bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract P120: Mir-338-3p is Down-regulated in Subcutaneous Small Arteries of Hypertensive Patients With Chronic Kidney Disease
Authors: Mahjoub Nada
Published: December 2018

Background:
Hypertension (HTN) and chronic kidney disease (CKD) are among the most prevalent global health conditions that cause millions of deaths per year. They are associated with vascular damage characterized by vascular remodeling, stiffening and endothelial dysfunction. miRNAs (miRs) are a class of small non-coding RNA that regulate gene expression by binding to their target messenger RNAs (mRNAs), thereby leading to mRNA degradation or translational repression. Their implication in vascular injury remains unclear. We aimed to identify differentially expressed (DE) miRs in small arteries of HTN and CKD human subjects to gather insight into pathophysiological molecular mechanisms in these conditions.

Methods and Results:
Normotensive, HTN [systolic blood pressure (BP) >135 mmHg or diastolic BP of 85-115 mmHg with BpTRU] and CKD subjects (eGFR<60mL/min/m
2
) (n=15-16) were studied. Small arteries were isolated from subcutaneous gluteal biopsies and RNA extracted for small and total RNA sequencing using Illumina HiSeq-2500. EdgeR identified DE miRs (
P
<0.05) uniquely associated with HTN (3 up and 6 down) or CKD (42 up and 39 down) or with both groups (2 down). Reverse transcription-quantitative PCR (RT-qPCR) was used to confirm miRNA differential expression. Correlation between RNA sequencing and RT-qPCR data was demonstrated for 3 miRs from 14 tested: mir-146a-5p (r=0.56,
P
<10
-4
), miR-338-3p (r=0.91,
P
<10
-16
) and mir-374a-3p (r=0.57,
P
<10
-4
). The best correlated DE miRNA, miR-338-3p was down-regulated by 76% (
P
<10
-4
) and uniquely associated with CKD. TargetScan predicted that the up-regulated mRNA encoding glutathione peroxidase 3 (
GPX3
, 1.57 fold,
P
<0.001) to be a miR-338-3p target. Both miR-338-3p and
GPX3
were found to be highly expressed in human aortic endothelial cells (HAECs) by RT-qPCR. Target prediction was validated by showing using RT-qPCR that
GPX3
expression was up-regulated 1.73±0.28 fold in HAECs transfected with anti-miR-338-3p (
P
<0.05).

Conclusion:
miR-338-3p down-regulation was found in small arteries, uniquely associated with CKD.
GPX3
, which may be important for preservation of endothelial function, is a miR-338-3p potential target. miR-338-3p and associated target may play a role in vascular remodeling in CKD.
Abstract P123: Plasma Endocan Level as a Predictor of Cardiovascular Composite Outcomes in Patients With End-Stage Renal Disease
Authors: Lee Ji Yung , Ihm Hyung Seok , Hwang Hyeon Seok , Jeong Kyung Hwan , Ihm Chun Gyoo , Lee Sang Ho , Lee Tae Won , Kim Yang Gyun , Moon Ju Young
Published: December 2018

Background:
Endocan, a proteoglycan which is a potential biomarker of endothelial dysfunction, has been shown to be associated with increased cardiovascular risk. We investigated plasma levels of endocan in patients with end-stage renal disease (ESRD) on hemodialysis to predict the risk of cardiovascular diseases.

Methods:
A total of 400 adult patients with ESRD undergoing hemodialysis were prospectively enrolled in 4 tertiary hospitals of South Korea from June 2016 to May 2018. They were observed for development of the cardiovascular composite outcomes. We compared clinical characteristics and the plasma levels of endocan between 47 patients with cardiovascular composite outcomes and 353 control patients without cardiovascular composite outcomes, and developed the predictive markers of cardiovascular diseases using Cox proportional-hazard analysis.

Results:
Previous histories of diabetes, acute coronary syndrome, arrhythmia and congestive heart failure were higher in ESRD patients with the cardiovascular composite outcomes than control patients. Patients with cardiovascular composite outcomes showed lower levels of hemoglobin, albumin, and high-density lipoproteins compared with control patients. Higher level of plasma endocan and white blood cells were associated with patients with cardiovascular composite outcomes. Cox proportional-hazard analysis showed that previous histories of diabetes and plasma level endocan were significantly associated with cardiovascular composite outcomes: hazard ratios were 2.1 (95% confidential interval (CI), 1.1 to 4.3) (
p
= 0.03) for diabetes and 15.4 (95% CI, 3.2 to 75.2) (
p
= 0.001) for endocan (log pg/mL). The patients with level of endocan of 2.96 log pg per mL or more showed significantly higher cumulative incidence of the cardiovascular composite outcomes in Kaplan-Meier curve (
p
= 0.03).

Conclusions:
Plasma Endocan level can a useful biomarker for prediction of cardiovascular diseases in patients with ESRD on hemodialysis.
Abstract P125: Role of Circulating Endothelial Progenitor Cell Levels Predict Cardiovascular Events in Patients on Maintenance Hemodialysis
Authors: Ihm Hyung-seok , Ihm Chun-gyoo , Lee Tae-won , Jeong Kyung-hwan , Hwang Hyeon-seok , Lee Sang-ho , Kim Woo-shik , Lee Ji yung
Published: December 2018

Background:
The number of circulating endothelial progenitor cells (EPCs) has been identified as a surrogate biologic marker for vascular function and cumulative cardiovascular (CV) risk in the general population. Patients with end-stage renal disease (ESRD) on hemodialysis (HD) have markedly decreased EPC counts and function. We hypothesized that the number of circulating EPCs predicts death from all causes and CV events in patients with ESRD on HD.

Methods:
We quantified the EPCs in blood samples from 70 patients with ESRD on HD. Circulating EPCs were counted by flow cytometry as the number of CD45 low CD34 + VEGFR2 + cells. Death from all causes and CV events served as outcome variables over a median follow-up period of 20 months

Results:
It has been postulated that the number of circulating EPCs at baseline ranged from 1 to 350 cells per 200 μl, with a mean of ± standard deviation (SD) of 26.0 ± 48.2 cells per 200 μl. The median, lowest and highest tertiles of EPC counts were 11.0, 9.0, and 17.0 cells/200 μl, respectively. Patients with the lowest tertile EPC counts had significantly higher rates of CV events, but mortality was similar between the two groups. After adjusting for these risk factors, HbA1c and the lowest tertile EPC count remained as independent predictors of CV events. A cutoff value of 9.5 cells per 200 μl maximized the power of the EPC count to predict future CV events as determined by ROC curve analysis.

