bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 067: Endothelial mTORC1 Signaling Regulates Vascular Function and Mediates Angiotensin II Actions
Authors: Reho John J , Guo Deng-Fu , Rahmouni Kamal
Published: September 2019

The mechanistic target of rapamycin complex 1 (mTORC1) serves as an intracellular energy sensor through its downstream signaling components the S6-kinase and the ribosomal S6 protein with recent implications for hypertension. In this study, we tested the hypothesis that endothelial mTORC1 signaling is critical for the regulation of vascular endothelial function. We generated tamoxifen-inducible endothelial-specific
Raptor
deleted mice (Tek
CreERT2
/Raptor
F/F
) to disrupt mTORC1 signaling in the endothelium. Tamoxifen-treated wildtype littermates were used as controls. We confirmed endothelial-specific deletion of Raptor by RT-PCR in MACS-sorted endothelial cells. Interestingly, homozygous
Raptor
deletion cause endothelial dysfunction as indicated by the reduced vasorelaxation of tamoxifen-treated Tek
CreERT2
/Raptor
F/F
mice aortic rings evoked to acetylcholine (Max relax: 34±11 vs 61±7% in controls; p<0.05), but not sodium nitroprusside (Max relax: 82 ±3 vs 84±2%). Restoring mTORC1 signaling in tamoxifen-treated Tek
CreERT2
/Raptor
F/F
aortic rings with an adenoviral S6-kinase constitutively active construct (Ad-S6KCA; 24hrs) rescued the reduction in acetylcholine-mediated relaxation (Max relax: 54±6 vs 31±10% in Ad-GFP; p<0.05), demonstrating that the endothelial dysfunction is Tek
CreERT2
/Raptor
F/F
mice is due to loss of mTORC1 signaling. Notably, aortic rings from heterozygous Tek
CreERT2
/Raptor
F/+
mice which have intact acetylcholine-induced vasorelaxation responses were protected from angiotensin II (Ang II)-induced endothelial dysfunction. Indeed, incubation of aortic rings from wildtype mice with Ang II (100nM; 24hrs) reduced acetylcholine-mediated vasorelaxation responses (Max relax: 3±8 vs 57±4 % in vehicle group; p<0.05). In contrast, aortic rings of tamoxifen-treated Tek
CreERT2
/Raptor
F/+
mice were partially protected from the endothelial dysfunction evoked by Ang II (Max relax: 36±9; p<0.05 vs wildtype controls). Our data indicate that endothelial mTORC1 signaling is required for the regulation of endothelial function. Our findings also highlight the importance of mTORC1 signaling in mediating the vascular endothelial effects of Ang II.
Abstract 069: Local and Circulating Sphingosine-1-Phosphate in Blood Pressure Regulation
Authors: Del Gaudio Ilaria , Rubinelli Luisa , Schwab Susan , Wadsack Christian , Di Lorenzo Annarita
Published: September 2019

Sphingosine-1-phosphate (S1P), a bioactive lipid, regulates different biological processes in health and disease. Once synthetized, endothelial S1P is transported outside the cells via Spinster-2 (Spns-2), and can signal mainly through the sphingosine-1-phosphate receptor-1 (S1P1) in autocrine/paracrine manner. Alternatively, S1P can bind circulating carriers, including ApoM of HDL or albumin. Recently, we reported a key role of endothelial S1P-S1P1 autocrine signaling in vascular function and blood pressure (BP) homeostasis. To dissect the contribution of endothelium-derived S1P versus circulating HDL-bound S1P on BP regulation, we used ApoM knockout mice (ApoM-/-) and developed mice lacking the endothelial Spinster-2 (ECKO-Spns2). At baseline, systolic BP (SBP) was markedly increased in both ECKO-Spns2 (120.8± 2.1 vs. 106.6±1.3 mmHg, n=8) and ApoM-/- (122.6±1.0 vs. 109.0±1.4 mmHg, n=8) vs control mice. Vasodilation of mesenteric arteries (MA) to acetylcholine was preserved in both mouse models, whereas flow-induced vasodilation was significantly reduced in ECKO-Spns2 (Emax 36.0±1.5 vs. 53.6±3.6 % vasodilation, n=5) but not in ApoM-/- MA, suggesting that autocrine S1P signaling requires the S1P transporter Spns-2. However, both ECKO-Spns2 and ApoM-/- showed a reduced NO production compared to control MA. Moreover, chronic infusion of AngII resulted in higher BP in ECKO-Spns2 (156.0± 1.2 vs. 142.0±1.6 mmHg, n=8) and ApoM-/- (158.5±1.1 vs. 144.1±1.3 mmHg, n=8) vs control mice. Interestingly, chronic AngII strongly reduced vasodilation in response to flow in both ECKO-Spns2 (Emax 19.3±2.5 vs. 35.4±7.1 % vasodilation, n=5) and ApoM-/- (Emax 22.9±1.1 vs. 34.8±6.8 % vasodilation, n=5) compared to control MA, suggesting a protective role of local and circulating S1P. Additionally, basal NO production was further impaired by the AngII infusion (ECKO-Spns2 -32.7% and ApoM-/- -42.8%, vs control MA), confirming the pivotal role of S1P-S1P1-NO pathway in vascular and BP homeostasis.
Abstract 070: The Prostaglandin Ep4 Receptor in Endothelium Modulates eNos Expression and Aortic Integrity
Authors: Yang Ting , Sparks Matthew A , Koller Beverly H , Coffman Thomas M
Published: September 2019

Prostaglandin EP4 receptor (EP4R) plays pivotal role in mediating PGE2-induced vasodilation and blood pressure homeostasis. Our previous work demonstrated that absence of EP4R in all tissues was associated with salt sensitivity and exaggerated Ang II-dependent hypertension. While cell-specific elimination of EP4R from vascular smooth muscle cells (VSMCs) attenuated EP4-dependent vasodilation, but the severity of Ang II-dependent hypertension was not affected. These findings indicate the effects of EP4R to resist hypertension are not attributable to the compensatory vasodilation mediated by EP4R in VSMCs and must be mediated by other cell lineages. Within the vasculature, endothelial cells (ECs) also express high levels of EP4R. To assess the role of EP4R in ECs, we generated conditional EC-specific EP4R knockout mice mice using Tie2-CreERT2 and

