bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract P025: Clinical Experience on Combination Therapy With Olmesartan and Amlodipine in Treatment of Resistant Hypertension
Authors: James Cyril , Tisheva Snezhana , Gaidarova Diana , Stancheva Nadia , Yakova Dilyana , Hristov Martin , Ohri Shreya , Gospodinov Konstantin , Yanakeva Asiq , Chakalova Tatqna , Jose Soumya
Published: March 2022

Objective:
To analyse the effect of fixed dose combination therapy with Olmesartan/Amlodipine in the treatment of Resistant Hypertension.

Design:
Questionnaire based cross sectional study. Method: The study was carried out among 364 patients admitted with history and 24 hour holter blood pressure monitor evidence for Resistant Hypertension in the Department of Cardiology between 1
st
January 2012 and 31
st
January 2015. Patients with history of Resistant Hypertension were screened with 24 hour holter blood pressure monitoring and those who fullfilled the criteria for Resistant Hypertension according to the ESC guideline for Management of Arterial Hypertension based on the treatment history and 24 hour holter blood pressure monitoring were included in the study after obtaining informed consent. Patients included in the study were started on therapy with Olmesartan/Amlodipine at fixed dose combination along with diuretic and Beta-Blocker. Patients were continued on this fixed dose combination therapy for 3 months. After the 3 month period patients were reassessed with control holter blood pressure monitoring to access the efficacy of the treatment and the circadian control of arterial blood pressure.

Result:
From the study it was observed that 68.68% patients (250 of 364) had reached optimal control of arterial blood pressure by the fixed dose combination therapy with Olmesartan/Amlodipine. 17.04% patients (62 of 364) had non optimal control of arterial blood pressure and 8.79% patients (32 of 364) continued to be with resistant hypertension in-spite of the maximal dosage of the fixed dose combination therapy with Olmesartan/ Amlodipine. 5.49%- patients (20 of 364) discontinued the treatment due to pedal edema. We observed from the screening holter blood pressure monitoring that 29.94% - patients (109 of 364) with resistant hypertension were non-dippers and after the fixed dose combination therapy with Olmesartan/Amlodipine only 10.44% patients (38 of 364) were non-dippers.

Conclusion:
The study revealed that fixed dose combination therapy with Olmesartan/Amlodipine in patients with Resistant Hypertension has high efficacy and minimal side effects with good circadian control of arterial blood pressure.
Abstract P028: Intravenous Anti-hypertensives Used for Non-urgent Hypertension in Inpatients
Authors: Rahman Faisal , Weinberg Janice , Bernard Sheilah , Arnold Lindsay
Published: March 2022

Background:
Asymptomatic hypertension is common in hospitalized patients but there are no clear treatment guidelines. Due to the concern for harm from elevated blood pressures (BP) physicians often prescribe rapidly acting intravenous (IV) agents, which may cause dangerous BP drops. We evaluated if the use of IV hydralazine or labetalol were associated with adverse events.

Methods:
We retrospectively identified patients >18 years of age admitted to Boston Medical Center over 1 year who had a recorded systolic BP > 160 mmHg or diastolic BP > 120mmHg during their hospitalization. Patients with hypertensive emergency, or admitted to intensive care or neurology services were excluded. We examined adverse events over the subsequent 24 hours comparing the use of IV labetalol and/or hydralazine versus use of neither IV anti-hypertensive in Cox proportional hazard models adjusted for age, gender, race, systolic and diastolic BP.

Results:
5,431 patients met inclusion criteria of which 203 patients received IV anti-hypertensives (78 labetalol only, 107 hydralazine only, 18 hydralazine and labetalol). In adjusted analyses, patients who received IV anti-hypertensives had increased risk of requiring IV fluids (hazard ratio [HR] 1.44; 95% confidence interval [CI] 1.03-2.01; P<0.03), and developing tachycardia (HR 1.82; 95% CI 1.18-2.82; P<0.007) compared to patients who did not receive IV agents. There was no difference in orders for computed tomography head, neurology or critical care consult, electrocardiogram, and troponin. The use of IV hydralazine compared with no IV drugs was associated with increased IV fluid use (HR 2.03; 95% CI 1.35-3.05; P<0.001) and tachycardia (HR 1.81; 95% CI 1.04-3.16; P<0.04), with no association of adverse events with IV labetalol use. There was no significant difference in 20% or 50% systolic blood pressure drop at 2 or 6 hours between IV hydralazine, IV labetalol or no IV anti-hypertensive groups.

Conclusions:
In contrast to previous concerns, our data suggest that the use of IV hydralazine or labetalol was not associated with severe adverse outcomes. Our retrospective study at a single center has several limitations and did not include all possible confounders, and future studies need to replicate these findings in other centers.
Abstract P035: Endothelin-1 Overexpression Preserves Endothelial Function in Mice with Vascular Smooth Muscle Cell-specific Deletion of Ppar-gamma
Authors: IdrIis Khodja Noureddine , Ouerd Sofiane , Trindade Michelle , Gornitsky Jordan , Rehman Asia , Offermanns Stefan , Gonzalez Frank J , Barhoumi Tlili , Paradis Pierre , Schiffrin Ernesto
Published: March 2022

Objective:
Peroxisome proliferator-activated receptor γ (PPARγ) agonists reduce blood pressure (BP) and vascular injury in hypertensive rodents and humans. Pparγ inactivation in vascular smooth muscle cells (VSMC)using a tamoxifen inducible Cre-Lox system enhanced angiotensin II-induced vascular injury. Transgenic mice overexpressing endothelin (ET)-1selectively in the endothelium (eET-1) exhibit endothelial dysfunction, increased oxidative stress and inflammation. We hypothesized that inactivation of Pparγ in VSMC(smPparγ-/-)will exaggerate ET-1-induced vascular damage.

