±0.2 fold) & activation of MAPK14 and ERK1/2 signaling. Taken together, our results show that SNPs in the hAGT Hap -6A favor high fat diet induced binding of transcriptional factors GR, MR, CEBP-β and STAT3 that lead to elevated expression of the hAGT in expanded mass of the adipose tissue. This also activates the RAAS with pathophysiological implications including, phosphorylation of kinases such as MAPK14 and ERK2, increase in tissue pro-inflammatory and oxidative stress molecules.
Abstract 087: Human Stomach Cell Gastrin Inhibits Renal NHE3 and NaKATPase in Concert With the Renal D1R
Authors: Gildea John J , Xu Peng , Zhang Chi , Bigler Wang Dora , Tran Hahn T , Jose Pedro A , Felder Robin A
Published: July 2021

The digestive track secretes gastrin in response to sodium ingestion stimulating increased renal sodium excretion. We previously showed that gastrin secreted by human colon cancer cells (SW626) can bind to cell surface cholecystokinin receptors on renal proximal tubule cells (RPTC), modulating the natriuretic dopaminergic system. We tested the hypothesis that a similar gastro/renal axis exists in humans. Novel human stomach antrum G-cell lines from 6 separate individuals were isolated and each shown to express gastrin mRNA and protein, and bind Phaseolus vulgaris Leucoagglutinin (PHA-l) by fluorescence lectin affinity (marker for gastrin secreting cells). It was determined that the D1 receptor (D1R) is also found on G-Cells. In human stomach tissue, gastrin expression was increased by Fenofibrate treatment, (PPARα agonist, p= 0.07, in 3 live ex-vivo cultured human stomachs). Moreover, we tested the effect of gastrin on CCKB2 receptors in human RPTC. Gastrin (100 nM 15 min.) increased RPTC phospholipace-C (PLC) activity by 1.07± 0.01 fold, N=35, p<0.0001, using a PLC-FRET biosensor, CYPHR, but did not increase cAMP levels using the specific cAMP-FRET biosensor, ICUE-YR. Both gastrin and SKF83822 (100 nM, a cAMP specific D1R agonist, 15 min) alone reduced sodium influx into RPTC via NHE3 (from VEH 100± 4.7% to 73.5± 6.2% or 74.1± 4.3% respectively N=6, p<0.05), but gastrin along with SKF83822 decreased sodium influx more than either alone (57.2 ± 5.8% N=6, p<0.05). Sodium efflux via NaKATPase was reduced by SKF83822 (from VEH 100± 3.9% to 84.2± 3.3%, N=6, p<0.05), but not gastrin alone, however SKF83822 along with gastrin reduced sodium efflux more than SKF83822 alone (72.3±5.1% vs N=6, p<0.05). Additionally we found that Angiotensin II (AngII, 10 nM, 15 min.) increased NHE3 activity (12.3± 3.6% N=6, p<0.01) and this increase was completely blocked by gastrin (N=6, p<0.01). The PLC inhibitor U73122 reversed the inhibitory effect of gastrin on NHE3 and NaKATPase. Gastrin was also found to decrease the amount of fluorescent AngII binding to RPTCs (27.4± 6.1% N=6, p<0.01) and this decrease was completely blocked by U73122. Thus, both stomach gastrin and the renal D1R inhibit NHE3 and NaKATPase, to increase sodium elimination from the body after salt is ingested.
Abstract 090: Outcomes of Hypertensive Crises as Predicted by Red Cell Distribution Width
Authors: Raheja Suraj , Patel Kush , Patel Ruchir , Mawri Sagger , Michaels Alexander , Gibbs Joseph , McCord James
Published: July 2021

Background:
Red cell distribution width (RDW) is a measure of the variability in size of erythrocytes. A high RDW value indicates greater variation in size between individual erythrocytes and has been shown to be an independent predictor of mortality in patients with coronary artery disease, heart failure and in patients undergoing percutaneous coronary intervention (PCI). The aim of this study was to evaluate the prognostic value of RDW in predicting clinical outcomes in patients with hypertensive crisis.