Conclusions:
Reduced circulating EPC counts independently predicted CV events in 70 patients with ESRD on maintenance HD. Circulating EPCs may play a role in vascular repair, thereby affecting the clinical course of CV events.
Abstract P128: Intensive Blood Pressure Treatment and Renal Outcomes in Diabetic Patients
Authors: Wang Ling , Rayamajhi Supratik , Wang Donna
Published: December 2018

Background:
There is no certain evidence to support a strategy of lowering systolic blood pressure in persons with type 2 diabetes. We conducted a post-hoc analysis on the data set from Action to Control Cardiovascular Risk in Diabetes blood pressure trial (ACCORD-BP) to test the hypothesis that intensive blood pressure treatment prevents or alleviates adverse renal outcomes among diabetic patients.

Methods:
Patients were randomly assigned to a systolic blood pressure (SBP) treatment target of <120 (intensive treatment) or <140 (standard treatment). To rule out the effect of glycemic treatment, we only included patients with standard glycemic control (glycated hemoglobin of 7.0-7.9%) that resulted in 1,184 patients on intensive arm and 1,178 patients on standard arm. Four renal outcomes at the end of the study are considered: 1) Doubling of serum creatinine (SCr) or >20 mL/min decrease in eGFR; 2) UAlb>=300; 3) Renal failure or End-Stage Renal Disease (dialysis) or SCr>3.3; 4) UAlb>=30. Generalized Estimating Equation method is employed.

Results:
After 1 year, the mean SBP was 120.8 in intensive group and 134.6 in standard group. SCr doubling or >20 mL/min decrease in eGFR attributed to intensive treatment occurred in 701 of the 1184 participants in the intensive-therapy group (57.0%) and 529 of the 1178 participants in the standard-therapy group (44.9%) (OR=1.83, 95% CI= 1.76-1.90). We further categorized BP over time into three tiers, by low (SBP<120 and DBP <70), medium (SBP=120-140 and DBP=70-85) and high (SBP>140 and DBP>85). Both high SBP (>140) and medium SBP (120-140) as compared to low SBP (<120) level, significantly reduced the chance of serum creatinine doubling or >20 ml/min decrease in eGFR (OR=0.79, p<0.0001). There are no significant differences in other three renal outcomes between intensive and standard group patients.

Conclusion:
In patients with type 2 diabetes, targeting a systolic blood pressure of less than 120, as compared with less than 140, increased the chance of having Scr doubling or >20 mL/min decrease in eGFR. These results, consistent with our previous study using SPRINT data from non-diabetic patients, provide evidence against the notion that intensive blood pressure treatment leads to beneficial renal outcomes in diabetic patients.
Abstract P131: Transdermal Detection of Low Concentrations of Hydrogen Sulfide
Authors: Wiest Elani F , Rios Lynnette , Friedrichsen Debra , Silaski Grayson , Mowry Curtis , Shekarriz Reza , Kanagy Nancy
Published: December 2018

We report here a novel approach to measure circulating hydrogen sulfide (H
2
S) non-invasively as a potential way to diagnose and monitor endothelial dysfunction and peripheral artery disease (PAD). PAD is a life-threatening condition caused by arterial constriction and obstruction of blood flow leading to limb ischemia. Current methods to diagnose and monitor PAD lack sensitivity, are expensive, and technically difficult. Recent studies indicate that decreased H
2
S production is an underlying cause of PAD. Also, reduced plasma H
2
S correlates with endothelial dysfunction in individuals with untreated hypertension, diabetes, sleep apnea and other cardiovascular diseases. The TAGS device was designed to measure transdermal H
2
S to test the hypothesis that the diffusion rate (and therefore gas phase concentration) of H
2
S is directly proportional to dermal blood flow. Healthy volunteers between the ages of 21-65 were recruited. Exclusion criteria included subjects currently treated for hypertension, hyperlipidemia, and diabetes. Smokers and pregnant women were also excluded. We demonstrate that H
2
S can be detected in healthy volunteers (n=11) at 10.8 part per billion (ppb). Interestingly, there is a positive correlation between age and TAGS readings in male volunteers (r=0.614; n= 6) and a negative correlation in female volunteers (r=0.482, n=5). The TAGS device has the potential, therefore, to serve as a more sensitive, economical, and easy to use diagnostic tool to detect H
2
S and predict dermal blood flow.
Abstract P134: Sex Differences in the Effects of Chronic Muscadine Grape Extract Treatment on Cardiac Function Parameters and Aortic Pulse Wave Velocity (PWV) in Hypertensive Transgenic (mRen2)27 Rats
Authors: Ryalat Fatima , Cruz-Diaz Nildris , Varagic Jasmina , Gallagher Patricia , Tallant Ann , Diz Debra , Yamaleyeva Liliya
Published: December 2018