EP4
flox/flox

mouse lines (ECKO). After pre-treatment with tamoxifen, we measured the blood pressure in conscious mice using radiotelemetry. At baseline, there were no significant differences in blood pressure between ECKO and controls 107.9±1.7 VS 109.3±1 mmHg). The response of ECKOs to Ang II infusion was variable. In one cohort, MAP during Ang II infusion was significantly lower than controls (142.8±3.1 vs 133.2±1.9 mmHg, p=0.02) whereas in a second cohort there was no difference in MAP with Ang II infusion between groups (131.7±2.3 vs 135.6±2.1 mmHg). Interestingly, the incidence of aortic aneurysm was significantly higher in ECKOs (12/27) compared to controls (2/25; p=0.01 by Fisher exact test). As urine sodium excretion during Ang II infusion was increased in ECKOs, we considered the possibility that aneurysm formation may have impacted the blood pressure. We also found the ECs isolated from ECKOs have significantly higher eNOS mRNA expression compared to control (fold change: 1.35±0.13; p=0.046). To determine whether this difference in eNOS expression have a functional impact, we compared blood pressure responses to L-NAME following Ang II infusion. ECKOs showed exaggerated response to L-NAME compared to controls (MAP increases +5.2±1.4 mmHg in ECKO vs. +0.5±1.5 mmHg in control; p=0.03). Thus, EP4R in ECs play important role in regulating EC function, impacting blood pressure and aortic integrity.
Abstract 072: Medroxyprogesterone Prevents the Decline in Renal Health Due to Estradiol
Authors: Zimmerman Margaret , Ogola Benard , Lindsey Sarah
Published: September 2019

We previously published that estradiol (E2) treatment initiated immediately after midlife ovariectomy (OVX) for 80 days in female Long Evans rats exerted negative effects on renal health. Medroxyprogesterone (MPA) is a progesterone derivative commonly used in menopausal hormone therapy to oppose the uterotrophic effects of E2. Therefore, our goal was to determine if coadministration with MPA opposed the negative effects of E2 on renal health. Female retired breeders underwent OVX at 11 months of age and received an implant of E2, E2+MPA or vehicle for 40 days. Body weights, blood pressure and urine were measured pre- and post-treatment along with tissue weights. Systolic blood pressure was not altered from baseline in any treatment group (Veh: 144±4 vs.135±5; E2: 143±4 vs.143±6; E2+MPA: 147±4 vs.143±5 mmHg; ns). Kidney hypertrophy was significantly increased following E2+MPA treatment (Veh: 2.73±0.1; E2: 3.03±0.9; E2+MPA: 3.42±0.2 g/kg body wt; P=0.002), while body and heart weights were comparable between groups. Coadministration of MPA prevented the E2-induced increase in proteinuria (Veh: 0.27±0.07; E2: 3.53±1.16; E2+MPA: 1.20±0.58 mg/mg creatinine; P=0.01) and decline in glomerular filtration rate (Veh: 0.51±0.02; E2: 0.24±0.05; E2+MPA: 0.39±0.05 ml/min; P=0.001). Our results show that even short-term E2 exerts a negative effect on renal health in midlife, but coadministration with MPA prevents E2-mediated damage, possibly via compensatory hypertrophy of the kidney. Therefore, midlife hormone therapy containing MPA may protect the kidneys against unopposed E2-mediated damage and provide a beneficial effect for menopausal women.
Abstract 077: Proximal Tubule-Specific Deletion of the NHE3 (na
+
/h
+
Exchanger 3) in the Kidney Attenuates Angiotensin II-Induced Hypertension in Mice
Authors: Li Xiao C , Zhu Dongmin , Chen Xu , Zheng Xiaowen , Zhao Chunling , Zhou Xinzhou , Soleimani Manoocher , Rubera Isabelle , Tauc Michel , Zhuo Jia L
Published: September 2019

The Na
+
/H
+
exchanger 3 (NHE3), a ~85 kDa protein encoded by the
SLC9A3
gene, is the most important Na
+
transporter in the proximal tubules of the kidney. Angiotensin II (ANG II) is well-recognized to increase proximal tubule Na
+
reabsorption by stimulating NHE3 expression and activity in the proximal tubules. However, a direct cause and effect relationship between ANG II and NHE3 in the proximal tubules in ANG II-induced hypertension has not been determined previously. The present study directly tested the hypothesis that NHE3 in the proximal tubules of the kidney is required for the development of ANG II-induced hypertension using proximal tubule-specific NHE3 knockout mice (PT-

Nhe3
-/-

). Specifically, PT-

Nhe3
-/-

mice were generated using the
SGLT2-Cre
/

Nhe3
loxlox

approach, whereas ANG II-induced hypertension was studied in 12 groups (n=5-12 per group) of adult male and female wild-type (WT) and PT-
Nhe3
-/-
mice. Under basal conditions, systolic (SBP), diastolic (DBP), and mean arterial blood pressure (MAP) were significantly lower in male and female PT-

Nhe3
-/-

than WT mice (
P
<0.01). A high pressor, 1.5 mg/kg/day, i.p., or a slow pressor dose of ANG II, 0.5 mg/kg/day, i.p., for 2 weeks significantly increased SBP, DBP, and MAP in male and female WT mice (
P
<0.01), but the hypertensive response to ANG II was markedly attenuated in male and female PT-

Nhe3
-/-

mice (
P
<0.01). ANG II impaired the pressure-natriuresis response in WT mice, whereas proximal tubule-specific deletion of NHE3 improved the pressure-natriuresis response in ANG II-infused PT-

Nhe3
-/-

mice (
P
<0.01). AT
1
receptor blocker losartan completely blocked ANG II-induced hypertension in both WT and PT-

Nhe3
-/-

mice (
P
<0.01). However, inhibition of nitric oxide synthase with L-NAME had no effect on ANG II-induced hypertension in WT or PT-