Methods:
We performed a retrospective study of 465 consecutive patients from January 2007 to March 2010 who presented with hypertensive crisis. Hypertensive crisis was defined as systolic BP >180 and/or diastolic BP >110mmHg with impending or progressive end organ dysfunction requiring inpatient hospitalization. The study sample consisted of 465 patients (38.9% men (181 of 465); mean age 59.6 ± 15.9). Baseline levels of RDW were measured at time of admission and analyzed as continuous and categorical variables (elevated RDW was defined as >14.5%). Multivariable regression analysis was performed for development of all-cause mortality, myocardial infarction, new-onset heart failure (defined as first time hospital admission for heart failure), stroke and MACE (MI, new-onset heart failure and stroke) at 2 years.

Results:
RDW > 14.5% was a strong independent predictor of all-cause mortality at 2 years (OR: 1.90, 95% CI: 1.1-3.3,
p
<0.05). Elevated RDW was also found to be an independent predictor of new-onset heart failure at 2 years (OR: 1.97, 95% CI: 1.1-3.7,
p
<0.05). Elevated RDW was not a predictor of MI, PCI or stroke at 2 years.

Conclusions:
Elevated RDW level in patients with hypertensive crisis was an independent predictor of all-cause mortality and new-onset heart failure in patients with hypertensive crisis.
Abstract 098: MicroRNA Regulation of NADPH Oxidase Mediates the Antioxidant Effect of Renal Paraoxonase 2
Authors: Cuevas Santiago , Yang Yu , Asico Laureano D , Jones John , Armando Ines , Jose Pedro A
Published: July 2021

Increased renal generation of reactive oxygen species (ROS) is important in the pathogenesis of hypertension caused by absent or dysfunctional dopamine receptor subtype. Germline deletion of the dopamine 2 receptor in mice increases renal NADPH oxidase (NOX) activity and decreases expression paraoxonase 2 (PON2) and results in ROS-dependent hypertension. We determined if microRNA (miR) is involved in PON2-mediated regulation of NOX. Silencing PON2 in human renal proximal tubules cells decreased PON2 (-60±4%, n=3,*P<0.05) and increased NOX2 (110±15%, n=3,*P<0.05) and NOX4 (80±10%, n=3,*P<0.05) proteins, NOX activity (50±6%, n=3,*P<0.05), and ROS production (57±3%, n=4,*P<0.05). Inhibition of NOX activity by diphenyleneiodonium normalized the increase in ROS caused by PON2 silencing. Renal-selective silencing of Pon2 in mice by the renal subcapsular infusion of Pon2 siRNA decreased PON2 (~50%), and increased NOX2 (191±11%,n=3, P<0.05), NOX4 (60±4%,n=3, P<0.05), NOX activity (94±23%, n=3, P<0.05), and blood pressure (BP) (+41±6 mmHg, n=3, P<0.05). Pon2-/- mice also had higher BP than wild-type littermates (+15±2 mmHg,n=3/4,*P<0.05) but less than observed with renal-selective silencing indicating extrarenal compensation. Renal NOX2 (220±64%, n=3/4,*P<0.05) and NOX activity (195±77%, n=3,*P<0.05) were also increased in Pon2-/- mice. However, the renal expression of NOX4 was similar in Pon2-/- and wild-type littermates. The renal expressions of miR-23b, miR-34a and miR-155 (reported to regulate NOX expression) were also similar in Pon2-/- mice and wild-type littermates. However, renal miR-146a expression was decreased (-25±4%, n=3/4,*P<0.05) while miR-204 (150±12%, n=3/4,*P<0.05) and NFAT expressions (21±7%, n=3/4,*P<0.05) were increased in Pon2-/- mice. The increase in miR-204 could be a compensatory response because miR-204 has been shown to decrease NFAT expression. It is known that NFAT and NOX2 can positively regulate each other’s expression while miR-146a negatively regulates NOX4 expression and inflammation. We conclude that PON2 by increasing miR-146a and decreasing NFAT expression negatively regulates NOX activity and reduce ROS production that would contribute to the maintenance of normal BP.
Abstract 105: High Salt Activates Immune Cells to Promote Hypertension
Authors: Kirabo Annet , Barbaro Natalia R , Foss Jason D , Norlander Allison E , Dikalov Sergey , Madhur Meena S , Harrison David G
Published: July 2021

