bluuebunny/crossref_metadata_2025 huggingface dataset:
Nanoceria distribution, biotransformation, and safety/toxicity in the rat
Authors: Yokel Robert A , Wu Peng , Tseng Michael T. , MacPhail Robert C. , Graham Uschi M. , Dan Mo , Unrine Jason M. , Sultana Rukhsana , Hardas Sarita S. , Butterfield D. Allan , Grulke Eric A.
Published: June 2021

Little is known about engineered nanomaterial (ENM) physico‐chemical properties that influence distribution across the blood‐brain barrier (BBB) and associated CNS effects. Ceria ENM is being investigated as an anti‐oxidant for neurodegenerative disorders.

Objective

Determine the distribution, biotransformation, persistence and effects of ceria ENMs.

Methods

In‐house synthesized, characterized, citrate‐coated ~ 5, 15, 30 and 55 nm polyhedral/cubic, and a low‐aspect ratio, ceria were given iv to rats; terminated 1 h to 90 days later for determination of BBB integrity, cerium concentration and valence, oxidative stress markers, and histological changes.

Results

Very little of any of these ceria ENMs crossed the BBB. Less than 1% was eliminated in feces and urine in 2 weeks, with most (98%) in feces. Nanoceria accumulated in the mononuclear phagocyte system with no significant decrease over 90 days. Initial antioxidant changes were followed by pro‐oxidant effects in the brain and liver. Granulomatous formations were seen in liver and spleen. Valence of freshly prepared ceria ENM did not significantly change in situ.

Conclusions

Nanoceria as small as 5 nm does not readily cross the BBB. Nanoceria persists in the mammal, without change in valence, causing toxicity. These results support concern that highly biopersistent ENMs are associated with adverse responses. Support: US EPA STAR Grant RD‐833772.
An experimental modelling of shock lung
Authors: Alipbekova Aigul , Dalenov Ebolat
Published: June 2021

We consider, that at intrapleural introduction of oleic acid there occurred a sufficient hypoxic clear effect. The purpose of the study was working out the model of shock lung creating in experiment. For this purpose there was injected an oleic acid directly into tissues of both lungs. The dosage of a substance was 0,27 ml per 100 gr of animal's mass. The material of the study made 240 white rats ‐ males of 160–200 gr. The performed studies revealed sufficient changes in organism of experimental animals which testified the forming of respiratory distress syndrome. It influenced on: a) development of animal organism's lymphocytary ‐ macrophagal system dysfunctions, particularly, changes of some populations quantity and lymphocytes subpopulations, exhaustion of oxygen dependent systems of polymorpho‐nuclear leucocytes and blood macrophages and broncho‐alveolar lavage, oppression of their capacity to phagocytosis that may be a basis of immunodeficiency forming at this pathology; b) changing metabolic activity of ferments: succinate dehydrogenase, alkaline and acid phosphates, myeloperoxidase in lymphocytes and neutrophiles of peripheral blood in dependence on terms of shock lung development; c) sufficient activation of kinin system.
Real‐time contrast‐enhanced ultrasound (RT‐CEU) for the determination of flowmotion within skeletal muscle
Authors: Rattigan Stephen , Bradley Eloise M , Richards Stephen M , Keske Michelle A
Published: June 2021

Flowmotion within skeletal muscle in vivo is difficult to determine and has depended upon invasive studies with implanted Laser Doppler flow (LDF) probes. This study reports the novel use of RT‐CEU to determine flowmotion in rat muscle in vivo.

Anaesthetised rats were infused with Definity® microbubbles and the microvascular flow monitored by an US transducer (L9‐3) on the upper thigh. Contrast imaging was performed with a iU22 US machine and analysis of regions of interest by QLab (Philips) Wavelet analysis was used to determine the contribution by endothelial (0.006–0.02Hz), neurogenic (0.02–0.06Hz) and myogenic (0.06–0.3Hz) vasomotion. RT‐CEU data was compared to simultaneous subcutaneous data made with LDF surface probes (Moor Instruments).

Both RT‐CEU and LDF data demonstrated that muscle and subcutaneous tissues displayed flowmotion with a strong neurogenic rhythmical variation. Phentolamine treatment abolished the neurogenic flowmotion and lead to a recruitment of microvascular flow to the same extent as muscle contraction.

RT‐CEU provides a new technique that can be used to determine in vivo flowmotion within skeletal muscle of experimental animals and humans. This technique has the potential to provide novel information about defects in the vascular regulation in pathological conditions (insulin resistance, sepsis) that are associated with poor nutrient delivery. Funded by NHMRC(Aust).
Mulltiple steady states in microvascular networks
Authors: Carr Russell T , Geddes John B , Bozzuto Lisa
Published: June 2021

The computation of steady state blood flow requires the simultaneous solution of three conservation laws at each junction point in the network. First the RBC mass balance must be satisfied. This is governed by nonlinear equations at diverging junctions – the plasma skimming rule. The second conservation law is the momentum balance (or pressure distribution). Again this momentum balance is governed by a nonlinear function – the Fahraeus‐Lindqvist effect for apparent viscosity. Finally, the total mass balance for volumetric blood flow is a linear system of equations that must be satisfied. This nonlinear system can result in multiple steady states as network parameters are varied. Many of these multiple steady states arise from what are called saddle node bifurcations in the nonlinear dynamics.

The same network can have steady states with different flow, hematocrit and pressure distributions. Some may be attractive, while others are repulsive. The system can exhibit hysteresis. These results can have significant physiological consequences on flow and hematocrit distributions in response to blood flow control action.