Nhe3
-/-

mice (n.s.). Furthermore, ANG II-induced hypertension was significantly attenuated by an orally absorbable NHE3 inhibitor AVE0657. In conclusion, NHE3 in the proximal tubules of the kidney is required for the full development of ANG II-induced hypertension. Our results suggest that NHE3 in the proximal tubules may be therapeutically targeted to treat hypertension induced by ANG II or associated with increased NHE3 expression in the proximal tubules.
Abstract 079: Elevated Fructose Intake Induces Hypertension in Dahl Salt Sensitive (SS) Rats on a Normal Na Diet: Role of SPAK (STK39) and NKCC2
Authors: Eaton Andrew J , Garcia-Pedraza Jose A , Ortiz Pablo A
Published: September 2019

Increased fructose consumption in sweetened beverages is associated to increased cardiovascular events and hypertension in patients. We found that in normal, salt resistant, Sprague Dawley (SD) rats, a moderately high fructose intake (20% fructose drinking water) induced salt sensitive hypertension whereas fructose in normal Na diet had little effect. The mechanisms by which fructose intake induced salt sensitive hypertension are unclear. Fructose directly increases the activity of the Na-K-2Cl cotransporter NKCC2 in the thick ascending limb (TAL), and when consumed for 7 days or more, increases phosphorylation of NKCC2, suggesting that this may contribute to fructose induced salt sensitive hypertension. The kinases upstream of NKCC2 phosphorylation, SPAK and OSR1 were also activated by fructose intake. The Dahl SS rat has higher NKCC2 and SPAK phosphorylation in the TAL. We hypothesized that elevated fructose intake causes hypertension in Dahl SS rats on normal Na, but not in SD rats, and that SPAK and NKCC2 phosphorylation are involved. We measured systolic BP (SBP) by radiotelemetry in conscious SD rats fed normal Na (0.4% Na) and 20% fructose in drinking water. SBP minimally increased by 7±2 mmHg (from 122±3 to 129±3 mmHg, p<0.05) after 4 weeks of high fructose and normal Na. In Dahl SS rats fed normal Na, SBP increased by 25±4 mmHg (from 148±3 to 173±4 mmHg, p<0.05 vs SD rat) after 4 weeks of high fructose intake. To study SPAK, we generated Dahl SS rats with genetic deletion of SPAK (SPAK KO). Baseline SBP was lower in SPAK KO (133±2 mmHg, p<0.05), and minimally increased by 10±3 mmHg (from 133±2 to 144±3 mmHg, p<0.05 vs Dahl SS) after 4 weeks of high fructose intake. The decreased effect of fructose in SPAK KO rats may be related to lower NKCC2 phosphorylation because phosphoNKCC2/total NKCC2 was 33 ± 7% lower (p<0.05) in TALs from SPAK KO compared to Dahl SS on high fructose for 14 days. We concluded that a moderately elevated fructose intake induces hypertension in Dahl SS rats on normal Na diet, but not in SD rats. Fructose-induced hypertension in Dahl SS is in part mediated by SPAK and related to higher NKCC2 phosphorylation. Our data suggest that in patients with genetic susceptibility to high salt, BP may also be enhanced by high fructose consumption.
Abstract 087: Impaired Brain Cystathionine Beta-Synthase-Derived H
2
S Production is Associated With the Development of Neurogenic Hypertension
Authors: Xia Huijing , Li Zhen , Lazartigues Eric , Kapusta Daniel R , Lefer David J
Published: September 2019

Hydrogen sulfide (H
2
S) is recognized as an important physiological signaling molecule for cardiovascular homeostasis. Decreased circulating H
2
S levels are associated with the development of hypertension, and central administration of H
2
S donors reduces blood pressure in animal models. Cystathionine β-synthase (CBS), one of the H
2
S producing enzymes, is highly expressed in the brain. However, little is known about the role of CBS and brain endogenous H
2
S in the regulation of blood pressure. We hypothesized that impaired brain H
2
S production is associated with the development of neurogenic hypertension. In the 1
st
experiment, male C57/BL6/J mice (10-12 wks, n=7) were infused with Angiotensin-II (Ang-II, 600 ng/kg/min, sc, mini-osmotic pump; neurogenic hypertension model) or saline for 4 weeks. Plasma and brain H
2
S levels were measured using gas chromatography-chemiluminesence. Following 4 wks of Ang-II infusion, mean arterial blood pressure (MAP) under anesthesia (carotid artery catheter) was significantly higher in Ang-II compared to saline-infused mice (108.5 ± 8 vs. 77.1 ± 1 mmHg, p < 0.01), confirming hypertension in the Ang-II group. Although plasma free H
2
S levels were not different between groups, brain free H
2
S level was significantly decreased in Ang-II-infused mice (3.3 ± 0.3 vs. 4.5 ± 0.4 pmol/mg, p < 0.05), suggesting that brain H
2
S production is impaired during neurogenic hypertension. In the 2
nd
experiment, Neuron-specific CBS knockout mice (NS-CBS KO) and CBS Flox controls (male, 10-12 wks, n=6) were implanted with telemetry probes for conscious blood pressure recording. Ang-II was infused after baseline recording. The basal level of MAP was not different between NS-CBSKO and CBS Flox mice (96.5 ± 3 vs. 99.3 ± 4 mmHg, p < 0.05). After 1 wk of Ang-II infusion, MAP was significantly higher in NS-CBS KO compared to CBS Flox mice (126.6 ± 10 vs. 106.8 ± 5 mmHg, p < 0.05). Taken together, these data suggest that CBS is involved in the central regulation of blood pressure, and impaired brain CBS-derived H
2
S production promotes the development of neurogenic hypertension. (Funding: NIH R01 HL146098-01; NIGMS P30GM106392)
Abstract 088: Overexpression of Neuronal Nitric Oxide Synthase Associated Protein in the Paraventricular Nucleus Contributes to Increased Sympathoexcitation in Male Mice by Modulating Subcellular Localization of Neuronal Nitric Oxide Synthase
Authors: Sharma Neeru , Katsurada Kenichi , Nandi Shyam , Gao Lie , Patel Kaushik
Published: September 2019