Methods and Results:
Elevenweek-old male control, eET-1, smPparγ-/-and eET-1/smPparγ-/- mice weretreated with tamoxifen (1 mg/kg/day, s.c.) for 5 days and sacrificed 4 weeks later. Systolic BP was higher in eET-1compared to control (123±5 vs 109±2 mmHg,P<0.05)and unaffected by Pparγ inactivation.Mesenteric artery (MA) vasodilatory responses to acetylcholine were impaired only in smPparγ-/- (P<0.05) compared to control (Emax: 52.0±6.7% vs 82.2±4.9%).Reactive oxygen species levels were increased in eET-1, smPparγ-/-and eET-1/smPparγ-/-compared to control (1.7±0.2, 2.2±0.2 and 2.8±0.4 vs 1.0±0.2, P<0.05).MA monocyte chemoattractant protein-1 expression was higher in smPparγ-/-compared to control (31.0±4.4 vs 18.7±2.5, P<0.05), and unaffected by ET-1 overexpression.Perivascular fat monocyte/macrophage infiltration was higher in eET-1 and smPparγ-/- compared to control (331±34 and 326±49 vs 140±8 cells/mm2, P<0.05), and further increased in eET-1/smPparγ-/- (557±77, P<0.05). Nitric oxide synthase (Nos)3 mRNA expression was increased only in eET-1 compared to WT (1.21±0.07 vs 1.00±0.02, P<0.05). Nos2 expression was increased in eET-1 and smPparγ-/-compared to WT (3.69±0.72 and 1.96±0.16 vs 1.00±0.13, P<0.05). The Ednra/EdnrbmRNA ratio was decreased in eET-1/smPparγ-/- compared to smPparγ-/-(0.76±0.04 vs 1.07±0.10, P<0.05).

Conclusion:
Increased ET-1 paradoxically preserves endothelial function in mice with smPparγinactivation, despite enhancedoxidative stress and inflammation.
Abstract P046: Interplay of Intronic and Promoter Region Polymorphisms Up-regulate Human Angiotensinogen and Cause Hypertension in Transgenic Mice
Authors: Kaw Meenakshi , Mopidevi Brahmaraju , Kindell Daniel G , Puri Nitin , Rana Anita , Jain Sudhir , Kumar Ashok
Published: March 2022

Angiotensinogen (AGT) is the substrate for the RAS-cascade and polymorphisms leading to its overexpression are linked to hypertension. We have shown that SNPs in linkage disequilibrium with -6A of the hAGT gene cause increased AGT expression and hypertension. Recently, two genome wide association studies (GWAS) have linked SNP at the +1164 in intron-I with increased blood pressure. The +1164A allele has stronger homology with HNF-3 binding site as compared to +1164G. HNF3 belongs to the family of “pioneer” transcription factors, which enable chromatin access for other tissue-specific transcription factors. Thus, we propose that a haplotype, where multiple SNP-groups occur together, is a better model to predict and study disease correlation with genetic variations than individual SNPs alone. Intronic SNPs could increase chromatin access thus modulating promoter-transcription factor (TF) interactions. To study these possible interactions between the intronic SNPs in the promoter and intron I of the hAGT gene, we have divided this gene in two major haplotypes: haplotype-I (containing -6A & +1164A) and II (containing -6G & +1164G). Reporter construct containing haplotype-I has increased promoter activity as compared to haplotype-II on transient transfection in human liver cells, without (hap-1: 73±2 vs. hap-II: 23.64±3.21 A.U; p<0.05) and with dexamethasone treatment (hap-1: 1174±68vs. hap-II: 333.3±37 A.U; p<0.05). Double transgenic mice (TG) containing human renin gene (hREN) and haplotype-I of the hAGT gene (containing 2.0 Kb of the promoter, all the introns, exons and 3’UTR of the hAGT gene at the HPRT locus) have increased basal and GR induced expression of the hAGT gene in liver, kidney and fat as compared to haplotype-II. Complementary CHIP analysis shows increased TF-chromatin binding in the liver of haplotype-I TG mice for HNF3β (4.35 folds), C/EBPβ (2.1 folds), STAT-3 (3.6 folds) and GR (4.2 folds). Crucially, haplotype-I TG mice have increased (p<0.05) SBP (137±4 mm Hg) when compared to haplotype-II mice (126±3 mm Hg). In conclusion, intronic SNPs regulate hAGT gene expression via complex interplay with SNPs in the gene-promoter and this underscores the need for a haplotype-based approach to genetic variability and disease correlation.
Abstract P057: Is Hypertension a Disease of the Bone Marrow?
Authors: Zubcevic Jasenka , Santisteban Monica M , Ahmari Niousha , Schmidt Jordan T , Martyniuk Christopher J , Marulanda Carvajal Jessica , Kim Seungbum , Raizada Mohan K
Published: March 2022

Decades of evidence have implicated involvement of inflammation in the development and establishment of hypertension (HTN); however, the central mechanisms remain elusive. We propose a hypothesis that dysfunctional bone marrow (BM) activity is critical in HTN, in view of the fact that BM is the predominant source of inflammatory and angiogenic cells. We provide the following evidence in support of this hypothesis: (1) BM from animal models of HTN is proinflammatory. Ablation of the spontaneously hypertensive rat (SHR) BM, and reconstitution with BM from the normotensive Wistar Kyoto (WKY), results in significant reduction in mean arterial pressure (MAP), as well as the decrease in proinflammatory and increase in angiogenic cells in the chimeric SHR; (2) Oral minocycline treatment attenuates MAP, restores autonomic balance, and decreases inflammation in both the SHR and Ang II rat HTN models; (3) Sympathetic nerve activity (SNA) and norepinephrine levels in the BM of the SHR and Ang II rat HTN models are elevated compared to normotensive rats; (4) C57-AdrB1.B2 knock-out (KO) chimera, generated by reconstitution of irradiated C57BL/6J mice with the BM cells of the adrenergic receptor beta 1/2 KO mice (Adrb1tm1Bkk Adrb2tm1Bkk/J), exhibits reduced peripheral inflammatory cell counts. Furthermore, transcriptomics analysis of the BM cells from these chimeric mice revealed significant changes in 67 signaling pathways, thirty-five of which were directly related to modulation of different immune system responses (P<0.01). Most notable were changes in the pathways involved in activation and migration of monocytes and T lymphocytes. These observations demonstrate that BM plays a critical role in regulation of the inflammatory status, and support our hypothesis that dysfunctional BM activity may be an important aspect in the development and establishment of HTN. AHA14SDG18300010.
Abstract P073: Chemerin Receptor Antagonism Improves Vascular Insulin Signaling in Diabetic Mice
Authors: Neves Karla B , Lobato Nubia S , Nguyen Aurelie D , Lopes Rheure A , Hood Katie Y , Mestriner Fabiola L , Oliveira Ana M , Montezano Augusto C , Touyz Rhian M , Tostes Rita C
Published: March 2022