Muscadine grape extract (MGE) polyphenolics have potential beneficial cardiovascular effects. Whether the effects are sex-specific during progression of hypertension is unknown. We measured aortic arch PWV and cardiac function parameters using transthoracic echocardiography (Vevo LAZR 2100) after 15 weeks of MGE-treatment (0.2 mg polyphenolics /mL in drinking water) in transgenic hypertensive (mRen2)27 rats (n = 4 each, control male and female, MGE-treated female; n = 3 MGE-treated male) starting at 15 weeks of age. As expected, systolic blood pressure (SBP) measured via tail cuff after acclimation was higher in males than females at 15 weeks of age (179 ± 3 vs. 169 ± 3 mm Hg, p = 0.04). By 30 weeks of age, SBP did not differ between male and female rats (171 ± 4 vs. 171 ± 3 mm Hg). Body weight was higher in male than female rats (589 ± 8 vs. 326 ± 5 g, p < 0.0001); thus, cardiac output (CO), stroke volume (SV) and left ventricular mass (cLVM) were normalized to body weight. Systolic cardiac function was lower in male compared with female rats: SV (0.34 ± 0.04 vs. 0.59 ± 0.03 μL/g, p < 0.001); CO (0.13 ± 0.001 vs. 0.26 ± 0.02 mL/min/g, p < 0.001); ejection fraction (46 ± 4 vs. 74 ± 5 %, p < 0.05); fractional shortening (24 ± 3 vs. 45 ± 4 %, p < 0.05). In contrast, there was no difference between male and female rats in cLVM (2.35 ± 0.20 vs. 2.85 ± 0.14); heart rate (367 ± 19 vs. 432 ± 22 bpm); E/e’ (12 ± 2 vs. 14 ± 0.4), or PWV (2.74 ± 0.21 vs. 2.85 ± 0.09 m/s). Sex differences in systolic cardiac function remained after 15 weeks of MGE treatment and MGE did not alter SBP or diastolic function (E/e’). However, aortic PWV was 36% lower in MGE treated males compared to control males (1.75 ± 0.13 vs. 2.74 ± 0.21 m/s, p < 0.005), which was also significantly lower than MGE treated females (2.40 ± 0.05 m/s, p < 0.05). Aortic PWV was 16% lower in MGE-treated females than control females, which did not reach significance (2.40 ± 0.05 vs. 2.85 ± 0.09 m/s). In conclusion, MGE intake improves arterial stiffness in male (mRen2)27 rats independent of differences in blood pressure or cardiac function. That aortic PWV in female rats did not derive benefit implies a potentially unique therapeutic profile for MGE in targeting mechanisms important for arterial stiffness in a sex-specific manner.
Abstract P137: Arterial Stiffness in Diabetic Patients on Oral Antidiabetic Agents V/s a Combination of Oral Antidiabetic Agents Plus Insulin
Authors: Cigarroa Joaquin , DiCicco Ignacio , Pham Son , Hecht Joan , Chilton Robert , Oliveros Rene
Published: December 2018

In a previous publication (Journal of the American Society of Hypertension 2014;8(4):e35-e36 we did a comparison study of arterial stiffness in diabetic pts taking oral antidiabetic agents (ODA) v/s those taking Insulin. The aim of the present study was to compare aortic stiffness determined from resting and exercise pulse pressure (PP) in a group of pts taking ODA v/s a group of pts taking a combination of ODA and insulin. We studied 86 diabetic pts referred to us for the performance of a maximal treadmill exercise test as part of a workup for chest pain. They were divided into 46 pts taking ODA (group 1) v/s 46 pts taking a combination of ODA plus insulin (group 2). Their age, heart rate(HR) both rest and exercise, PP both rest and exercise, exercise duration in seconds and HbA1C are noted in the table. There was no significant difference in the HR and PP both at rest and during exercise between both groups. Also, no significant difference was found in their ages and exercise duration although group 2 pts were older, had greater PP both at rest and exercise and had shorter exercise duration. The only significant difference was that goup 2 had a sinificantly higher A1C than group 1(p<0.003) reflecting the severity of their diabetes. We conclude from this small study, that the addition of Insulin to oral antidiabetic agents did not affect aortic stiffness as determined by PP which is a surrogate for other measures of aortic stiffness.
Abstract P138: The Arterial Wall of the Central Vascular Compartment a Novel Target of Iron-Overload Disorders: A Preliminary Study With Experimental Correlation in Animals
Authors: Duarte Mariano , Aquieri Analia , Coyle Javier , Yaryour Claudio , Gonzalez Ballerga Esteban , Reyes Toso Carlos
Published: December 2018

In Hemochromatosis (H) iron is stored in tissues, causing organ injury and failure. Patients with H have a shorter life expectancy and an increase in CVR. Our group could prove that arterial stiffness was increased and they had added endothelial dysfunction. After that we also were able to demonstrate that both improve by reducing iron levels with phlebotomies. Due to these findings we ask how iron-overload made it (getting inside or more likely inducing inflammation). Anyway either directly or indirectly AW could be a target of iron-overload. To test this hypothesis we did an experimental correlation by using a known model of H in rats and we analize histologically the AW Twelve young rats (3 months old), an average weight of 300 gr, fed as usual, in individual cages with dark light cycles of 12 h and controlled temperature (22±3°C), were ramdonly divided in 2 groups: 1) Iron-overoload group (IOG). They were daily administrated intraperitoneally 10 mg of elemental iron (0,1 ml of dextran iron, Venofer®) 12 injections in total 2) Control group (CG) the same injections scheme but with placebo. The handlings of animals was under the norms of the bioterium and standards designed for that purpose. At the end of experimental period (8 weeks) animals were sacrificed and studied. The histoarchitecture of the proximal aortic wall (fixed in 4 % formalin, paraffin-embedded, sectioned into thin slices, H&E, Masson′s trichrome and VVG staining) showed no significant differences in the histology (H&E) between IOG and CG. No evidence of iron inside the AW. On the other hand inflammatory component was significantly increased in IOG compared to CG, perivascular lymphocyte count 5-12/0 IOG/CG, perivascular macrophages, 3-5/0 IOG/CG. Elastic fibers were not fragmented but become thin and lost their disposition in IOG. In conclusions we can affirm that AW could represent a novel target in H and could may the nexus to cardiovascular disease. The indirect mechanism (inflammation and elastic fibers) explains why previously improved after phlebotomies. Our findings supports future research including inflammatory mediators and their tissue expression, and extend them to others iron overload disorders.
Abstract P142: Aberrant Notch2 Signaling in the Vascular Endothelium Leads to Increased Proliferation and Migration Operant in Pulmonary Arterial Hypertension
Authors: Sahoo Sanghamitra , Sembrat John C , Rojas Mauricio , Goncharova Elena A , Pagano Patrick J
Published: December 2018

Background:
Pulmonary arterial hypertension (PAH) develops when pulmonary vascular resistance rises sharply, leading to right heart failure and death. Most studies have focused on attenuating lung vessel constriction; however, little is known about the mechanisms controlling intimal proliferation and luminal obstruction. Numerous studies suggest that the Notch signaling influences vascular cell proliferation and migration, hallmark features of deleterious remodeling. Despite this, a role for Notch2 in PAH is entirely unknown. In this study, we hypothesize that hypoxia-induced repression of lung endothelial Notch2 results in increased EC proliferation/migration in PAH.

