bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 533: Vascular Smooth Muscle Na/Ca Exchanger-1 Modulates Myogenic Reactivity and Blood Pressure in Chronic Intermittent Hypoxia
Authors: Chen Ling , Wang Youhua , Wang Jingyu , Zhang Jin
Published: July 2021

Chronic intermittent hypoxia (CIH) results in systemic hypertension and altered vascular reactivity in human and rodents. We hypothesize that vascular smooth muscle (VSM) Na/Ca exchanger type-1 (NCX1) is a potential molecular mechanism mediating CIH-induced changes in vascular reactivity and blood pressure (BP). Wild type (WT) and transgenic (TG) mice with VSM-specific NCX1 overexpression were exposed to either CIH (1 min 5% nadir O
2
, followed by 1 min 21% O
2
; 8 hrs/day for 5 weeks), or normoxia (constant 21% O
2
). CIH resulted in greater BP elevation in TG (102 ± 3 mm Hg vs. 92 ± 6 mm Hg in normoxia, P < 0.05, n = 6 and 3, measured by intra-carotid artery catheterization under 1.5% isoflurane) than in WT (88 ± 4 mm Hg vs. 83 ± 2 mm Hg in normoxia, NS, n = 3 and 5) mice. Mesenteric small resistance arteries were then isolated and pressurized for diameter and wall thickness measurement. Compared to normoxia controls, CIH-exposed WT arteries exhibited increased resting myogenic tone (21 ± 2% vs. 16 ± 2% of PD in normoxia, P < 0.05, n = 5 each group). Moreover, myogenic reactivity in WT arteries of CIH-exposed mice shifted downward at lower range of intraluminal pressure (20-80 mm Hg), and upward at higher pressure (100-140 mm Hg). The effects of CIH were significantly greater in TG arteries compared to those in WT arteries, suggesting that VSM NCX1 augmented the myogenic response to CIH. Vasoconstrictions induced by phenylephrine (0.003-100 μM), greater in TG arteries, or high (60 mM) external K
+
solution, were similar between control and CIH groups in the same genotype, suggesting that arterial contractility to agonist stimulation was not affected by CIH. Arterial wall thickness or passive diameters under various intraluminal pressures in Ca
2+
-free solution were not different between control and CIH groups in either genotype, suggesting a lack of arterial stiffness or remodeling. In summary, 1) CIH results in greater BP elevation in TG mice; 2) CIH leads to biphasic changes in myogenic reactivity in small resistance arteries; 3) Overexpression of VSM NCX1 exaggerates BP elevation in CIH, likely by modulating myogenic reactivity, suggesting that VSM NCX1 is a potential mechanism mediating CIH-induced hypertension.
Abstract 552: Aldosterone Enhances Connecting Tubule Glomerular Feedback (CTGF)
Authors: Ren YiLin , D'Ambrosio Martin A , Garvin Jeffrey L , Leung Pablo , Wang Hong , Carretero Oscar A
Published: July 2021

Increasing Na delivery to the connecting tubule (CNT) stimulates epithelial Na channels (ENaC) and dilates the afferent arteriole (Af-Art), a process we call connecting tubule glomerular feedback (CTGF). We hypothesize that aldosterone (aldo) enhances CTGF
via
a nongenomic mechanism that stimulates CNT ENaC
via
GPR30 and/or mineralocorticoid receptors (MR). Rabbit Af-Arts and their adherent CNTs were microdissected and simultaneously perfused. Two consecutive CTGF curves were elicited by increasing luminal NaCl in the CNT. Addition of aldo 10
-8
M to the CNT potentiated CTGF, seen as a left-shift in the concentration of NaCl that elicited a half-maximal response (EC
50
), see Figure. The MR blocker eplerenone (10
-5
M) prevented the enhancement of CTGF by aldo (control EC
50
= 32.4 ± 2.3 mM; aldo + eplerenone EC
50
= 35.4 ± 1.7 mM; n = 7). Neither the transcription inhibitor actinomycin D (5x10
-6
M) nor the translation inhibitor cycloheximide (10
-5
M) prevented the effect of aldo (control EC
50
= 33.0 ± 2.0 mM; aldo + actinomycin D EC
50
= 15.4 ± 1.5 mM; n = 6;
P
< 0.001, control EC
50
= 33.2 ± 2.4 mM; aldo + cycloheximide EC
50
= 11.2 ± 1.3 mM; n = 6;
P
< 0.001). We conclude that aldo in the lumen of the CNT enhances CTGF
via
a nongenomic effect possibly involving MR and/or GPR30 receptors. Enhanced CTGF induced by aldosterone may contribute to renal damage by causing increases in Af-Art dilation and glomerular capillary pressure (glomerular barotrauma).

Figure. Control CTGF (○) seen as dilation of norepinephrine-preconstricted Af-Arts induced by increasing NaCl in the CNT. Aldo 10
-8
M (•) enhanced CTGF (n = 6; *
P
< 0.05; **
P
< 0.01; ***
P
< 0.001;
vs
. control). Vertical dashed lines indicate EC
50
.
Abstract 559: Augmented Renal Levels of PDGFß And PDGFß Receptors in Cyp1a1-Ren2 Transgenic Rats With Angiotensin II-Dependent Malignant Hypertension
Authors: Mitchell Kenneth D , Prieto Minolfa C , Seth Dale M , Davis Porcha D , Bourgeois Camille , Milani Carlo J
Published: July 2021