High salt intake and inflammation are implicated in the genesis of hypertension. We have previously shown that hypertension activates dendritic cells (DCs), by promoting the formation of immunogenic isoketal-protein adducts. Recently it has become clear that sodium can accumulate in the interstitial space and promote inflammation. We hypothesized that high salt activates antigen presenting cells including monocytes and DCs to produce immunogenic isoketals. Exposure of mouse splenic DCs to high salt (190 mM) for 24 hours led to an increase in superoxide production compared to regular RPMI media (146.3±9.5 vs. 100.0±5.0 % control, p<0.001). This was NADPH oxidase dependent because incubation with the gp91ds-tat peptide prevented this effect. High salt exposure also led to an increase in the activation markers CD80, CD86, and a 48% increase in DCs containing isoketal-protein adducts. Moreover, DCs exposed to high salt drove T cell proliferation (5198.2±2398.6 vs. 15.3±7.1 proliferated CD4+ cells and 25381.6±9495.6 vs. 9.8±4.1 proliferated CD8+ cells, p<0.05). This was not due to increased osmolality, as mannitol did not mimic these effects. Western blots of protein extracts from DCs indicated that all NADPH subunits (p47phox, p22phox, gp91phox and p67phox) were increased by exposure of cells to high salt, and that these effects were prevented by inhibition of the salt-sensing glucokinase (SGK1). In additional studies, we found that human monocytes, which are precursors of myeloid DCs, from hypertensive subjects possess higher CD86 and isoketal-protein adducts compared to normotensives, and that these are further increased by exposure to 190 mM salt for 48 hours. Taken together, our data indicate that antigen presenting cells are activated by exposure to the high sodium environment that can occur in pro-hypertensive states, and that this likely involves increased expression and activation of the NADPH oxidase. Our data also describe a previously undefined role of the SGK1 in this process.
Abstract 107: Cytochrome B5 Reductase 3 Sensitizes Soluble Guanylate Cyclase to Nitric Oxide in Vascular Smooth Muscle
Authors: Straub Adam C , Nguyen Anh T , Rahaman Mizanur , Mutchler Stephanie M , Miller Megan , Watkins Courtney , Gladwin Mark , Gordeuk Victor , Prchal Josef , Martin Emil
Published: July 2021