Methods and Results:
In a hypoxia model replicating the PAH phenotype in vitro, HPAECs were subjected to 24 hrs hypoxia (1% O
2
) compared to normoxia (21% O
2
). Hypoxia attenuated Notch2 mRNA (0.64±0.02-fold vs. normoxia, p<0.0001) but did not affect other endothelial Notch receptors - Notch1 and 4. Furthermore, hypoxia decreased Notch2 protein levels (0.65±0.05-fold vs. normoxia, p<0.0001). q-RTPCR of lysates of pulmonary arteries from PAH patients showed marked reductions in Notch2 mRNA (0.30±0.08-fold vs. control patients,
p
<0.05). Notch2 siRNA under normoxic conditions increased EC migration in a “wound healing-scratch” assay (1.5±0.01-fold vs. scrambled control, p<0.05) and EC proliferation (1.2±0.07-fold vs. scrambled control,
p
=0.05) measured by CyQuant assay. Western blot analysis of signaling intermediaries showed decreased cell cycle inhibitor p21
cip
(0.54±0.05 vs. scrambled control, p<0.05), and increased levels of phospho-ERK1/2 (1.3±0.04-fold vs. scrambled control, p<0.001).

Conclusions and Discussion:
In the present study, we found that hypoxia suppressed Notch2 expression in the endothelium which appears to derepress ERK1/2 and downregulate p21

cip

leading to EC proliferation and migration. These data are consistent with a key modulatory role for Notch2 dysfunction in EC proliferation/migration and vascular remodeling in PAH. Data from human patients substantiates this dysfunction. The current study identifies a new signaling axis mediated by endothelial Notch2 that could contribute to pathophysiological changes associated with human PAH.
Abstract P143: Nox1 Elicits Endothelial Cell Proliferation in a CREB- and Ref-1- Dependent Manner
Authors: de Jesus Daniel , Li Yao , DeVallance Evan , Pagano Patrick
Published: December 2018

Pulmonary arterial hypertension (PAH) is a complex and progressive disorder, characterized by an increase in vascular remodeling, a rise in pulmonary arterial pressure and right heart failure. Recently, we and others showed that BMP antagonist Gremlin1 elicits pulmonary endothelial cell (EC) proliferation in response to hypoxia and that Gremlin1 haplodeficiency attenuates PAH. NADPH oxidase (Nox)-derived reactive oxygen species (ROS) seem to play a pivotal role in PAH. However, the mechanisms by which ROS propagates the disease are unclear. Transcription factor CREB, which is activated by hypoxia, ROS, and interaction with Ref-1 (redox factor 1), mediates gene transcription related to proliferation and vascular remodeling. We postulated that Nox1/Ref-1-mediated CREB activation leads to Gremlin1-induced EC proliferation. Human pulmonary arterial EC (HPAECs, Lonza) were subjected to 24 hr hypoxia (1% O
2
vs. normoxia - 21% O
2
) to mimic the EC phenotype in PAH. Hypoxia upregulated Nox1 protein level (1.40±0.09-fold, p<0.05) and H
2
O
2
production level/mg protein (2.20±0.21-fold, p<0.0001) vs. normoxia. siNox1 (Nox1 gene silencing) abolished hypoxia-induced Nox1, and Nox1 inhibitor, NoxA1ds, decreased hypoxia-promoted H
2
O
2
. Moreover, siNox1 decreased hypoxia-induced pCREB (1.74±0.22-fold v. normoxia, vs. 1.07±0.08 siNox1+hypoxia, p<0.05), and siCREB decreased hypoxia-induced Gremlin1 (1.83±0.16 hypoxia vs. 0.35±0.01 siCREB+hypoxia, p<0.05). Further, hypoxia augmented nuclear pCREB/Ref-1 interaction (1.812±0.207-fold vs. normoxia, p<0.05) and pCREB association with its DNA binding motif. siRNA for Ref-1 impaired hypoxia-induced pCREB DNA binding (1.96±0.06 hypoxia vs. 0.815±0.219 siRef-1+hypoxia, p<0.01). Moreover, siCREB decreased hypoxia-induced EC proliferation (1.40±0.050 hypoxia vs. 0.96±0.04 siCREB+hypoxia). Finally, preliminary data show pCREB binding to the Gremlin1 promoter. Taken together, our data support a previously unidentified redox signaling pathway by which Nox1-derived ROS promote Gremlin1 expression and EC proliferation in PAH.
Abstract P145: The Bacterial Metabolite Trimethylamine (TMA), but Not Trimethylamine N-Oxide (TMAO), Causes Vascular Contraction
Authors: Restini Carolina B , Fink Gregory D , Watts Stephanie W
Published: December 2018

Circulating levels of TMAO are elevated in humans with cardiometabolic diseases. The gut microbiota-initiated TMA/flavin-containing monooxygenase 3 (FMO3)/TMAO pathway has been identified as a potential modulator of host cardiometabolic phenotypes. Like TMA/TMAO, FMO3 expression is positively correlated with body mass index and high-fat diet-induced obesity We hypothesized that TMA/TMAO stimulate vascular contraction which could be modified by perivascular adipose tissue (PVAT) expression of FMO3. Our model was the isolated thoracic aorta with (+) or without (-) PVAT, in presence (+) or in absence (+) of endothelium (E) of male Sprague Dawley rat to perform tissue bath studies to measure isometric tone using a large range of TMA/TMAO concentrations: 1 nM-10
-1
M. Immunohistochemistry (IHC) studies were done to identify the presence of FMO3 in aorta. TMA and TMAO, at any concentration, did not relax half maximally contracted tissues when compared to vehicle control. TMA and TMAO elicited arterial contraction independent of PVAT or E status. Interestingly, contractions stimulated by TMA were significantly higher (% PE max 123.7±15.6%) than by TMAO (% PE max 38.3±15.5%). IHC studies revealed FMO3 in aortic PVAT, but the FMO3 inhibitor methimazole (100 μM) did not affect aortic contraction stimulated by TMA (+PVAT). The L type Ca2+ channel antagonist nifedipine (100 nM), reduced the TMA-induced contraction (% PE max: +PVAT: 73±9%; -PVAT: 63±6.7%), compared to vehicle (+PVAT: 149.8±18%; -PVAT: 111.8±9%). These results support that TMA can induce arterial contraction with higher potency and efficacy compared to TMAO, independent of the conversion to TMAO. The contractile mechanism is ≈ 50% dependent of Ca
2+
influx through L-type VOC. Considering the high concentration needed to achieve the contraction it is unlikely that increasing vascular tone is a mechanism that could contribute to CVD but dependence of contraction on Ca
2+
channel activation suggests specific activity of TMA in the vessel wall that warrants further study.
Abstract P148: Muscarinic Acetylcholine Receptors are Localized to Primary Endothelial Cilia and Involved in Nitric Oxide Signaling
Authors: Saternos Hannah C , Gibson Hayley , Meqdad Mahmood , AbouAlaiwi Wissam A
Published: December 2018