PDGF receptor antagonism with imatinib mesylate prevents the renal injury, proteinuria and augmented urinary ANG II excretion independent of changes in blood pressure that occur in Cyp1a1-Ren2 rats with ANG II-dependent malignant hypertension. These findings suggest that arterial pressure-dependent increases in PDGFβ protein levels and PDGFβ receptor activation contribute importantly to the marked renal functional and morphological derangements that occur in ANG II-dependent malignant hypertension. To address this issue, the present study was performed to determine the protein levels of PDGFβ and PDGFβ receptors in renal cortical and medullary tissue from kidneys of hypertensive Cyp1a1-Ren2 transgenic rats. Male Cyp1a1-Ren2 rats were fed a diet containing indole-3-carbinol (I3C; 0.3%) for 10 days to induce malignant hypertension (n=5) or a non-I3C containing diet (controls, n=6). Rats induced with I3C developed malignant hypertension and exhibited increases in PDGFβ protein levels in both the renal cortex (0.046±0.002 vs.0.026±0.003 AU, P<0.05) and renal medulla (0.054±0.007 vs.0.026±0.003 AU, P<0.05), and elevated PDGFβ receptor levels in both renal cortex (0.26±0.03 vs.0.12±0.02 AU, P<0.05) and renal medulla (0.60±0.06 vs.0.21±0.02 AU, P<0.05). In a separate group (n=5), rats were chronically treated with the PDGF receptor antagonist, imatinib mesylate, by oral gavage (60 mg/kg/d) starting 3 days before initiating I3C induction and maintained on imatinib for the 10 day duration of I3C administration. Chronic PDGF receptor blockade prevented the increase in PDGFβ protein levels in both renal cortical and medullary tissues (0.027±0.004 vs. 0.026±0.003 and 0.029±0.005 vs. 0.026±0.003 AU, respectively) but did not influence the elevated PDGFβ receptor levels in either renal cortex (0.23±0.02 vs. 0.26±0.02 AU) or renal medulla (0.46±0.03 vs.0.60±0.06 AU). These data demonstrate that both PDGFβ protein and PDGFβ receptor levels are elevated in both renal cortex and medulla in ANG II-dependent malignant hypertension. Such elevated levels may contribute to the renal injury and the increased urinary ANG II excretion in ANG II-dependent malignant hypertension.
Abstract 564: Differential Expression and Regulation of Angiotensinogen in Renal Proximal Tubule Cells From S1, S2 and S3 Segments
Authors: Satou Ryousuke , Hering-Smith Kathleen S , Navar L G
Published: July 2021

In angiotensin II (Ang II)-dependent hypertension, intrarenal angiotensinogen (AGT) augmentation induced by Ang II and associated pathogenic factors including interleukin 6 (IL-6) cause further elevation of intratubular Ang II production, leading to the progression of hypertension and kidney injury. Recent studies have suggested that renal proximal straight tubules (S3 segment) are the main source of intrarenal AGT and that S1 and S2 segments do not express AGT mRNA under normal conditions. However, AGT expression and its regulation by Ang II and/or IL-6 in each proximal tubule segment have not been demonstrated an in vitro setting. The availability of specific cell lines derived from mouse S1, S2 and S3 segments provided an opportunity to decisively determine each segments’ capability to express AGT and respond to stimuli. Thus, this study was performed to determine AGT expression and its response to stimulation with Ang II and IL-6 in S1, S2 and S3 cell line. Basal AGT mRNA and protein levels were detected by RT-PCR and western blot analysis. Basal levels of Ang II type 1 receptor (AT1R) and STAT3, which is a transcription factor in IL-6 signaling pathway, were also measured. In addition, the cells were incubated with 100 nM Ang II and/or 400 nM IL-6 for 24 h. Basal AGT levels in S1 and S3 cells were lower than in mouse whole kidney (0.09-fold and 0.33-fold compared with mouse whole kidney). S2 cells exhibited the highest basal AGT levels (4.15-fold) among these cells. In S1 cells, AGT expression was stimulated by IL-6 (1.89 ± 0.32, ratio to control) and co-stimulation with Ang II and IL-6 (1.85 ± 0.28) although Ang II alone did not alter AGT levels. In S2 cells, only the co-stimulation increased AGT expression (1.35 ± 0.01). No changes were observed by similar treatments in S3 cells. Basal AT1R levels were lower in S3 than in S1 and S2 cells (0.97 ± 0.09 in S2, 0.32 ± 0.07 in S3, ratio to S1). S1 cells showed the highest basal levels of STAT3. Basal STAT3 levels in S3 cells were lower than that in S1 and S2 cells. These results indicate that S2 cells are main source of intrarenal AGT which can be augmented by Ang II and IL-6 during the development of Ang II-dependent hypertension. Furthermore, low basal levels of AT1R and STAT3 in S3 cells explain why these cells do not respond to Ang II and IL-6.
Abstract 603: Effective And Sustained ACE2 Delivery Using Minicircles Technology
Authors: Wysocki Jan , Haber Philipp K , Ye Minghao , Maier Christoph , Osborn Mark J , Batlle Daniel
Published: July 2021

Chronic and sustained amplification of ACE2 activity
in vivo
has required the development of

transgenic mice or the use of viral vectors. Minicircle is a new gene delivery technology

which is resistant to gene silencing, and therefore represents an attractive

platform for gene replacement strategies
in vivo
. Here we cloned cDNA of

soluble mouse ACE2 into a circular expression cassette and the resulting ACE2

minicircle (MC) was injected to female
FVB

mice using iv. hydrodynamic approach (10ug or 30ug/mouse). At 3-7d after MC

administration, serum ACE2 activity in mice that received 10ug ACE2MC (n=9) was

over 100-fold higher than in controls (n=9) (138±48 vs 0.7±0.2 RFU/uL/hr) and in

ACE2MC mice (30ug) (n=8) was almost 1000-fold higher than in controls (n=14) (480

±153 vs 0.5±0.1 RFU/uL/hr, respectively). Mice that received 10 ug ACE2MC were

followed for consecutive serum ACE2 activity monitoring, BP measurements and

plasma Ang levels. The increase in serum ACE2 activity was sustained until the

end of the study (up to 82 days) (Figure).

Despite such a marked increase in serum ACE2 activity in ACE2MC mice, conscious

SBP was not different from controls (137±8 vs 138±7 mmHg, respectively). At the

end of the study, when Ang II was infused acutely (0.2 ug/kg BW i.p.), the

increase in plasma Ang II in ACE2MC mice was significantly reduced compared to

control mice (915±154 vs 1420±131 fmoL/mL, p<0.05).

