bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 650: Folic Acid and Vitamin B12 Deficiencies are Cardiovascular Disease Risk Factors Which Can Be Prevented by the Recruitment of the Wnt10b Signaling Pathway
Authors: Schroeder Amanda , Kim DongHyun , Maheshwari Mani , Sodhi Komal , Drummond George , Abraham Nader G
Published: March 2022

Introduction:
Studies have shown that there is a deficiency of folate and vitamin B12 in obese individuals compared with normal BMI controls. Folic acid reduces oxidative stress while vitamin B12 protects against superoxide injuries in endothelial cells. This suggests that increased levels of folate and vitamin B12 might attenuate vascular disease.

Objective:
We examined the hypothesis that folic acid and vitamin B12 regulate mesenchymal stem cell-derived adipocytes by up-regulating Wnt10b and heme oxygenase-1 (HO-1), thereby promoting the growth of small healthy adipocytes and reducing the number of large inflamed adipocytes.

Methods:
Human bone marrow-derived mesenchymal stem cells were used as a source of adipocytes. MSC-derived adipocytes were treated with either vehicle or one of four concentrations (0.1, 1.0, 10, or 100 μM) of either folic acid or vitamin B12. SiRNA of Wnt10b, DKK-1, and other related genes were also studied.

Results:
There was a decrease in adipocytes in a concentration-dependent manner with an optimal level of 10 μM (p<0.05) in both folic acid and vitamin B12 treatment. Treatment of folic acid (10 μM) and vitamin B12 (10 μM) resulted in a decrease in the number of large, unhealthy adipocytes when analyzed using ImagePro Analyzer (p<0.05). The beneficial effects of folic acid and vitamin B12 were reversed when cells were treated with siRNA Wnt10b (p<0.05). The folic acid- and vitamin B12-mediated decreases in adipocyte differentiation were additionally prevented by protein siRNAs including HO-1 (p<0.05).

Conclusion:
These novel results demonstrate that increased levels of both folic acid and vitamin B12 attenuate adiposity via significant increases in Wnt10b and related proteins. Folic acid and vitamin B12 supplementation is deemed viable in the prevention and treatment of obesity and obesity-associated diseases.
Abstract 660: Inducible Ablation Of Emilin1 In Smooth Muscle Cells Increases Blood Pressure And Myogenic Tone By Enhancing Tgf-beta/alk5 Signaling.
Authors: Carnevale Daniela , Facchinello Nicola , Damato Antonio , Braghetta Paola , Bressan Giorgio , Lembo Giuseppe
Published: March 2022

Emilin1 (E1), a protein of the elastic extracellular matrix expressed in the cardiovascular system, regulates TGFβ bioavailability through proteolysis of the proTGFβ, precursor to the LAP/TGFβ complex, that releases the growth factor.
E1 null
mice are hypertensives, with an increased activation of TGFβ. As E1 is expressed in blood vessels starting from early embryonic life to adulthood, is still unknown whether the
E1 null
phenotype results from a developmental defect or lack of a homeostatic role exerted during the adult life.

To dissect this issue, we used a conditional gene targeting approach to inactivate
E1
in smooth muscle cells (SMCs) of adult mice, by the use of floxed
E1
and CreER
T2
[a tamoxifen (TAM) inducible
Cre
recombinase] under the control of the smooth muscle myosin heavy chain (
Smmhc
) promoter. TAM administration induced Cre activity selectively in SMCs, as shown in mice with the Rosa26R mutation.

When

E1
flox/flox

mice carrying the
Smmhc
-CreER
T2
(
Smmhc
-CreER
T2

E1
flox/flox

) were given TAM for 7 days, E1 completely disappeared in 10 days from start of treatment. After that, blood pressure significantly increased in
Smmhc
-CreER
T2

E1
flox/flox

as compared to

E1
flox/flox

(124±1 vs 106±1 mmHg, n=10), whereas no structural changes of both large and resistance arteries were detected. Interestingly, a treatment with anti-TGFβ antibodies rescued the hypertensive phenotype of
Smmhc
-CreER
T2

E1
flox/flox

mice.

Then, we found that isolated resistance arteries of
Smmhc
-CreER
T2

E1
flox/flox

displayed and increased myogenic response to perfusion pressure as compared to

E1
flox/flox

(7±1 vs 16±2 %, n=10). Moreover, we further clarified that the increased myogenic response was rescued by resistance arteries exposure to TGFβ neutralizing antibody. Finally, we found that the inhibition of TGFβ receptor ALK5, obtained transfecting isolated vessels with an adenovirus carrying a dominant negative ALK5mutant, rescued as well the increased myogenic response of
Smmhc
-CreER
T2

E1
flox/flox.

.

Our results demonstrate that E1 exerts a homeostatic control of blood pressure by regulating myogenic tone of resistance arteries, through TGFβ/ALK5 in SMCs, thus suggesting that this pathway should be carefully investigated in essential hypertension.
Abstract 661: Genetic Interference with Peroxisome Proliferator-Activated Receptor Gamma in Vascular Muscle Impairs Cerebrovascular Responses to Nitric Oxide and Enhances Myogenic Tone Via a Rho Kinase-Dependent Mechanism
Authors: De Silva Travice M , Modrick Mary L , Pelham Christopher , Ketsawatsomkron Pimonrat , Sigmund Curt D , Faraci Frank M
Published: March 2022