Funding provided by UNH and Cornell University
Gender differences in cardiac function of Kv1.5−/− mice during aging
Authors: Ohanyan Vahagn , Yin Liya , Logan Suzanna , Enrick Molly , Hakobyan Tatevik , Kolz Cristopher L , Pung Yuh Fen , Bratz Ian , Chilian William
Published: June 2021

Voltage‐gated potassium channels, Kv1.5 channels in particular, play a critical role in coupling myocardial blood flow to cardiac work. However, the long‐term effects of impaired coupling of flow to metabolism, termed microvascular dysfunction, are not known. Moreover, coronary microvascular dysfunction leading to cardiac pathology is more frequent in females than in males. Based on these observations, we hypothesized that Kv1.5−/− mice, with impaired coronary metabolic dilation will gradually develop cardiac systolic and diastolic dysfunction. For this study, Kv1.5−/− male (n=10), Kv1.5−/− female (n=10) and WT male (n=6) mice were used. Echocardiography was performed at 6, 12 and 18 months to assess cardiac function. 12 month survival rate was significantly decreased in Kv1.5−/− female (62%) compared to male (91%) and WT animals (100%). This disparity was increased at 18 months: survival rate ‐ female 18%, male 72%, WT 100%. Ejection fraction was significantly lower in Kv1.5−/− female and male mice at all ages compared to WT mice. Diastolic function (E/A ratio) was significantly lower in Kv1.5−/− female mice at 18 months of age. Taken together, we conclude that microvascular dysfunction in Kv1.5−/−mice impairs cardiac systolic function in aged males and females, however, diastolic dysfunction is more severe in females and this appears to be linked to higher mortality.
Chronic heart failure (CHF) alters nNOS‐mediated control of skeletal muscle contractile function
Authors: Copp Steven , Hirai Daniel M , Ferguson Scott K , Holdsworth Clark T , Poole David C , Musch Timothy I
Published: June 2021

Neuronal NO synthase (nNOS)‐derived NO modulates skeletal muscle contractile function. It is unknown if altered nNOS function contributes to the muscle contractile dysfunction in CHF. We tested the hypothesis that nNOS‐mediated control of skeletal muscle contractile function and economy is impaired in CHF rats. Spinotrapezius muscles were exteriorized and linked to a force transducer in 5 healthy (LVEDP: 6±1 mmHg) and 4 CHF (coronary artery ligation, LVEDP: 13±1 mmHg) rats. Force was measured during 3 min of electrically‐induced 1 Hz (~6–8 V) twitch contractions pre‐ and post‐nNOS inhibition with SMTC (0.56 mg/kg i.a.). Muscle VO2 was calculated (Fick equation) from measured blood flow (radiolabelled microspheres), arterial O2 content, and microvascular Po2 (phosphorescence quenching). Healthy: SMTC elevated the force‐time integral (+8%) and reduced contracting steady‐state VO2 (−17%). SMTC elevated the steady‐state force/VO2 ratio (60±20 g/ml/min, P<0.05). CHF: SMTC did not alter the force‐time integral or contracting steady‐state VO2. Moreover, SMTC did not change steady‐state Δ force/VO2 ratio (−10±60 g/ml/min, P>0.05). The nNOS‐derived NO modulation of skeletal muscle function present in healthy rats is attenuated in CHF rats indicating CHF‐induced alterations in the control mechanisms regulating contractile function. (AHA Midwest Affiliate, NIH‐HL‐108328)
Normal human IgG attenuates intracellular superoxide level and permeability in human aortic endothelial cells (HAECs)
Authors: Wang Xiuqing , Wang Qianyu , Sun Zhongjie
Published: June 2021

Background

The protective role of normal human IgG, a circulating antibody, in vascular endothelial cells is not poorly understood. The purpose of this study was to investigate our hypothesis that IgG attenuates intracellular superoxide (O2−) levels and permeability in HAECs via activation of AMPK.

Results

IgG significantly decreased the intracellular O2− levels and endothelial permeability. NADPH oxidase activity and Rac1 activity were attenuated by IgG, which may lead to the decreased O2− formation. In addition, IgG increased MnSOD expression and eNOS Ser1177 phosphorylation contributing to IgG‐induced decrease in O2− level. Notably, IgG activated AMPK as evidenced by increased phosphorylation of AMPK (pAMPK). Interestingly, suppression of AMPK activity by an AMPK inhibitor (AI) abolished IgG‐induced decreases in O2− levels, endothelial permeability, stress fiber formation, and cell migration, suggesting that the protective effects of IgG on HAECs are mediated by AMPK activation. The mechanistic investigation further revealed that inhibition of AMPK prevented IgG‐induced decreases in Rac1 and NADPH oxidase activities and IgG‐induced increases in MnSOD expression and eNOS activity.

Conclusion

IgG may protect HAECs by attenuating O2− levels by activation of AMPK and subsequent downregulation of NADPH oxidase activity and upregulation of MnSOD expression and eNOS activity.
Endothelium‐dependent dilation is inversely related to hematocrit among healthy young and older adults
Authors: Pierce Gary L , Donato Anthony J , LaRocca Thomas J , Jablonski Kristen L , Seals Douglas R
Published: June 2021

Hemoglobin (Hb) is a catalytic sink for endothelial‐released nitric oxide (NO) into the artery lumen. Because elevated hematocrit (Hct) within the “normal range” is related to cardiovascular (CV) events, we hypothesized that a higher normal Hct would be associated with lower endothelium‐dependent dilation (EDD) in both young and older adults. Forearm blood flow (FBF) responses to incremental brachial artery infusions of acetylcholine (EDD) and sodium nitroprusside (endothelium‐independent dilation, EID) were assessed in young (n=22; mean ± SE; age 24 ± 1 yrs) and older (n=58; 62 ± 1 yrs) healthy adults with normal Hct. EDD (peak FBF) was lower in young and older adults (Young: 15.1 ± 2.6 vs. 25.1 ± 3.7 ml/ml FAV/min; Older: 12.1 ± 1.4 vs.16.8 ± 1.8, both P<0.05) with Hct above vs. below sex‐adjusted median values, but EID was not different. EDD was inversely related to Hct (n=80, r=−0.43, P<0.01), and this remained significant after adjustment for age, sex and other clinical characteristics. In a subset (n=18), inhibition of NO (Ng‐monomethyl‐L‐arginine) reduced EDD more in adults with higher (−63%) vs. lower (−39%) Hct, abolishing group differences (P>0.05). High‐normal Hb is associated with lower EDD in both young and older healthy adults as a consequence of reduced NO bioavailability. Impaired endothelial function may contribute to the increased CV risks of high normal Hct. NIH AG013038, AG000279, UL1 RR025780
Sodium nitrite treatment restores vascular endothelial function in old mice with CKD
Authors: Sindler Amy L , Zigler Melanie C , Yoshizawa Mutsuko , Baylis Chris , Seals Douglas R
Published: June 2021