Mini-circle delivery of ACE2 results in a dose-dependent and sustained long-term

increase in serum ACE2 that efficiently degrades plasma Ang II. Extremely high

increases in serum ACE2 activity do not reduce BP probably due to activation of

non-ACE2 dependent compensatory Ang-hydrolyzing pathways.
Abstract 608: Circulating Soluble Insulin Receptor Level Is Associated With Left Ventricular Hypertrophy And Cardiac-dysfunction Independent Of Blood Pressure And Glucose
Authors: Takata Yasunori , Funada Junichi , Hiasa Go , Matsumoto Yuji , Sato Sumiko , Yoshii Toyofumi , Okayama Hideki , Kawamura Ryoichi , Onuma Hiroshi , Osawa Haruhiko
Published: July 2021

Recent studies have demonstrated that both insulin resistance (IR) and diabetes were the critical risk factors of heart failure (HF). Importantly, it is reported the high prevalence of previously unknown left ventricular dysfunction including the prevalence of HF preserved EF (HFPEF) in diabetic subjects. However, the underlying mechanism and a biomarker have not been well defined.

To identify underlying mechanism and candidate biomarker of HF in subjects with type 2 diabetes (T2DM), we performed retrospective study of 143 T2DM and 181 non-diabetic subjects. We evaluated cardiac function by echocardiography measuring left ventricular ejection fraction (LVEF), left ventricular mass index (LVMi) and the ratio of early transmitral flow velocity to tissue doppler early diastolic mitral annular velocity (E/e’).

Here we demonstrate that plasma soluble insulin receptor (sIR), a novel biomarker of diabetes, was associated with left ventricular hypertrophy (LVH) and cardiac-dysfunction. Simple regression analysis revealed that circulating sIR was positively correlated with fasting glucose and negatively correlated with fasting insulin. Furthermore, sIR was positively correlated with LVMi and E/e’, and inversely correlated with LVEF (
R
=0.25;
p
<0.01,
R
=0.22,
p
=0.03,
R
=-0.19;
p
=0.02, respectively). Moreover, multivariable regression analysis revealed that sIR was associated with LVMi, E/e’ and LVEF after adjustment for age, gender, BMI, plasma fasting glucose, systolic blood pressure, and eGFR in subjects with T2DM (
β
= 0.26;
p
<0.01,
β
=0.25;
p
=0.02,
β
=-0.23;
p
<0.01,respectively). It is reported that circulating sIR is associated with cleavage of the extracellular domain of insulin receptor and sIR binds to circulating insulin. Thus, our results suggest the possibility that the influence of sIR on insulin signaling may be involved in LVH and cardiac-dysfunction in subjects with T2DM.
Abstract 639: Fructose Potently Stimulates NKCC2 Trafficking in rat Thick Ascending Limbs
Authors: Ares Gustavo R , Ortiz Pablo A
Published: July 2021

Fructose consumption has increased dramatically in the last 30 years and is directly linked to the epidemic of obesity, type-2 diabetes and also hypertension. A moderately high fructose intake induces salt-sensitive hypertension and enhances urinary excretion of fructose. Despite the potential implications of fructose in renal salt handling, very little is known about its effect along the nephron. The thick ascending limb (TAL) reabsorbs 25% of filtered NaCl, primarily
via
the apical Na/K/2Cl cotransporter (NKCC2). Enhanced NKCC2 activity and trafficking is related to salt-sensitive hypertension in genetic animal models. Thus, we hypothesized that fructose acutely stimulate NKCC2 trafficking to the apical surface of TALs. To test this hypothesis we measured membrane and total NKCC2 expression as well as phosphorylation of NKCC2 at Ser126 and Thre96,101 in suspensions of rat TALs. We found that acute treatment (20 minutes) of TALs with fructose potently enhanced apical surface NKCC2 in a concentration-dependent manner (Baseline: 100%; fructose 1 mM= 122 ± 5%, fructose 5 mM= 49 ± 10%, fructose 10 mM = 201 ± 59%;
p
<0.05). Acute fructose treatment did not increase NKCC2 phosphorylation at Ser126 or Thre96,101 and did not affect total NKCC2 expression. To test if the effect of fructose on apical surface NKCC2 was specific to this hexose, we studied the effect of glucose and fructose 1,6 bisphosphate (1,6BP). We found that 20 minute-incubation with glucose (5 mM) and fructose 1,6BP (5 mM) did not affect baseline surface NKCC2 levels (baseline = 100, glucose (5 mM)= 101 ± 7%, fructose 1,6BP (5 mM)= 92 ± 6%, not significant). We concluded that acute fructose treatment, at physiologically relevant concentrations, increases membrane NKCC2 expression without enhancing its phosphorylation. Compared to β-adrenergic receptor stimulation and vasopressin (which enhance apical surface NKCC2 in TALs by 25-40%, fructose (but not glucose) is a very potent agonist of NKCC2 trafficking to the apical membrane of rat TALs. Our data suggest that fructose-stimulated TAL NaCl reabsorption could contribute to the salt-sensitive hypertension caused by a high fructose diet.
Abstract 85: Low-grade Inflammation Participates on the Enhanced Vasoconstriction to the Endogenous Endothelin-1 in Small Vessels From Obese Patients
Authors: Virdis Agostino , Dell'Agnello Umberto , Duranti Emiliano , Rossi Chiara , Anselmino Marco , Chiarugi Massimo , Solini Anna , Taddei Stefano
Published: July 2021

Objective:
Obesity is characterized by vascular low-grade inflammation, with an augmented endogenous endothelin(ET)-1-mediated vasoconstriction (VC) and a blunted tonic nitric oxid(NO)-mediated relaxation (VD). We evaluated whether TNF-α, localized in the vascular wall and in perivascular adipocytes (PVA), contributes to the VC induced by endogenous ET-1 and whether this action is mediated by an effect on tonic NO release in small resistance arteries from obese patients (Ob) and controls (Ctrl).

Method:
Sample of visceral fat were obtained in 14 Ob (BMI:42.1±3.7) and 14 Ctrl (BMI:25.4±3.6), matched for metabolic profile and blood pressure values, undergoing laparoscopic surgical procedure. Small arteries were investigated on a pressurized micromyograph. Endogenous ET-1 was assessed by vascular response to BQ-123. TNF-α and NO were assessed by Infliximab (IFX) and L-NAME, respectively. Gene and protein expression of TNF-α, ET-1 and ETA receptors were determined by RT-PCR (gene/reference) and IHC (AU) on arterial wall and in PVA.