Cerebral arteries are important resistance vessels in brain and play a crucial role in regulating cerebral blood flow. Nitric oxide (NO) plays a major role in the cerebral circulation influencing vascular structure and blood flow, mediating endothelium-dependent vasodilation, and contributing to neurovascular coupling. Peroxisome proliferator-activated receptor-gamma (PPARγ) is a transcription factor that may play a protective role in the vasculature by enhancing NO signaling and inhibiting increases in vascular tone. We tested whether cell-specific interference with PPARγ in smooth muscle would alter responses to NO and increase myogenic tone in cerebral arteries. We studied mice expressing either a dominant negative mutation of human PPARγ (P467L) or increased expression of WT PPARγ (S-WT) in smooth muscle (S-P467L). In basilar arteries, vasodilation to nitroprusside (NO donor) was markedly impaired in S-P467L compared with S-WT or non-transgenic mice (NT) (e.g. 1 μM nitroprusside: NT, 60±6% vs. S-WT, 63±2% vs. S-P467L, 16±3%, n=6-11). As PPARγ may suppress rho kinase signaling, we examined its role in regulating responses to NO. Impaired vasodilation to nitroprusside in S-P467L mice were restored to normal by an inhibitor of rho kinase (Y27632, 3 μM). Angiotensin II (Ang II) activates rho kinase via AT1 receptors. As AT1-mediated effects may also be suppressed by PPARγ, we examined effects of an AT1 receptor antagonist (losartan). Losartan (1 μM) completely inhibited vasoconstriction to Ang II and increased responses to nitroprusside to ∼70% of normal in S-P467L mice. In response to intraluminal pressure (75 mmHg), middle cerebral arteries (MCA) from S-P467L mice constricted 31±4% compared with 9±2% for NT (n=7-8). Dilation of MCA from S-P467L mice to acetylcholine was significantly impaired compared with controls, whereas dilation to papaverine was similar between genotypes. Treatment of the MCA from S-P467L mice with Y27632 abolished the augmentation of myogenic tone (control, 34±5% vs. Y27632, 9±4%, n=7). These findings suggest that PPARγ in vascular muscle normally inhibits rho kinase, influencing responses to NO and myogenic tone in small resistance arteries.
Abstract 68: Diminazene, an Angiotensin Converting Enzyme2 (ACE2) Activator, Improves Right Heart Function in Experimental Models of Pressure Overload
Authors: Shenoy Vinayak , Qi Yanfei , Patel Jawaharlal , Katovich Michael , Raizada Mohan
Published: March 2022

BACKGROUND:
Right ventricular (RV) failure due to pressure overload is a major cause of morbidity and mortality in pulmonary hypertensive patients. We and others have shown that ACE2 exerts cardiopulmonary protection against lung diseases. This has led us to hypothesize that activation of ACE2 would improve RV hemodynamics and attenuate cardiac remodeling in animal models of pressure-overload.

Methods:
Hypoxia and Monocrotaline (MCT) models of pressure-overload were employed to evaluate the effects of Diminazene (DIZE), an ACE2 activator. Rats were exposed to hypoxia for 28 days with simultaneous DIZE treatment (15mg/Kg). In the other model, rats were challenged with MCT (50mg/Kg).Both prevention and reversal protocols were studied in this model. In the prevention protocol, rats were co-treated with DIZE for 4-weeks, while for the reversal protocol, DIZE was injected after 3-weeks of MCT-challenge and the drug treatment continued for additional 2-weeks.

Results:
Hypoxia exposed rats exhibited increased ratio of right ventricular to left ventricular plus septum weights, an index of RV hypertrophy (Con:0.26+0.01; Hypoxia: 0.48+0.03), along with cardiac dysfunction in terms of higher right ventricular end diastolic pressure (RVEDP; Con:3.28+0.65; Hypoxia:9.08+0.72mmHg), elevated +dP/dt (Con:2039+230; Hypoxia:2815+156mmHg/s) and increased -dP/dt (Con:-1476+98; Hypoxia:-2663+150mmHg/s). However, DIZE improved all these cardiovascular alterations (RVH:0.38+0.01; RVEDP:7.03+0.3mmHg; +dP/dt:2086+137mmHg/s;-dP/dt: -1945+100mmHg/s). Likewise, MCT injection induced RV hypertrophy (Con:0.27+0.01; MCT:0.50+0.03) and cardiac dysfunction (RVEDP; Con:3.98+0.46; MCT: 8.54+0.56mmHg; +dP/dt; Con:2107+189; MCT:3718+222mmHg/s; -dP/dt; Con:-1545+95; MCT:-2918+238mmHg/s), which was prevented by DIZE (RVH:0.29+0.02; RVEDP:4.76+0.44mmHg; +dP/dt:2525+122mmHg/s; -dP/dt:-2076+150mmHg/s). Furthermore, MCT caused a 3-fold increase in myocardial fibrosis, which was reduced by DIZE. In the reversal protocol, DIZE attenuated RV hypertrophy and improved cardiac hemodynamics.

CONCLUSIONS:
Collectively, our results identify a therapeutic potential of DIZE for RV dysfunction induced by pulmonary pressure-overload.
Abstract 12: Cox2 Signaling During the Peri-implantation Period Plays an Important Role in Fetoplacental Heath in a Mouse Model of Preeclampsia, BPH/5
Authors: Sones Jenny L , Cha Jeeyeon , Isroff Catherine E , Dey Sudhansu K , Davisson Robin L
Published: March 2022

Preeclampsia (PE), a hypertensive proteinuric disease of pregnancy, is a leading cause of perinatal and maternal morbidity/mortality. Although abnormal placentation is widely accepted as a cause for PE, the underlying mechanism is unclear. The BPH/5 mouse model spontaneously develops the cardinal features of PE with poor fetoplacental outcomes. These mice exhibit deferred implantation coincident with a delayed rise in serum estrogen and defective decidualization. We tested the hypothesis that aberrations in peri-implantation events influence fetoplacental health in BPH/5 mice. Real time PCR,
in situ
hybridization (ISH), and Western blotting were performed to characterize the expression of critical molecular mediators during the peri-implantation period in BPH/5 vs C57 control females. Implantation sites (IS) of BPH/5 females show reduced levels of
Lif
(2-fold),
Bmp2
(8.5-fold) and
Ptgs2
(encoding Cox2, 4-fold) on day 4.5 of pregnancy (n=6-9, p<0.05 vs C57). By day 5.5,
Ptgs2
is upregulated in BPH/5 IS (2-fold; n=7, p<0.05 vs C57), which persists through day 7.5 and is consistent with upregulated Cox2 protein levels (2.75-fold±0.1; n=4, p<0.05 vs C57). ISH results are consistent with those of RT-PCR and demonstrated appropriate expression patterns of
Lif
,
Bmp2
, and
Ptgs2
. BPH/5 mice demonstrate three increasingly severe phenotypic classes of fetoplacental health as seen by ultrasound at day 10.5: healthy, compromised, and resorbed. Since Cox2-derived prostaglandins are crucial for decidualization, placentation and placental angiogenesis, we tested the effect of a selective Cox2 inhibitor (celecoxib) on fetoplacental development in BPH/5 females. Administration of celecoxib by oral gavage on day 6.5 (10mg/kg BW/dose) increased the percentage of healthy embryos (65.9±6.8 vs 45.1±7.9 veh) with reduced percentage of compromised embryos (20.7±2.5 vs 35.4±6.1veh) within BPH/5 litters (n=4-5, p<0.05). These results provide evidence that Cox2 plays a key role in fetoplacental health in a mouse model of PE, and that celecoxib improves fetoplacental outcomes in BPH/5 females. This highlights the significance of peri-implantation events in pregnancy outcome and suggests that aberrations in early pregnancy may give rise to PE.
Abstract 152: Development of a Chronic Angiotensin II (AngII)-stimulated Neuronal Cell Culture Model
Authors: Basu Urmi , Case Adam J , Madayiputhiya Nandakumar , Zimmerman Matthew C
Published: March 2022