Nitric oxide (NO•) bioavailability within the paraventricular nucleus (PVN) plays a pivotal role in the pathogenesis of many cardiovascular diseases including hypertension. Aligning neuronal nitric oxide synthase (NOS1) with
N
-methyl-D-aspartate (NMDA) receptor by tethering to PSD95 (postsynaptic density 95 protein) allows the production of NO• initiated by an influx of Ca
2+
in neurons within the PVN. NOS1 associated protein (Nos1ap) competes with PSD95 for NOS1 binding and thus influences the production of NO•. We investigated the effect of specifically overexpressing Nos1ap in the PVN on sympathetic outflow using cre recombinase-conditional
Nos1ap
over-expression transgenic(Tg) mice injected with lentivirus expressing Cre-GFP or GFP into the PVN. Two weeks of post-viral administration, the sympathetic outflow was evaluated in both male and female mice. Male-Cre-Nos1ap mice displayed a significantly elevated urinary excretion of norepinephrine, an index of overall sympathetic outflow compared to GFP-Nos1ap (0.64±0.11* vs. 0.17±0.03 ug/day) while female littermates showed no difference (0.44±0.06 vs. 0.36±0.03 ug/day). Basal renal sympathetic nerve activity assessed as a percent of maximal activity was elevated 1.5-fold in Cre-Nos1ap male mice (46.1 ± 3.6* vs.31.0 ± 3.2) while there was no significant difference in Cre-Nos1ap female mice(22.6± 1.2 vs. 24.8 ± 2.6) compared to respective GFP controls. Interestingly, female mice had higher levels of basal NOS1 in the PVN compared to male littermates (0.79 ±0.01* vs. 0.37±0.05). Further, a robust overexpression of Nos1ap in the PVN of Cre-Nos1ap (0.49±0.04* vs. 0.21±0.03 male; 0.53±0.08* vs. 0.28±0.04 female) compared to GFP-Nos1ap mice does not lead to any change in NOS1, NMDA receptor(NR)2A, NR2B, while NR1 expression tended to increase in both sexes. Overexpression of Nos1ap in NG108 neurons,
in vitro,
led to relocalization of 46% of membrane-associated NOS1 to the cytosolic compartment, suggesting that the decrease in NMDA receptor-mediated NOS1 activity leads to reduced NO• production. Taken together, our data indicate that overexpression of Nos1ap within the PVN reduces the levels of NO• generated by NOS1 leading to increased sympathetic outflow in male mice but not in female mice.
Abstract 090: Intergenerational Inheritance of Obesity-Induced Diastolic Dysfunction With Preserved Ejection Fraction
Authors: da Silva Alexandre A , Gava Fabio N , Omoto Ana C , Moak Sydney P , Dai Xuemei , Hall John E , Docarmo Jussara M
Published: September 2019

Obesity is strongly associated with cardiomyopathy and heart failure. We examined the impact of paternal and maternal obesity on systolic and diastolic function in male adult offspring. The following groups were studied: 1) offspring fed normal diet (ND) born from ND parents (ND-F1-ND, n=5); 2) offspring fed high fat diet (HFD) born from ND parents (HFD-F1-ND, n=6); 3) offspring fed ND born from HFD parents (ND-F1-HFD, n=7); and 4) offspring fed HFD born from HFD parents (HFD-F1-HFD, n=7). Parents were fed HFD or ND from weaning until the end of lactation, while the offspring were placed on ND or HFD from weaning until the end of experiments. Body weight, fat and lean mass, measured using EchoMRI4-in-1 System, were measured weekly. Cardiac function including ejection fraction (EF), isovolumetric relaxation time (IVRT), late filling velocity (A’), E and E’ early filling velocities were determined at 22 weeks of age using long and short axis left ventricle dimensions, pulse wave and tissue Doppler (VisualSonics 30 MHZ transducer, VEVO3100®). Compared to lean control ND-F1-ND mice, ND-F1-HFD mice had similar body weight (28.9±1.0 vs. 27.9±0.4 g) but higher fat mass (4.5±0.9 vs. 3.0±0.3 g) and lower lean mass (22.9±0.4 vs. 24.6±0.3 g). Both groups of mice fed HFD after weaning were heavier than offspring fed ND. However, HFD-F1-HFD mice were heavier than HFD-F1-ND mice (47.3±0.8 vs. 42.5±1.9 g) due to greater fat mass (19.4±0.6 vs. 15.7±1.6 g) and lean mass (28.3±0.6 vs. 26.9 vs. 0.3 g). Despite no major differences in ejection fraction among all 4 groups, mice fed HFD or born from parents fed HFD exhibited diastolic dysfunction; worse indicators of diastolic function were observed in mice from HFD parents as indicated by increased IVRT (30.4±1.8, 25.0±1.8, 22.1±2.0 vs. 21.4±0.9 ms) and E/E’ ratio (36.6±3.6, 32.3±1.8, 31.8±1.2 vs. 27.9±2.5) and reduced E’/A’ ratio (0.9±0.1, 0.7±0.1, 1.5±0.2 vs. 1.4±0.1) for HFD-F1-HFD, ND-F1-HFD, HFD-F1-ND and ND-F1-ND, respectively. These results indicate that diet-induced parental obesity is associated with diastolic dysfunction and preserved EF in the offspring, which is exacerbated by HFD-induced obesity. (NHLBI-PO1HL51971, NIGMS P20GM104357 and U54GM115428)
Abstract 091: Leptin-Thyroliberin Pathway in Left Ventricle Hypertrophy
Authors: Aisicovich Maia , Peres Diaz Ludmila S , Schuman Mariano L , Toblli Jorge E , Landa Maria S , García Silvia I
Published: September 2019