Results:
In Ctrl, L-NAME-induced VC (15.5±0.6%) was not affected by IFX (15.1±0.4%). In contrast, Ob showed a blunted VC to L-NAME (6.0±0.7%; P<0.01 vs Ctrl) which was potentiated (P<0.01) by IFX (12.5±0.8%). In Ob, the VD to BQ-123 (47.0±1.5%) was attenuated (P<0.01) by IFX (29.1±2.4%) and not affected by L-NAME (43,3±0.6%). During IFX co-infusion, L-NAME further reduced the VD to BQ-123 (19.4±3.0%; P<0.01 vs BQ-123+IFX). In Ctrl, VD to BQ-123 was blunted (26.3±1.3%; P<0.01 vs Ob), not affected by IFX (24.1±0.6%) and significantly reduced by L-NAME (12.3±1.1%). Ob showed a significant overexpression of TNF-α respect to Ctrl, either at the level of arterial wall (24.9±19.6 vs 2.8±2.5 AU, P<0.001) or in PVA (2.9±1.8 vs 1.2±0.7 gene/reference, P<0.005). These results were paralleled by a higher arterial expression of ET-1 (45.8±10.3 vs 24.3±15.0 AU, P<0.01) and ETA receptors (69.4±6.0 vs 9.6±2.8 AU, P<0.001) in Ob vs Ctrl.

Conclusion:
Small vessels of Ob show an enhanced ETA-mediated VC and a blunted NO-mediated VD. An excess of vascular and perivascular TNF-α, coupled with an increased expression of ET-1 and ETA in the vasculature of Ob, contributes to the enhanced ET-1-mediated VC tone partly by an impairment of tonic NO release.
Abstract 91: Inhibition of Toll-like Receptor 4 Suppresses Reactive Oxygen Species Generation in Diabetic Vasculature
Authors: Carrillo-Sepulveda Maria Alicia , Wenceslau Camilla , Webb R. Clinton
Published: July 2021

Reactive oxygen species (ROS) are augmented in diabetes and play a central role in vascular dysfunction. TLR4, a key component of the innate immune system, is elevated during diabetes; however, the mechanistic link between TLR4, ROS, and vascular dysfunction remains elusive. Our previous results show that under high glucose (HG), TLR4 is upregulated in vascular smooth muscle cells (VSMCs) and mediates HG-induced ROS production. We hypothesized that TLR4 augments ROS in diabetic resistance vessels, which contributes to vascular dysfunction. To test our hypothesis, streptozotocin (STZ)-induced diabetic Sprague-Dawley rats (65mg/kg; 5weeks) were treated with 50ug/day of CLI-095, an inhibitor of TLR4, over 14 days. Glucose levels were increased (390±13 vs. 97±8.2mg/dL control) and body weight decreased (310±38.4 vs. 385±21g control) in the STZ group. These effects were unchanged by CLI-095 treatment. Mesenteric resistance arteries (MRA) from STZ rats exhibited impaired acetylcholine-induced relaxation (35.26±4.38% vs. 81.83±2.61% control, p<0.05), and CLI-095 treatment ameliorated this effect (71.99±3.48%, p<0.05). Moreover, dihydroethidium staining showed that the increase in ROS due to STZ (1.8 fold vs. control, p<0.05) was attenuated after CLI-095 treatment (1.6 fold vs. STZ, p<0.05). To test if TLR4 signaling is activated in MRA from the STZ rats, MyD88 protein levels, a downstream adaptor molecule, were measured. Arteries from the STZ rats showed increased expression of MyD88 (1.6 fold vs. control, p<0.05) which was reduced after CLI-095 treatment (1.2 fold vs. STZ, p<0.05). Finally, we assessed if TLR4 signaling in VSMCs following HG treatment is altered. Immunoprecipitation showed that HG did not alter the interaction between TLR4 and MyD88 in endothelial cells. In contrast, HG conditions increased TLR4 and MyD88 interaction in VSMCs. Concurrently, HG-induced activation of NFKB in VSMCs was diminished in cells pre-treated with CLI-095. Together our data suggest that activation of TLR4 signaling in VSMCs leads to ROS generation thereby contributing to a reduction in nitric oxide bioavailabitity contributing to endothelial dysfunction in diabetes. Thus, TLR4 is a putative target for the treatment of diabetic vascular complications.
Abstract 99: Ac-SDKP Protects the Heart From Post-myocardial Infarction Remodeling and Dysfunction in Mice
Authors: Peng Hongmei , Xu Jiang , Yang Xiao-Ping , Dai Xiangguo , Petersion Edward , Carretero Oscar , Rhaleb Nour-Eddine
Published: July 2021

N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) has anti-inflammatory, anti-fibrotic and pro-angiogenic effects. We previously observed that Ac-SDKP reduces incidence of cardiac rupture in mice with acute myocardial infarction (MI), which is associated with its anti-inflammatory and pro-angiogenic properties. We hypothesized that Ac-SDKP,
via
its anti-fibrotic and pro-angiogenic actions, ameliorates left ventricular (LV) dilatation and fibrosis, thus improving cardiac function post-MI. C57BL/6J mice were divided into three groups: 1) sham MI, 2) MI + vehicle (VEH), and 3) MI + Ac-SDKP. Ac-SDKP (1.6 mg/kg/day) was given immediately after MI induction (
i.p.
via
osmotic pump) for 5 weeks. Cardiac remodeling and function were assessed by echocardiography, and interstitial collagen fraction (ICF) and capillary density was determined histologically. We found that Ac-SDKP 1) reduced LV remodeling, evidenced by decreased LV chamber dimensions and areas, and reduced ICF and increased capillary density, with no changes in posterior wall thickness and LV weight (LVW), and 2) improved cardiac function, demonstrated by increased fractional shortening (FS) and ejection fraction (EF) (Table). We conclude that in murine models of MI, Ac-SDKP protects the heart from more severe remodeling and dysfunction, possibly
via
its anti-fibrotic and pro-angiogenic actions. Thus, the use of Ac-SDKP or its analogues could be a new and effective alternative in restoring cardiac function in patients after MI.
Abstract 010: Adipocyte-derived Aldosterone And Cortisol Are Nox1/4 Dependent: Implications In Obesity-associated Vascular Dysfunction.
Authors: Even Sarah E , Rios Francisco J , Nguyen Dinh Cat Aurélie , Antunes Tayze T , Briet Marie , He Ying , Callera Glaucia E , Jandeleit-Dahm Karin , Cooper Mark , Szyndralewiez Cedric , Page Patrick , Montezano Augusto C , Touyz Rhian M
Published: July 2021