We tested the hypothesis that sodium nitrite treatment would improve endothelial function in older mice with chronic kidney disease (CKD). Old (~27 mo) C57 male mice that underwent 5/6 nephrectomy (OCKD, n=4) had greater circulating plasma creatinine vs. old control (O, n=5) (0.067 ± 0.03 vs. 0.017 ± 0.002 mg/dl, p ≤ 0.01). Endothelium‐dependent dilation (EDD) (carotid artery dilation to acetylcholine [ACh]) was impaired in O vs. young (~6 mo, Y, n=4) (80 ± 1 vs. 96 ± 3%, p < 0.05), and was impaired further with CKD (71 ± 2%), which was mediated by reduced NO (EDD to ACh alone minus EDD to ACh + L‐NAME: Y 62 ± 9% vs. O 43 ± 2% vs. OCKD 32 ± 6%, p < 0.05). Sodium nitrite added to drinking water (50 mg/l) for 8 weeks completely restored EDD in OCKD (92 ± 4%) by restoring NO bioavailability (61 ± 19%). Incubation with the superoxide dismutase mimetic, TEMPOL, increased EDD in O and OCKD (96 ± 1 and 95 ± 3%), but had no effect in Y or OCKD mice treated with nitrite. Endothelium‐independent dilation did not differ among groups/conditions. Superoxide production (EPR spectroscopy) from aortic rings was greater in O vs. Y (4139 ± 406 vs. 1734 ± 368 AU), further increased in OCKD (5858 ± 1509 AU), and normalized by sodium nitrite treatment (2697 ± 265 AU). These results indicate that nitrite supplementation may be an effective therapy for enhancing vascular endothelial function in age‐associated CKD by improving NO bioavailability and oxidative stress.

Work supported by NIH AG013038, HL007822, AG000279
Impaired fasting blood glucose‐related exacerbation of age‐associated vascular endothelial dysfunction: protective effect of regular aerobic exercise
Authors: DeVan Allison E. , Eskurza Iratxe , Pierce Gary L. , Walker Ashley E. , Jablonski Kristen L. , Kaplon Rachelle E. , Seals Douglas R.
Published: June 2021

We tested the hypothesis that impaired fasting blood glucose (IFG) exacerbates vascular endothelial dysfunction with aging, and that this effect is smaller or prevented by habitual exercise. Brachial artery flow‐mediated (endothelium‐dependent) dilation (FMD) was lower (5.14±0.38 vs. 8.06±0.35 %Δ, P<0.001) in older adults with normal fasting blood glucose (NFG, ≤99 mg/dl, 62±1 yrs, n=37, 31M/6F) vs. young NFG controls (24±1, n=29, 26/3). FMD was further impaired (3.62±0.38 %Δ, P<0.01 vs. older NFG) in otherwise healthy older adults with IFG (100–125 mg/dl, 64±1, n=31, 26/5). Older adults with NFG who regularly (≥3d/w) performed moderate to vigorous aerobic exercise (62±1, n=23, 21/2) had largely preserved FMD (6.38±0.35 %Δ, P<0.05 vs. older NFG). Most importantly, FMD also was well preserved (7.02±0.65 %Δ) in older exercising adults with IFG (65±1, n=17, 13/4). These overall group differences remained significant (P<0.05) after controlling for endothelium‐independent dilation and clinical characteristics. In the pooled sample (n=137), FMD was inversely related to FG (r=−0.32, P<0.001), plasma insulin (r=−0.23, P<0.05) and an index of insulin resistance, HOMA‐IR (r=−0.28, P<0.01). IFG, a prediabetic state, exacerbates age‐associated vascular endothelial dysfunction. This adverse effect is prevented by regular aerobic exercise.

NIH AG013038, AG000279, AG031617, AG033994, AG027910, RR025780
Biologic nanoparticles promote mineralization of vascular smooth muscle cells in vitro
Authors: Hunter Larry W , Lieske John C , Miller Virginia M
Published: June 2021

Biologic nanoparticles (NPs) ‐ protein/mineral complexes isolatable from human calcified, but not uncalcified arteries and from plasma ‐ propagate and accumulate calcium phosphate when in cell culture conditions. IV injection of NPs into animals results in vascular calcification at sites of endothelial injury. Our aim was to determine if NPs promote mineralization by localizing to vascular smooth muscle cells (VSMCs), then sequestering calcium from their environment. Porcine VSMCs were cultured in 24‐well plates to sub‐confluency (day 0). 45Ca (1 μCi/well) to label deposited mineral was added to media containing vehicle (control), NPs (10 or 100 nephelometric turbidity units; NTU; average diameter 210 nm), or 2 mM Pi (positive‐control). After 15 days, VSMCs formed multicellular nodules. VSMCs in control media were negative for calcification by von Kossa staining; whereas micron‐sized to smaller particles stained positive for calcium in nodules from NP‐treated and Pi‐treated cells. Cells treated identically were demineralized with 0.6N HCl and the liberated 45Ca was quantified by scintillation counting. CPM/well (x103) were 1.5 in controls and 4.6, 28.2 and 64.7 in cells treated with 10 NTU NPs, Pi, and 100 NTU NPs, respectively. NPs localize to VSMCs in vitro and promote mineralization through sequestration of Ca from the environment. (Supported by NIH grant HL88988, and Mayo Clinic).
Increment in nNOS and Akt‐eNOS pathway in coronary arteries post‐myocardial infarction can prevent the onset of heart failure
Authors: Couto Gisele Kruger , Britto Luiz Roberto Giorgetti , Mill José Geraldo , Rossoni Luciana Venturini
Published: June 2021