Adipose tissue releases many adipokines, including chemerin, which produces its effects through activation of chemokine-like receptor 1 (ChemR23). Chemerin influences major aspects of the metabolic syndrome, including adipogenesis and insulin sensitivity in adipocytes and skeletal muscle cells. Considering that chemerin impairs vascular reactivity and that amongst its many actions, insulin also influences vascular function, we postulate that chemerin decreases insulin-induced vasodilatation, by reducing PI3K/Akt signaling, and impairs glucose uptake by vascular smooth muscle cells (VSMC). Mesenteric arteries, endothelial cells (EC) or VSMC from C57Bl6 mice were incubated with chemerin (0.5 ng/mL, 1 h) or vehicle (veh). Chemerin decreased insulin-induced relaxation (0.1 - 3000 ng/mL, pD2: chemerin: 94.5±0.1 vs. veh: 23.6±0,1), which was prevented by the ChemR23 antagonist CCX 832 (pD2: veh= 16.9±0.1; chem= 174.1±0.1, CCX+chem= 37.4±0.1) and a PI3K activator (YS-49) (pD2: veh= 23.0±0.1; chem= 108.5±0.1, YS-49+chem= 28.5±0.1). Chemerin also decreased PI3K (0.8±0.1 vs. veh 1.1±0.1), Akt (0.8±0.1 vs. veh 1.2±0.1) and AMPK [0.7±0.2 vs. veh 1.0 ±0.1] phosphorylation in VSMC. IRS1 and IRS2 gene expression was decreased in VSMC stimulated with chemerin. In addition, chemerin decreased insulin-stimulated NO production in EC through activation of ChemR23 and PI3K and inhibited insulin-stimulated glucose uptake by VSMC (counts per minute: 5668±729.0 vs. veh 9923±662.7), which was mediated by ChemR23, AMPK and PI3K. Importantly, ChemR23 antagonism in diabetic (db/db) mice partially reversed decreased insulin-induced vasodilatation (mesenteric resistance arteries; pD2: db/m: 6.1±1.0; db/db: 7.3±0.2; db/db+CCX832: 6.9±0.3). In conclusion, chemerin decreases vascular insulin responses by reducing PI3K/Akt and AMPK signaling. Moreover, chemerin/ChemR23 system plays a crucial role in impaired vascular insulin signaling in diabetes, suggesting its involvement in the pathogenesis of vascular insulin resistance. Our study may contribute to a better understanding of the role of chemerin in vascular insulin dysfunction in diabetes- and obesity-associated diseases. Financial Support: FAPESP, Brazil.
Abstract P134: Identification of Tissue Inhibitor of Metalloproteinases (TIMP)-4 as a Novel PPARγ Target in Smooth Muscle Cell
Authors: Ketsawatsomkron Pimonrat , Lu Ko-Ting , Stump Madeliene , Keen Henry L , Sigmund Curt D
Published: March 2022

Patients with PPARγ mutations develop severe early onset hypertension, type 2 diabetes and lipodystrophy. Our recent findings demonstrated that transgenic mice expressing a dominant negative (DN) PPARγ mutant in vascular smooth muscle (S-P467L) exhibited exacerbated DOCA-salt induced hypertension and vascular remodeling in conduit and resistance arteries. Here, we hypothesized that predisposition to vascular injury during hypertension in S-P467L mice is through altered expression of PPARγ target genes in smooth muscle cells (SMC). Gene expression profiling in aorta and mesenteric arteries revealed a significant loss of Tissue Inhibitor of Metalloproteinases (TIMP)-4 in S-P467L compared to non-transgenic (NT) littermates. Interference with PPARγ activity either by treatment with PPARγ inhibitor, GW9662 or expressing P467L PPARγ markedly suppressed TIMP-4 in primary SMC, suggesting that loss of TIMP-4 in S-P467L arteries is a direct result of PPARγ inhibition. Downregulation of TIMP-4 in SMC by GW9662 was correlated with a significant increase in total MMP activity (Fluorescent signal subtracted from negative control; vehicle: 1±0.5, GW9662: 2.2±0.7, p<0.05), consistent with TIMP-4 function as an endogenous inhibitor of MMPs. Overexpressing TIMP-4 in SMC significantly blunted cell migration compared to those with empty plasmid (change in open area after 8 hr; Empty: 1±0.05, TIMP-4: 0.8±0.051, p<0.05), whereas decreased TIMP-4 expression caused by mutant PPARγ resulted in increased migration (change in open area after 10 hr; GFP: 1±0.25, P467L: 1.63±0.26, p<0.05). The significance of TIMP-4 was also underscored during hypertension since the compensatory increase in TIMP-4 during DOCA-salt in NT mesenteric arteries was lost in S-P467L. We identified two highly conserved potential PPAR response elements (PPREs) close to TIMP-4 promoter region using a sequence-based model. Chromatin immunoprecipitation assay showed strong binding of PPARγ at one of the suggested PPREs in SMC (% input±sem; GFP: 37±0.6, P467L: 66±0.7, p<0.05 vs. IgG), suggesting that TIMP-4 is a direct target of PPARγ. Our findings highlight one of protective mechanisms of PPARγ during hypertension and provide a novel link between PPARγ and TIMP-4.
Abstract P148: Ambulatory Blood Pressure in Hypertensive Patients Treated with Different 2-Drug and 3-Drug Combinations
Authors: De la Sierra Alejandro , Gorostidi Manuel , Banegas Jose R , Segura Julian , Vinyoles Ernest , De la Cruz Juan J , Ruilope Luis M
Published: March 2022

An elevated proportion of patients require 2-drug and 3-drug combinations to achieve BP control. There is scarce evidence regarding if there are differences in such BP control among different types of combinations in daily practice. We aimed to assess office and ambulatory BP values achieved, as well as the proportion of controlled patients, depending on the type of drug combination used.