Dysregulation of AngII-mediated intra-neuronal signaling contributes to the pathogenesis of hypertension. Previous studies using AngII-sensitive neuronal cell culture models suggest that AngII induces chronic changes in the expression of various cell signaling proteins, including its own receptors. However, many of these studies stimulated cultured neurons with a single administration of exogenous AngII and harvested neurons 3-24 hours later to investigate changes in mRNA and protein levels. One limitation of these “chronic” AngII stimulation studies is the lack of evidence indicating the stability of AngII in neuronal cell culture media. Here, we tested the hypothesis that exogenous AngII is rapidly metabolized in neuronal cell culture media. Further, we hypothesized that AngII receptors are differentially expressed depending on the chronicity of stimulation. AngII (100nM) was added to catecholaminergic (CATH.a) neurons in culture, media was collected 15 min - 24 hr later, and AngII levels were determined by liquid chromatography-tandem mass spectrometry. AngII was rapidly metabolized with media levels of AngII returning to near baseline within 3 hr of administration (baseline: 2.2 nM AngII; 15 min: 50.6 nM; 30 min: 42.9 nM; 1hr: 28.8 nM ; 3hr: 4.0 nM, n = 3-4). To begin investigating the relevance of this observation, we measured mRNA levels of AngII type 1 (AT1R) and type 2 (AT2R) receptors using real time quantitative RT-PCR in CATH.a neurons 24 hr after a single administration of AngII (100nM) or repeated administration every 3 hr. While a single administration of AngII did not alter AT1R mRNA levels, replenishing media every 3 hr with fresh AngII did induce a 20% decrease. In addition, fresh AngII every 3 hr resulted in a 41% decrease in AT2R mRNA levels; whereas, a single dose of AngII increased AT2R mRNA by 19%. These data indicate that AngII is quickly metabolized in neuronal cell culture media, and that supplementing fresh AngII every 3 hr to maintain elevated levels chronically yields different changes in AngII receptor mRNA levels as compared to a single administration. In conclusion, we suggest that to accurately model hypertension, where AngII is chronically elevated, the metabolism of AngII in a given cell culture model should first be determined.
Abstract 167: Age and Gender Differences in Pulse Pressure Response to Repeat Blood Pressure Measurement After Brief Patient Rest Period
Authors: Karumbaiah Keerthana , Omar Bassam A
Published: March 2022

BACKGROUND:
Increased pulse pressure (PP) is an independent determinant of cardiac disease. The effect of rest period on office-based systolic blood pressure has been previously reported, but not on PP in particular.

OBJECTIVE:
To assess the effect of age and gender on repeat PP measurement after a brief rest period in an outpatient cardiology clinic.

METHODS:
Patient charts reviewed in University-based cardiology clinic identified 170 encounters which contained BP re-measurement data due to elevated initial BP of > 130/80 mmHg. BP was measured initially by a nurse, with the patient in a sitting position and the arm resting at the level of the heart. If BP was > 130/80 mmHg, it was repeated by physician after resting the patient for 15 minutes. There were 112 (66%) elderly patients ≥ 60 years of age and 58 (34%) younger patients < 60 years of age. Among the elderly patients, there were 51 males (46%) and 61 females (54%).

RESULTS:
Among all encounters, after a brief rest period, initial PP of 67 ± 2 mmHg decreased to 62 ± 1 mmHg (5 mmHg; P < 0.01). PP decreased by 8 mmHg in the elderly (72 ± 2 to 64 ± 2 mmHg; P < 0.01) but did not significantly change in the young (56 ± 3 to 58 ± 3 mmHg; P = 0.3). PP decrease among the elderly was more pronounced in females (11 mmHg; 76 ± 4 to 65 ± 2 mmHg; P < 0.01) compared with males (4 mmHg; 68 ± 3 to 64 ± 2 mmHg; P = 0.03).DISCUSSION: Hypertension is a challenging public health problem. JNC 7 guidelines recommend that prior to BP measurement, persons should be seated quietly for at least 5 minutes in a chair, with feet on the floor, and arm supported at heart level; which resulted in significant decrease in pulse pressure in our patients. We show that while a decrease in pulse pressure was confined to the elderly, elderly females had a more pronounced PP decrease compared to males; both, however, fell to the same level. White coat hypertension may explain this observation, since it is more frequent in elderly females. The implication of this observation is that physicians should take extra care in remeasuring the blood pressure and pulse pressure, especially in elderly females, in whom a more pronounced drop in pressure may be observed after a brief rest period, and thereby, result in reclassifying their risk and need for treatment.
Abstract 172: Angiotensin II in Combination With High Salt Causes Increased Prostaglandin D2 Levels and Down-regulation of Neuronal Prostaglandin D1 Receptor Expression in Rostral Ventrolateral Medulla
Authors: Asirvatham-Jeyaraj Ninitha , Jones Daniel A , Fink Gregory D
Published: March 2022

