bluuebunny/crossref_metadata_2025_split huggingface dataset:
Design of Novel Oxime Reactivators and Direct Characterization of Their Interaction With OP‐ChE Conjugates
Authors: Radic Zoran , Kalisiak Jaroslaw , Cochran Rory , Zhang Limin , Fokin Valery V , Sharpless K Barry , Taylor Palmer 
Published: June 2021

Oximes are commonly used nucleophilic reactivators of phosphylated acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). Their reactivation rates are several orders of magnitude slower than the rate of ChE inhibition by OPs or rate of acetylcholine hydrolysis, catalyzed by the same enzyme. The rate of oxime reactivation can therefore be substantially enhanced. Using “click chemistry” we prepared more than 800 new oximes and tested them for ability to reactivate phosphylated human ChEs. Interaction constants for three of oximes with native, diethylphosphorylated, VX and cyclosarin phosphonylated and aged human ChEs were determined. Stopped‐flow kinetics of oxime interaction was monitored using quenching of intrinsic tryptophan fluorescence of ChEs as an indicator of oxime binding. We developed mutant AChE resistant to aging and screened the capacity of oximes for reactivation of the mutant. Combination of the mutant enzyme with oxime HI6 proved most efficient for reactivation with rate constant in 10^5 1/(M*min) range being the most efficient ChE based aging resistant catalytic scavenger developed, to date. We also screened our oxime libraries for reactivation of human BuChE and identified candidate oxime reactivators capable of enhancing reactivation rates to an efficient catalytic scavenger of nerve agents.Supported by U01 NS58046 grant to PT.
Affinity of dopamine receptor antagonists determines the time to maximum effect on cocaine self‐administration
Authors: Fey Brittney K , Tabet Michael R , Norman Mantana K , Tsibulsky Vladimir L , Millard Ronald W , Norman Andrew B 
Published: June 2021

The time course of the dopamine receptor antagonist‐induced acceleration of cocaine self‐administration may correspond to the time to achieve maximum fractional occupancy. If so, the time to maximal response (Tmax) will be proportional to antagonist affinity. Male Sprague‐Dawley rats self‐administered cocaine (unit dose of 3 μmol/kg) at regular intervals. The cocaine concentration at the time of each self‐administration (agonist Cmin) was calculated. A series of 9 antagonists spanning a 10,000‐fold range of affinities for D2‐like dopamine receptors was administered i.v. and the effect of each antagonist on the calculated cocaine Cmin was determined.All of the antagonists accelerated cocaine self‐administration with a concomitant increase in the calculated cocaine Cmin. The antagonist doses that produced approximately equivalent increases in the maximum cocaine Cmin ranged from 0.015 to 4 μmol/kg. The mean Tmax values across this series of antagonists ranged from approximately 5 min to 120 min. Furthermore, the antagonist Tmax values were inversely related to their KD values across this set of antagonists.These data are consistent with the hypothesis that the agonist Tmax reflects the time course of the increase in antagonist fractional occupancy of D2‐like dopamine receptors. It is concluded that the Tmax is related to antagonist affinity.Supported by DA018538 and ASPET SURF program.
Role of Growth factor receptor transactivation in high glucose‐induced increased levels of Gq/11α and signaling in vascular smooth muscle cells
Authors: Descorbeth Magda , Anand‐Srivastava Madhu B. 
Published: June 2021