The inability nitric oxide (NO) to stimulate soluble guanylate cyclase (sGC) has been linked to numerous cardiovascular diseases (CVD) including hypertension. While several studies have defined the importance of sGC expression in the cardiovascular system, the basic mechanisms that regulate sGC activity remain incompletely understood. Here, we report for the first time that sGC heme iron redox state, which is essential for NO-induced sGC activation, is regulated by cytochrome B5 reductase 3 (CyB5R3). Genetic knockdown and pharmacological inhibition of CyB5R3 in primary rat vascular smooth muscle cells resulted in a 60% loss in cGMP production. Conversely, the sGC activator Bay 58-2667, which activates oxidized or heme free sGC, reversed these effects. Consistent with our cell culture work, purified protein studies demonstrate that CyB5R3 can directly reduce oxidized sGC heme iron and sensitize sGC to NO. To test the functional importance of Cyb5R3 activity, we cultured mouse thoracodorsal arteries with a pharmacological inhibitor of Cyb5R3 (ZINC 747) and performed vascular reactivity studies using pressure myography. Arteries treated with ZINC 747 showed decreased responsiveness the NO donor DETA-NONOate but increase sensitivity to Bay 58-2667. We then treated mice with 10mg/kg/day of ZINC 747 using osmotic mini pumps, which caused an increase in mean arterial blood pressure (107.5±3.4 vs 131±13.16) measured via radio telemetry. Lastly, translational studies reveal that the CyB5R3 T116S polymorphism with allele frequency 0.23 only in African Americans is unable to reduce sGC and correlates with increased blood pressure. Considering the defining role of sGC in NO signaling and the fact that the oxidation state of sGC may predict responses to NO therapies and new classes of sGC activator medications, we anticipate that these studies may significantly impact our understanding of biology, precision therapeutics (right drug for the right patient) and pharmacogenetics (T117S SNP based drug selection).
Abstract 113: Renal Phenotype of Inwardly Rectifying Potassium Channel Kcnj16 (Kir 5.1) Knockout in the Dahl Salt-Sensitive Rats
Authors: Palygin Oleg , Levchenko Vladislav , Ilatovskaya Daria V , Barnett Jessica L , Geurts Aron M , Jacob Howard J , Staruschenko Alexander
Published: July 2021

The inward-rectifying channels play an important role in the control of resting membrane potential and tubular homeostasis in the kidney. Kcnj16 (Kir 5.1) form a heteromeric channel with Kcnj10 (Kir 4.1) at the basolateral membranes of aldosterone-sensitive distal nephron (ASDN); mutations in the human KCNJ10 gene result in SeSAME)/EAST syndrome, a complex disorder that includes salt wasting and hypokalemic alkalosis. To illuminate the importance of Kcnj16 (Kir 5.1) in the context of a disease state in vivo, we generated a Kcnj16 knockout rat model in Dahl salt-sensitive (SS) background by using ZFN technology. ZFN against Kcnj16 caused a 18-bp in-frame deletion that occurred in the second protein transmembrane domain. IHC analysis demonstrated highly specific expression of Kcnj16 on the basolateral membranes of ASDN in the control kidneys of SS rats, which was completely abolished in Kcnj16-/- rats. The electrophysiological recording of K+ channels in the CCD basolateral membrane revealed activity of only homomeric Kcnj10 channels (21 pS channel in Kcnj16-/- rats compared to both 41 and 21 pS channels in SS rats). Thus, these data provide evidence of successful knock out of this protein and consequent degradation of the channel in renal tubules. The Kcnj16-/- knockout in SS rat induces electrolyte imbalance, epileptic seizures and result in changes in development (37% reduction in body and 54% in kidney mass). The mean arterial pressure was significantly lower in Kcnj16-/- compared to SS rats (91.3±1.8 to 104.7±5.5 mmHg) when animals were fed a low salt (0.4%) diet. Knockout of Kcnj16 resulted in hypokalemia (4.25±0.09 vs 2.08±0.12 mmol/L in serum of control vs KO rats), hypermagnesemia (0.49±0.02 vs 0.63±0.01 mmol/L in serum of control vs KO rats), and FSGS. Urea electrolyte balance was also disturbed compared to control animals. Importantly, change of the diet to high salt (4%) caused mortality of KO rats within 1-2 days. These data demonstrate critical role of Kcnj16 channels in renal salt handling and in the development of salt-sensitive hypertension.
Abstract 114: K+ Rich Diet During Angiotensin II Hypertension Reduces Renal Na-Cl Cotransporter and Phosphorylation, but Not Blood Pressure
Authors: Veiras Luciana C , Han Jiyang , Ralph Donna L , McDonough Alicia A
Published: July 2021

During Ang II hypertension distal tubule Na-Cl Cotransporter (NCC) abundance and its activating phosphorylation (NCCp), as well as Epithelial Na+ channels (ENaC) abundance and activating cleavage are increased 1.5-3 fold. Fasting plasma [K+] is significantly lower in Ang II hypertension (3.3 ± 0.1 mM) versus controls (4.0 ± 0.1 mM), likely secondary to ENaC stimulation driving K+ secretion. The aim of this study was to test the hypothesis that doubling dietary K+ intake during Ang II infusion will lower NCC and NCCp abundance to increase Na+ delivery to ENaC to drive K+ excretion and reduce blood pressure.