Neurogenic hypertension (HTN) is characterized by increased sympathetic activity, but the specific causes remain to be determined. Using the angiotensin II-high salt model (AngII-salt) of neurogenic HTN in rats, we observed that chronic central prostaglandin D synthase (L-PGDS) inhibition caused attenuation of HTN development and neurogenic pressor activity. As L-PGDS promotes prostaglandin D2 (PGD2) synthesis in the brain, we hypothesized that increased PGD2 levels acting on DP-1 receptors in key cardio-regulatory brain regions participate in AngII-salt HTN development. To test this hypothesis adult male Sprague Dawley rats on high salt (2% NaCl) diet received either vehicle or angiotensin II (150ng/kg/min, sc.), using a mini-osmotic pump. On day 4, the developmental stage of HTN, brains from 5 control and 5 AngII treated rats were collected for PGD2 measurement using liquid chromatography coupled with mass spectrometry. In another set of control and treated rats, brains were perfusion fixed and then collected for DP1R expression measurement in neurons using immunofluorescence. We found that AngII-salt treated rats had significantly increased PGD2 levels (34.5±13.4ng/region) in the rostral ventrolateral medulla (RVLM) compared to vehicle-treated rats (12.4±4.8ng/region; p<0.05). Relative expression of neuronal DP1 receptors in RVLM was less in AngII-salt treated rats (163.8±11.1 relative intensity units) than that of the control rats (200.6±14). Similar changes were not seen in any other brain region. These data suggest that AngII and salt increase PGD2 in the RVLM that acts on neuronal DP1 receptors to cause sympathoexcitation and hypertension.
Abstract 182: Medullary Endocannabinoid Content Contributes to the Differential Resting Baroreflex Sensitivity in Rats with Altered Brain Renin-Angiotensin System Expression
Authors: Schaich Chris L , Howlett Allyn C , Thomas Brian F , Grabenauer Megan A , Diz Debra I
Published: March 2022

Activation of CB1 receptors in the solitary tract nucleus (NTS) over a 100-fold dose range elicits biphasic effects, with high doses inhibiting baroreflex sensitivity (BRS) for control of heart rate and lower doses facilitating BRS. There is growing evidence for signaling interactions between the endocannabinoid system (ECS) and pathogenic actions of angiotensin (Ang) II at AT1 receptors in hypertension and the metabolic syndrome. Hypotensive ASrAOGEN (AS) rats, overexpressing an angiotensinogen antisense in glia, have significantly elevated medullary tissue glutamate levels and enhanced BRS compared to normotensive Sprague-Dawley (SD) rats (1.42 ± 0.11 vs. 1.04 ± 0.05 ms/mmHg; n = 5 - 6; P = 0.02). Transgenic (mRen2)27 rats, a monogenetic Ang II-dependent model of hypertension, have markedly impaired BRS compared to SD rats (0.43 ± 0.06 vs. 1.04 ± 0.05 mmHg; n = 5; P < 0.0001). There is no effect of the CB1 receptor antagonist SR141716 on BRS when microinjected into the NTS of SD rats; however, SR141716 injected into the NTS of hypertensive (mRen2)27 rats dose-dependently improved BRS. In contrast, SR141716 injected into the NTS of hypotensive AS rats dose-dependently reduced BRS. Microinjection of 0.36 pmol SR141716 (n = 3) reduced BRS from a baseline of 1.23 ± 0.05 ms/mmHg to 0.81 ± 0.14 (P = 0.01) and 36 pmol SR141716 (n = 3) decreased BRS from 1.60 ± 0.17 to 0.48 ± 0.06 ms/mmHg (P = 0.001), a level comparable with baseline BRS of hypertensive (mRen2)27 rats. The effects of CB1 receptor blockade on BRS parallel the endogenous levels of the endocannabinoid 2-arachidonoylglycerol (2-AG) as measured by mass spectrometry with lower 2-AG in AS compared to (mRen2)27 rats (1.17 ± 0.09 vs. 2.70 ± 0.28 ng/mg tissue; n = 4 - 5; P = 0.002). Normotensive SD rats had intermediate 2-AG levels (1.85 ± 0.27 ng/mg tissue; n = 5; n.s. vs. other strains). These observations reveal that an upregulated ECS contributes to RAS-dependent suppression of BRS. That there is a reduction in BRS in response to NTS CB1 receptor blockade in hypotensive AS rats exhibiting elevated glutamate levels suggests differential regulatory functions of the brain ECS that may involve alterations in transmitters known to influence BRS and that parallel the Ang II:Ang-(1-7) ratio in these animals.
Abstract 193: Urinary Angiotensinogen Could be a Prognostic Marker of Renoprotective Effects of Alogliptin in Patients with Type 2 Diabetes
Authors: Mizushige Tomoko , Kobori Hiroyuki , Nishijima Yoko , Yano Yuichiro , Sakata Koji , Hayakawa Manabu , Nishiyama Akira
Published: March 2022

The development of dipeptidyl peptidase-IV (DPP-4) inhibitors, such as alogliptin, has led to the improvement in glycemic control in type 2 diabetes (T2D) by preventing the degradation of the incretin hormones glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide. However, the renoprotective effects of alogliptin have not been addressed yet. This 12-week study in Japanese patients with T2D was performed to address the renoprotective effects of alogliptin. In addition, urinary angiotensinogen (AGT), a marker of intrarenal renin-angiotensin system (RAS) activity, was examined to demonstrate the clinical usage as a prognostic marker.

Forty-three patients with T2D (18 women, age: 66.1+/-11.2) were recruited in Miyazaki University and its affiliated hospitals, and alogliptin (25 mg/day) was added on the top of the traditional hypoglycemic agents. The urinary concentrations of albumin (Alb) and AGT were measured using commercially available ELISA kits before and after the alogliptin treatment, and normalized by the urinary concentration of creatinine (Cr) (UAlbCR and UAGTCR, respectively).

The alogliptin treatment tended to decrease UAlbCR (99.6 +/- 26.8 vs. 114.6 +/- 36.0, mg/g Cr). However, this change was not statistically significant (p = 0.1976). Then, we defined good responders to the alogliptin treatment in terms of %change in UAlbCR less than -25% after the 12-week treatment, and a logistic analysis of UAGTCR before the treatment showed the area under the curve (AUC) as 0.644. When we set the cutoff value of UAGTCR as 20.8 μg/g Cr, the maximum specificity (17/27 = 63.0%) and sensitivity (10/16 = 62.5%) were obtained (Youden index = 0.255). Based on this cutoff value of UAGTCR before the treatment, we divided all patients into 2 groups as higher (group H, N = 20) and lower (group L) values of UAGTCR at the baseline. %Change in UAlbCR was significantly lower in the group H compared with the group L (-14.6% +/- 8.6% vs. +22.8% +/- 16.8%, p = 0.0327). These data indicate that the T2D patients with the higher UAGTCR before the treatment would show more decrease in UAlbCR by the alogliptin treatment.