We have recently shown that high glucose (HG) increased the expression of Gq/11α, PLCβ and mediated signaling in A10 vascular smooth muscle cells (VSMC). Since HG has been shown to increase growth factor receptor (GFR) activation, we investigated the role of epidermal growth factor receptor (EGF‐R) and platelet‐derived growth factor receptor (PDGF‐R) transactivation in HG‐induced enhanced expression of Gq/11α and PLCβ. Pre‐treatment of A10 VSMC with HG (26 mM) for 3 days, increased the levels of Gqα, G11α, PLCβ‐1 and PLCβ‐2 proteins as determined by Western blotting which were restored to control levels by AG1478, an inhibitor of EGF‐R, AG1295, an inhibitor of PDGF‐R and PP2, an inhibitor of c‐Src but not by PP3. In addition, endothelin‐1 (ET‐1)‐stimulated production of IP3 that was enhanced by HG was also restored towards control levels by AG1478, AG1295 and PP2. HG also increased the phosphorylation of EGF‐R and PDGF‐R which was abolished by AG1478, AG1295 and PP2. Furthermore, HG‐induced enhanced levels of Gqα, G11α and PLCβ were also attenuated by inhibition of MAP Kinase, PI3‐Kinase and c‐Src. In addition, GFR inhibitors also attenuated HG‐induced enhanced phosphorylation of ERK1/2 and AKT. These results suggest that c‐Src‐induced transactivation of growth factor receptor contributes to the high glucose‐induced enhanced expression of Gq/11α/PLC and ‐mediated cell signaling through MAPK/PI3K pathway.This work was supported by Canadian Institutes of Health Research grant
Contamination of the Norepinephrine Pro‐Drug Droxidopa by Dihydroxyphenylacetaldehyde
Authors: Holmes Courtney S , Goldstein David S 
Published: June 2021

BackgroundDroxidopa (3,4‐threo‐L‐dihydroxyphenylserine, L‐DOPS) is a norepinephrine (NE) pro‐drug under development to treat orthostatic hypotension. 3,4‐Dihydroxyphenylacetaldehyde (DOPAL) is a toxic catecholaldehyde.MethodsThirteen subjects took 400 mg Droxidopa orally, with blood sampled at baseline and 1, 2, 3, 6, 24, and 48 hours after drug administration and assayed for plasma L‐DOPS, NE, and DOPAL. Droxidopa in acidic solution was vacuum centrifuged for 1 hour at 30 degrees centigrade.ResultsDroxidopa contained 0.01% DOPAL. At 6 hours, all subjects had DOPAL detected in plasma (1.89 nmol/L, p=0.0001). Across the sampling times plasma DOPAL correlated with plasma L‐DOPS (r=1.00). The mean increment in plasma DOPAL averaged more than 4 times the mean increment in plasma NE (0.39 nmol/L). In 2 patients with Parkinson disease and orthostatic hypotension, DOPAL was detected at baseline (0.12 nmol/L) and increased by about 70‐fold after Droxidopa. After 1 hour of vacuum centrifugation, L‐DOPS was converted completely to DOPAL.InterpretationDroxidopa is contaminated with DOPAL. After Droxidopa ingestion DOPAL is detectable in plasma. Droxidopa is converted completely to DOPAL under conditions resembling those in the stomach.
Endothelin‐A Receptor Blockade Improves Angiogenic Signaling in the Stenotic Kidney
Authors: Chade Alejandro R , Kelsen Silvia , Bailey James E , Fails Fredrick 
Published: June 2021

BackgroundChronic renal artery stenosis (RAS) induces endothelin (ET)‐1 up‐regulation and via the ET‐A receptor can promote renal injury. Chronic RAS promotes renal microvascular (MV) damage and loss but the role of the ET/ET‐A is unknown. Thus, we hypothesized that ET‐A blockade in RAS will preserve the MV architecture by improving the angiogenic signaling in the stenotic kidney.MethodsRAS was induced in 12 pigs and observed for 6 weeks. In 6 of them, ET‐A blocker was chronically administered (RAS+ET‐A, 0.75 mg/kg/day). Additional animals were used as normal controls (n=6). Single‐kidney MV architecture and function was quantified using CT and renal angiogenic and apoptotic signaling determined by western blot.ResultsET‐A blockade in RAS improved the renal expression of VEGF and augmented downstream mediators such as angiopoietin‐1 and 2, its receptor Tie‐2, and eNOS. Interestingly, it also decreased Bax/Bcl‐2 ratio, suggesting attenuated apoptotic signaling in the stenotic kidney. Consequently, MV density and function in the RAS+ET‐A kidney was improved.ConclusionChronic ET‐A blockade improved angiogenic and decreased apoptotic signaling in the stenotic kidney. These ultimately preserved the renal MV architecture and function in the stenotic kidney, supporting a novel renoprotective role of ET‐A blockade in chronic RAS.Support: AHA‐SDG‐0830100N
Meningeal vascular response to estrogen‐based therapy in post ovariectomized pigs
Authors: Glinskii Olga , Huxley Virginia , Rubin Leona , Abraha Tsghe , Glinsky Vladislav 
Published: June 2021