Cardiac TRH (cTRH) induces LVH and fibrosis. The adiponectin Leptin induces TRH in CNS. We hypothesized that in obesity, the increase of cTRH induced by hyperleptinemia is responsible for the LVH, mostly attributed to pressure load. We studied obese hyperleptinemic with mild hypertension Agouti mice, and found LVH with higher (p<0.05) cTRH, fibrotic (I/III col,TGFβ) and hypertrophic (BNP,βMHC) marker genes vs lean (BL/6J). We normalized pressure in Agouti mice with hydroclorothiazide (20 mg/kg/day) since weaning (n=9) vs non treated mice. Still both developed (p<0.05) LVH, higher cTRH, fibrotic and hypertrophic marker genes expression, suggesting that LVH would not be due to hypertension. Opposite to Agouti model, we used obese normotensive ob/ob mice, lacking Leptin expression, and without LVH despite their obesity. We treated them with Leptin (80 ug/kg/day) or saline since weaning for 15 days. Only the treated group developed LVH (p< 0.05, n=7) demonstrating that it is Leptin dependant. Also we found increase (p<0.05) in cTRH, col III, TGFβ and BNP suggesting that leptin-TRH interaction is required in obesity-induced LVH. Finally, we evaluated whether Leptin administration, (n=6, 10 ug/kg/day) could induce hypertrophy in lean C57 mice with and withouth native cTRH system, by previous siRNA injection (5ug). Diuretic was given to Leptin groups, to avoid its hypertensive effect. Leptin induced cTRH expression, not observed in the groups with siRNA-TRH (p<0.05).This probably induce fibrotic and hypertrophic markers, demonstrating that the novel interaction is functional also in mildy hyperleptinemia at normal weight status. To confirm TRH's Leptin induction we used cardiomyocytes cell line H9C2 (n=6) stimulated with Leptin (10 and 100 ng/ml). TRH expression and precursor content increased (p<0.05) post Leptin treatment with both concentrations. Moreover in primary cardiomyocytes culture from neonate rats, Leptin stimulus (80 ng/ml, 24 hs) increased (p<0.05) TRH content vs controls, confirming the direct TRH-Leptin induction in heart cells. We described a novel Leptin-cTRH pathway which explains Leptin-induced LVH, highly likely since early stages.
Abstract 098: Susceptibility of Mice Lacking Renin-B to Chronic Angiotensin II Infusion
Authors: Nakagawa Pablo , Nair Anand , Zhang Shao Yang , Lu Ko-Ting , Zanaty Mario , Gomez Javier , Grobe Justin L , Sigmund Curt D
Published: September 2019

Prior experience to various hypertensive conditions enhances the response to subsequent hypertensive challenges and the activation of the brain renin angiotensin system (RAS) is required for the sensitization to these hypertensive challenges. We previously showed that the brain-specific renin alternative isoform (Ren-b) tonically inhibits the activation of the brain RAS and that ablation of Ren-b (Ren-b KO) results in brain RAS disinhibition leading to elevated blood pressure (BP). Retrospective analysis of five cohorts of mice indicated that Ren-b KO model exhibits a high degree of variability in BP, which may be attributed to the susceptibility to stress. Therefore, we hypothesized that Ren-b KO are sensitized to hypertensive challenge such as angiotensin (Ang) II. Wildtype (WT) or Ren-b KO were infused with either vehicle or Ang II (400 ng/kg/min; sc.) via osmotic minipumps for 4 weeks. Ang II significantly increased BP in both WT and Ren-b KO to the same degree (WT+veh: 103±4 vs WT+Ang: 120±4 mmHg; p = 0.01 (n=10-12) and KO+veh: 104±3 vs KO+Ang: 118±5 mmHg; p = 0.03 (n=10-14)). Although the BP between Ren-b KO and WT mice was elevated equally, Ang II induced an exaggerated suppression of plasma renin in Ren-b KO compared to WT (WT+Ang: 14.8±2.1 vs KO±Ang: 7.2±2.6 ng/ml; p = 0. 03; n=7-9). Ren-b KO exhibited enhanced Ang II-induced dipsogenic response (WT+Ang: 3.7±0.3 vs KO+Ang: 4.6±0.4 ml/day; p = 0. 05; n=10-12) and elevated heart weight (WT+Ang: 4.8±0.2 vs KO±Ang: 5.2±0.1 mg/g of body weight; p = 0.03; n=10-12) concomitant with an increased cardiac collagen type I alpha-2 mRNA expression (p = 0.03; n=7-9). Altogether, this data indicates that in the presence of Ang II, Ren-b KO exhibits higher susceptibility to cardiovascular disease. After successive breeding and backcrossing Ren-b KO developed a milder and more varied baseline BP elevation compared to our initial cohorts. This suggests that epigenetic silencing or other physiological and molecular mechanisms might compensate for some of the cardiovascular alterations driven by the genetic deletion of renin-b. Studies of a conditional Ren-b KO model will be used to elucidate the physiological and/or pathophysiological consequences of an acute downregulation of renin-b in adulthood.
Abstract 099: Novel Angiotensin-Converting-Enzyme 2 (ACE2) Truncates to Target the Kidney Renin-Angiotensin-System (RAS)
Authors: Wysocki Jan , Schulze Arndt , Zhao Ming , Haney Chad , Batlle Daniel
Published: September 2019

ACE2 is an enzyme with a molecular size of more than 100 kDa which produces Angiotensin (1-7) from Angiotensin II (1-8). Attempts to target kidney RAS using ACE2 to treat kidney disease are hampered by its large molecular size which precludes its glomerular filtration and subsequent tubular uptake. We tested two ACE2 truncates (1-605 and 1-619AA) that we generated through truncation from the C terminus. ACE2 enzyme activity measured using Mca-APK-Dnp substrate was as high for each of them as native rACE2 (1-740 AA). The two truncates had an apparent MW of ~70 kDa, as expected from the amino acid sequence, as shown by western blot. For radioimaging each purified truncate was labeled with
99m
Tc to study kidney uptake. After i.v. injection of
Tc99m
-labeled rACE2, there was kidney cortex retention for
Tc99m
1-605 [8.6±0.7% of whole body (WB) radioactivity] and for
Tc99m
1-619 (4.2±2.1% WB) as compared to only trace retention after injection of native
Tc99m
-rACE2 1-740 (1.2±0.2% WB)(figure). When infused to mice with genetic ACE2 deficiency, a single i.v. injection of 1-619 resulted in detectable ACE2 activity in urine and kidneys (23.1±4.3 and 1.96±0.73 RFU/ug prot/hr, respectively). In addition, the kidneys of ACE2-nul mice infused with 1-619 studied ex vivo formed more Ang (1-7) from exogenous Ang II than those of uninfused animals (p<0.05).