From the Spanish ABPM Registry we selected 17187 patients treated with the 6 most common types of 2-drug combinations and 9724 treated with the 6 most common types of 3-drug combination. We looked for differences in achieved office and ambulatory BP, as well as nocturnal dip, and the proportion of controlled patients among types of combinations, after adjusting for confounders (age, gender, BMI, smoking, diabetes, dyslipidemia, and previous CV disease).

With respect to the combination of renin angiotensin system (RAS) blockers and diuretics (reference), none of the other combinations achieved lower BP values or better BP control. Ambulatory BP control (including 24-hour, daytime and nighttime) was worse with combinations of RAS blockers/beta blockers (OR: 1.06; 95%CI: 1.01-1.11), with combinations of calcium channel blockers (CCB)/beta blockers (1.1; 1.04-1.16) and with combinations of RAS blockers and CCB (1.38; 1.23-1.55). Nondipping was also more frequent in combinations other than RAS blockers/diuretics. In patients receiving 3-drug combinations, and with respect to RAS blockers/CCB/diuretic combinations (reference), ambulatory BP were higher and non dipping more frequent in other types of combinations. Ambulatory BP control was worse in RAS blockers/CCB/alpha blockers, RAS blockers/diuretics/alpha blockers, and CCB/beta blockers/diuretics combinations. No differences in office BP control were observed among types of 2-drug or 3-drug combinations.

We conclude that RAS/diuretic combinations and RAS/diuretic/CCB combinations are associated with better ambulatory BP control and more pronounced dipping in comparison with other types of 2-drug or 3-drug combinations, even with same rates of office BP control. These results can be helpful in deciding the way to combine antihypertensive agents in patients who require combination therapy.
Abstract P158: Increased Circulating Cathepsin K in Patients with Chronic Heart Failure
Authors: Cheng Xian Wu , Zhao Guangxian , Han Xoingyi , Fang Ennan , Li Xiang , Zhou Dongmei , Jiang Haiying , Li Yuzi , Cui Lan
Published: March 2022

Background:
Cysteinyl cathepsin K (CatK) is one of the most potent mammalian collagenases involved in cardiovascular disease. We investigated the clinical predictive value of serum CatK levels in patients with chronic heart failure (CHF).

Methods and Results:
We examined 134 patients with CHF, measuring their serum CatK, troponin I, high-sensitive C-reactive protein, and pre-operative N-terminal pro-brain natriuretic peptide levels. The patients were divided into two groups: the 44 patients who showed a left ventricular (LV) ejection fraction (LVEF) < 40% (the “lowLVEF” group) and the 90 patients showing LVEF values ≥ 40% (the “highLVEF” group). The lowLVEF patients had significantly higher serum CatK levels compared to the highLVEF patients (58.4 ± 12.2 vs. 44.7 ± 16.4, P < 0.001). Overall, a linear regression analysis showed that CatK levels correlated negatively with LVEF (r = –0.4, P < 0.001) and positively with LV end-diastolic dimensions (r = 0.2, P < 0.01), LV end-systolic dimensions (r = 0.3, P < 0.001), and left atrial diameters (r = 0.2, P < 0.01). A multiple logistic regression analysis showed that CatK levels were independent predictors of CHF (odds ratio, 0.90; 95% confidence interval, 0.84-0.95; P = 0.001).

Conclusions:
These data indicate that elevated levels of CatK are closely associated with the presence of CHF and that the measurement of circulating CatK provides a noninvasive method of documenting and monitoring the extent of cardiac remodeling and dysfunction in patients with CHF.
Abstract P159: Effect of Pharmacological Kinin Receptor Activation on Brain Damage and Mortality in Experimental Cerebral Ischemia in Non-diabetic and Diabetic Mice
Authors: Desposito Dorinne , Taveau Christopher , Zadigue Georges , Adam Clovis , Bouby Nadine , Alhenc-Gelas Francois , Roussel Ronan
Published: March 2022

Brain ischemia is a major complication of arterial diseases and has a poorer prognosis in diabetic patients. As activation of the kallikrein-kinin system has been shown to enhance cardiac and renal tolerance to ischemia we tested effect of kinin receptor activation by pharmacological agonists, selective B1R or B2R, in a mouse model of transient middle cerebral artery occlusion [C57bl6 male, 10 week-old, anaesthesia, occlusion 60 min (MCAO)]. Treatment with the B1R agonist NG29 (B1Rago) or the B2R agonist NG291 (B2Rago) was started at reperfusion using osmotic micropumps. Neurological deficit (ND) was evaluated at 1 and 2 days using a panel of 8 established tests combined in a 0-30 deficit score. Brain infarction was quantified at day 2 using TTC and hematoxylin-eosin staining. In some mice diabetes was induced by streptozotocin 8 weeks before MCAO.

MCAO induced bradychardia, mild hypotension (mean -11.3 mmHg), ND (19.7 ± 2.4) and resulted in partial brain infarction (18 ± 1.4 %), all p<0.05 compared to sham, n=10/group. B2Rago (720 nmol/kg.day-1) increased ND to 27 ± 1.8 at day 1 and mortality to 60% at day 2 (both p<0.05, n=10) while decreasing brain infarct size by 66% (p<0.01). B2Rago reduced BP by 16 mmHg and increased plasma creatinine (73 ± 25 μmol/l, p<0.05). Although B1R mRNA level increased by 1.3 fold in the ischemic brain B1Rago had no effect on ND, mortality or brain infarction.

In diabetic mice MCAO increased ND (28 ± 1), mortality (25%) and infarct size (40 ± 3 %) more than in non-diabetic mice (n=8, p<0.05). B2Rago increased mortality to 80% (p<0.05, n=9). B1Rago, tested at two different dosages (720 or 240 nmol/kg.day-1, n= 8/group) reduced ND (22 ± 2 at day 2 for the low dosage, p< 0.05) and did not increase mortality or alter renal function. B1Rago reduced infarct size by 66 and 71 %, at the two dosages, respectively (p<0.01).