Methods:
Male Sprague Dawley rats (225-250 g; n= 7-9/group) were treated over 2 weeks: 1) Control 1% K diet fed (C1K); 2) Ang II infused (400 ng/kg/min) 1% K diet fed (A1K); or 3) Ang II infused 2% K diet fed (A2K). Blood pressure (BP) was determined by tail cuff, electrolytes by flame photometry and transporters’ abundance by immunoblot of cortical homogenates.

Results:
As previously reported, Ang II infusion increased systolic BP (from 132 ± 5 to 197 ± 4 mmHg), urine volume (UV, 2.4 fold), urine Na+ (UNaV, 1.3 fold), heart /body weight ratio (1.23 fold) and clearance of endogenous Li+ (CLi, measures fluid volume leaving the proximal tubule, from 0.26 ± 0.02 to 0.51 ± 0.01 ml/min/kg) all evidence for pressure natriuresis. A2K rats exhibited normal plasma [K+] (4.6 ± 0.1 mM, unfasted), doubled urine K+ (UKV, from 0.20 to 0.44 mmol/hr), and increased CLi (to 0.8 ± 0.1 ml/min/kg) but UV, UNaV, cardiac hypertrophy and BP were unchanged versus the A1K group. As expected, NCC, NCCpS71 and NCCpT53 abundance increased in the A1K group to 1.5 ± 0.1, 2.9 ± 0.5 and 2.8 ± 0.4 fold versus C1K, respectively. As predicted by our hypothesis, when dietary K+ was doubled (A2K), Ang II infusion did not activate NCC, NCCpS71 nor NCCpT53 (0.91 ± 0.04, 1.3 ± 0.1 and 1.6 ± 0.2 fold versus C1K, respectively). ENaC subunit abundance and cleavage increased 1.5 to 3 fold in both A1K and A2K groups; ROMK was unaffected by Ang II or dietary K.

In conclusion, evidence is presented that stimulation of NCC during Ang II hypertension is secondary to K+ deficiency driven by ENaC stimulation since doubling dietary K+ prevents the activation. The results also indicate that elevation in BP is independent of NCC activation
Abstract MP02: Angiotensin II Mediates Microvascular Rarefaction
In Vivo
and Exacerbates Endothelial-To-Mesenchymal Transition
In Vitro
Authors: Nather Katrin , Flores-Muñoz Mónica , Touyz Rhian M , Loughrey Christopher M , Nicklin Stuart A
Published: July 2021

Cardiac fibrosis accompanies numerous cardiovascular diseases (CVD) such as hypertension and myocardial infarction and increases myocardial stiffness leading to contractile dysfunction. Recently, endothelial-to-mesenchymal transition (EndMT) has been shown to contribute to myocardial fibrosis. EndMT describes a process by which endothelial cells transform into mesenchymal cells such as fibroblasts and has been implicated in many fibrotic diseases. Angiotensin II (AngII) plays a key role in myocardial fibrosis and has been associated with the activation of fibroblasts to myofibroblasts and an increase in myocardial collagen deposition. Here, we assessed the role of AngII in capillary loss and EndMT
in vivo
and
in vitro
.