We conclude that our novel ACE2 truncates undergo glomerular filtration which is associated with kidney uptake of enzymatically active protein that can enhance formation of Angiotensin 1-7 from Angiotensin II (1-8). These truncates may offer a potentially useful approach to target kidney RAS overactivity to combat kidney disease.
Abstract 103: The Effects of s Lower Blood Pressure Treatment Goal on Orthostatic Hypotension in Adults With Recent Stroke
Authors: Juraschek Stephen P , Beach Jennifer L , Ishak Anthony , Mukamal Kenneth J
Published: September 2019

Background:
Lower systolic blood pressure (SBP) treatment goals reduce risk of orthostatic hypotension (OH). Whether a lower goal also reduces orthostatic symptoms is unknown.

Methods:
We examined the effect of SBP goal on OH in SPS3-BP (Secondary Prevention of Small Subcortical Strokes - Blood Pressure), an open label, randomized trial. Adults with a recent stroke (<6 months) were assigned to either a SBP goal of 130-149 mm Hg or <130 mm Hg. During follow-up, BP was measured 3 times in a seated position and once after 2 minutes of standing. OH was defined as a drop in SBP ≥20 mm Hg or DBP ≥10 mm Hg. We also examined extreme standing BP values (SBP ≥190 mm Hg or ≤90 mm Hg; DBP ≥110 mm Hg or ≤40 mm Hg). Participants were asked about dizziness and lightheadedness in the process of standing up. We used generalized estimating equations to account for repeat measurements.

Results:
During 36,342 visits, involving 2,876 participants (mean age 62.8 ± 10.7 yrs, 37% women, 16% black) with a mean of 15 follow-up visits, OH was present 2,591 times in 1,165 participants. Similarly, dizziness was present 1,343 times (in 592 participants) and lightheadedness was present 941 times (in 893 participants). A lower SBP goal (compared to standard) was associated with a lower risk of OH (OR 0.86; 95% CI: 0.75, 0.99), a higher risk of SBP ≤90 mm Hg (3.79; 2.35, 6.11) or DBP ≤40 mm Hg (2.42; 1.41, 4.15), and a lower risk of SBP ≥190 mm Hg (0.40; 0.27, 0.60) or DBP ≥110 mm Hg (0.31; 0.19, 0.50) (
Table
). BP goal did not affect dizziness or lightheadedness.

Conclusions:
A lower SBP goal decreased OH without affecting orthostatic symptoms. These findings support recent observations that more intensive SBP treatment does not increase risk of falls.
Abstract 104: Amplification of Systolic Blood Pressure and Pulse Pressure is Higher in Resistant Hypertensive Patients Irrespective of Blood Pressure Control
Authors: Valaiyapathi Badhma , Dudenbostel Tanja , Siddiqui Mohammed , Zhang Bin , El Hachem Maria , Oparil Suzanne , Calhoun David A
Published: September 2019

Background:
Systolic blood pressure (SBP) is higher in peripheral than in central arteries, a phenomenon referred to as BP amplification. A greater amplification of SBP and pulse pressure (PP) is associated with increased cardiovascular disease (CVD). The objective of this study is to investigate whether BP amplification is different in patients with uncontrolled resistant hypertension (uRHTN) when compared to those with controlled RHTN (cRHTN) and controlled non-RHTN (cnRHTN).

Methods:
In this prospective study, 149 patients were recruited from The University of Alabama at Birmingham’s HTN clinic. Brachial BP was measured using in-office BpTRU. Aortic BP was determined using pulse wave analysis by SpygmoCor. BP amplification was calculated as the difference between brachial and aortic BP.

Results:
The cohort comprised 50.0% women and 53.6% African Americans. Mean baseline values were: age 58.6 ± 10.3 years, BMI 34.2 ± 6.7 Kg/m
2
, SBP amplification 11.0 ± 4.4 mmHg and PP amplification 12.0 ± 4.7 mmHg. Multiple regression analysis showed a significant correlation of gender and increasing BMI with amplification of SBP (p<0.0001 and 0.003, respectively) and PP (p=0.001 and 0.007, respectively). Amplification of SBP and PP were significantly higher in patients with uRHTN compared to patients with cRHTN and similarly, patients with cRHTN had higher amplification than those with cnRHTN (p=0.046 and 0.021, respectively).

Conclusion:
This prospective study showed that the amplification of SBP and PP remains high in patients with RHTN regardless of BP control, indicating that patients with RHTN retain a higher risk of CVD morbidity and mortality in spite of successful BP control.
Abstract 106: The Association Between Overnight Rostral Fluid Shift and Sleep Disordered Breathing in Patients With Treatment Resistant Hypertension
Authors: Ishiwata Sayaki , Matsumoto Hiroki , Sato Akihiro , Shitara Jun , Suda Shoko , Kasai Takatoshi
Published: September 2019

Background:
Sleep disordered breathing (SDB) is an important modifiable risk factor of treatment resistant hypertension (TRH). Overnight rostral fluid shift from lower extremities is associated with the severity of SDB in patients prone to fluid accumulation (e.g. TRH, heart failure, and end-stage renal disease). However, the association between overnight rostral fluid shift and SDB in Japanese patients with TRH remains unclear.

Methods:
All patients with TRH underwent polysomnography. Neck circumference (NC) and leg fluid volume (LFV) were measured by tape measure and bioelectrical impedance just before sleep and after waking up. Correlation between apnea-hypopnea index (AHI), overnight change of LFV (ΔLFV), and overnight change of NC (ΔNC) were assessed by regression analysis.

Results:
Total 11 patients were examined (7 men [63%], Age: 62.6±13.4 years, body mass index: 28.1±4.2 kg/m
2
). Mean AHI, ΔLFV, and ΔNC were 30.5±24.1/h. -267.5±156.1 ml, and 0.4±0.5 cm, respectively. ΔLFV correlated significantly with ΔNC (r=-0.63; P=0.04). Besides, both ΔLFV and ΔNC were significantly associated AHI respectively (ΔLFV, r=-0.92; P<0.01, ΔNC, r=0.67; P=0.02). In the stepwise regression analysis including estimated glomerular filtration rate, B-type natriuretic peptide along with ΔNC and ΔLFV, onlyΔLFV was the significant independent correlate with AHI,

Conclusion:
In Japanese patients with TRH, overnight rostral fluid shift contributes to the worsening of SDB, in association with the increase in NC.
Abstract 111: Active Time-Restricted Feeding Potently Protects Blood Pressure Daily Rhythm in Diabetic Mice
Authors: Hou Tianfei , Su Wen , Duncan Marilyn J , Gong Ming C , Guo Zhenheng
Published: September 2019

