bluuebunny/crossref_metadata_2025_split huggingface dataset:
Fluorine Magnetic Resonance Assessment of siRNA Delivery to Endothelial Cells via Perfluorocarbon Nanoparticles
Authors: Myerson Jacob Wheatley , Kaneda Megan M , Lanza Gregory M , Wickline Samuel A 
Published: June 2021

RNA interference mediated by the introduction of exogenous siRNAs has been a powerful tool for the modulation of gene expression. The ability to track siRNA delivery would be advantageous for future in vivo studies.To design a synthetic vehicle to serve as both an siRNA delivery agent and an imaging agent, perfluorocarbon nanoparticles (PFC NP) were loaded with the cationic lipid DOTAP in the lipid monolayer to form transfection complexes with siRNA to the VCAM‐1 gene. siRNA loading onto nanoparticles was measured via PAGE gel. Mouse 2F2B endothelial cells were incubated with PFC NP/siRNA for 4h. 19F MR spectroscopy was performed at 11.7 T to determine the number of NP bound to each cell. mRNA levels were measured 48h after transfection to assess knockdown.The selected loading conditions resulted in 645 siRNA molecules per nanoparticle. Incubation with 2F2B cells led to a roughly linear increase in bound nanoparticles over the range of concentrations tested. In one condition which resulted in a 72% reduction in VCAM‐1 mRNA levels, 19F spectroscopy data reported 5563 particles per cell.For future in vivo studies, a trackable delivery agent would aid in the determination of localization of siRNA delivery to specific tissues. To this end, we have developed a nanoparticle which is able to deliver siRNA to mouse endothelial cells and whose signal can be detected by 19F MR spectroscopy and imaging.
Human CYP3A Expression under Hypertonic Stimuli In Vivo
Authors: Chuang Andrew I , Yang Mingdong , Dalvi Pooja , Gonzalez Frank J , Harper Patricia A , Ito Shinya 
Published: June 2021

We previously showed that human CYP3A expression in vitro is under the transcriptional control mediated by NFAT5, which is a tonicity responsive transcriptional factor. Interestingly, hypertonicity increases CYP3A expression, while hypotonicity decreases it, suggesting constitutive regulation roles of the NFAT5‐CYP3A pathway. In contrast, mouse Cyp3a was unresponsive to ambient hypertonicity in vitro. In the present study, our objective was to show if human CYP3A expression can be induced by hypertonic stimuli in vivo. The humanized CYP3A4–3A7 transgenic mice were subjected to 4 cycles of 24h water deprivation every other day, or to a 7‐day high‐salt diet (8% salt content, HSD), that mimics the dehydration and hypertonic luminal content in the intestine, respectively. In the kidney of mice undergoing the cyclic dehydration, NFAT5‐regulated BGT1 mRNA was significantly increased compared to control (5.9±1.1 fold: M±SEM; p=0.003). Fold‐changes of the CYP3A4 and 3A7 were 2.5±0.5 (p=0.02) and 0.7±0.2 (p=0.13), respectively. In mice after HSD, fold‐induction of the duodenal BGT1, CYP3A4 and 3A7 expressions were 25.2±8.2 (p=0.02), 4.7±0.9 (p=0.01) and 2.0±0.3 (p=0.08), respectively. Our findings suggest that human CYP3A transcription is under the influence of ambient tonicity. Direct involvement of NFAT5 in vivo awaits further studies.CIHR supported
Modulation of the human polymorphic hs1,2 enhancer within the 3′IgH regulatory region by TCDD
Authors: Sulentic Courtney E.W. , Fernando Tharu M. , Ochs Sharon 
Published: June 2021