Primary cilia are considered sensory hubs housing a variety of mechanosensory proteins, chemosensory receptors, and ion channels to translate extracellular stimuli into an intracellular biochemical signals such as the synthesis and release of nitric oxide (NO). Malformations in cilia structure or defects in cilia function lead to ciliopathies such as Polycystic Kidney Disease (PKD) and hypertension. Muscarinic receptors 1 and 3 play an essential role in regulating cardiovascular function by mediating both dilation and constriction in the vasculature. However, nothing is known about the relationship between primary cilia and muscarinic receptors. Supported by our exciting discovery of the mAChRs in the cilia, we hypothesize that

primary cilia in the vascular system play important roles in regulating blood pressure through NO biosynthesis.

Our objective is to unravel the mechanism by which cilia dysfunction contribute to hypertension and to introduce cilia as a novel therapeutic target for PKD. To explore the relationship between mAChRs and primary cilia, the effects of muscarinic modulators on cilia length and function in wildtype, and cilia mutant endothelial cells,
Pkd1
–

/

–
(dysfunctional cilia) and
Tg737

orpk/orpk

(cilia-less) were examined. We show that both AChM1R and AChM3R localize to primary endothelial cilia. AChM1R and AChM3R activation lead to a significant increase in cilia length in endothelial cells treated with cdd0102a, an AChM1R and AChM3R agonist (2.84±0.02 vs. 3.47± 0.04 for wildtype, 2.31± 0.03 vs. 2.53± 0.03 for
Pkd1
–

/

–
, and 0.024 ± 0.005 vs. 0.3 ± 0.004 for
Tg737

orpk/orpk

) compared to control cells. Treatment with pirenzepine, an AChM1R antagonist, led to a significant decrease of cilia length (2.66± 0.02 vs. 2.48± 0.03 in wildtype, and 2.12± 0.02 vs. 1.93± 0.02 in
Pkd1
–

/

–
) compared to control cells. Treatment with cdd0102a also significantly upregulated the expression of AChM1R, AChM3R and phosphorylated eNOS in wildtype and
Pkd1
-

/

-
cells. More importantly, cdd0102a treatment rescued NO response in
Pkd1
-

/

-
cells in response to fluid shear stress. Our data clearly suggest that modulating cilia sensory function could have a positive influence on nitric oxide generation and blood pressure regulation.
Abstract P150: Chronic Exercise Provides Antihypertensive Effects With Upregulation of Oxidative Stress-Induced Nitric Oxide Upregulation in The Kidney of High-Fructose Fed Rat
Authors: Hu Gaizun , Xu Lusi , Kohzuki Masahiro , Ito Osamu
Published: December 2018

Object:
The fructose consumption was reported to correlated with increase of oxidative stress, which thereby impair nitric oxide system. Chronic exercise (Ex) can provide antihypertensive effects though nitric oxide upregulation. The mechanisms of Ex in nitric oxide upregulation are various and remains unclarified.

Method:
A total of 24 Sprague Dawley rats at 5-week age were allocated to 3 groups as controlled diet group (CON), high-fructose diet group (HFr) and HFr underwent 12-week chronic exercise (HFr-Ex). The systolic blood pressure (SBP) and urinary albumin excretion were taken every 2 weeks. At the final week, after the insulin tolerance test, the rats were euthanized. The plasma and kidneys were obtained. The plasma and renal oxidative stress markers were measured. The nitric oxide related parameters including renal endothelial nitric oxide synthase (eNOS) expression were measured.

Result:
The SBP was significantly increased in HFr than CON, which Ex significantly attenuated the SBP elevation (115 vs. 141 mmHg, p<0.01). Ex reversed albuminuria induced by HFr without alteration in creatinine clearance ratio (474 vs. 1446μg/day, p<0.01). Ex significantly lowered uric acid, and attenuated the insulin resistance (uric acid, 1.41 vs. 2.05 mg/dL, p<0.01; HOMA-IR, 0.64 vs. 0.50, p<0.01). Ex significantly inhibited renal cortex xanthine oxidase activity (XO), meanwhile the NADPH oxidase activity (NADPH) was even enhanced (XO, 15.7 vs. 11.8 units/mg protein, p<0.05; NADPH, 897 vs. 1370 c.p.m, p<0.01). The hydrogen peroxide was not changed. Renal eNOS expression was increased in HFr (228%, p<0.01), and Ex significantly enhanced the expression higher than HFr (316%, p<0.01). The phosphorylation of eNOS in serine 1177 were inhibited by HFr and reversed by Ex (87%, p<0.01). AMPK was not changed by HFr, and Ex significantly increased AMPK expression (266%, p<0.01). The phosphorylation of AMPK at threonine 172 were increased by HFr (306%, p<0.01), and Ex even enhanced the expression (473%, p<0.01).