Thus, B2R activation reduces brain infarction but paradoxically increases mortality by mechanisms that may involve brain oedema and renal insufficiency. B1R activation has no effect in non-diabetic mice but in diabetic animals it reduces infarct size and improves ND without adverse effect on renal function and survival. Longer follow-up studies are in progress for further evaluating interest and limitation of B1R activation in MCAO.
Abstract P201: Divergent Angiotensin Receptor Signaling in a Mouse Model of Post-Traumatic Stress Disorder (PTSD)
Authors: Swiercz Adam , Moshfegh Cassandra , Desai Amishi , Hopkins Lauren , Marvar Paul J
Published: March 2022

Independent of their beneficial effects on hypertension and cardiovascular related disease, angiotensin receptor type 1 (AT1R) blockers can improve stress-related symptoms. AT1R receptor-mediated actions can be counteracted directly or indirectly by the angiotensin receptor type 2 receptor (AT2R). Our recent studies in a mouse model of PTSD have shown that AT1R blockade increases the extinction (learned inhibition) of a traumatic fear memory and that AT1R mRNA expression is reduced in fear related brain regions of animals treated with the AT1R antagonist losartan. These data imply that downstream AT1 signaling events maybe important in consolidation of fear memory extinction. Therefore we investigated the acute effects of AT2R inhibition and AT2R stimulation on fear memory and baseline anxiety in mice. We performed classical Pavlovian fear conditioning pairing auditory cues with foot shocks and examined fear extinction (% freezing to conditioned stimulus) in the presence of the AT2R antagonist PD 123319 and agonist Compound 21. Twenty-four hours following fear conditioning, PD 123319 (15 mg/kg IP), Compound 21 (10 mg/kg IP) or vehicle was administered prior to fear memory extinction. The PD treated group exhibited significantly less percent freezing (68%; 68 of 100) compared to vehicle control (47%; 47 of 100) during fear expression (F10, 300 = 1.9; p<0.05, n=15) while no effect was observed during extinction retention, an index of long-term fear memory. On the other hand, Compound 21 had no effect on fear expression, extinction or basal levels of anxiety. Moreover, following fear conditioning, qPCR data revealed that mRNA expression of AT2R and angiotensin converting enzyme 2 (ACE2) in the central amygdala were elevated (5 fold and 3 fold respectively, P<0.05, n=6) compared to home cage control, however the AT1R and angiotensin converting enzyme (ACE) gene expression pathways were unaltered. PD 123319 and Compound 21 had no effect on basal levels of anxiety as determined by open field testing. These data indicate that AT1R and AT2R may have divergent effects on short and long-term fear memory formation. Further studies are required to understand the differential regulation of angiotensin receptor signaling in PTSD.
Abstract P205: Angiotensin II Metabolic Pathways In Isoproterenol-Induced Heart Failure: Myocytes vs Non-myocytes
Authors: Ahmad Sarfaraz , Li Tiankai , Zhang Xiaowei , Varagic Jasmina , Groban Leanne , Louis Dell'Italia J , Cheng Che Ping , Ferrario Carlos M
Published: March 2022

Although myocytes represent three-fours of the ventricular volume in mammalian hearts 90% of the remaining myocardial cells are interstitial cardiac fibroblasts. To shed light into the paracrine mechanisms that contribute to the expression and function of the cardiac angiotensins, we investigated the comparative activity of chymase, ACE, and ACE2 in freshly isolated myocytes and non-cardiac myocytes (NCM) from the left ventricle of rats in which heart failure (HF) was induced by two injections of isoproterenol (Iso) (170 mg/kg body weight, s.c.) spaced 24 h apart. Consistently, chymase enzymatic activity was approximately 10-fold higher in NCM (8.37 ± 0.66 in normal and 6.76 ± 2.7 fmol/min/mg in Iso-induced HF, P < 0.05) compared to myocytes obtained from normal (0.43 ± 0.11 fmol/min/mg) or Iso-induced HF rats (0.65 ± 0.17 fmol/min/mg). Compared to chymase, ACE activity was several folds lower in both NCM (1.37 ± 0.21 in normal and 1.45 ± 0.49 fmol/min/mg in Iso-induced HF) and myocytes (0.28 ± 0.01 fmol/min/mg in normal and 0.37 ± 0.07 fmol/min/mg in Iso-induced HF). As illustrated in the Figure, both chymase and ACE2 activity tended to be higher in myocytes from HF rats. These data suggest that Iso-induced HF causes selective changes in the activity of chymase and ACE2 in myocytes but not in NCM. The clinical significance of these novel findings suggest that chymase rather than ACE inhibitors will have a greater benefit in the management of adverse cardiac remodeling.
Abstract P209: RAS-Equilibrium Analysis: Simplified Biochemical Characterization of the Renin-angiotensin-system and Implications for Diagnosis and Treatment of Hypertension
Authors: Poglitsch Marko , Schwager Cornelia , van Oyen Dunja , Aigner Claudia , Domenig Oliver
Published: March 2022

The concentrations of angiotensins are maintained by equal rates of formation and degradation. The resulting steady-state angiotensin levels are affected by local molecular factors including tissue, endothelium or blood cell associated angiotensin receptors and processing enzymes as well as plasma soluble RAS components. The high concentration of the pro-hormone angiotensinogen in human plasma in combination with reported ranges for plasma renin activity result in a long-lasting and stable Ang I formation rate without significantly reducing angiotensinogen levels within several hours. This phenomenon can be utilized for the generation of an ex vivo situation that is characterized by significantly higher but stable angiotensin peptide levels. These ex vivo equilibrium angiotensin levels provide an integrated picture about plasma angiotensinase activities and therefore represent a powerful diagnostic tool for analyzing the systemic RAS in clinical samples. Moreover, these ex vivo levels remain stable over hours of incubation at 37°C and show a very high correlation (>90%) with angiotensin levels obtained by a state-of-the art sample collection procedure using complex inhibitor cocktails for stabilizing angiotensin peptides during blood collection.