C57BL/6J mice were infused with H
2
O (control) or 24μg/kg/hr AngII for 4 weeks. Mice infused with AngII developed significant cardiac fibrosis characterised by the deposition of collagen I (2.5-fold
vs.
control; p<0.05) and III (1.9-fold
vs.
control; p<0.05). Capillary density was assessed by CD31 immunohistochemistry and revealed significant vascular rarefaction (control 2161±111
vs
. AngII 838±132 capillaries/mm
2
; p<0.05). To investigate whether AngII can induce EndMT
in vitro
, human coronary artery endothelial cells were stimulated with 10ng/mL TGFβ
1
alone or in combination with 1μM AngII for 10 days. AngII significantly enhanced TGFβ
1
-induced gene expression of α-smooth muscle actin (TGFβ
1
1.8-fold; TGFβ
1
±AngII 4.3-fold
vs
. control; p<0.05) and collagen I (TGFβ
1
9.2-fold; TGFβ
1
+AngII 30.2-fold
vs
. control; p<0.05). Concomitantly, AngII significantly increased α-smooth muscle actin protein expression (TGFβ
1
3.9-fold; TGFβ
1
+AngII 23.6-fold
vs
. control; p<0.05) and significantly decreased CD31 expression (TGFβ
1
0.9-fold; TGFβ
1
+AngII 0.7-fold
vs
. control; p<0.05), suggesting AngII acts in concert with TGFβ
1
to enhance conversion of endothelial cells to myofibroblasts. Further studies investigating the underlying mechanism, including the role of the Smad pathway, are ongoing.

These results demonstrate that AngII induces vascular rarefaction
in vivo
and potentiates TGFβ
1
-induced EndMT
in vitro.
Understanding the molecular basis for these observations may help to identify new therapeutic options in CVD.
Abstract P014: Poor Blood Pressure Control and Severe Coronary Artery Lesions Are Risks of Cerebral Microbleeds in Patients With Coronary Artery Disease During Antiplatelet Therapy
Authors: Iwamoto Yoshiko
Published: July 2021

Background and Purpose:
Brain hemorrhage is a serious complication of antiplatelet therapy, particularly dual antiplatelet therapy (DAPT), in patients with coronary artery disease (CAD) who undergo percutaneous coronary intervention (PCI). Cerebral microbleeds (CMBs) detected on MRI is a surrogate of symptomatic brain hemorrhage. However, little is known regarding CMBs in CAD patients during long-term antiplatelet therapy. Therefore, we investigated the temporal change of CMBs during antiplatelet therapy in CAD patients after PCI.

Method:
Brain MRI was performed at baseline and after 8-month follow-up in consecutive 14 patients who underwent PCI and antiplatelet therapy (DAPT in 13 patients).

Results:
Baseline MRI revealed CMBs in 2 patients (14%). New CMBs were detected by follow-up MRI in 2 other patients (14%). Although blood pressure (BP) at baseline did not differ between the CMB-positive and CMB-negative patients, BP after 8 months was significantly higher in CMB-positive patients than CMB-negative ones (systolic BP: p=0.03, diastolic BP: p=0.02). Moreover, CMB-positive patients had greater number of coronary artery lesions and higher SYNTAX score at baseline than CMB-negative patients (Figure).

Conclusions:
CAD patients with poor BP control and severe coronary artery lesions would be at higher risk for CMBs and eventually brain hemorrhage during antiplatelet therapy. Accordingly, strict coronary risk control, especially BP control, is necessary in CAD patients receiving long-term antiplatelet therapy.
Abstract P018: Periodic Water Intake Exacerbates Hypertension, Decreases Renal Function and Increases Renal Inflammation in Spontaneously Hypertensive Rats
Authors: Hilliard Lucinda M , Bulmer Louise , Mirabito Katrina M , Vinh Antony , Denton Kate M
Published: July 2021

Water is essential for life. The kidneys are major regulators of fluid balance but also appear susceptible to adverse effects of periodic water intake since epidemiological studies have linked low water intake to chronic kidney disease. However, the impact of periodic water intake on renal health has largely been ignored. This study investigated the chronic effects of periodic drinking on blood pressure, kidney function, and immune cell infiltration in rats with pre-existing hypertension.