Conclusion:
Ex provided antihypertensive effects with nitric oxide upregulation. The mechanism of nitric oxide upregulation may mediate with the improve of phosphorylation and alteration in oxidases activities.
Abstract P156: Toll-Like Receptor 2 Deficiency Exacerbates Deoxycorticosterone Acetate-Salt-Induced Hypertension
Authors: Uijl Estrellita , Severs David , Danser A.H. Jan , Zietse Robert , Hoorn Ewout J
Published: December 2018

The early stage of hypertension increases so-called damage-associated molecular patterns (DAMPs). In turn, DAMPs can activate Toll-like receptors (TLRs) and thereby exacerbate hypertension. Although various TLRs have been implicated in the pathogenesis of salt-sensitive hypertension, the role of TLR2 and TLR4 remains unclear. Here, we hypothesize that TLR2 or TLR4 deficiency protects against salt-sensitive hypertension. To address this, we treated wild-type (WT, n=3), TLR2
-/-
(n=5) and TLR4
-/-
(n=5) mice with deoxycorticosterone acetate and 0.9% NaCl as drinking water (DOCA-salt) for four weeks. Baseline telemetric mean arterial blood pressure (MAP) was slightly but non-significantly higher in TLR4
-/-
(108±2 mmHg) and TLR2
-/-
mice (111±3 mm Hg) compared with WT mice (99±5 mmHg). MAP peaked similarly after two weeks in WT and TLR4
-/-
mice (111±1 vs. 116±3 mmHg; both P<0.05 vs. baseline), but reduced after four weeks (108±3 vs. 110±2 mmHg). In contrast, TLR2
-/-
mice displayed a much greater rise in MAP to DOCA-salt and remained hypertensive after two weeks (134±5 mmHg) and after four weeks (128±4 mmHg; P<0.0001 vs. baseline; P<0.05 for delta MAP vs. TLR4
-/-
). Kidney sodium transporter analysis showed that the phosphorylated (activated) form of the sodium-chloride cotransporter (NCC) was 2.4-fold higher in TLR2
-/-
compared with WT mice (P<0.05). In wire myographs of iliac arteries, angiotensin II-induced constriction was non-significantly lower in iliac arteries of TLR4
-/-
mice (53±9%) than in TLR2
-/-
(84±9%) and WT mice (84±15%). Addition of the nitric oxide synthase blocker L-NAME increased the contractile response to Ang II in WT mice, but not in TLR2
-/-
mice, indicating NO deficiency in the latter mice. The Ang II type 2 (AT2) receptor blocker PD123319 blunted the effects of Ang II in both WT and TLR2
-/-
mice, suggesting that DOCA-salt upregulates constrictor AT2 receptors, but not in TLR4
-/-
mice. In conclusion, TLR2 deficiency unexpectedly exacerbates experimental salt-sensitive hypertension. This effect appears to involve increased renal sodium reabsorption through NCC, decreased vascular availability of nitric oxide and a constrictive rather than a vasodilatory response to AT2 receptor stimulation.
Abstract P161: Effect of Rapamycin on the Expression of T Regulatory Cells, Foxp3, and Toll-Like Receptors in the Kidneys of
Npr1
Gene-Knockout Mice
Authors: Gogulamudi Venkateswara R , Mani Indra , Subramanian Umadevi , Pandey Kailash N
Published: December 2018

The alterations in the inflammatory system contribute to the pathogenesis of hypertension leading to end-organ damage. Guanylyl cyclase/natriuretic peptide receptor-A (GC-A/NPRA) gene (
Npr1
) disruption elevates blood pressure and activates the immunogenic responses in mice. The goal of this study was to determine the effect of rapamycin on the expression of T regulatory (Treg) cells, Foxp3, and Toll-like receptors (TLRs) in
Npr1
gene-disrupted and -duplicated mice. The adult 0-copy (

Npr1
-/-

), 2-copy (
Npr1
+/+
), and 4-copy (

Npr1
++/++

) female mice were pre-treated with rapamycin (1mg/kg/day). Kidneys, spleen, and blood were collected, qRT-PCR, Western blot, immunofluorescence, and flow cytometry analyses were done to assess the expression of Tregs, Foxp3, and TLRs. The disruption of
Npr1
led to significant reductions in Foxp3
+
mRNA expression in 0-copy (77.5%) and 1-copy (71.5%) mice but not in 2-copy and 4-copy mice. Similarly, CD25
+
expression was reduced in 0-copy mice by 75% and in 1-copy by 60% compared to 2-copy and 4-copy mice. In contrast, the total CD4
+
count was up-regulated by 40% in 0-copy and 31% in 1-copy mice. Treatment with rapamycin up-regulated Foxp3
+
cells in 0-copy (17.38%,
P
<0.001) and 1-copy (8%,
P
<0.05) mice, and CD25
+
T cell levels were increased by 62.4% (
P
<0.001) in 0-copy mice and 38.6% (
P
<0.05) in 1-copy mice compared to vehicle-treated control mice. In contrast, CD4
+
cell expression was significantly reduced by 15.4% (
P
<0.001) in 0-copy and 12.2% (
P
<0.05) in 1-copy mice. Interestingly, rapamycin significantly (
P
< 0.001) increased renal Foxp3
+
protein expression to 85% in 0-copy mice and 68% in 1-copy mice compared to untreated controls. The expression of TLR2/TLR4 was significantly increased by 3.4- to 4.5-fold in 1-copy and 0-copy mice than 2-copy and 4-copy mice; however, rapamycin greatly decreased the expression of both TLRs in both 1-copy and 0-copy mice. In summary, rapamycin treatment upregulated the expression of Foxp3
+
, CD25
+
cells, and TLRs while suppressing pathogenic CD4
+
T cell expression in 0-copy and 1-copy mice kidneys. The present results implicate that
Npr1
and immune cell interactions could provide new therapeutic targets for the treatment and prevention of hypertension and kidney dysfunction in humans.
Abstract P164: Inflammatory Cell Populations in a Mouse Model of Cardiorenal Syndrome
Authors: Koid Suang Suang , Shand Francis H , Ueha Satoshi , Matsushima Kouji , Shimosawa Tatsuo
Published: December 2018

Objective:
Renal dysfunction frequently coexists with heart disease, manifesting in a condition called cardiorenal syndrome (CRS). CRS results in accelerated failure of the heart and kidney via a complex combination of hemodynamic, neurohormonal, immunological and biochemical feedback pathways. Preliminary evidence suggests that inflammatory cells such as monocytes and macrophages may contribute to the pathogenesis of CRS. However, animal models of CRS remain limited and the roles of specific inflammatory cell subsets in CRS remains unclear. Here, we describe the establishment of a mouse model of CRS.