The quantification of equilibrium angiotensin levels does not require any special sample collection procedures and can be applied to frozen serum and heparin plasma samples. It turned out that ex vivo angiotensin levels represent a measure for the consumption of angiotensin metabolites by the organism, therefore providing a powerful and versatile tool for assessing in vivo angiotensin signaling on the patient specific level enabling new diagnosis based rationales for anti-hypertensive therapies. The application of ex vivo RAS-Fingerprinting in clinical studies could substantially enhance our understanding of the regulation and physiology of the human RAS and could further lead to the development of powerful personalized approaches in the future treatment of hypertension.
Abstract P604: Equivalency Validation of a Novel Freely Moving Animal Based Telemetry System versus a Stationary Cage Bound System for Blood Pressure Recordings
Authors: Knot Harm , Miller Gary
Published: March 2022

Traditional rodent blood pressure (BP) monitoring required single housing, a known stressor for rodents. We developed a microprocessor based long range wireless telemetry system with large internal memory that allows rodents to live in groups, interact, exercise and be housed in large enriched environments. BP is recorded with a solid-state sensor along with core temperature and 3D axial activity within the implant. The measurement protocol is transmitted once to the programmable implant via a single room antenna mobile base station. The implant becomes an autonomous recording device until contacted again by the base station with an updated protocol. The implanted animals are thus allowed to undergo procedures, leave the room or be subject other tests while the recording is running. Data can be automatically downloaded periodically within the protocol.

In order to objectively validate this system we contracted a study with the Michigan State University INVIVO facility. 8 male SD rats of 275g were implanted with a model PTA-M TSE Stellar telemetry implant. After 8 days of recovery the animals entered a crossover study and were dosed with vehicle, 3, 10, and 100 mg/kg L-NAME or Verapamil at t=0 hrs. BP was recorded for 10s hourly for 24hrs sampled at 200Hz following drug administration. The BP responses were compared to a control group of 8 animals implanted with a similar conventional implant from a competitor. Drug responses were qualitatively the same between the groups. Mean arterial pressures (MAP) at +12hrs in the 100 mg/kg LNAME group with the TSE implant increased by 26±11 versus 26±9 mmHg in the control group. MAP in the 100 mg/kg Verapamil group at +12 hrs dropped by 31±9 versus 20±6 mm Hg in the control group. Baseline MAP was significantly elevated to 118±5 versus 92±11 in the control group with the conventional implant which may account for the blunted verapamil response.

We conclude that there are no significant differences in quantitative blood pressure responses to L-NAME or Verapamil in animals implanted with a TSE Stellar device versus a leading competing device. We also conclude that the short hourly recordings used for the TSE device produced equivalent MAP values while significantly prolonging implant battery life, thus extending possible study length.
Abstract P629: Effect of Circulating ACE2 Activity on plasma Angiotensin Peptides
Authors: Wysocki Jan , Ye Minghao , Khattab Ahmed M , Serfozo Peter D , Osborn Mark , Batlle Daniel
Published: March 2022

Circulating ACE2, an enzyme that degrades Ang II, has been reported to be increased in rodent models of diabetes. Upregulation of ACE2 by increasing hydrolysis of Ang II could help downregulate RAS overactivity in a compensatory way and/or reflect increased Ang II metabolism. ACE2 activity in plasma, however, is relatively low even in diabetic conditions. Therefore, further increases in ACE2 effected by exogenous ACE2 administration could have a therapeutic action as a result of enhanced Ang II degradation and formation of Ang 1-7.

We examined the effect of a massive increase in circulating ACE2 activity on plasma levels of Ang II and other Ang peptides in mice rendered diabetic by STZ. Circulating ACE2 was amplified using ACE2 mini-circle (ACE2MC) gene delivery. Multiple Ang peptides were evaluated in plasma from STZ-mice that received ACE2MC and sham STZ-mice (n=6/group).

Plasma Ang II in STZ-ACE2MC mice were markedly lower than in STZ sham mice (3±1 vs. 22±13, pg/mL, respectively), but the difference was not significant owing to high variability. Since the seemingly lower Ang II levels could be influenced by either lower levels of the precursor peptide Ang I, increased degradation of Ang II, or both, a ratio between Ang II and Ang I was calculated. Ang II/Ang I ratio was significantly lower in STZ-ACE2MC as compared to STZ sham mice (0.15±0.05 vs. 0.35±0.06, p<0.05) indicating that per given amount of Ang I, there is significantly less Ang II in STZ-ACE2MC mice. This is consistent with enhanced Ang II degradation. Plasma levels of Ang 1-7 and Ang 1-9, the direct products of ACE2 cleavage of Ang I and Ang II, respectively, were very low and mostly below the level of detection. In plasma, Ang 1-7 is quickly converted to Ang 1-5, by ACE. Therefore, as an index of Ang II conversion to Ang 1-7, and as a surrogate for this conversion, we used Ang 1-5/Ang II ratio. This ratio was markedly higher in STZ-ACE2MC than in sham STZ-mice (3.15±1.5 vs. 0.38±0.07, p<0.05, respectively) suggesting an increased conversion of Ang II to Ang 1-7, and later to Ang 1-5 as a result of plasma ACE2 overexpression.

We conclude that a massive increase in circulating ACE2 is associated with an accelerated metabolism of Ang II. The increase in Ang II degradation seems best reflected by an increase in Ang 1-5/Ang II ratio.
Abstract 002: Neuropilin1: New Splenic Mechanisms Mediating the Guidance of Neural Cues in Immune Cells Relevant for Hypertension
Authors: Carnevale Daniela , Perrotta Marialuisa , Piacenti Maria , Iacobucci Roberta , Cifelli Giuseppe , Lembo Giuseppe
Published: March 2022