Diabetes is associated with high prevalence of non-dipping blood pressure (BP), which is the most common form of disruption in BP daily rhythm. Non-dipping BP is emerging as an independent risk factor of cardiovascular diseases and target organ damages. A large body of human and animal evidence demonstrated that active-phase time-restricted feeding (ATRF) is highly metabolic beneficial. However, the effects of ATRF on BP daily rhythm are unknown. The current study investigates whether, and how ATRF protects mice from non-dipping BP in a type 2 diabetic
db/db
mouse model. The results demonstrated that ATRF potently prevented the disruption, as well as restored the already disrupted BP daily rhythm in
db/db
mice (BP oscillation amplitude (mmHg):
ad libitum
feeding (ALF)
vs.
ATRF: 4.95±0.83
vs.
12.74±1.22,
p
<0.001). The loss of BP dipping in
db/db
mice was associated with an increased portion of daily food intake during the inactive light-phase, which preceded the development of non-dipping BP (light-phase food intake (g): control
vs. db/db
: 0.33±0.19
vs.
3.38±0.07,
p
<0.0001). Moreover, feeding normal mice at the wrong time with inactive-phase time-restricted feeding (ITRF) compromised BP daily rhythm (BP oscillation amplitude (mmHg): ALF
vs.
ITRF: 7.69±0.44
vs.
4.59±0.44,
p
<0.01), suggesting the time of feeding plays an important role in BP daily rhythm. While multiple mechanisms likely contribute to the powerful protection of ATRF on BP daily rhythm in
db/db
mice, we found that improved autonomic nervous system mediates, at least in part, the protective effects of ATRF. At the molecular level, we found that among the multiple clock genes examined, the mRNA oscillation of
Bmal1
, an obligatory key core clock gene, was disrupted in diabetes and restored by ATRF in multiple peripheral tissues in
db/db
mice. Using inducible global Bmal1 knockout mice, we further demonstrated that
Bmal1
was partially required for ATRF to protect the BP daily rhythm. Taken together, our results revealed that ATRF potently protects BP daily rhythm in diabetic
db/db
mice in part by modulating autonomic nervous system and clock gene Bmal1. These novel findings suggest ATRF may serve as a novel and effective chronotherapy against the disruption of BP daily rhythm in diabetes.
Abstract 116: BMI is Not a Risk Factor for All-Cause Mortality and Cardiovascular Disease Among African American Women
Authors: Kidambi Srividya , Agarwal Priyanka , Yang Almira , Chelius Thomas , Mattson David , Cowley Allen , Laud Purushottam , Wang Tao , Liang Mingyu , Kotchen Theodore
Published: September 2019

Background:
Across the age spectrum, obesity rates are higher among African Americans (AA) than Whites, particularly women. Previous findings from a large cohort study indicated that risk of all-cause mortality is only moderately elevated with increased weight among AA, but this relationship is a much stronger finding in Whites. In this study, we examined the relationship of baseline body mass index (BMI) to incident all-cause mortality and fatal and non-fatal cardiovascular and renal disease (CVRD) events.

Methods:
Between 1994 - 2006, 1723 AA (52% women, age: 44±7 yrs., BMI: 29 ± 6 kg/m
2
, 51% hypertensive) were recruited for genetic studies of hypertension. Height and weight were obtained at baseline in a standardized manner. Mortality data were obtained from the National Death Index in 2018 on all subjects. Incidence of fatal/non-fatal CVRD events was obtained and confirmed by review of paper and electronic medical records between 2015-2019 in a sub-sample. Subjects who had either a CVRD event or at least 10 years of follow-up data available with no event were included (n=788). Logistic regression (LR) was performed to determine if BMI predicted mortality or a CVRD event.

Results:
There were 325 deaths (19%, time-to-death: 12±3 yrs.) from all causes. During a follow-up of 14±4 yrs., 154 subjects had a CVRD (20%) event/death. There was no difference in age between sexes, but women had a higher BMI compared to men (30±7 vs. 27±5 kg/m
2
, p<0.0001). Men had a higher mortality (23% vs. 15%, p<0.0001) and more CVRD events (24% vs. 16%, p = 0.008). The odds ratio (OR) for death was 1.62 (CI: 1.3-2.1, p=0.0002) in men compared to women. Baseline BMI was not predictive of all-cause mortality in the total sample or in men or women separately. BMI predicted CVRD events in men with an OR of 1.08 (CI: 1.03-1.13), i.e. one-unit increase in BMI in men led to 8% increase in odds for CVRD events. However, BMI was not predictive of CVDR events in women (OR: 1.01 [0.98-1.05]).

Conclusion:
This is the first study to evaluate the effect of BMI longitudinally on incident CVRD events among AA. Although BMI is the most common metric to define obesity, it is not predictive of CVRD and/or mortality risk among AA women. This factor should be considered while determining cardiovascular disease risk status among AA.
Abstract 117: Transplantation of Bone Marrow Cells from miR-150-KO Mice Improves Immune Function and Attenuates Arterial Stiffness and Hypertension in Senescence-Accelerated Mice
Authors: Fan Jun , Wang Shirley , Sun Zhongjie
Published: September 2019

Background and Objective:
Aging is associated with increased prevalence of arterial stiffness and hypertension. The senescence-accelerated mouse (SAMP1) demonstrates early and spontaneous aging phenotypes around 6-8 months of age. In contrast, microRNA-150 knockout (miR-150 KO) slows down the aging process and extends lifespan in mice by 20-30%. The object of this study is to investigate whether transplantation of bone marrow cells (BMCs) from miR-150 KO mice attenuates arterial stiffening and hypertension in SAMP1 mice.