The imbalance in NO modulation in coronary artery (CA) has been observed in cardiovascular diseases. We tested the hypothesis that NO modulation in CA could be differently adjusted after myocardial infarction (MI) in the presence or absence of heart failure (HF). Male Wistar rats were submitted to CA ligation (MI) or sham operation (SHAM). After 4 weeks, the infarcted rats were divided in two groups: without (INF) or with HF according to hemodynamic parameters and the septal CA was mounted in a wire myograph. The acetylcholine (ACh, 100ñM‐100μM)‐induced relaxation was decreased (35%) in HF, while it was enhanced (61%) in INF CA. The ACh‐induced NO production evaluated by 4,5‐diaminofluorescein diacetate was reduced in CA of HF, while it was increased in INF. The NO synthase (NOS) (L‐NAME, 100μM), neuronal (n) NOS (7‐NI, 100μM) and PI3‐kinase (LY294002, 50μM) inhibitors abolished the enhanced ACh‐induced relaxation observed in CA of INF. In addition, endothelial (e) and nNOS isoforms and Akt protein expression were greater in CA of INF than SHAM. The endothelium‐independent relaxation was similar among groups. CA from MI showed increased NO modulation due to nNOS and Akt‐eNOS pathway, although CA from HF presents endothelial dysfunction. These results support the idea that NO can prevent the onset of HF after MI. Supported: FAPESP/CNPq.
Time course of doxorubicin accumulation and dysfunction in the rat aorta
Authors: Gibson Noah M. , Hydock David S. , Hayward Reid
Published: June 2021

Doxorubicin (DOX) is a highly effective anthracycline antibiotic used to treat a wide array of cancers. Its use is limited due to dose‐dependent acute and chronic cardiovascular toxicity. However, little is known about the mechanisms of DOX induced vascular dysfunction. The purpose of this study was to determine if DOX accumulates in vascular tissue and contributes to the onset and progression of vascular dysfunction over time. Male Sprague‐Dawley rats were randomly assigned to receive 15 mg DOX/kg of body mass or saline (SAL); DOX rats were further randomized to be sacrificed 1, 3 or 5 days post treatment. Isolated aortic rings were used to examine vascular function, and aortic DOX accumulation was quantified using high performance liquid chromatography. While DOX did in fact accumulate in vascular tissue (204±89 ng DOX/g LV) within 24 hours, there was no significant difference in accumulation over time (183±107 ng DOX/g LV at 3 days; 156±60 ng DOX/g LV at 5 days). There were significant differences between SAL and DOX groups (p<0.05) in vasocontriction, endothelium‐dependent and ‐ independent vasodilation, however there were no functional differences over time with DOX treatment. Our results show that there is early‐onset, sustained vascular dysfunction during the 5 days following DOX treatment, which is accompanied by a sustained accumulation of DOX in vascular tissue.
Losartan abolished hyperglycemic effect of chronic mild and unpredictable stress in rats
Authors: Marcondes Fernanda Klein , Sanches Andrea , Costa Rafaela , Cunha Tatiana Sousa , Casarini Dulce Elena
Published: June 2021

We've shown previously that chronic mild and unpredictable stress (CMUS) induced endothelium dysfunction and increased renin‐angiotensin system (RAS) activity in rats. Since endothelial dysfunction and high RAS activity has been associated with insulin resistance the aim of this study was to evaluate the effect of losartan on the response to oral glucose tolerance test (OGTT) of rats submitted to CMUS. Male Sprague‐Dawley rats were divided into four groups: control, CMUS, losartan (50mg/Kg) and CMUS + losartan (n=12/group). CMUS protocol consisted in applying different stressor stimuli (immobilization, overnight illumination, water and food deprivation, wed bedding and reversed light/dark cycle) 7 days/wk during three consecutive weeks (from the 3rd to 5th wk) of the experimental protocol, 7 weeks long. OGTT has been performed one week after CMUS. Data were analysed by ANOVA bifatorial (p<0.05). All procedures were approved by Institutional Ethics Committee. CMUS and CMUS+losartan groups presented higher corticosterone plasma level in comparison with non stressed groups. CMUS group presented higher area under the curve of glucose during OGTT in comparison with control group (885,2 ± 10,07 vs. 754,4 ± 16,08 mmol x min/L), without difference between losartan (745,9 ± 15,40) and CMUS+losartan group (816,4 ± 17,01 mmol x min/L). These data show that increased RAS activity is involved in the CMUS hyperglycemia.
Mineralocorticoid Receptors Activation in Choroid Plexus and PVN Exaggerates Salt‐Induced Development of Hypertension in Stroke‐Prone Spontaneously Hypertensive Rats
Authors: Nakano Masatsugu , Hirooka Yoshitaka , Matsukawa Ryuichi , Ito Koji , Sunagawa Kenji
Published: June 2021

Background

In salt sensitive models, the increases in cerebrospinal fluid Na+ concentration (CSF [Na]) derived from choroid plexus (CP) play a pivotal role in high salt (HS) induced hypertension. Mineralocorticoid receptors (MR) contribute to Na+ handling. Therefore, we investigated whether MR in CP and paraventricular nucleus of hypothalamus (PVN) exaggerates salt‐induced development of hypertension in stroke‐prone spontaneously hypertensive rats (SHRSP).

Methods and Results

Serum glucocorticoid inducible kinase 1 (sgk1) expression levels in CP and PVN were greater in SHRSP than those in Wistar‐Kyoto (WKY) rats (N=6, P<0.05) associated with increases in CSF[Na] (155.6 ± 0.81 vs. 152.8 ± 0.49 mM, P<0.01). The depressor response to intracerebroventricular (ICV) infusion of MR blocker, eplerenone (10 ng) was greater in SHRSP than in WKY (Δarterial pressure (AP) −25.2 ± 1.4 vs. −8.2 ± 1.4 mmHg, N=5, P<0.001). In SHRSP, HS intake (8%) increased AP and urinary norepinephrine excretion (uNE) compared with those by regular salt diet (2 weeks after ΔAP 41.7 ± 3.2 mmHg, ΔuNE 0.24 ± 0.07 μg/day, N=5, P<0.05) associated with increases in MR and sgk1 expression in CP and PVN. These alterations were attenuated by ICV infusion of eplerenone (10 μg/kg/day).