Investigation of immune mechanisms involved in hypertension (HTN) is one of the most emerging research area in the field. On this note, we found that hypertensive hits activate nervous system in splenic marginal zone (MZ) that represents the boundary between lymphoid white pulp and innate red pulp. The fact that both innate and adaptive immunity contribute to HTN strongly suggests that neuroimmune mechanisms couple myeloid and lymphoid cells in transducing hypertensive hits into increased blood pressure (BP). We previously showed that Placental Growth Factor (PlGF), a VEGF homolog, released in the splenic MZ upon Angiotensin II (AngII), mediates this interaction. Here we dissect the downstream pathway conveying nervous outflow into immune activation. PlGF is expected to act through its major receptor VEGFR1, its only known tyrosine kinase receptor. First we found that it is mainly expressed in splenic innate immune cells and vasculature, both in basal conditions and upon AngII. To test VEGFR1 in HTN, we challenged transgenic mice with defective VEGFR1 signaling (Flt1-TK) with AngII. To our surprise, they raised BP upon 28-days AngII as WT mice did (SBP: Flt1-TK 139±2; WT 138±2; ***p< 0.001), thus suggesting the existence of alternative pathways. PlGF may also act on a co-receptor, Neuropilin1 (Nrp1), known to enhance responses to ligands. In addition, Nrp1 has unique functions in neurons’ guidance during development and in the maturation of dendritic cells from monocytes. We found Nrp1 basally expressed and strongly increased upon AngII, especially in MZ macrophages. To test the hypothesis that PlGF/Nrp1 pathway could mediate a neuroimmune activation of dendritic cells relevant for HTN, we selectively deleted the receptor in myeloid lineage, generating Nrp1
fl;fl
;LysM-Cre transgenic mice. After assessing Nrp1deletion in splenic monocyte/macrophages by FACS, we infused AngII and found a protection from T cells costimulation and egression from the spleen. More important, they were protected from HTN (SBP: Nrp1
fl;fl
;LysM-Cre
+/-
140±3; Nrp1
fl;fl
;LysM-Cre
-/-
105±2; ***p< 0.001). Our data indicate a crucial role of Nrp1 in HTN, mediating non-canonical PlGF signaling in innate immunity that serves as a guidance of nerves priming adaptive immune responses.
Abstract 024: Angiotensin-(1-7) Attenuates Diet-induced Obesity in Mice by Browning White Adipose Tissue to Enhance Energy Expenditure
Authors: Czyzyk Traci A , Cooper Timothy K , Bingaman Sarah S , Arnold Amy C
Published: March 2022

Overactivation of the renin-angiotensin (Ang) system, and in particular Ang II, contributes to obesity-induced insulin resistance via multiple mechanisms. Obesity is also associated with deficiency of Ang-(1-7), a beneficial hormone that opposes Ang II actions. In this study, we tested the hypothesis that Ang-(1-7) could prevent development of diet-induced obesity in mice. To test this hypothesis, adult male C57BL/6J mice received 12-week Ang-(1-7) (400 ng/kg/min; n=7) or saline (n=6) infusion via subcutaneous osmotic mini-pumps. Mice were placed on 60% high fat diet (HFD) immediately following mini-pump implantation. Body composition, energy balance, and insulin sensitivity were measured during the last week of treatment. Ang-(1-7) attenuated HFD-induced weight gain (39.7±1.3 vs. 43.9±0.7 g saline; p=0.023) and adiposity (32±1 vs. 29±1% saline; p=0.050). This weight-reducing effect was due to enhanced average energy expenditure [0.55±0.02 vs. 0.42±0.06 kcal/hour saline, p=0.038], with no changes in locomotor activity or food intake. Ang-(1-7) attenuated HFD-induced hyperinsulinemia (1.4±0.4 vs. 5.3±1.6-ng/mL saline; p=0.032) and improved the ability of intraperitoneal insulin to decrease blood glucose levels (20% reduction in area under the curve vs. saline; p=0.015), consistent with insulin-sensitizing effects. Given effects of Ang-(1-7) on energy expenditure, we performed blinded histological and semi-quantitative real-time PCR analysis of brown and white adipose tissue. Ang-(1-7) reduced HFD-induced lipid droplet accumulation in brown adipose, but did not improve thermogenesis markers (e.g. uncoupling protein-1). Ang-(1-7) improved inflammation in visceral white adipose (mean number crown-like structures: 3±1 vs. 8±2 saline; p=0.046). Ang-(1-7) reduced adipocyte size and increased gene expression of the browning marker PRDM16 (1.7±0.2 vs. 1.1±0.1; p=0.024) in subcutaneous white adipose. These data suggest that Ang-(1-7) attenuates diet-induced obesity in mice by enhancing energy expenditure, in part by browning of white adipose tissue. These overall findings provide rationale for targeting Ang-(1-7) to prevent development of obesity and related pathology such as insulin resistance and adipose inflammation.
Abstract 039: Blockade of Macula Densa NOS1 Decreases GFR and Promotes the Development of Hypertension During Pregnancy
Authors: Wei Jin , Zhang Jie , Intapad Suttira , Wang Lei , Jiang Shan , Granger Joey , Liu Ruisheng
Published: March 2022