Methods:
Three-week-old C57BL/6J mice were subjected to sham uninephrectomy and normal-salt (SHAM/NS) diet or left uninephrectomy and high-salt (UNIX/HS) diet. After 4, 6 or 8 weeks, urine was collected for salt and protein analyses. The heart and kidney were collected for histochemical staining and analyses of inflammatory cell distribution by flow cytometry.

Results:
UNIX/HS mice had a higher kidney weight/body weight ratio and higher water intake than that of SHAM/NS mice. As early as 4 weeks post-surgery, UNIX/HS mice had higher urine creatinine levels (0.22 ± 0.02 mg/day c.f. 0.13 ± 0.02 mg/day) and higher proteinuria (0.46 ± 0.18 mg/day c.f. 5.37 ± 1.19 mg/day) than SHAM/NS mice. These results suggest the development of kidney dysfunction. Histological analysis revealed increased fibrosis in the heart and kidney at 8 weeks after UNIX/HS. Using flow cytometry, we characterized changes in the inflammatory cell populations in the heart and kidney that occur as CRS progresses. A significant increase in Ly6C
hi
monocyte (0.29 ± 0.06% c.f. 0.15 ± 0.02% of mean 1.0 x 10
6
total live cells) and neutrophil (1.00 ± 0.27% c.f. 0.41 ± 0.09%) proportions in the heart was observed 8 weeks after UNIX/HS.

Conclusion:
Therapeutic strategies targeting inflammatory signaling pathways and modulating specific inflammatory cell populations may reduce organ damage and improve prognosis in patients with CRS.
Abstract P167: Altered Gut Microbiome Contributes to Metabolite Biomarkers in Patients With Pulmonary Hypertension
Authors: Kim Seungbum , Walejko Jacquelyn M , Rigatto Katya , Jordan Ray , Shapiro Brian , Pepine Carl J , Richards Elaine M , Raizada Mohan K
Published: December 2018

Objectives:
Pulmonary arterial hypertension (PAH) is a cardiopulmonary disease without effective cure. The objective of this study is to test the hypothesis that PAH patients have a distinct gut microbiome composition acting as a driving force for the increased PAH metabolic biomarkers, acyl carnitines and urate.

Methods:
Fecal samples from 18 type 1 PAH patients (mean pulmonary arterial pressure [mPAP] 57.4±16.7 mmHg) and 13 reference subjects in Brazil were collected for comparison of microbiome using shotgun metagenomics. Plasma samples from 33 type I PAH patients (mPAP 51.7± 13.5 mmHg) and 19 reference subjects in the USA were used for untargeted metabolomic analysis.

Results:
More than 80 bacterial taxa were differentially enriched in the microbiomes of the PAH cohort. Random forest machine-learning algorithm predicted PAH patients with 82.4% accuracy from their gut microbiome composition. The most increased taxa in the PAH microbiome were trimethylamine (TMA) producing bacteria like Collinsella (P=0.0037). The PAH metabolome also showed increases of gut microbiome-derived short chain acyl carnitines like isubutyrylcarnitine (P=0.010) and valerylcarnitine (P=0.030), which are the precursors of TMA. In addition, increased urate (P=0.0004) and allantoin (P=0.027) in PAH plasma correlated with significantly increased enzymes producing urate in the PAH microbiome (purine metabolism (P=0.017) and xanthine oxidase (P<0.05)).

Conclusion:
These findings suggest that the random forest modeling of the gut microbiome composition can be used for prediction of PAH and that the unique gut microbiome of PAH influences acyl carnitines and urate metabolism to contribute to pathophysiology.
Abstract P169: Characterization of the Peak Exercise Blood Pressure Response During Cardiopulmonary Exercise Testing in African American and Caucasian American Men
Authors: Sabbahi Ahmad , Arena Ross , Kaminsky Leonard , Myers Jonathan , Fernhall Bo , Phillips Shane
Published: December 2018

Objective:
It has well been established that African American (AA) individuals have higher overall incidence and prevalence of hypertension compared to their Caucasian American (CA) counterparts. However, the maximum BP response of AA to exercise has not been characterized. The objective of this study is to investigate this peak BP response across both ethnic groups.

Patients and Methods:
A total of 9363 apparently healthy men from the FRIEND database who underwent maximum cardiopulmonary exercise tests on a cycle ergometer were included in this analysis. All subjects had a respiratory exchange ratio (RER) of ≥ 1.0 and had no reports of diagnosed disease.

Results:
There were significant differences between age and ethnic groups for peak SBP and peak DBP starting from the 30-39 years age group to the 60-69 years age group (
P
<.001). There were also significant differences in delta SBP and DBP between age and ethnic groups (
P
<.001).

Conclusion:
These data indicate that the physiologic BP response to exercise in BA men is exaggerated compared to their WA counterparts, irrespective of baseline resting BP values.
Abstract P172: A Correlative Study of Cuff Hypertension and Anthropometric Variables in Obese and Non Obese Subjects
Authors: Saini Abhishek , Krishnamurthy Ajoy , Bohra Chandrashekar
Published: December 2018

Introduction:
According to the National Health And Nutrition Examination Survey (NHANES), 65 million Americans have hypertension and one-quarter of adults have blood pressure in the “pre hypertension” range. In such scenario accurate BP measurement is very important. This study focused on prevalence of hypertension among Obese individuals.

Methods:
400 patients ( 200 patients each with < and > BMI 25 ) were studied in rural tertiary hospital in India. Inclusion criteria - patients aged 20-50 yrs. Obese ( defined by BMI > 25) . Exclusion criteria, Subjects on anti- hypertensive medication
,
Secondary causes of clinically identifiable hypertension Diabetic subjects who are known hypertensives or with complications of DM
,
Pregnant women, Known cases of Ischemic heart disease. Enrolled patient were evaluated for Height , weight. Two cuff sizes 12 cm/15 cm used to measure BP. Statistical analysis was done with SPSS package and MS Excel, Student’s t-test and linear correlation coefficient used. p values < 0.05 were considered t significant. SBP - systolic Blood pressure and DBP - Diastolic Blood pressure.