Conclusion

MR activation in CP might contribute to increases in CSF[Na], thereby exaggerates salt‐induced development of hypertension via MR activation in PVN in SHRSP.
Enhanced endothelin‐1 response in mesenteric arteries in acquired and inherited forms of hypertension
Authors: Pulgar Victor M , Jeffers Anne B , Diz Debra I , Aileru Azeez A
Published: June 2021

Both endothelin and the ET‐1 receptor are altered in ouabain sensitive and hypertensive (mRen‐2)27 transgenic rats, consistent with the lowering of blood pressure in both models of hypertension with either AT1 or ET‐1 receptor blockade. We hypothesized that responses to ET‐1 receptor activation are enhanced in the vasculature of the animals with acquired and genetic hypertension. Sprague‐Dawley rats were implanted subcutaneously with a 60‐ day 1.5 mg ouabain (OUA) pellet (n = 6 control, n = 6 treated). Blood pressure (BP) was recorded daily via tail cuff for 8 weeks. Transgenic hypertensive (mRen‐2)27 rats (n = 6) were also used. Third branch mesenteric arteries were mounted in a wire myograph (DMT‐USA). Maximal responses to KCl and ET‐1 were expressed as KMAX (mN/mm) and ET‐1MAX (% KMAX) and sensitivity as pD2 (pD2=‐Log[EC50]). Pre‐incubation with indomethacin (10−5M) or LNAME (10−4M) were performed in parallel. OUA increased systolic BP by 30% (p<0.05) whereas age‐matched (mRen‐2)27 rats had a 55% higher BP vs control (p<0.05). ET‐1MAX was similar in the three groups [ET‐1MAX (mRen‐2)27 136 ± 5%; OUA 128 ± 14% vs control 137 ± 8 % KMAX]. However sensitivity to ET‐1 was increased in hypertensive vs control rat vasculature [(mRen‐2)27 8.6 ± 0.1; OUA 8.5 ± 0.2 vs control 8 ± 0.1, p<0.05]. A greater sensitivity to ET‐1 may serve as mediator of enhanced vascular tone in both models of hypertension.

Supported by NIMHD MD002303
Atorvastatin, sildenafil and their combination down‐regulate matrix metalloproteinases (MMPs) in 2K1C hypertension
Authors: Guimaraes Danielle Aparecida , Rizzi Elen , Ceron Carla Speroni , Oliveira Alisson Martins , Gerlach Raquel Fernanda , dos Santos José Eduardo Tanus
Published: June 2021

Atorvastatin (ATORVA) and sildenafil (SILD) may downregulate MMPs, especially when both drugs are combined (ATORVA+SILD), and therefore attenuated the vascular changes found in hypertension. We evaluated whether these drugs attenuate vascular MMP‐2 levels/activity in 2kidney‐1clip (2K1C) hypertension. Sham or 2K1C hypertensive rats were treated with vehicle, ATORVA (50 mg/Kg), SILD (45 mg/Kg) or both for 8 weeks. Systolic blood pressure (SBP) was monitored weekly. Aortic MMP‐2 levels/activity was evaluated by gelatin and in situ zymography and by immunofluorescence. We studied whether these drugs directly inhibit human recombinant MMP‐2 (hrMMP‐2) in vitro activity. All treatments attenuated 2K1C hypertension. Aortas from 2K1C rats showed higher MMP‐2 levels (75, 72 and 64 kDa) compared with sham. All treatments attenuated 2K1C hypertension‐induced increases in MMP‐2 levels. All treatments attenuated the increases in total MMP activity and in MMP‐2 levels without additive effects. However these drugs had no effects on hr‐MMP‐2 activity. ATORVA and SILD reduced the SBP and vascular proteolytic activity in 2K1C hypertension, without additive effects. Our results suggest that both drugs can prevent, at least in part, the vascular dysfunction of hypertension by downregulating vascular MMP levels. Supported by: Fundação de Amparo a Pesquisa do Estado de São Paulo (FAPESP), Conselho Nacional de Desenvolvimento Cientifico e Tecnológico (CNPq).
MicroRNA‐98 negatively regulates hypoxia‐induced endothelin‐1 expression in human pulmonary artery endothelial cells
Authors: Kang Bum-Yong , Park Kathy K. , Sutliff Roy L. , Hart C. Michael
Published: June 2021

Endothelin‐1 (ET‐1) plays a critical role in endothelial dysfunction and contributes to the pathogenesis of hypoxia‐induced pulmonary hypertension (PH). We previously reported that PPARγ activation attenuates hypoxia‐induced ET‐1 expression and pulmonary vascular cell proliferation. The contribution of posttranscriptional mechanisms to these findings has not been defined. We hypothesized that PPARγ regulates microRNA (miR)‐98 to modulate ET‐1 expression and endothelial proliferation. Human pulmonary artery endothelial cells (HPAECs) were exposed to hypoxia (1% O2) or normoxia (21% O2) for 72 h. PPARγ was activated in HPAEC by treatment with rosiglitazone (RSG, 10 μM) during the last 24 h. Hypoxia reduced miR‐98 expression, and RSG attenuated reductions in miR‐98 expression. PPARγ knockdown with siRNA (100 nM) reduced miR‐98 expression and increased expression of NF‐κB‐p50 and ET‐1. Transfection with a miR‐98 mimic (30 nM) decreased ET‐1 mRNA levels and HPAEC proliferation, but did not alter PPARγ expression. These findings suggest that PPARγ activation during hypoxia restores reduced levels of miR‐98 to suppress ET‐1 expression and HPAEC proliferation. PPARγ‐mediated regulation of microRNAs may represent a novel therapeutic target to modulate PH pathogenesis. Funding: Atlanta VAMC Research Service, NIH DK074519/HL102167 (CMH), Emory/Children Healthcare of Atlanta CEB F16788‐00 (BYK).
Synergistic effects of prostacyclin analogs and PDE5 inhibitors on ATP release from erythrocytes (RBCs): implications for the treatment of pulmonary hypertension
Authors: Sprague Randy S , Elrick Mollisa M , Bowles Elizabeth A , Zdanovec Amber K , Stephenson Alan H , Ellsworth Mary L
Published: June 2021