We hypothesized that neuronal nitric oxide synthesis (NOS1) β in the macula densa promotes the renal hemodynamic changes during normal pregnancy and inhibition of macula densa NOS1 induces hypertension during pregnancy. First, we measured the protein levels of NOS1β in the renal cortex, which increased by 11.5±1.7 folds in pregnant mice (n=5) compared with virgin mice (n=7, p<0.01 vs virgin), while there was no change in NOS1α. Then, in isolated and perfused juxtaglomerular apparatus, we measured tubuloglomerular feedback (TGF)-induced NO generation by the macula densa, which increased by 66.7±6.1% in pregnant mice (n=5) compared with the virgin mice (n=5, p<0.01 vs virgin). We next measured TGF
in vivo
using micropuncture, which was 3.6±0.3 mmHg in mice at day 19 of pregnancy (n=3) and 5.5±0.7 mmHg in virgin mice (n=7, p<0.05 vs virgin). To determine the significance of the macula densa NOS1, we measured glomerular filtration rate (GFR) and mean arterial pressure (MAP) with telemetry in macula densa specific NOS1 knockout (KO) mice (NKCC2
cre
; NOS1
flox/flox
) and WT (NOS1
flox/flox
) mice. In the WT mice, GFR raised by 25.7±1.1% while the MAP decreased by 6.1±2.7 mmHg at day 18 of pregnancy (n=7, p<0.01). The elevations in GFR were largely blunted in KO mice with only a 13.7±2.8% increase while MAP gradually rose by 28.2±3.3 mmHg above basal level at day 18 of pregnancy (n=5, p<0.05 vs baseline). Finally, we examined macula densa NOS1 in a mouse model of preeclampsia, a reduced uterine perfusion pressure (RUPP) model. At day 19 of pregnancy, the protein levels of NOS1β decreased 71.4±9.6% (n=5), NO generation reduced 26.7±5.8% (n=5) and TGF
in vivo
increased 41.6±12.2% (n=3) in RUPP mice, compared with normal pregnant mice (p<0.05). Moreover, the MAP or GFR was no significant different between the pregnant KO mice with RUPP and the pregnant KO mice. In conclusion, upregulated macula densa NOS1β blunts TGF response and promotes elevation of GFR during normal pregnancy. In contrast, downregulation of NOS1β in the macula densa enhances the TGF response, decreases GFR and promotes hypertension during pregnancy. These results suggest that inhibition of macula densa NOS1β could be an important mechanism mediating the decrease in GFR and elevation in MAP in preeclampsia.
Abstract 044: Sex Differences in Relative Contributions of Hemodynamic Parameters to Blood Pressure: A Population-based Study of Adolescents and Middle-aged Adults
Authors: Syme Catriona A , Shin Jean , Abrahamowicz Michal , Leonard Gabriel , Perron Michel , Richer Louis , Veillette Suzanne , Gaudet Daniel , Paus Tomas , Pausova Zdenka
Published: March 2022

Hypertension is a leading cause of death. Sex differences in blood pressure (BP) and hypertension prevalence emerge during adolescence and remain throughout reproductive age. Sex differences in BP-underlying hemodynamics, i.e., in the relative contributions of stroke volume (SV), total peripheral resistance (TPR) and heart rate (HR) to BP, have not been investigated in these age categories in a population-based setting. We studied a cohort of 1,347 individuals, including 911 adolescents (12-18 years, 52 % female) and 426 young to middle-aged adults (36-65 years, 56 % female). Beat-by-beat systolic BP (SBP) and diastolic BP (DBP), together with HR, SV, and TPR, were measured with a Finometer throughout a 52-min protocol; the protocol was intended to “mimic” daily-life activities, such as changes in posture and mental stress. It is well established that BP during regular daily activities (ambulatory BP monitoring) is a better predictor of target-organ damage than standard office BP. The relative contributions of HR, SV and TPR to SBP and DBP were determined by decomposing the model-explained variance into non-negative contributions. The relative contributions of SV, TPR and HR to SBP and DBP showed marked sex differences in young and middle-aged adults. The main determinant of higher SBP was SV in females (55 [50-60] % in females vs. only 35 [30-40] % in males), whereas it was TPR in males (47 [41-52] % in males vs. only 30 [26-34] % in females). The main determinant of higher DBP was TPR in both sexes, but its contribution was higher in males than females (58 [52-63] % vs. 41 [36-45] %, respectively). These sex differences were seen across most of the 52-min protocol, being most prominent during standing and least evident during mental stress. Similar sex differences were observed in adolescents, but they were less pronounced, being significant only during standing. The present population-based study of adolescents and young to middle-aged adults, suggest that marked sex differences exist in BP-underlying hemodynamics, with BP being driven mainly by SV in females and by TPR in males. These results underscore the need for sex-specific treatments of hypertension (not recommended currently), which affects 3-5 % of adolescents and 23-58% of young to middle-aged adults.
Abstract 053: Urinary Extracellular Messenger RNA Transcripts as Biomarkers of Mineralocorticoid Receptor Activity
Authors: Byrd J. Brian , Bazzell Brian G , Rainey William E , Auchus Richard J , Hummel Scott L
Published: March 2022

Objectives:
Mineralocorticoid receptor (MR) activation regulates sodium homeostasis & blood pressure. Our objective was to identify a non-invasive biomarker of MR activation in humans.

Background:
MR is a ligand-activated transcription factor that alters expression of target genes in the distal renal tubule, as shown in cell & animal models. Renal tubular cells secrete extracellular vesicles which carry RNA into urine. Whether human urinary extracellular mRNA reflects changes in gene expression in the renal tubule & thus might provide insight into MR activation is unknown.

Methods:
Pre-hypertensive participants consumed a low-sodium diet followed by sodium loading. We designed qPCR assays for MR gene targets & control genes not expected to respond to MR activation. Using samples collected during low-sodium diet & after sodium infusion, we assayed plasma renin activity (PRA), serum aldosterone, urinary sodium & urinary MR-responsive & control extracellular mRNA.

Results:
Eighteen participants’ low-sodium & 17 participants’ sodium-loaded urine samples were available. PRA & serum aldosterone were higher (
P
<0.001) & urinary sodium excretion was lower (
P
<0.001) during low-sodium diet. Four of 14 target gene qPCR assays (29%) changed after sodium loading, including assays for
SCNN1A
,
SCNN1G
&
TSC22D3
(
P
=0.006 to 0.01), whereas an assay for
SGK1
approached significance (
P
=0.07). In contrast, only 1 out of 10 control gene assays (for
NR3C2
, encoding MR itself) was significantly different after sodium loading (
P
=0.003). Log serum aldosterone inversely associated with C
t
values for 7 of 14 target gene assays (including assays for
SCNN1A
,
SCNN1G
,
SGK1
&
TSC22D3
[r= -0.46 to -0.52,
P
=0.003 to 0.048]) & for control gene assays for
NR3C2
&
UMOD
. C
t
values for 6 of 14 target gene qPCR assays associated with log urinary sodium/creatinine ratio including assays for
SCNN1A
,
SCNN1G
,
SGK1
&
TSC22D3
associated with log urinary sodium/creatinine ratio ([r= 0.36 to 0.52,
P
=0.003 to 0.048]) but not with control gene assays.

Conclusions:
Perturbations in human endocrine physiology can be detected as changes in urinary extracellular mRNA. Our findings suggest a new strategy to screen for mineralocorticoid excess & to assess pharmacologic or dietary changes in MR activity.