Mean arterial pressure (MAP) was measured continuously in spontaneously hypertensive rats (SHR) that were only given access to water for 2 hours per day versus control rats given free access to water over a 4-week period (n=>8 per group). Glomerular filtration rate (GFR) was measured via transcutaneous FITC-sinistrin clearance at baseline and at the end of the 4-week protocol. Renal immune cell infiltration and cytokine production was assessed by flow cytometry.

Basal MAP and renal function were similar between the control and water-restricted (WR) groups. Water restriction led to a significant increase in MAP from baseline (8.8±4.5mmHg versus 0.6±5.5mmHg in WR and control SHR, respectively; P<0.05). Moreover, GFR decreased from baseline by 29±6% in the WR SHR (P<0.001). In comparison, GFR did not change significantly over time in the control SHR (-5.3±2.6% from baseline; P>0.05). The increase in MAP and reduction in renal function in the WR-SHR was associated with periodic increases in urine osmolality (69±26% versus baseline, P<0.05), indicative of cycles of dehydration and replenishment. Renal immune cell infiltration was similar between the groups. However, cytokine analyses revealed a phenotypic shift towards a pro-inflammatory Th1 phenotype in kidney T cell infiltrate from WR versus control SHR. This was indicated by a greater proportion of T cells producing IFN-γ in SHR subjected to water restriction than control SHR (9.5±3.3% versus 2.9±0.9%, respectively; P<0.05).

In conclusion, recurrent dehydration associated with periodic drinking exacerbates hypertension, renal dysfunction and inflammation in male SHR. This highlights the importance of regular water intake for the maintenance of kidney health, particularly in populations with existing disease.
Abstract P031: Cyclooxygenase Inhibition Does Not Augment Reno-protective Effect of Soluble Epoxide Hydrolase Inhibitor in Streptozotocin-induced Diabetic Rats
Authors: Katary Mohamed A , Pye Chelsey , Elmarakby Ahmed A
Published: July 2021

Cyclooxygenase (COX)-derived prostaglandins and cytochrome P450 epoxygenase/soluble epoxide hydrolase (sEH) derived epoxyeicosatrienoic acids (EETs) play important role in the regulation of vascular tone. The aim of this work is to examine whether COX inhibition exacerbates the reno-protective effects of increased EETs levels, via inhibiting EETs degradation by sEH using sEH inhibitor, in streptozotocin-induced type 1 diabetes. Diabetes was induced in 12 week old Sprague Dawley rats using streptozotocin (50 mg/kg, i.v). Rats were divided into 5 groups (n=6); control non diabetic, diabetic, diabetic treated with the sEH inhibitor trans-4-[4-(3-adamantan-1-yl-ureido)-cyclohexyloxy]-benzoic acid (t-AUCB, 10 mg/L in drinking water), diabetic treated with COX inhibitor meloxicam (5 mg/kg/day in drinking water) and diabetic treated with both inhibitors for two months. Inhibition of sEH reduced the elevation in proteinuria and creatinine clearance in diabetic rats (P less than 0.05) and meloxicam did not exacerbate t-AUCB effects. Glomerular permeability to albumin and albuminuria were elevated in diabetic rats and were reduced with t-AUCB treatment; however, meloxicam did not augment t-AUCB effects on reducing glomerular injury. Inhibition of sEH with t-AUCB also restored the decrease in glomerular integrin and nephrin expression levels in diabetic rats without synergestic effect from meloxicam. Renal collagen deposition and the inflammatory marker monocyte chemoattractant protein-1 (MCP-1) excretion levels were significantly elevated in diabetic rats and were reduced with t-AUCB treatment without further significant protective effects from meloxicam treatment. The number of TUNEL positive cells, a marker of apoptosis, was higher in the kidney sections from diabetic rats versus control (P less than 0.05), an effect that was significantly mitigated by t-AUCB treatment but not with meloxicam treatment. In conclusion, increasing EETs levels in type 1 diabetic rats, using sEH inhibition, reduced renal apoptosis, inflammation and injury whereas inhibition of COX failed to provide renal protection or to augment reno-protective effect of sEH inhibition in diabetic rats.