Idiopathic pulmonary arterial hypertension (IPAH) is a disease of uncertain etiology that leads to right ventricular failure and death. IPAH treatment includes the vasodilators: prostacyclin (PGI2), other PGI2 receptor agonists and phosphodiesterase 5 (PDE5) inhibitors. We reported that: 1) RBC‐derived ATP vasodilates isolated‐perfused lungs, 2) RBCs possess PGI2 receptors and 3) PGI2 analogs stimulate ATP release from RBCs via a signaling pathway that requires increases in cAMP. Here we investigated the hypothesis that PDE5 inhibitors can enhance PGI2 agonist‐induced ATP release from human RBCs. ATP release from RBCs was measured in the absence and presence of the PGI2 agonist, UT‐15C or its vehicle, saline, alone or in combination with the selective PDE5 inhibitor, zapranist (ZAP, 10 μM). Compared to control cells, UT‐15C (1 uM) treated RBCs (n=7) demonstrated an increase in ATP release that was potentiated by ZAP pretreatment (p>0.05). When RBCs were incubated with UT‐15C at 30 (n=7) or 100 nM (n=7), no ATP release was detected. However, when the same cells were pretreated with ZAP, ATP release was stimulated by both concentrations of UT‐15C. These results provide an alternative mechanism by which PGI2 agonists and PDE5 inhibitors interact to stimulate pulmonary vasodilation making RBCs novel targets for development of new therapies for IPAH. Supported by ADA, NIH and United Therapeutics.
Genetic regulation and functional relevance of the p67phox gene in salt‐sensitive hypertension
Authors: Feng Di , Yang Chun , Lazar Jozef , Mattson David , OˈConnor Paul , Cowley Allen W.
Published: June 2021

A narrow region on rat Chr 13 was identified to harbor salt‐sensitive genes. p67phox, a cytosolic subunit of NAD(P)H oxidase, is located in this region. We have previously found that on a high salt diet, the renal outer medulla (OM) of Dahl salt‐sensitive (SS) rats exhibited higher levels of p67phox expression and NAD(P)H oxidase activity than salt‐resistant congenic rats that contain the p67phox allele from the salt‐resistant Brown Norway rat. We generated the first p67phox null mutant (p67phox−/−) SS rat in which we observed significantly reduced salt‐sensitive hypertension. In our present study, we sequenced a 1650 bp promoter region and found that the SS allele of p67phox had a 204 bp deletion and four SNPs compared to the BN allele. The activity of the SS p67phox promoter was 1.7 fold higher than that of the BN p67phox promoter. We further characterized p67phox−/− rats and showed that they had a 40% reduction in OM H2O2 levels measured from interstitial fluid collected by microdialysis. Respiratory burst responses of peritoneal macrophages to phorbol 12‐myristate 13‐acetate were abolished in p67phox−/− rats. p67phox−/− rats also showed reduced renal injury, including reduced OM fibrosis, infiltrated T cells and macrophages, and glomerulosclerosis. These data provide new insights into the genetic regulation and functional relevance of p67phox in salt‐sensitive hypertension. (HL‐82798; HL‐29587)
Endoplasmic reticulum (ER) stress mediates nuclear factor‐κ‐B (NFκB) activation in the subfornical organ (SFO) during slow‐pressor angiotensin‐II (Ang‐II) hypertension
Authors: Young Colin N. , Davisson Robin L.
Published: June 2021

We recently demonstrated activation of ER stress pathways and the transcription factor NFκB in the SFO of the brain during slow‐pressor Ang‐II hypertension (HTN). Emerging evidence suggests these molecular mechanisms are closely linked. Here we tested the hypothesis that ER stress is upstream of NFκB activation in the SFO during slow‐pressor Ang‐II HTN. C57 mice underwent SFO targeted injection of an adenovirus encoding firefly luciferase downstream of NFκB response elements along with installation of an intracerebroventricular (ICV) cannula. After recovery, osmominipumps were implanted for infusion of slow‐pressor Ang‐II (600 ng/kg/min). NFκB activity was monitored in vivo by daily bioluminescence imaging during concomitant ICV administration of the chemical ER stress inhibitor TUDCA or vehicle (Veh). NFκB activity in the SFO increased gradually during Ang‐II infusion, with a pre‐hypertensive surge at day 5. This response was significantly reduced by ICV TUDCA (Veh 119744 ± 27568 vs. TUDCA 57312 ± 8164 photons/s; n=8–9, p<0.05). In line with these findings, acute induction of ER stress, via ICV administration of the chemical ER stress inducer thapsigargin (TG; 1ug), caused a robust increase in SFO‐NFκB activity (Veh Δ11919 ± 4501 vs. TG Δ194445 ± 46882 photons/s; n=4–5, p<0.05). These findings suggest that ER stress is upstream of NFκB activation in the SFO during slow‐pressor Ang‐II HTN. HL063887, HL864624
Potential Proteomic Biomarkers of Secondhand Smoking‐ Induced Cardiovascular Disease
Authors: Zweier Jay L. , Tewari Arun K , El-Mahdy Mohamed A , Abdelghany Tamer M , Hemann Craig , El-Sherbiny Gamal A.
Published: June 2021

Identification and characterization of proteomic changes that can serve as predictive markers of the onset and progression of secondhand smoke (SHS)‐induced disease are now in the forefront of cardiovascular (CV) research. Our objective was to identify a differential pattern of protein expression in the plasma of SHS‐exposed mice through a time course study. C57BL/6 mice were exposed for up to 48 weeks to SHS generated from 3R4F reference research cigarettes using the Teague smoking machine. Utilizing 2D‐DIGE and MS/MS, we have found a differential expression of 35 proteins in SHS‐exposed mice as compared to controls. Some of those are involved in vascular function, coagulation, or metabolism. Expression of ceruloplasmin, haptoglobin, fibrinogen, apolipoprotein E and murinoglobulin was ~2.5, 7, 3, 2 and 3.5‐fold lower than controls, respectively. The expression of alpha‐1 antitrypsin and complement C3 was ~6 and 13‐fold higher than controls, respectively. The alterations in fibrinogen, alpha‐1 antitrypsin, haptoglobin, and complement C3 are of particular interest since these have been linked to CV and lung pathology mediated by oxidative stress. Our data suggest that SHS exposure leads to changes in plasma protein expression. These changes can potentially serve as predictive markers of the onset and/or temporal progression of CV and pulmonary disease in secondhand smokers. (NIH # HL63744 to JLZ)
Epigenetically‐induced vascular dysfunction and hypertension by in vitro fertilization; prevention by addition of melatonin to the culture media
Authors: Rexhaj Emrush , Rimoldi Stefano , Giacobino Ariane , Sartori Claudio , Allemann Yves , Scherrer Urs
Published: June 2021