We previously demonstrated that aldosterone (aldo) is produced by adipocytes, an effect associated with reactive oxygen species (ROS) production and adipokine production, which influences vascular function. These processes are exaggerated in obesity. Whether ROS themselves play a role in adipocyte-derived aldo is unclear. Studies were performed in db/m (lean) and db/db (obese) mice, treated with low (20mg/kg/day) or high dose (60mg/kg/day) GKT137831 (GKT, Nox1/4 inhibitor, 16 weeks). Epididymal (EVAT) and perivascular (PVAT) fat were collected. Human adipocytes (SW872) were also studied. Aldo and corticosterone levels were measured by ELISA. Gene expression was assessed by qPCR. ROS generation was assessed by chemiluminescence and amplex red. Plasma aldo levels in db/db (pg/mL: 518 vs. 272g) and aldo levels in culture media from db/db adipocytes were increased (pg/mL/μg RNA: 1964 vs. 388), p<0.05. All effects were decreased by high dose GKT. In PVAT, CYP11B2 gene expression was increased in db/db (2.6±0.8 vs control 1.1±0.1, p<0.05), an effect blocked by Nox1/4 inhibition. Corticosterone levels in culture media from db/db adipocytes were also increased. Gene expression of adipocyte differentiation marker, AP2, was increased (3.5±1.1 vs control 1.4±0.4) in obese mice. GKT decreased AP2 levels. In human adipocytes, AngII stimulation increased aldo (6 fold) and cortisol (4 fold) production, as well as superoxide (1 fold) and H2O2 (2 fold) levels (p<0.05 vs vehicle). Increased levels of superoxide by Ang II were blocked by GKT and ML171 (Nox1 inhibitor); while Ang II-induced H2O2 production was inhibited only by GKT. Ang II-induced aldo production was blocked by tempol (SOD mimetic), GKT and ML171. In contrast, cortisol was only blocked by tempol and GKT. In conclusion, aldo production in adipocytes is dependent on ROS formation and involves Nox1 and Nox4. Nox4 also influences adipocyte-derived cortisol. These data suggest that Nox1/4 may play a role in adipocyte-derived aldosterone and cortisol production, effects that are amplified in obesity. Our findings suggest that adipocyte Nox1/4 may be a putative therapeutic target in obesity-associated hyperaldosteronism and cardiovascular damage.
Abstract 016: Bbs1 Gene Deletion From The Leptin Receptor Neurons Causes Obesity And Hypertension In Mice
Authors: Guo Deng F , Morgan Donald A , Grobe Justin L , Nishimura Darryl , Searby Charles , Sheffield Val C , Rahmouni Kamal
Published: July 2021

Bardet-Biedl syndrome (BBS) is a pleiotropic autosomal recessive disorder associated with several features including obesity and hypertension. Deletion of BBS genes globally or in the nervous system recapitulated many of the BBS phenotypes including obesity and hypertension. Here, we assessed the effect of ablating the
Bbs1
gene from the neurons expressing the long signaling form of the leptin receptor (LepRb). Breeding Bbs1
flox
with LepRb
Cre
mice created mice deficient in the
Bbs1
gene only in LepRb-positive neurons (visualized by tdTomato expression) as indicated by loss of leptin activation of Stat3. Importantly, Bbs1
flox
/LepR
Cre
mice display an obesity phenotype as indicated by the increased (P<0.05) body weight (39±2 vs. 30±1 g in controls) and fat mass measured by MRI (14±3 vs. 4±1 g in controls) associated with increased (P<0.05) food intake (3.4±0.1 vs. 2.9±0.1 g in controls) in 25 weeks old mice. However, body weight and fat pads of pair-fed LRb
Cre
/Bbs1
fl/fl
mice remained significantly elevated compared to controls suggesting that LRb
Cre
/Bbs1
fl/fl
mice have reduced energy expenditure. Consistent with this possibility, LRb
Cre
/Bbs1
fl/fl
mice displayed decreased (P<0.05) O
2
consumption (2.6±0.1 vs. 3.1±0.1 mL/100g/min in controls) and heat production (8.1±0.3 vs. 9.6±0.3 kcal/kg/h in controls). These results indicate that hyperphagia and decreased energy expenditure contribute to the development of obesity in Bbs1
flox
/LepR
Cre
mice. Next, we assessed the effect on arterial pressure (AP) and sympathetic nerve activity (SNA) of ablating the
Bbs1
gene from the LepR-containing neurons. Interestingly, deletion of the
Bbs1
gene in LepR neurons recapitulates the hypertension phenotype of BBS as indicated by elevated mean AP (125±4 vs 109±3 mmHg in controls, P=0.03). Conscious renal SNA was also elevated in LRb
Cre
/Bbs1
fl/fl
mice relative to controls (97±8 vs 62±10 spikes/sec, P<0.05). Finally, the depressor effect of ganglionic blockade (hexamethonium) was exaggerated in Bbs1
flox
/LepR
Cre
mice (-57±5 vs -38±5 mmHg in control, P=0.01). These findings demonstrate that the
Bbs1
gene in LepR neurons is critical for energy homeostasis and arterial pressure regulation.
Abstract 036: Pomc Neurons, But Not Agrp Neurons, Are Required For Leptin-induced Sympathetic Activation
Authors: Bell Balyssa B , Harlan Shannon M , Morgan Donald A , Rahmouni Kamal
Published: July 2021