Method and Results:
Pulse wave velocity (PWV) and blood pressure (BP) levels were increased significantly in the SAMP1 mice compared with the age-matched control (AKR/J) mice (10 months), indicating that SAMP1 mice developed arterial stiffening and hypertension. Interestingly, transplantation of bone marrow cells (BMCs) from miR-150-KO mice decreased PWV and BP in SAMP1 mice to nearly control levels within 3 months. BMC transplantation decreased expression of collagen1, TGFβ1, Scleraxis (collagen gene transcription factor), MMP-2 and MMP-9 but increased expression of elastin in aortas of SAMP1 mice, indicating improved arterial remodeling. BMC transplantation from miR-150-KO mice partially rescued the downregulation of B cells lymphocytes and increased the serum level of IgG and IgM, abolished infiltration of T cells and macrophages, and decreased inflammatory cytokine expression in aortas in SAMP1 mice. In the cell culture study, IgG deficiency increased inflammatory factor cytokine expression in endothelial cells, which can be abolished by addition of IgG protein.

Conclusion:
BMC transplantation from miR-150-KO mice reversed arterial stiffness and hypertension in SAMP1 mice likely via modulation of the immune function. This study demonstrates, for the first time, that BMC transplantation may be an effective therapeutic strategy for aging-related arterial stiffness and hypertension.
Abstract 118: Mitochondrial Fission in Vascular Cells Mediates Hypertensive Vascular Remodeling and Inflammation
Authors: Cooper Hannah , Miyao Masashi , Kawai Tatsuo , Preston Kyle , Elliott Kathy , Scalia Rosario , Rizzo Victor , Eguchi Satoru
Published: September 2019

Worldwide, high blood pressure affects nearly 1/3 of adults and is the most common cause of cardiovascular disease-related deaths. While there is effective blood pressure reduction therapy, the underlying pathophysiology leading to the hypertensive end-organ damage including the mechanism of vascular remodeling remains poorly understood. Mitochondrial dysfunction has been implicated in various cardiovascular diseases. Currently, there is little understanding on mechanism and role of mitochondrial quality control in hypertension. Our data shows that activation of mitochondrial fission via a dynamin-related GTPase, Drp1, is involved in hypertensive vascular remodeling. In vivo, 8-week old male C57BL/6 mice were infused with AngII (1000ng/kg/min) for 2 weeks with or without treatment with a Drp1 inhibitor, mdivi1, (25 mg/kg ip every other day). Mdivi1 suppressed vascular hypertrophy and perivascular fibrosis induced by AngII in aorta (thickness Mean±SEM: 0.291±.012 AngII only vs. 0.275±.006 AngII + mdivi1; p=0.07) and heart (LV wall thickness Mean±SEM: 0.423±.034 AngII only vs. 0.219±.017 AngII + mdivi1; p<0.01). Mdivi1 also inhibited AngII-induced left ventricular hypertrophy but not the elevated blood pressure measured continuously by telemetry. Reduced KDEL and nitro-tyrosine staining in coronary and renal arteries suggests attenuation of vascular ER stress and oxidative stress, respectively, in mdivi1 treated mice. To assess the role of acute signaling in this model C57BL/6 mice went through the same protocol for only 3 days. Several inflammatory targets (VCAM1, MMP2, HMGB1) were upregulated in aortas of AngII stimulated mice and attenuated with mdivi1 treatment. In addition, inhibition of mitochondrial fission (mdivi1) attenuated AngII-induced leukocyte adhesion at mesenteric micro-circulation assessed by intravital microscopy.
In vitro
, Ang-II induced inflammatory response in rat aortic endothelial cells (ECs) with THP-1 adhesion assay was attenuated by mdivi1 and an ER stress inhibitor (4-PBA). In conclusion, these data suggest that inhibition of mitochondrial fission prevents AngII-induced cardiovascular remodeling and inflammation through attenuating VSMC as well as EC signaling events.
Abstract 119: Ccl18 as a Potential Mediator of the Pro-Fibrotic Actions of M2 Macrophages in the Vessel Wall During Hypertension
Authors: Zhu Mingyu , Lewis Caitlin V , Finemore Meghan J , Lieu Maggie , Widdop Robert E , Samuel Chrishan S , Drummond Grant R , Kemp-Harper Barbara K
Published: September 2019

M2 macrophages contribute to vascular fibrosis and stiffening in hypertension and may mediate these actions via release of the pro-fibrotic chemokine, CCL18, which they release to a greater extent than TGF-β1. The role of CCL18 and its cognate receptor, CCR8 in hypertension and vascular fibrosis has not been investigated. This study aimed to identify vascular targets of CCL18 and explore its ability to promote fibrosis.

Blood pressure (BP) of saline or angiotensin II (0.7 mg/kg/day, 28 days;
s.c.
)-treated male C57BL/6J mice was measured by tail cuff. Localisation and expression of CCR8 and the murine functional analogue of CCL18, CCL8, were assessed in the thoracic aorta by immunohistochemistry and qRT-PCR, respectively. Human aortic adventitial fibroblasts (AoAFs) and endothelial cells were treated with the pro-fibrotic agent TGF-β1 (10 ng/ml) or CCL18 (3-300 ng/ml) for 3-72h or 7d, respectively. In human AoAFs, expression of pro-collagen I, mature collagen I and α-SMA and were measured (qRT-PCR, Western blotting). Endothelial-mesenchymal transition was measured via VE-cadherin and α-SMA protein expression.

Angiotensin II infusion significantly elevated systolic BP (151±5 mmHg, p<0.05) as compared to saline treated mice (111±5 mmHg, n=8). In aortas from hypertensive mice, mRNA expression of CCL8 was elevated 3-fold (p<0.05, n=4-8), and CCL8 expression in the aortic wall was co-localised with the M2 macrophage marker CD206. CCR8 expression was evident in the endothelium, adventitia and perivascular fat. In human AoAFs, TGF-β1 caused a 2-fold increase in protein expression of collagen I (24h, p<0.01) and α-SMA (24-72h, p<0.05). CCL18 (300 ng/ml) did not change α-SMA expression, but increased the protein expression of pro-collagen I by 2-fold (24h; p<0.01), and elevated mature collagen I by 3.6-fold (72h; p<0.05). In human aortic endothelial cells, CCL18 (10 ng/ml) induced a trend towards reduced VE-cadherin, while increasing α-SMA expression (1.5-fold, p<0.05).

CCL18 stimulates collagen synthesis from human AoAFs and may promote endothelial-to-mesenchymal cell transition. Expression of its receptor in the vascular wall suggests that the CCL18-CCR8 axis may serve as a potential target for the treatment of vascular fibrosis during hypertension.