Loss of gonadal function in women is associated with increased incidence of stroke, cranial aneurysms, and neurodegenerative disease. Those sequelae infer that intracranial vascular adaptation at the onset of reproductive senescence confers enhanced cerebrovascular risk. Our data from in vivo pig model show that following ovariectomy (OVX), meningeal vasculature undergoes substantial remodeling characterized by loss of capillaries, increase in average microvessel size and augmented permeability associated with estrogen receptor (ER) alpha dependent decrease in angiopoietin‐1 expression. Even 2 months post OVX, vascular networks remain destabilized and prone to further remodeling. Mechanistically, this process is regulated in part by ER‐related pathways and, therefore, is estrogen‐dependent. However, effects of estrogen (E2) based therapy on such remodeling depend strikingly on the treatment regimen. We show that flat‐dose E2 therapy fails to prevent vascular remodeling, while E2 regimens mimicking natural estrous cycle‐associated E2 fluctuations prevent post OVX microvascular alterations. It appears that, as opposed to conventional flat‐dose regimens, pulsed E2‐based therapy could present a compelling strategy for preventing massive intracranial vascular remodeling during reproductive senescence compromising functional and structural integrity of meningeal microvasculature.Supported by AHA National SDG 0830287N, NIHHL078816, NASA NNJ05HF37G, NIHC06RR017353
Aging‐induced changes in angiotensin II‐induced contractions and tachyphylaxis of isolated carotid arteries
Authors: Koller Akos , Toth Peter , Rozsa Bernadett , Cseplo Peter , Hamar Janos , Vamos Zoltan 
Published: June 2021

Angiotensin II (Ang II) is a potent vasoconstrictor, yet the potential aging‐ induced changes in the magnitude of responses have not yet been explored. We hypothesized that aging induces changes in the magnitude of vasomotor responses to Ang II. Thus carotid arteries from newborn (8 days: 8d), young (2 months: 2m) and old (24 months: 24m) rats were isolated and placed in a wire myograph to measure changes in their isometric tension. Two dose response curves to Ang II were obtained in a sequential manner (1‐, and 2‐administration (Adm). Contractions of vessels to 1‐Adm of Ang II in 8d vessels were significantly less compared to those of 2m and 24m, which were similar in magnitude (1.40±0.2 mN and 5.2±0.6 and 5.7±0.39 mN, respectively). Compared to the 1‐Adm, contractions of vessels to 2‐Adm of Ang II were significantly reduced in 2m and 24m vessels (4.4±0.54 and 0.5±0.19mN), whereas it did not change in vessels of 8d rats (1.28±0,2 mN).These findings suggest that in carotid arteries 1) Ang II‐induced contractions increase with age, 2) repeated administration of Ang II elicits tachyphylaxis and the magnitude of tachyphylaxis increases with aging.(Hungarian Nat. Sci. Res. Fund.‐OTKA K71591, K67984, AHA Founders Aff., 0555897T)
The essential role of inducible nitric oxide synthase for mobilization of bone marrow stem cells to the cochlea
Authors: Shi Xiaorui , Nuttall Alfred L. 
Published: June 2021