Leptin acts in the brain to decrease food intake and promote energy expenditure by increasing sympathetic nerve activity (SNA) to thermogenic brown adipose tissue. Leptin also increases SNA to other beds including kidney with implications for obesity-induced hypertension. We previously demonstrated the importance the arcuate nucleus (Arc) of the hypothalamus in mediating leptin-induced increases in regional SNA, but the specific neuronal population within the Arc that mediates these responses is unknown. We hypothesized that agouti-related peptide (AgRP) and/or proopiomelanocortin (POMC) neurons of the Arc are critical for the increases in SNA in response to leptin. To test this, we generated mice lacking the leptin receptor specifically in AgRP or POMC neurons by crossing the LepR
flox/flox
mice with AgRP
Cre
or POMC
Cre
mice. Consistent with previous reports, both AgRP
Cre/
LepR
flox/flox
mice and POMC
Cre
/LepR
flox/flox
mice have a slightly elevated body weight relative to littermate controls. Next, we used multifiber sympathetic nerve recording to assess the SNA effects of leptin. Intracerebroventricular (ICV) injection of leptin (2 μg) led to a comparable increase in renal SNA in control mice (210±93%) and AgRP
Cre
/LepR
flox/flox
mice (191±53%). AgRP
Cre/
LepR
flox/flox
mice also displayed a normal lumbar SNA response to ICV leptin (384±86%) relative to controls (325±46%). In contrast, POMC
Cre
/LepR
flox/flox
mice exhibited a significantly reduced renal SNA response to ICV leptin (5±17%) as compared to controls (174±45%). Lumbar SNA response to leptin was also blunted in POMC
Cre
/LepR
flox/flox
mice (24±21%) as compared to controls (358±53%). Thus, deletion of the leptin receptor from POMC neurons, but not AgRP neurons, interferes with the ability of leptin to increase sympathetic traffic. These results demonstrate that the sympathetic nerve responses evoked by leptin emanate from leptin action on POMC neurons.
Abstract 047: 20-SOLA, A Novel Water-soluble 20-HETE Antagonist, Elicits Natriuresis And Abrogates Hypertension In Cyp4a14 Knockout Mice.
Authors: Pandey Varunkumar G , Garcia Victor , Joseph Gregory , Zhang Frank F , Shkolnik Brian , Mishra Priyanka , Manthati Vijaya L , Falck John R , Capdevila Jorge , Schwartzman Michal L
Published: July 2021

20-HETE (20-hydroxyeicosatetraenoic acid) is a cytochrome P450 (CYP) 4A-derived arachidonate metabolite. In the vasculature, 20-HETE increases smooth muscle contraction, endothelial dysfunction/activation and sensitivity to constrictor stimuli; all of which contribute to hypertension. In renal tubules, 20-HETE inhibits ion transport and has pro-natriuretic functions in salt-sensitive models of hypertension. The pro-natriuretic effect of 20-HETE has not been reported in mouse models of 20-HETE-dependent hypertension. The CYP4a14 knockout (KO) male mice exhibit androgen-dependent upregulation of Cyp4a12, the sole 20-HETE synthase in mice, and display 20-HETE-dependent hypertension; female KO mice show no increase in Cyp412-20-HETE and are normotensive. We examined the effect of 20-SOLA (2,5,8,11,14,17-hexaoxanonadecan-19-yl-20-hydroxyeicosa-6(Z),15(Z)-dienoate), a water-soluble 20-HETE antagonist, and determined its effect on blood pressure (BP) and natriuresis in the KO mice. Administration of 20-SOLA (10mg/kg/day in drinking water) normalized BP in male KO mice; a significant reduction was observed on day 4 (146±1 vs153±2 mmHg in untreated littermates) reaching BP of the normotensive female KO mice (124±1 and 126±1 mmHg for treated and untreated, respectively) at day 10 of treatment (124±1 vs. 153±2 mmHg in untreated male KO; p<0.05). 20-SOLA decreased vascular reactivity to phenylephrine in male KO renal microvessels by 7-fold (EC50: 1.58±0.23 vs 0.22±0.03 μM). Moreover, treatment with 20-SOLA increased urine output in KO male mice as compared to untreated littermates (1.25±0.04 vs. 1.06±0.04 mL; p<0.05). The urine output to water intake ratio was significantly elevated in 20-SOLA-treated KO vs. untreated (0.25±0.01 vs. 0.19±0.01, p<0.05). Importantly, treatment with 20-SOLA increased urinary sodium excretion in the KO male mice (12.33±0.44 vs. 8.33±0.63 μmol/gr BW/24Hrs; p<0.05). These results suggest that 20-HETE has an anti-natriuretic effect in the kidneys of KO mice and that antagonism of 20-HETE’s action using 20-SOLA elicited natriuresis concurrent with reduction in blood pressure. The potential vascular and tubular mechanisms underlying 20-HETE anti-natriuretic action are being currently explored.
Abstract 050: Endothelin-1 Overexpression In Endothelial Cells Increases Blood Pressure In An Endothelin Type A Receptor-Dependent Manner
Authors: Coelho Suellen C , Rautureau Yohann , Rehman Asia , Offermanns Stefan , Paradis Pierre , Schiffrin Ernesto
Published: July 2021

Introduction:
The mechanisms of blood pressure (BP) regulation by endothelin (ET)-1 produced by endothelial cells (EC) are complex and remain unclear. Transgenic mice with constitutive EC-specific human ET-1 (
EDN1
) overexpression presented vascular injury but no change in BP, which could be due to adaptation to life-long high ET-1 exposure. We have now generated an inducible EC-restricted
EDN1
overexpressing mouse (ieET-1) in order to demonstrate the effects of ET-1 on BP regulation independent of developmental effects.

Method:
Two transgenic mouse lines (C134 and C170) expressing chloramphenicol acetyltransferase (
cat
) and
EDN1
before and after Cre-mediated excision, respectively, were crossed with mice expressing tamoxifen-inducible CreER
T2
under the control of
Tie2
promoter (ieCre) to generate ieET-1 mice. Mice were treated with tamoxifen (1 mg/kg/day, SC) or vehicle for 5 days and sacrificed 16 days later. Additional mice were treated with 5 or 10 mg/kg/day PO of ET type A receptor blocker, atrasentan, from day 10. BP was determined by telemetry. Plasma ET-1 levels were assessed by ELISA. ET type A and B receptors expression,
Ednra
and
Ednrb
, were evaluated in the kidney by quantitative PCR.

Results:
Tamoxifen increased plasma ET-1 in ieET-1 C134 and C170 (7.1±0.7 and 13.2±2 pg/mL, respectively,
P
<0.01), when compared to control ieET-1 C134 and C170 (0.7±0.2 and 1.0±0.3 pg/mL) and tamoxifen-treated ieCre

(0.8±0.1 pg/mL). ET-1 overexpression increased night systolic BP in ieET-1 C134 and C170 (137±4 and 132±4 mmHg) compared to tamoxifen-treated ieCre (116±7 mmHg), which was reversed partially or completely with 5 or 10 mg/kg/day of atrasentan, respectively (122±3 and 115±4 mmHg,
P
<0.01). ET-1 overexpression in ieET-1 C134 was accompanied by increased
Ednra
expression in renal cortex and medulla (2.9±0.6 and 8.2±3.0 fold,
P
<0.05) and
Ednrb
levels in the renal cortex (2.0±0.3 fold,
P
<0.05) and a trend to increased levels in renal medulla (3.8±1.3 fold) compared to control ieET-1 C134.