In conclusion, urinary AGT could be a prognostic marker of renoprotective effects of alogliptin in T2D patients.
Abstract 207: Angiotensin 1-7 Reduces Mortality and Aneurysm Rupture in a Mouse Model of Intracranial Aneurysms
Authors: Pena Silva Ricardo A , Kung David K , Mitchell Ian J , Alenina Natalia , Bader Michael , Santos Robson A , Heistad Donald D , Hasan David M
Published: March 2022

Inflammation has been linked to progression and rupture of intracranial aneurysms in humans and animal models. Angiotensin II drives vascular inflammation in several models of vascular disease. Angiotensin 1-7, the product of metabolism of angiotensin II by ACE2, acts on Mas receptors and counteracts the deleterious effects of angiotensin II. We previously demonstrated that ACE2 deficiency is associated with cerebrovascular dysfunction. We now explore the hypothesis that angiotensin 1-7 attenuates formation and rupture of intracranial aneurysms in mice. In wild type (WT) C57Bl/6 mice an intracranial injection of elastase was made, and then angiotensin II (Ang II) or angiotensin II + angiotensin 1-7 (Ang II + Ang 1-7) were infused for 3 weeks. Increase in systolic pressure was similar in mice receiving Ang II (146±5 mmHg, mean ±SE) or Ang II + Ang 1-7 (147±5 mmHg). Mortality was higher in Ang II-treated mice (64% (16/25)) than in Ang II + Ang 1-7-infused mice (33% (7/21) p<0.05). Aneurysm formation was similar in both groups (88% (22/25) Ang II vs. 81% (17/21) Ang II + ang 1-7). Incidence of subarachnoid hemorrhage (SAH) was lower (43% (9/21)) in Ang II + Ang 1-7 than in Ang II treated mice (68% (17/25), p<0.05). Similar studies were performed in Mas receptor knockout mice (Mas-KO). Increase in systolic pressure was similar (136±4 vs. 137±8 mmHg) in Mas-KO mice that received Ang II or Ang II + Ang 1-7 respectively. There was no significant difference in aneurysm formation (83% 10/12 vs. 100% 14/14) or SAH incidence (50% 6/12 vs. 64% 9/14) in Mas-KO mice treated with Ang II or Ang II + Ang 1-7 respectively (p>0.05). In conclusion, angiotensin 1-7 does not attenuate the increase in systolic blood pressure induced by Ang II in WT or Mas-KO mice. Angiotensin 1-7 does not affect the incidence of aneurysm formation in this mouse model of intracranial aneurysms. Importantly, angiotensin 1-7 decreases mortality and intracranial aneurysm rupture. Protective effects of angiotensin 1-7 on aneurysm rupture are not seen in Mas-KO mice, confirming that these effects of angiotensin 1-7 are mediated by its receptor Mas. We speculate that angiotensin 1-7 may be of therapeutic value for prevention of rupture of intracranial aneurysms.
Abstract 248: Leptin-mediated Cardiovascular Disease Involves Aldosterone in Female Mice and Adolescent Women
Authors: Huby Anne-Cecile , Zhu Haidong , Tremblay Michel L , Bollag Wendy , Dong Yanbin , Belin De Chantemele Eric
Published: March 2022

Premenopausal women have a better endothelial function than men but are more sensitive to obesity induced endothelial dysfunction (ED) and more reactive to mineralocorticoid receptor (MR) blockade. Due to its control of sympathetic activity, leptin has been reported to be the link between metabolic and cardiovascular disease (CVD) in men. Whether a similar correlation exists in females that secrete 4 times more leptin than men is unknown. Using a population based study and mice deficient (KO) in protein tyrosine phosphatase 1B (PTP1B), a leptin sensitizer, we tested the hypothesis that leptin mediated CVD involves sex specific mechanisms. Plasma leptin were 4 times higher in female mice (KO
F
:1648±271
vs
KO
M
: 347±47 pg/ml; p<0.05) indifferently of the genotype (WT
F
: 2386±765
vs
WT
M
: 375±37 pg/ml; p<0.05). PTP1B deletion increased leptin sensitivity and raised blood pressure (BP) to a similar extent in males and females (124±4 vs 126±4 respectively; WT 102±5 mmHg) but induced ED in females KO only (p<0.01
vs
WT and males). Sympathetic tone was assessed by measuring BP response to ganglionic blockade. Leptin sensitized males only exhibit an increased sympathetic tone, but leptin sensitized females only exhibit high plasma aldosterone levels (aldo; KO
F
:1234±173
vs
WT
F
: 545±71and KO
M
: 624±70 pg/ml). Both leptin receptor antagonist and MR blockade restored BP in KO females.
In vivo
and
ex vivo
(aortic ring) MR blockade restored endothelial function in females KO suggesting direct aldo effects on the vessels. Neither corticosterone, potassium nor angiotensin II was increased in KO females. However aldosterone synthase expression was impressively increased in females KO only. Aldo levels are low in obese leptin receptor deficient females (333±89 pg/ml), and leptin directly stimulates aldosterone secretion in glomerulosa cells. In parallel we demonstrated that African American adolescent females secrete 4 times more leptin than males, and established a correlation between leptin and aldosterone(r=0.464, p=0.011) in female subjects only. These human and animal data suggest that leptin regulates aldo secretion in females. All together these data suggest that obesity mediated CVD is leptin dependent and involves aldo dependent mechanisms in females.
Abstract 261: Relationship of Arginine Vasopressin and Blood Pressure in Patients with Orthostatic Intolerance and Nausea
Authors: Wagoner Ashley L , Shaltout Hossam A , D'Agostino Ralph B , Diz Debra I , Fortunato John E
Published: March 2022