Inducible nitric oxide synthase (iNOS) produces an essential signal for homing of circulating bone marrow derived stem cells (BMDSC) to target tissues. We find that the iNOS responsible for homing of BMDSC in the vascular “lateral wall” of the cochlea is from resident macrophages. The de novo macrophages appear to be essential for vascular repair in a model of inner ear ischemia produced by loud sound. The data show that mice deficient in iNOS (Nos2−/−) or mice treated with an iNOS‐inhibitor had a reduction of the mobilization of bone marrow progenitor cells to the cochlea and less hearing function recovery after acoustic trauma. Mechanistically, stromal cell‐derived factor‐1(SDF‐1) and its chemokine (CXC motif) receptor 4 (CXCR4) is required for stem cell mobilization, and was found to be reduced in the bone marrow of Nos2−/− mice and in animals treated with iNOS inhibitor. Loud sound induces iNOS expression in local perivascular cells (macrophages) which then becomes a major cell for NO production in the local microenvironment. These findings indicate that iNOS expressed by local resident macrophages influences the recruitment of BMDSC. The homing is abrogated by inhibition of SDF‐1 a/CXCR4 productions. Movement of BMDSC into the vascular area could be a basis of vascular remodeling and repair following loud sound damage.Support: NIDCD DC 00888, NIDCD DC 00105, NIDCD DC 005983 (p30)
In vivo inhibition of Nuclear Factor of Activated T‐cells (NFAT) results in enhanced levels of anti‐inflammatory IL‐10 in the retina of diabetic mice
Authors: Zetterqvist Anna V , Nilsson‐Öhman Jenny , Kotova Olga , Nilsson‐Berglund Lisa M , de Frutos Garcia Sergio , McGuire Paul , Gonzalez‐Bosc Laura V , Gomez Maria F 
Published: June 2021

Hyperglycemia is an important risk factor for the development of diabetic retinopathy. Recently we showed that high glucose activates NFAT in cerebral artery smooth muscle through local release of extracellular nucleotides, acting on P2Y receptors, leading to increased [Ca2+]i and calcineurin activation. NFAT activation promotes cell proliferation and the expression of inflammatory markers, such as IL‐6, COX‐2 and osteopontin, which are elevated in diabetic retinopathy. Here we investigate whether: 1) NFAT is activated by hyperglycemia in the wall of retinal microvessels (RM) and 2) inhibition of NFAT signaling may have a protective effect on the retina of diabetic mice. Using confocal immunofluorescence microscopy we show that NFATc3 is expressed in the endothelium of RM and readily activated by high glucose ex vivo and in vivo (intra peritoneal glucose tolerance test). This activation is inhibited by the NFAT blocker A‐285222 and by the ecto‐nucleotidase apyrase. Chronic hyperglycemia in STZ‐treated NFAT‐luc mice leads to increased NFAT transcriptional activity in isolated RM. Interestingly, two weeks after the first STZ injection, levels of the potent anti‐inflammatory cytokine IL‐10 were decreased in retina and this was prevented by treatment with A‐285222. Results suggest that NFAT inhibition may protect the retina of diabetic mice.Funding: Swedish Heart & Lung Foundations, RAC HSC‐UNM 624195
Cardiovagal baroreflex sensitivity is not correlated to sympathetic baroreceptor sensitivity within healthy, young individuals
Authors: Dutoit Andrea , Hart Emma , Charkoudian Nisha , Wallin BG , Curry Tim , Joyner Michael 
Published: June 2021