Conclusion:
Our results demonstrate that this new inducible EC-restricted
EDN1
overexpressing mouse exhibits ET-1-dependent elevated BP mediated by ET type A receptors. The increased expression of renal ET receptors could play a role in ET-1-induced BP rise.
Abstract 053: The Role of Adventitial Sca-1+ Progenitor Cells in Angiotensin II-induced Aortic Stiffening
Authors: Wu Jing , Montaniel Kim , Hoglund Virginia , Dong Xiurong , Chen Wei , Majesky Mark W
Published: July 2021

The aortic adventitia contains progenitor cells that express the surface marker stem cell antigen-1 (Sca-1). Sca-1+ cells have been reported to transform into smooth muscle cells and promote neo-intimal formation. We have previously found that hypertension is associated with striking adventitial collagen deposition that contributes to aortic stiffening. We hypothesized that Sca-1+ progenitor cells serve as a source of adventitial collagen. Using confocal microscopy we found that Sca-1+ cells represent approximately one fourth of total adventitial cells and that chronic angiotensin II infusion increases their presence by approximately 3-fold. Flow cytometry confirmed that aortic Sca-1+CD31- cells are tripled in angiotensin II-induced hypertension (5.6±1.6 х103 vs. 15.9±2.4 х103 cells/per aorta). We isolated aortic Sca-1+ cells using magnetic-activated cell sorting and found collagen 1a1, 3a1, 5a1 and fibronectin-1 genes were significantly upregulated from 2.7 to 5.4-fold in these cells in angiotensin II-induced hypertension. These fibrotic genes were also upregulated by 2-3 fold in CD31+ cells suggesting endothelial to mesenchymal transition. Immunofluorescence staining demonstrated colocalization of Sca-1 and markers of fibroblasts such as vimentin, fibroblast specific protein-1 (FSP-1) and collagen I, indicating differentiation of these progenitor cells into fibroblasts during the onset of hypertension. Using bone marrow transplantation of EGFP+ cells, we find that angiotensin II stimulates mobilization of these cells from the marrow, and that they home to the vascular adventitia, where they shed Sca-1 and c-kit markers as they begin to produce collagen I. In conclusion, aortic Sca-1+ progenitor cells, both resident and bone marrow-derived, differentiate into fibroblasts and contribute to adventitial collagen deposition and promote aortic stiffening in response to hypertensive stimuli like angiotensin II. Our data also show that the bone marrow is responsive to angiotensin II through mechanisms that remain to be defined.
Abstract 064: Toll-like Receptor 2 Signaling Contributes To Cerebrovascular Dysfunction And Decreased Cerebral Blood Flow In Type-1 And Type-2 Diabetes.
Authors: Hardigan Trevor , Hoda Nasrul , Abdelsaid Mohammed , Ergul Adviye
Published: July 2021

We have shown that type 2 diabetic Goto-Kakizaki (GK) rats develop cerebrovascular endothelial dysfunction, impaired cerebral blood flow (CBF) and mild cognitive deficits. It is increasingly recognized that vascular cognitive impairment may be a new complication of the disease in both type 1 and type 2 diabetes. Toll-like receptor-2 (TLR2) plays a role in cardiovascular complications of diabetes but its involvement in diabetic cerebrovascular disease is unknown. Since brain function heavily depends on constant perfusion, and decreased CBF precedes development of inflammation and cognitive deficits, we hypothesized that enhanced TLR2 signaling in both type 1 and 2 diabetes would contribute to cerebrovascular dysfunction and decreased CBF. Endothelium-dependent relaxation was assessed by measuring acetylcholine (ACh, 10-9 -10-4 M) induced dilatory response in basilar arteries from GK rats in the presence and absence of an anti-TLR2 (1μg) antibody. Vascular contractility to serotonin (10-9 -10-5 M) stimulation was also assessed. Area under the curve (AUC) and maximal effective concentration (Emax as % of max KCl response) were calculated as indices of total relaxation and total contraction, respectively. Basilar artery relaxation was significantly improved in the vessels preincubated (30’) with antiTLR2 (184.8± 24.0 vs. 87.7± 4, p =0.007). The EMax in response to serotonin stimulation in the anti-TLR2 treated vessels was not significantly different than the vessels from the untreated diabetic GK rats (110.6± 4.9% vs 99.4.±4.5%). To further assess the in vivo functional effects of TLR2 signaling, CBF (relative intensity) was measured using laser speckle imaging in wild type and TLR2-knockout (KO) mice using an STZ induced diabetes model. Six weeks after induction of diabetes, wild-type diabetic mice exhibited a significant decrease in CBF vs. control (210±22.5 vs. 300.3±18.4, p<0.05). This decrease in cerebral perfusion was attenuated in the TLR2-KO diabetic mice compared to TLR2 KO control (322.6±10.1 vs. 344. 5±11. 04). These findings suggest that TLR2 signaling leads to vascular dysfunction through decreased endothelium dependent relaxation, and could contribute to decreased CBF in diabetes predisposing to vascular cognitive impairment.
Abstract 077: Renal Denervation Attenuates Salt-Sensitive Hypertension and Oxidative Stress in Rats with Unilateral Hydronephrosis
Authors: Peleli Maria , Al-Mashhadi Ammar , Yang Ting , Persson Erik G , Carlstrom Mattias
Published: July 2021