Children with chronic unexplained nausea have a high incidence of cardiovascular instability (~ 75%) manifesting as orthostatic intolerance (OI), with exaggerated suppression of baroreflexes upon standing. Because vasopressin (AVP) causes nausea and may be triggered by a fall in blood pressure (BP), we aimed to determine the relationship between AVP and tilt-induced changes in BP in children with OI. Forty-eight patients (mean age [range] of 15.2 [10-18] years) (36 females, 12 males) underwent tilt table testing for symptoms of OI and nausea. Subjects were maintained supine for 15 min before 45 min upright tilt (from 0 to 70 degrees). Plasma AVP was measured in blood sampled immediately before and 15 min into upright tilt. Of the 48 tilt tests: 9 were normal and 39 abnormal. Abnormal tests were classified as: postural orthostatic tachycardia syndrome (POTS, heart rate >120 bpm or increase by 30 bpm during first 10 min of tilt; n = 15), neutrally mediated hypotension (NMH, decrease in systolic BP [SBP] >25 mmHg during first 10 min of tilt; n = 2), and neurocardiogenic syncope (NCS, hypotension followed by syncope; n = 22). Mean supine SBP of normal subjects was 129 ± 9 mmHg and diastolic BP [DBP] was 64 ± 3 mm Hg (mean ± SEM). There were no differences in supine BPs among groups. Supine AVP levels were 4 ± 2 and 2 ± 1 in abnormal vs normal subjects (p = NS) There was a positive correlation between SBP and AVP in the supine position before tilt (0.34, p = 0.02). AVP increased by 44 ± 13 pg/mL vs. 1 ± 1 pg/mL for subjects with abnormal vs. normal tilt tests (p = 0.002), by 43 ± 17 pg/mL for NCS alone vs. normal (p = 0.02), and by 37 ± 20 pg/mL for POTS vs. normal (p = NS). There was a negative correlation between AVP and both SBP and DBP during tilt (-0.3, p = 0.035 and -0.4, p = 0.005, respectively). Hypotension and syncope upon tilt are associated with acute increases in AVP in subjects with OI and nausea. The increase in AVP may serve as a compensatory mechanism in the face of impaired autonomic reflexes. It is not known whether the increase in AVP contributes to the nausea observed during tilt, but better understanding of this relationship may allow for more effective treatment options for the management of nausea in these subjects.
Abstract 264: Serotonin Signaling Through Nox1 in Human Pulmonary Artery Smooth Muscle Cells - Implications in Vascular Remodeling in Pulmonary Arterial Hypertension
Authors: Hood Katie Y , Montezano Augusto C , Morecroft Ian , MacLean Margaret R , Touyz Rhian M
Published: March 2022

Serotonin (5HT) and NADPH oxidases (Nox) have been linked to the pathology of pulmonary arterial hypertension (PAH). However, it is unclear whether 5HT plays a role in PAH through Nox-dependent mechanisms. We hypothesised that 5HT through Nox1-induced reactive oxygen species (ROS) induces PASMC damage. Human PASMC primary cultures were stimulated with 5HT (1μM) in the absence/presence of tempol (antioxidant, 10μmol/L), ML171 (Nox1 inhibitor; 1μM), and EHT1864 (Rac1 inhibitor; 100μM). ROS generation was assessed by chemiluminescence. Markers of fibrosis (fibronectin), growth (PCNA), inflammation (TRAIL, DcR2), Nrf2 regulation (Nrf2, BACH1, HO1) and MAPKs activation (ERK1/2 and p38MAPK) were analyzed by immunoblotting. Proliferation was assessed by [3H]-thymidine incorporation in the presence/absence of Nox1 inhibitor. 5HT induced increased in ROS production at 4h of stimulation (224 ± 49% vs vehicle; p<0.05) in PASMC, an effect blocked by tempol, EHT1864 and ML171. Fibronectin (192 ± 26%), PCNA (124 ± 2%), TRAIL (122 ± 4%) and DcR2 (133 ± 2%) protein levels were increased by 5HT in PASMCs (p<0.05, vs vehicle). In addition, 5HT increased phosphorylation of ERK1/2 (169 ± 10%) and p38MAPK (157 ± 15%) (p<0.05, vs vehicle). Levels of Nrf2 and its downstream target, HO-1, were decreased by 5HT stimulation (90 ± 1%; 84 ± 6% respectively vs vehicle 100%, p<0.05). BACH1 expression, a protein that negatively modulates antioxidant regulator Nrf2, was increased by 5HT in PASMCs (153 ± 8% vs vehicle; p<0.05). Nox1, but not Nox2, expression was increased in PASMC after stimulation with 5HT (139 ± 11% vs vehicle, p<0.05). 5HT induced a 3.95±0.39 fold increase in proliferation PASMCs (P<0.05), which was blocked by ML171. In conclusion, Nox1 may play a role in 5HT-induced pro-proliferative, fibrotic and inflammatory responses. These data highlight some molecular mechanisms whereby 5HT may exert deleterious vascular effects in PAH.
Abstract 265: Plasma Lactate and Incident Hypertension in the Atherosclerosis Risk in Communities (ARIC) Study
Authors: Juraschek Stephen P , Bower Julie K , Selvin Elizabeth , Shanta Ghanshyam P , Hoogeveen Ron C , Ballantyne Christie M , Young J. H
Published: March 2022

Insufficient oxidative capacity, or mitochondrial dysfunction, may contribute to the development of high blood pressure by stimulating increased blood flow. To test this hypothesis, we examined the prospective association of lactate, a product of glycolysis and marker of oxidative capacity, with incident hypertension in 5,554 participants from the Atherosclerosis Risk in Communities (ARIC) Study. Participants had no diagnosed hypertension and blood pressures <140/<90 mm Hg at baseline (1996-1998). Incident hypertension was assessed during annual telephone calls and defined as a self-reported physician diagnosis of hypertension or hypertension medication use. Analyses were performed with Cox proportional hazards models. The mean age was 61.9 years, and the mean lactate was 0.08 mmol/L. The population was 45.1% male and 12.8% black. During a median follow-up period of 11.9 years (range: 26.9 days to 13.4 years), there were 3,849 new cases of hypertension. The unadjusted 10-yr cumulative incidences across quartiles 1-4 of lactate were 14.6, 18.3, 21.5, and 32.3%. After adjustment for traditional hypertension risk factors, including baseline systolic and diastolic blood pressure, the highest quartile of baseline lactate was associated with 1.18 times the hazard of the lowest quartile (95% CI: 1.07, 1.31; P-trend = 0.001) (Figure). This association was stronger (HR 1.42; 95% CI: 1.23, 1.63) in participants with blood pressures <120/<80 mm Hg. Mild elevations in lactate precede hypertension even in a normotensive population. These findings have important implications for early detection of hypertension risk in healthy adults.
Abstract 284: P21 is Essential for the Beneficial Effects of Renal Ischemic Preconditioning in Ischemic Acute Kidney Injury
Authors: Nishioka Satoshi , Nakano Daisuke , Kitada Kento , Ohsaki Hiroyuki , Sofue Tadashi , Kohno Masakazu , Nishiyama Akira
Published: March 2022