In vitro fertilization (IVF) induces vascular dysfunction in mice and humans that may be related to an epigenetic mechanism. We speculated that this problem is induced during blastocyst culture. We therefore tested the effects of the addition of melatonin – a pineal gland hormone that acts on epigenetic regulation ‐ to the culture media on DNA‐methylation of genes in vascular tissue and vascular function in vitro and in vivo in 12 weeks‐old IVF mice. We found that addition of melatonin (10−6 M) to the culture media prevented the dysmethylation of imprinted genes H19 (P=.05, vs. vehicle), Gtl2 (P=.04, vs. vehicle), Peg3 (P<.01, vs. vehicle) and of the eNOS promoter (P=.02, vs. vehicle) in IVF mice. Prevention of vascular DNA dysmethylation in IVF mice by melatonin was accompanied by normalization of the mesentery‐artery responsiveness to acetylcholine in vitro (P<.0001 vs. vehicle) and arterial blood pressure in vivo in vivo (105±4 vs. 109±4 mmHg, X±SD, P=.04 vs. vehicle). Here we show for the first time that IVF induces vascular dysfunction by an epigenetic mechanism that is triggered during blastocyst culture. We speculate that in humans IVF induces vascular dysfunction by a similar mechanism. If so, modification of the culture media may allow to prevent this problem in humans.
Mapping a genetic biomarker of blood pressure to <807.3kb using two genetically hypertensive rats
Authors: Kumarasamy Sivarajan , Gopalakrishnan Kathirvel , Yerga-Woolwine Shane , Farms Phyllis , Liu Jiang , Joe Bina
Published: June 2021

Two of the most widely used strains for mapping quantitative trait loci (QTLs) for blood pressure (BP) are the Dahl Salt‐sensitive (S) rat and the Spontaneously Hypertensive Rat (SHR). To map a previously identified BP QTL that differentiate these two strains, we continued substitution mapping studies on a BP QTL on chromosome 9 identified by a linkage study. A total of 11 new congenic substrains were developed and characterized. BP and heart weights of two of these substrains with introgressed segments of 2.05Mb and 807.3Kb were both significantly (p<0.001) lower than that of the S by 56 ± 6.61 and 41 ± 8.55 mm Hg (p<0.001)), respectively. The mRNA and protein levels of Tmeff2 were lower in the congenic substrain spanning 2.05Mb. Further, because Tmeff2 was the only protein‐coding candidate gene within the substrain spanning 807.3kb, which also had a BP effect, it was prioritized as a candidate gene. Tmeff2 is known to be upregulated in patients from multiple cohorts with cardiac hypertrophy. Because Tmeff2 is also known to affect cell proliferation, we assessed the proliferation rates of cells obtained from S and the congenic substrains with BP effect. Proliferation of cardiac fibroblasts was significantly slower in the congenic compared with S (p<0.05). Taken together, these results suggest that variants within the 807.3Kb congenic segment that alter the expression of Tmeff2 can be prioritized as candidate biomarkers controlling blood pressure and associated cardiac hypertrophy in both rats and in humans.
RNAi Silencing of TNFα Attenuates Cold‐induced Pulmonary Hypertension (CIPH)
Authors: Crosswhite Patrick , Sun Zhongjie
Published: June 2021

Background

Exposure to a moderate cold temperature (5ºC) causes pulmonary hypertension. Tumor necrosis factor‐alpha (TNF‐α) was increased in lungs and pulmonary arteries of rats prior to cold‐induced elevation of pulmonary arterial blood pressure (PABP). The purpose of this study was to investigate if the upregulation of TNFα plays a role in CIPH.

Methods and Results

Three groups of male SD rats (8 rats/group) received IV delivery of adeno‐associated virus (AAV)‐carrying TNFα small hairpin RNA (TNFαshRNA), AAV‐carrying scrambled shRNA, and PBS, respectively, 3 days before exposure to a cold environment (5°C). Three additional groups with the same treatments were housed in a warm chamber (25°C) and serve as controls. Eight weeks after injection, right ventricular (RV) systolic BP (equivalent of PABP) was recorded and PA smooth muscle cells were isolated for in vitro studies. TNFαshRNA significantly decreased cold‐induced elevation of RV BP but not RV hypertrophy. TNFαshRNA prevented cold‐induced upregulation of TNFα in lungs and PAs, confirming effective silencing of TNFα. TNFα inhibition decreased cold‐induced increases in intracellular levels of O2−, calcium, and cGMP in PA SMCs. RNAi silencing of TNFα also prevented the cold‐induced increase in PCNA, a marker of cell proliferation.

Conclusions

The upregulation of TNFα may play a key role in the development of CIPH and PASMC proliferation.
Chronic and intermittent hypoxia differentially regulate the left ventricular inflammatory and extracellular matrix responses
Authors: Ramirez Trevi Ann , Jourdan-LeSaux Claude , Joy Anne , Zhang Jianhua , Dai Qiuxia , Mifflin Steve , Lindsey Merry L
Published: June 2021

While hypoxia effects on the lung and right ventricle have been well characterized, no one to date has used chronic sustained (CSH) and chronic intermittent hypoxia (CIH) models to determine how changes in the lungs and right ventricle feed forward to alter the structure and function of the left ventricle (LV). In this study we assessed the LV response to hypoxia by comparing male Sprague‐Dawley rats exposed to normoxia (control, n=18), to CSH (n=12), or to CIH (n=12) for 7 days. Right ventricle and lung weights increased in CSH compared to control and CIH, while the LV weights were similar among all three. Of 168 inflammatory and extracellular matrix genes evaluated through RT2‐PCR, only 7 changed in CSH LV compared to control. Ltb, Cdh4, Col5a1, Ecm1, Mmp11, and Timp2 all increased, while Tnfrsf1a decreased, indicating an overall increase in inflammatory status of the CSH LV. 103 genes in the LV were significantly altered by CIH and revealed a decrease in inflammatory gene profiles. Genes that decreased include Ltb, Spp1, Ccl5, Mmp9, and Fn. Consistent with a decreased inflammatory status, Hif‐2α and eNOS protein levels were decreased in CIH LV. In conclusion, our results indicate that the inflammatory response is increased in CSH and decreased in CIH. This report is the first to demonstrate specific and differential changes seen in the LV during CSH vs CIH. This work was supported by funding from the NIH/NHLBI and the VA.
Melanocortin 4 receptors in the paraventricular nucleus of the hypothalamus do not mediate chronic metabolic or cardiovascular effects of leptin after established obesity in mice
Authors: Hamza Shereen M. , Dubinion John H. , Adi Ahmad , Hall John E.
Published: June 2021