Hydronephrotic rats and mice have impaired renal function and develop salt-sensitive hypertension, which are associated with oxidative stress. Increased sympathetic nerve activity and oxidative stress in the kidney may play important roles in renovascular hypertension. This study aimed to investigate the contribution of renal sympathetic nerve activity in the development of hypertension in hydronephrosis. A partial unilateral ureteral obstruction (PUUO) was created in 3-weeks old rats to induce hydronephrosis. Surgical denervation, or sham operation, of the PUUO kidney was performed at the time of ureteral obstruction and again 4-weeks later during implantation of a telemetric blood pressure device. Hydronephrotic animals had higher blood pressure (115±3 mmHg) compared with controls (87±1 mmHg), and the blood pressure elevation to a high salt diet was more pronounced (15±2 vs 5±1 mmHg) (p<0.05). Hydronephrosis was also associated with increased urine production (40±4 μl/24h/gBW) and lower urine osmolality (1242±109 mOsm/kg H2O) compared with controls (28±3 μl/24h/gBW and 1751±83 mOsm) (p<0.05). Renal denervation in rats with PUUO attenuated hypertension (97±3 mmHg) and normalized salt-sensitivity (5±1 mmHg), urine production (32±2 μl/24h/gBW) and urine osmolality (1586±127 mOsm/kg H2O) (p<0.05). NADPH oxidase activity in renal cortex from PUUO rats was increased compared with controls (4608±396 vs 3373±217 CLU/min/mg protein) (p<0.05). This was associated with increased cortical mRNA expression of Nox2 (2.3±0.43), p22phox (2.65±0.67) and p47phox (1.39±0.23) compared with controls (p<0.05). Remarkably, denervation in PUUO rats normalized both NADPH oxidase activity (3363±258 CLU/min/mg protein) and mRNA expression of Nox2, p22phox and p47phox (p<0.05). Interestingly, also myocardial tissue from PUUO displayed increased mRNA expression of Nox2 (1.68±0.23) and p22phox (2.82±0.51) compared with control rats, and this was normalized by renal denervation (0.81±0.43) (p<0.05). In conclusion, renal denervation of the hydronephrotic kidney attenuates hypertension and salt-sensitivity, and restores renal excretion pattern. Mechanistically, this is associated with reduced renal NADPH oxidase activity and expression.
Abstract 084: Activation Of Trpv1-expressing Renal Sensory Nerves With N-oleoyldopamine Attenuates High Fat Diet-induced Impairment In Renal Function
Authors: Yu Shuang-Quan , Wang Donna H
Published: July 2021

Renal injury occurs in obesity. Accumulating evidence indicates that activation of the transient receptor potential vanilloid 1 (TRPV1) protects tissues from injury albeit the mechanisms are largely unknown. We test the hypothesis that high fat diet (HFD) intake impairs afferent renal nerves expressing TRPV1 channels, leading to increased renal sympathetic nerve activity (RSNA), decreased renal function, and hypertension, and that chronic activation of TRPV1 positive afferent renal nerves attenuates HFD-induced impairment. N-oleoyldopamine (OLDA, 1 ng/kg, daily), a selective TRPV1 agonist, was administrated intrathecally (T8-L3) via a indwelled catheter to chronically activate TRPV1 positive afferent renal nerves in rats fed a HFD or normal fat diet (Con) for 8 weeks. HFD decreased renal TRPV1 expression and afferent renal nerve activity (ARNA, Con: 133±8, Con+OLDA: 127±15, HFD: 84±5, HFD+OLDA: 115±7, p<0.05) in response to intra-pelvis perfusion of capsaicin (4 μM, 3 min, 20 ml/min), a selective TRPV1 agonist, which were prevented by OLDA. HFD increased RSNA responses to intrathecal injection of muscimol (3 nmol/kg), a GABA-A receptor agonist, and urinary norepinephrine levels, which were prevented by OLDA. HFD decreased creatinine clearance and increased urinary albumin levels, which were prevented by OLDA (Creatinine clearance, Con: 0.52±0.09, Con+OLDA: 0.54±0.12, HFD: 0.24±0.04, HFD+OLDA: 0.40±0.06 ml/min/100 gbwt, p<0.05). HFD increased levels of collagen deposition, connective tissue growth factor (CTGF), and matrix metalloproteinase-2 (MMP-2) in the kidney, which were prevented by OLDA. HFD increased systolic blood pressure by the week of 6 after HFD, which was prevented by OLDA. Thus, HFD impairs TRPV1-positive afferent renal nerves, increases renal sympathetic nerve activity, and leads to renal injury and hypertension. Segment-specific intrathecal injection of OLDA protects against HFD-induced impairment in afferent renal nerves and prevents HFD-induced renal injury and hypertension. Our data illustrate that preservation of TRPV1 positive afferent renal nerves may be a therapeutic strategy in preventing obesity- or type 2 diabetes-induced renal injury and hypertension.
Abstract 203: Adoptions of Smartphones, Tablets, and Mobile Applications Among Patients with Hypertension
Authors: Benvenuto Victor , Baquero Giselle , Scher David L , LaPine Stacey , Fragin Jason , Heron Kristin , Douglas Leslie , Nickolaus Michelle , Curry William , Banchs Javier E
Published: July 2021

optimize management, monitoring, and therapy compliance in patients with hypertension (HTN). It could potentially prevent unnecessary emergency room visits and hospital admissions.

Methods:
We surveyed patients with HTN in cardiology and primary care clinics regarding their use of mobile technology.

Results:
148 patients were included (79 female; age 16-64: 47%, >65: 53%, 70 male; 18-64:41%, >65: 59%). Associated diagnosis were coronary artery disease (21%), myocardial infarction (13%), arrhythmias (36%), heart failure (6%), or other forms of heart disease (3%). 83% own a cell phone, 29% are smartphones. 78% own a personal computer, laptop, iPod, or tablet. 35% report using APPS, of which 26% use health-related applications. 2% used APPS for blood pressure monitoring or management. 17% used APPS for diet or calorie counters, 13% for exercise, fitness or heart monitoring. 63% use APPS on a smartphone, 12% on iPod Touch, 62% on Tablet. Among all HTN patients surveyed, 64% report looking up medical information online on a computer, 53% more than once per month. When asked about willingness to pay, 44% were not willing to pay for a smartphone or tablet that would help monitor their condition. 61% were not willing to pay if an ER visit and its copayment could be prevented. 26% were not willing to use these devices if they were free or covered by insurance.

Conclusions:
A majority of HTN patients surveyed had access to technology with 22% using smartphones with APPS. However, there was limited use of health-specific APPS, especially those related to HTN. Limiting factors may include lack of outcomes-based research, structured programs that incorporate APPS and devices, or lack of familiarity with APPS and technological devices. A comprehensive strategy to develop, market, and demonstrate benefits of this rapidly growing technology is urgently needed.