We previously reported that p21, a cell cycle regulator, plays important roles in chronic hypertensive renal injury. However, the roles of renal p21 in acute kidney injury, a life-threatening disease that can occur independently of the pathological background of patients (whether renal p21 is up-regulated or not), have not been fully clarified yet. In the present study, we evaluated the role of p21 in acute kidney injury and the effects of ischemic preconditioning (IPC). The mice lacking functional p21 (p21-KO) and its wild-type control underwent renal ischemia followed by reperfusion (I/R) with or without ischemic preconditioning. IPC attenuated I/R injury in wild-type mice, but not in p21-KO mice. Moreover, IPC increased the renal expression of p21 prior to I/R compared with sham surgery (p21/β-actin, sham: 1.00±0.18 fold, IPC: 4.12±0.32 fold, n=6, p<0.01). Immunohistochemisty (IHC) for p21 showed that there was an increase in the number of p21-positive tubular cells in mice that underwent I/R with and without IPC, and that the p21 immunoreactivity was increased in the nuclei of tubular cells, suggesting the effects on cell cycle. IPC decreased the number of proliferating tubular cells before I/R (0.24±0.11 fold vs. sham group, p<0.01) and increased it at 24 h after I/R (1.66±0.03 fold vs. sham group, p<0.01) in the kidney of wild-type mice. In p21-KO mice, IPC did not change the number of proliferating cells before I/R (0.99±0.10 fold, vs. sham group), and decreased it after I/R (0.53±0.08 fold, vs. sham group, p<0.01). Flow cytometry showed that IPC increased the number of cells in the G1 phase of cell cycle before I/R compared with sham surgery. Additionally, intravital imaging in combination with the mice that express mKO2, a fluorescent protein, at G1 phase of cell cycle showed that IPC increased the number of mKO2-positive cells in the proximal tubules. In conclusion, renal p21 is essential for the beneficial effects of renal IPC. Transient cell cycle arrest by IPC through a p21-dependent pathway seems to be important for recovering tubular cell proliferation after I/R.
Abstract 29: Hydrogen Peroxide Impairs Myogenic Response of Afferent Arterioles from Mice with the Reduce Renal Mass Model of Chronic Kidney Disease
Authors: Li Lingli , Lai En Yin , Welch William J , Wilcox Christopher S
Published: March 2022

Background:
Impaired autoregulation and systemic hypertension underlie progressive kidney injury in chronic kidney disease (CKD). We reported that the myogenic response (MR) was severely impaired in afferent arterioles (Affs) from mice with reduced renal mass (RRM) and oxidative stress, yet superoxide (O
2
.-
) enhanced MRs of normal mice. Therefore, we tested the hypothesis that an increase in perfusion pressure (PP) enhances the generation of H
2
O
2
selectively in Affs from mice with RRM and that H
2
O
2
causes the impaired MR.

Methods and Results:
Groups of sham and RRM C57BL/6 mice were fed a high salt (6% NaCl) diet for 3 months. Alternative groups were treated with 2 mM of the redox cycling nitroxide tempol in drinking water for 3 months, which reduced the excretion of both 8-isoprostane (marker of O
2
.-
) and H
2
O
2
. MRs were assessed from the % change in luminal diameter of isolated, perfused Affs after increasing PP from 40 to 80 mmHg, O
2
.-
from PEG-SOD inhibitable ethidium:dihydroethidium fluorescence and H
2
O
2
from PEG-catalase (CAT) inhibitable H2DCFDA fluorescence.

Results:
1. Compared to sham, increasing PP in Affs from RRM mice increased O
2
.-
more (21 ± 2% vs 11 ± 2%, P<0.01), but this was prevented by tempol, and increased H
2
O
2
more (29 ± 8% vs 4 ± 1%, P<0.01), which also was prevented by tempol. 2. Compared to sham, Affs from mice with RRM mice had severely impaired MRs (sham, -14 ± 4% vs RRM, -1 ± 4%, P<0.05), yet this defect was corrected in mice drinking tempol (sham + tempol, -9 ± 6% vs RRM + tempol, -11 ± 2%, NS). 3. Bath addition of PEG-SOD (200 units/ml) attenuated MRs from sham mice (sham + vehicle, -14 ± 4%; sham + SOD, -6 ± 1%; P<0.05), but did not affect responses in RRM mice. 4. Bath addition of PEG-CAT (1000 units/ml) selectively enhanced the MRs in RRM mice (RRM + vehicle -1 ± 4%; RRM + CAT -19 ± 2%; P< 0.01) without effect in sham mice (sham + vehicle, -14 ± 4%; sham + PEG-CAT, -14 ± 2% NS).