Results:
SBP and DBP increased significantly with increase of BMI in both men and women ( p < 0.005 ). There is statistically significant standard and large cuff differences in SBP with P value 0.003 and 0.01 and DBP with p values were 0.004 and 0.00 for male and female respectively. There was also a statistically significant ( p value < 0.05) standard-large cuff difference in both SBP and DBP in W/H ratio categories ( <1 and >1 ) in males

Discussion:
Cuff characteristics, i.e. the cuff-bladder width and length, can bias measurement of blood pressure in obese. Our study explained the potential for overdiagnosis of BP with using a wrong size cuff for people with BMI > 25 and W/H > 1. The prevalence of Systolic Hypertension based on JNC guidelines in the obese population with a regular cuff was 28% and with large appropriate size cuff it was 12%, a difference of 16% and similarly a difference of 12.5 % with diastolic BP. The prevalence of obesity in the US is 37% and this concept of Cuff Hypertension highlighted in our study is very important is such a setting as it can lead to lot of mis-diagnosed hypertension with unnecessary treatment burden and side effects.
Abstract P173: Sex Differences in the Diurnal Natriuretic Response to Benzamil in Sprague Dawley Rats
Authors: Soliman Reham H , Johnston Jermaine G , Gohar Eman Y , Pollock David M
Published: December 2018

Epithelial sodium channel (ENaC) expression follows a circadian pattern, but how this impacts activity is not known. In addition to the well-established sex differences in renal Na handling, recent data show that ENaC expression is higher (Western blot) in female rats. Further, we do not know if there are time of day differences or sex differences in ENaC activity. Therefore, the aim of this study was to determine if the diuretic response to ENaC inhibition (benzamil) is different between sexes at different times of day. SD rats (12-16 wk old) were placed in metabolic cages, where 12-hour urine collections were obtained to measure baseline urine volume and Na excretion as well as water and food intake. On day 3, benzamil was given at a dose of 1mg/kg (i.p.) either at the beginning of their inactive period/lights on (Zeitgeber Time 0, ZT0) or their active period/lights off (ZT12). The natriuretic response to benzamil was significantly greater in male compared to female rats (909 ± 302 vs 523 ± 83 μEq/kg/hr n=8) at ZT0 and (934 ± 94 vs 714 ± 151 μEq/kg/hr n=8) at ZT12. The diuretic response followed natriuresis being more prominent in male than female rats regardless of time of day (4.2 ± 0.7 vs 3.3 ± 0.5 ml/kg/hr n=8) at ZT0 and (3.6 ± 0.6 vs 2.5 ± 0.3 ml/kg/hr n=8) at ZT12. However, the larger response to benzamil given at the beginning of the inactive period (ZT0) compared to active period (ZT12) was not statistically significant (311 ± 98 vs 354 ± 30 μEq/hr and 120 ± 17.75 vs 174 ± 37 μEq/hr n=8) in male and female rats respectively. Given that endothelin-1 (ET-1) is an upstream inhibitor of ENaC, we measured urinary ET-1 levels to assess intrarenal production. ET-1 excretion significantly increased following benzamil administration in both sexes but was significantly greater in females. ET-1 excretion increased from 0.06 ± 0.01 to 0.29 ± 0.05 pg/hr in males (n=8) and from 0.10 ± 0.01 to 0.57 ± 0.28 pg/hr in females (n=8) at ZT0. At ZT12, ET-1 increased from 0.09 ± 0.02 to 0.23 ± 0.06 pg/hr in males (n=8) and from 0.14 ± 0.05 to 0.34 ± 0.06 pg/hr in females (n=8). These results demonstrate that the response to ENaC inhibition is less prominent in females independent of renal ET-1. This suggests less ENaC activity independent of expression in females or differences in non-ENaC related effects of benzamil.
Abstract P175: Influence of Age and Estradiol on Blood Pressure Responses to Exercise
Authors: Wenner Megan M , Matthews Evan L , Kaur Jasdeep , Greaney Jody L , Fadel Paul J
Published: December 2018

An exaggerated pressor response to exercise is associated with increased risk of cardiovascular events. The exercise pressor reflex, comprised of metabolic and mechanically sensitive skeletal muscle afferents, contributes importantly to blood pressure control during exercise. Data in preclinical animal models suggest that estradiol (E2) attenuates the pressor responses to exercise pressor reflex activation, yet it remains unclear if these findings translate to humans. Accordingly, we tested the hypothesis that pressor responses during exercise are augmented in post-menopausal women (PMW) compared to young women (YW), and that E2 administration will attenuate these responses in PMW.

Methods:
Mean arterial pressure (MAP, Finometer) and heart rate (ECG) were continuously measured in 12 PMW (age 59±2 years) and 16 YW (age 22±1 years) during 2-min of isometric handgrip performed at 30% of maximal voluntary contraction (MVC). Handgrip was immediately followed by 3-min of post-exercise ischemia (PEI), which isolates the muscle metaboreflex. Separately, BP responses during isometric handgrip at 35% MVC and PEI were measured in 6 PMW (age 53±1 years) before and after 1 month of transdermal E2 administration (100 μg/day).

Results:
Resting MAP was similar between PMW (77±3 mmHg) and YW (80±3 mmHg;
P
>0.05). During handgrip, the increase in MAP was greater in PMW (Δ20±2mmHg) compared to YW (Δ12±1 mmHg;
P
<0.05), and this was maintained during PEI (Δ14±1 mmHg PMW vs. Δ9±1 mmHg YW;
P
<0.05). Estradiol administration decreased resting BP in PMW (MAP 93±3 mmHg vs. 89±3 mmHg;
P
<0.05). Moreover, the increase in MAP during exercise in PMW was attenuated following estradiol administration (Δ31±8 mmHg vs. Δ20±6 mmHg;
P
<0.05). There was also a tendency for this reduction in MAP during PEI (Δ22±6 mmHg vs. Δ18±7 mmHg;
P
=0.08).

Conclusions:
These preliminary data suggest that PMW exhibit an exaggerated BP response to isometric exercise, due in part to heightened metaboreflex activation, and that estradiol administration can attenuate such responses.