The purpose of this study was to evaluate the relationship between the cardiac and sympathetic baroreflex sensitivities within healthy, young individuals. The gains in cardiac (as measured by changes in R‐R interval, RRI, and heart rate, HR) and sympathetic (as measured by changes in muscle sympathetic nerve activity, MSNA) baroreflex during rapid changes in arterial blood pressure were compared in 27 normotensive individuals (18 men and 9 women; age 24 yrs ±1.24; body mass index 24 ±0.51 cm/kg2, mean ±SEM). Heart rate, arterial blood pressure (brachial artery catheter), and peroneal MSNA were recorded under resting conditions (HR 58 ±1.6 bpm; SBP 124 ±2.7 mmHg; DBP 71 ±1.7 mmHg; MAP 87 ±1.7 mmHg; MSNA BF 17 ±1.6 bursts/min) as well as during rapid changes in arterial blood pressure induced by sequential boluses of nitroprusside and phenylephrine (Modified Oxford technique). Integrated baroreflex sensitivities for MSNA and HR were analyzed using the slopes of the linear portions of the MSNA‐diastolic blood pressure and RRI‐systolic blood pressure relationships, respectively. When individual cardiac baroreflex sensitivity was compared to sympathetic baroreflex sensitivity, no correlation (R2=0.0003) was found. These results indicate that although both cardiac and sympathetic efferents function in baroreflex control of the autonomic system, there does not appear to be a direct correlation in their function within healthy normotensive individuals.Research Support: NIH HL083947 and Swedish Medical Council grant 12170
Early‐staged electro‐cardiac conduction in larvae and embryonic zebrafish
Authors: YU FEI , Wolla Christopher David , Chen Jaunian , Hsiai Tzung 
Published: June 2021

Zebrafish (Danio rerio) is an emerging model organism for pharmacological studies, tissue engineering, and genetically modified cardiomyocytes. However, early‐staged cardiac function in zebrafish has not been well understood. We proposed to monitor electrocardiography (ECG) in zebrafish at embryonic and larva stage. Larvae were sedated with 0.01% tricane methanesulfonate and placed in E3 medium (5 mM NaCl, 0.17 mM KCl, 0.4 mM CaCl2, and 0.16 mM MgSO4) under microscope. A 5μm micropipette filled with 300% Danieau solution were in direct contact with the larva heart for voltage signal recording, and a second micropipette electrode was connected to the ground. Signal processing and wavelet transform enhanced the signal‐to‐noise ratios. ECG signals displaying distinct P wave and QRS complex were acquired from 7 days post‐fertilization (dpf) to 42 dpf. Larva ECG patterns started to transform to the adult features at 35 dpf. When the larvae were treated with Amiodarone (1μM), a Class III anti‐arrhythmic potassium channel blocker, single‐peak QRS complex degenerated into multiple‐peak and wide QRS duration. Thus, we provide a unique in vivo zebrafish model to assess developmental‐staged cardiac conduction phenotype with implication for high throughput research.Research supported by NHLB 083015 and NHLBI 068689.
The functional differences between biventricular (BiV) endocardial and epicardial pacing in a dyssynchronous left ventricle are dependent on atrioventricular (AV) delay
Authors: Howard Elliot Jacob , Covell James W , Mulligan Lawrence J , Omens Jeffrey H , McCulloch Andrew D , Kerckhoffs Roy C.P. 
Published: June 2021

Biventricular (BiV) pacing is used to reduce electromechanical dyssynchrony in patients with left bundle branch block (LBBB). LV lead placement is limited by the patient's anatomy and condition, yielding controversy over selecting an optimal pacing site. We hypothesized that the addition of a LV stimulus that restores transmural activation sequence at late‐activated regions is an important determinant of improved LV function.To test this hypothesis, dyssynchrony (simLBBB) was produced in open‐chest dogs (n=6). We compared the effects of an additional LV stimulus at late (94.1 ms) activated regions during endocardial (Endo) and epicardial (Epi) pacing at varying atrioventricular (AV) delay. Regional deformation was assessed by biplane cineradiography of implanted beads.Compared to baseline, simLBBB significantly reduced LV function. Compared to simLBBB, BiV Endo pacing (AV delay of 40 ms) significantly improved systolic function: dPdt max (51%), LVESP (48.8 %), and E33 (0.55). BiV Epi pacing (AV delay of 40 ms) did not differ from simLBBB. At higher AV delays (80–130 ms), BiV Epi pacing significantly improved systolic function, however this improvement was less than BiV Endo pacing.We conclude that Endo LV pacing has a functional advantage over Epi LV pacing. Endo LV pacing in late‐activated regions may serve to reduce electrical and mechanical dyskinesis, thereby improving ventricular performance.
NO negatively regulates cell surface expression of CD73 in sheared endothelial cells
Authors: Fan Fan , Ohta Masanori , Nandedkar Sandhya , Li Weidong , Jo Hanjoong , Miura Hiroto 
Published: June 2021