Leptin modulates metabolic and cardiovascular regulation via the central melanocortin system and the melanocortin 4 receptor (MC4R) causing a reduction in appetite and increases in blood pressure, heart rate and oxygen consumption (VO2). The specific contribution of MC4R in the paraventricular nucleus (PVN) in mediating these actions is unknown. We used Cre/loxP technology to create mice with total deletion of MC4R (LoxTB) or selective rescue of MC4R in the PVN (LoxTBSIM1). LoxTB female mice were heavier than wild type (WT) littermates (44±3.2 vs 27±1.5g, p<0.05). Selective rescue of MC4R in the PVN (LoxTBSIM1) did not restore normal body weight (41±4.2g). Mice were instrumented for telemetric blood pressure (BP)/heart rate (HR) measurement and i.v. infusion. After 5 days of control, leptin was infused (2μg/kg/min i.v.) for 7 days. Leptin reduced food intake in WT (− 1.2±0.3g, p<0.05; −32%) but not in LoxTB (−0.5±0.3g) or LoxTBSIM1 (−0.3±0.7g). Leptin‐induced increases in BP and HR were greater in WT (5±1mmHg, 21±9 bpm) compared to LoxTB (− 1±2 mmHg, −27±5 bpm) or LoxTBSIM1 (−2±1 mmHg; −23±4 bpm, p<0.05). VO2 also increased more in WT (13.9±1.5) than in LoxTB (6.2±2.5) or LoxTBSIM1 (6.0±3.0 mL/kg/min, p<0.05). Rescue of MC4R in the PVN does not restore normal body weight and in established obesity PVN MC4R do not appear to mediate chronic metabolic or cardiovascular effects of leptin in female mice. NIH HL51971, CIHR.
Caffeic acid and ferulic acid improves diet induced metabolic syndrome in mice
Authors: da Costa Bocco Barbara Miranda Leite , Lorena Fernanda Beraldo , Fernandes Gustavo Werpel , Cysneiros Roberta Monterazzo , Ribeiro Miriam Oliveira
Published: June 2021

The use of plants for medicinal benefits has been widely explored. Analysis of chloroform extract of Baccharis articulate indicated the presence of fenolic compounds as caffeic acid (CA) and ferulic acid (FA) that act as an antioxidant and anti‐inflammatory and anti‐diabetic, among others. The aim of this study was to analyze the effect of CA and FA on the metabolic syndrome. To this end, C57 mice were treated for 60 days with high fat diet (HFD) (40%) to induce the metabolic syndrome and concomitantly with CA plus FA. Our results showed that the combination of CA (0.9 mg / kg / day) and FA (25 or 50 mg/kg/day) reduced body weight gain induced by HFD (9.3±3.7 HFD; 3.7±0.7 (CA0,9+FA25) and 3.5±0.7 (CA0.9+FA50); p<0.001), fasting hyperglycemia (258±39 HFD, 208±28 (CA0.9+FA25); p<0,05 and 94±14 (CA0.9+FA50); p<0.001), but not hypercholesterolemia (100 ±14 HFD vs. 128±14 (CA0.9+FA25) and 104±7 (CA0.9+FA50)) induced by high fat diet. Surprisingly, all the parameters worsen when treated with 1.8 mg/kg/day of CA. In conclusion, the combination of caffeic acid and ferulic acid improves the abnormalities observed in metabolic syndrome induced by high fat diet.
Enhancement of cellular Na+K+ATPase activity in the mouse renal tissue in‐vitro with low concentration of peroxynitrite
Authors: Maiti Arpan Kumar , Islam Mohammed Toriqul , Majid Dewan S A
Published: June 2021

Endogenous production of peroxynitrite (ONOO°−) in normal condition has been suggested to play a renoprotective role. However, the exact mechanism associated with this protection is not yet clearly defined. ONOO°− at higher concentrations (μM to mM; in pathology) can inhibit cellular Na+K+ATPase (NKA) activity. However, the effect of ONOO°− on NKA activity at low conc. (nM; which is physiologically more relevant) remains unknown. To examine the hypothesis that ONOO°− at low conc. enhances NKA activity, preparations of cellular membrane fraction from mouse renal tissue (n=5 in each case) were incubated for 10 min each with different conc. of ONOO°− (10 nM to 200 μM). NKA activity in these samples was measured via a colorimetric assay capable of detecting inorganic phosphate. At high conc. (1–200 μM), ONOO°− caused dose‐dependent inhibition of NKA activity (−3.0±0.6 % to – 36.4±1.4 %) as reported previously. However, NKA activity remained unchanged at 100 and 500 nM ONOO°− conc., but interestingly, at lower conc.(10 and 50 nM), ONOO°− caused a small but significant increases in the NKA activity (3.3±1.1 % & 3.1±0.6 %). These responses were prevented at preparations co‐incubated with ONOO°− scavenger, mercapto ethyl guanadine (200 μM). These data indicate that NKA activity remains unaffected or rather increased at the physiological conc. of ONOO°− that facilitates the reno‐protective role for this oxidant molecule.
New Opportunities and Challenges in Network Biology
Authors: Ideker Trey
Published: June 2021

The past decade has seen an explosion in “genome‐era” technologies which profile genes, proteins, metabolites and the intricate web of interconnections among them. Much of bioinformatics and functional genomics is now focused on methods to assemble these diverse measurements into models of functional networks and pathways within the cell.

Mapping the genetic network underlying the response to DNA damage. We will describe our ongoing efforts to apply ChIP‐sequencing and synthetic‐lethal screens to map how the cellˈs transcriptional network is remodeled by DNA‐damaging conditions.

Network‐based biomarkers for disease diagnosis and personalized medicine. Recently, we have shown that networks can also serve as powerful biomarkers and in many cases are more predictive than any individual gene. Our approach is to project gene and/or protein expression profiles of each patient onto the known human genetic network map to identify pathways that are predictive of disease.

Protein network comparative genomics. We are developing a library of standard approaches for comparing protein interaction networks across species, conditions, and network types.

We are developers of the Cytoscape platform, an Open‐Source software environment for visualization and analysis of biological networks and models (http://www.cytoscape.org/).