Conclusions:
Excessive H
2
O
2
is released by increased PP of the afferent arterioles only from mice with RRM and accounted for the impaired myogenic response. Tempol, or other drugs that metabolize H
2
O
2
, may preserve renal autoregulation and prevent progressive loss of GFR in CKD.
Abstract 292: AT2 Receptor Activation Prevents High Sodium-induced Increase in Blood Pressure and Insulin Resistance in Obese Rats
Authors: Ali Quaisar , Nag Sourashish , Hussain Tahir
Published: March 2022

High sodium intake is a risk factor for the pathogenesis of hypertension and also implicated in insulin resistance. Recently, renal angiotensin AT2 receptor has been shown to promote natriuresis, especially in obese Zucker rats, a model of insulin resistance with modest high blood pressure. However, the role of AT2 receptor in high sodium-induced hypertension and insulin resistance in obesity is not known. Male obese Zucker rats were treated with AT2 receptor agonist C21 (1mg/kg/day oral) while maintained on either normal sodium (NSD, 0.4%) or high sodium diet (HSD, 4%) for 2-weeks. Telemetry monitoring of blood pressure (BP) demonstrated an increase in systolic blood pressure (SBP) by 20 mm Hg in obese HSD compared to NSD group. This increase in SBP was prevented by C21 in rats placed on HSD, but C21 had no effect on SBP in rats on NSD. The rats on HSD had increase in body weight gain (NSD: 45±8 HSD: 74±5 gm) and kidney weight (HSD: 4.7 ±.09, NSD 2.9±.02 gm); the increases were lowered by C21 treatment in HSD group. Compared to NSD rats, HSD rats had significantly reduced GFR (NSD: 654±82, NSD: 302±89 μl/min) which was modestly improved by C21 treatment in HSD group. C21 treatment did not affect food intake in any of the experimental groups. Interestingly, fasting blood glucose and plasma insulin measured at the end of 2-weeks was significantly increased in HSD compared to NSD (glucose: NSD 94±2, HSD 137±8 mg/dL; insulin: NSD 4.3±.3 HSD 8.5±0.6 ng/ml); the increases in both glucose and insulin were prevented by C21 treatment. Similarly, oral glucose tolerance was impaired in HSD obese rats, and the C21 treatment completely normalized the glucose tolerance in HSD rats. Treatment with C21 did not affect blood glucose, insulin levels and GTT in obese rats on NSD. C21 treatment also modestly reduced the epididymal white adipose tissue weight (eWAT) by 15% in HSD+C21 group. This study clearly demonstrate that long-term AT2R agonist treatment prevents the high sodium-induced increase in blood pressure and insulin resistance in in obese rats indicating a potential role in improving metabolic syndrome.
Abstract 314: Arterial Stiffness and Pulse Pressure Variability
Authors: Bahrainwala Jehan , Patel Ami , Diaz Kieth M , Veerabadrappa Praveen , Cohen-Stein Debbie , Cuchiarra Andrew , Townsend Raymond R
Published: March 2022

Blood pressure variability (BPV) is associated with adverse cardiovascular outcomes. Factors determining BPV are unclear. Arterial stiffness manifested by pulse pressure (PP) may be related to BPV, and is reflected in the pulse wave velocity (PWV), as well as the regression of diastolic on systolic pressure determined by the Ambulatory Arterial Stiffness Index (AASI). We hypothesized that arterial stiffness was associated with PP variability (PPVAR) and determined arterial stiffness using AASI and PWV.

Ambulatory blood pressure (BP) data from untreated subjects (n=56, 28 men) with SBP >130 to <160 mmHg with a mean age 49 ±11 yrs were obtained in our Research Center. We calculated AASI as 1-regression slope of diastolic on systolic BP and PWV by carotid-femoral tonometry (Sphygmocor). We calculated PPVAR by the Standard Deviation (SD) and the Average Real Variability (ARV) of PP over 24h, day, and night periods (by patient diary).

AASI correlated positively with day PPVAR (r=0.57, p <0.000) and night PPVAR (r=0.34, P = 0.01) by SD. AASI correlated positively with day PPVAR (r=0.53, P <0.000 Fig1A) and night PPVAR (r=0.43 and p<0.001 Fig1B) by ARV. AASI was not significantly correlated with 24h PPVAR by SD or ARV. PWV was not correlated with any PPVAR estimate.

The relationship between AASI and PPVAR was only evident when the blood pressure periods were broken down into day and night. The 24 hour regression relationship of diastolic on systolic blood pressure with PPVAR appears different between day or night compared with the 24 hour aggregate, suggesting circadian differences in factors which modify PPVAR. Poor PWV correlation with PPVAR is likely due to relatively young age of our subjects.
Abstract 35: PI3K-mTOR Axis is a Critical Node for Hypothalamic Insulin Receptor Signaling and Sympathetic Activation Induced by Insulin
Authors: Muta Kenjiro , Morgan Donald A , Rahmouni Kamal
Published: March 2022

Insulin action in the hypothalamus produces sympathetic nerve activation with pathological implications in obesity and type 2 diabetes. However, the molecular events mediating the sympathetic effects evoked by hypothalamic action of insulin are poorly understood. Mammalian target of rapamycin (mTOR) which can be activated by the insulin receptor has emerged as an important regulator of sympathetic nerve activity (SNA). Phosphatidylinositol 3-kinase (PI3K) pathway is an important upstream regulator of mTOR signaling; and insulin stimulates PI3K in the hypothalamus. Thus, we postulate that the PI3K-mTOR axis is critical for insulin-induced sympathetic nerve activation. In hypothalamic GT1-7 neurons, we found that insulin (100 nM) activated mTOR signaling as indicated by the increase in phosphorylated levels of the downstream effectors, S6K (1.5-fold) and S6 (1.4-fold), measured by Western blot. In GT1-7 neurons, insulin also increased the phosphorylation levels of Akt (2.7-fold), a downstream effector of PI3K. Importantly, inhibition of PI3K (LY294202, 10 μM) abolished insulin-induced activation of S6K and S6. To study further the contribution of PI3K signaling to the insulin action, we used p110αD933/WT mice which carry a heterozygous mutation disrupting PI3K signaling. In wild type (WT) control mice, ICV administration of insulin (100 μU) significantly elevated the number of phospho-S6 positive cells in the hypothalamic arcuate nucleus (29±1, n=3, p<0.05 vs. vehicle), assessed by immunohistochemistry. This response was significantly attenuated in p110αD933/WT mice (12±3, n=3, p<0.05 vs. WT mice). Moreover, the increase in renal SNA, measured directly using multifiber recording, induced by ICV insulin (100 μU) was blunted (p<0.05) in the p110αD933A/WT mice (-15±25%, n=6) as compared to littermate controls (327±115%, n=6). In WT mice, ICV pre-treatment with an mTOR inhibitor (rapamycin, 1 ng) also attenuated the renal SNA response induced by ICV insulin (-16±14%, n=6), mimicking the effect of PI3K inhibition. These results suggest that hypothalamic PI3K-mTOR axis is a critical node in the control of renal sympathetic outflow by insulin.