Both NO and adenosine produced from eNOS and cell surface CD73 in endothelial cells (EC), respectively, are potent vasodilator and anti‐inflammatory molecules. Adenosine increases eNOS expression and activity in EC. We examined if NO regulates cell surface expression of CD73 in EC under shear stress.In human coronary artery EC, 24‐hour exposure to laminar shear stress (LS; 15 dyne/cm2) increased CD73 mRNA expression (2.1±0.3‐fold vs. static condition, p<0.05, n=10–17), whereas oscillatory shear (OS; ± 5 dyne/cm2) had no effect (0.9±0.2 fold, n=6). However, there was no difference in total CD73 protein levels between LS and OS, since LS caused cleavage of CD73 into the medium (static 22±0.3 and LS 37±0.2‐fold increase vs. fresh medium density, n=4) leading to less surface CD73 expression. Addition of an NO donor DETA‐NONOate abolished surface CD73. This was consistent with less surface expression of CD73 in LS‐exposed regions of mouse aortas than in OS‐regions. On the other hand, an NOS inhibitor LNAME augmented the mRNA up‐regulation (3.7±1.1 fold, n=5) and total and surface protein expressions in LS‐exposed EC. This was consistent with increased CD73 protein expression in eNOS−/− mouse aortas.These results indicate that NO negatively regulates EC surface expression of CD73 by down‐regulating the mRNA expression and by facilitating cleavage of surface CD73 protein under LS.
Expression of iNOS is reduced by an inhibitor of fatty acyl CoA synthetase.
Authors: Weis Margaret Teresa , Maliha George , Chen Xiong 
Published: June 2021

ObjectiveTo test the hypothesis that fatty acyl CoA derivatives of fatty acids modulate cytokine stimulated iNOS expression in C6 astrocytes.RationaleTriacsin C, a fungal metabolite, inhibits fatty acyl CoA synthetase in many cell types. The compound inhibits fatty acid induced iNOS expression and apoptosis in isolated islets of Langerhans (Shimabukuro et al 1996), suggesting that fatty acyl CoA derivatives regulate iNOS expression.MethodsC6 cells were stimulated with a cytokine mix consisting of TNFα (60 ng/mL), ILβ (2 ng/mL) and IFNγ (100 units/mL) in the presence or absence of triacsin C. Expression of iNOS was estimated by measuring iNOS protein by immunoblotting. In vivo iNOS activity was estimated by measuring media nitrite (Greiss reaction) and in vitro iNOS activity was measured by conversion of [14C]arginine to [14C]citrulline in the absence of added calcium.ResultsIn C6 cell homogenates, triacin C inhibited FACS activity in a concentration dependent manner, ranging from 31.9 ± 1.5% at 300 nM to 71.1 ± 3.8% at 5 μM. The effect of 5 μM triacsin C on cytokine‐stimulated media nitrite, catalytic activity and immunoreactive protein were measured at 18 and 24 hours:

% of Cytokine Stimulated Activity

Media Nitrite
Catalytic Activity
Immunoreactivity

18 hours
62.5 ± 6.9%
103 ± 11.1%
25.1 ± 13.2%

24 hours
80.6 ± 1.39%*
373 ± 72%*
36.6 ± 5.81%

= p< 0.05 as compared to 18 hours by Student's ttest; n=3 independent experiments for each parameter, each done in duplicate or triplicate.

ConclusionsThese results suggest that triacsin C inhibits cytokine stimulated NO formation, and that iNOS protein is significantly less in triacsin C treated cells. Surprisingly, not all of the immunoreactive iNOS protein seems to be catalytically active. The data suggest that triacsin C may delay cytokine‐stimulated expression of iNOS and thus its overall activity in C6 cells.