2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD) is a potent environmental toxin known to inhibit immunoglobulin (Ig) gene expression. Transcriptional regulation of the Ig heavy chain (IgH) involves the 3′IgH regulatory region (3′IgHRR) and its enhancers (hs3, hs1,2, and hs4) which contain DNA binding sites for several transcription factors including NF‐κB and dioxin response elements (DRE). TCDD induces binding of the aryl hydrocarbon receptor complex to DREs within the 3′IgHRR and inhibits murine 3′IgHRR activation in a well characterized mouse B‐cell line (CH12.LX). In humans, a polymorphism of the hs1,2 enhancer resulting in varying numbers of a 53 bp sequence has been correlated with autoimmune diseases like IgA nephropathy and Celiac disease. The repeated sequence contains κB and DRE binding sites. The objective of this study was to comparatively evaluate the effect of TCDD and LPS on human and mouse hs1,2 enhancers in the CH12.LX model. In transient luciferase studies, an increased number of repeats in the human hs1,2 enhancer increased the sensitivity to LPS. Interestingly, TCDD also markedly enhanced human hs1,2 activity and even augmented LPS‐induced activation. This starkly contrasted with TCDD‐induced inhibition of the mouse hs1,2 and 3′IgHRR in LPS‐stimulated CH12.LX cells suggesting a species difference in IgH transcriptional regulation.(Supported by NIEHS R01ES014676)
Amelioration of the acute cardiovascular effects of cocaine by a longer acting mutant cocaine esterase in freely moving rhesus monkeys
Authors: Collins Gregory Thomas , Carey Kathy Ann , Ko Mei‐Chuan 
Published: June 2021

Cocaine is the most common illicit drug related to emergency room admissions, with the majority of visits due to cocaine‐associated chest pain resulting from increases in blood pressure (BP) and heart rate (HR). These studies were aimed at characterizing the capacity of a longer acting mutant form of cocaine esterase (T172R/G173Q; DM CocE) to ameliorate the cardiovascular effects of cocaine in four freely moving rhesus monkeys implanted with radio‐telemetric probes (DSI; D70‐PCT) capable of collecting continuous measures of BP and HR. DM CocE (0, 0.032, 0.1, 0.32, 1.0, or 3.2 mg/kg; IV) was administered 10 min after a dose of 3.2 mg/kg; IV cocaine (M2 was tested at 1.0 mg/kg; IV), and BP and HR were recorded for the 120 min following cocaine administration. DM CocE dose‐dependently, and rapidly decreased cocaine‐induced elevations in BP in each of the four monkeys, with saline‐like levels observed within the first 5 min. Similarly, DM CocE dose‐dependently decreased cocaine‐induced elevations in HR in three of the four monkeys, although over a slightly longer timecourse. These results suggest that DM CocE may provide a novel, and effective therapeutic for the treatment of acute cocaine toxicity.Supported by NIDA grant DA 023213
Adolescent Maturation of D1 and D2 Dopamine Receptor Interactions
Authors: Dwyer Jennifer Buenzle , Martinez Lesly , Ganddini Gabriela , Perez Mayra , Leslie Frances M 
Published: June 2021

Adolescence is a developmental period during which neuropsychiatric disorders often emerge. This onset is likely influenced by the continued maturation of mesocorticolimbic dopamine (DA) systems. Consistent with this development, adolescents respond uniquely to drugs that target DA systems, with blunted behavioral responses to indirect agonists like amphetamine. We have previously shown using direct agonists, that the functional sensitivity of D1 and D2‐like receptors changes with age. To evaluate age differences in D1/D2 behavioral interactions, adolescent and adult rats were given combinations of second messenger‐selective direct D1 and D2 agonists. Drug‐induced locomotor and stereotypic behaviors were measured, and brains were collected for analysis of immediate early gene (IEG) expression via in situ hybridization for c‐fos and arc mRNA. Behavioral data suggests that D1/D2 interactions are immature during adolescence, as adults show synergistic behavioral responses to D1/D2 combinations while adolescents do not. IEG analysis suggests age differences in the neural circuitry underlying these behaviors. Interregional IEG correlation further suggests that dopaminergic drugs alter functional connectivity in the limbic system, with distinct effects in adolescence. These data have important implications for pharmacotherapy design and use in this population.Support: DA19138, PhRMA Foundation
Development of behavioral and neuronal responses to social stress: early adolescence as a sensitive period
Authors: Bingham Brian , McFadden Kile , Zhang Xiaoyan , Bhatnagar Seema , Beck Sheryl , Valentino Rita 
Published: June 2021

The research goal was to determine whether social stress differentially impacts on behavior and brain physiology during critical windows in adolescent development. Early adolescent (EA), mid‐adolescent (MA) and adult rats were exposed to 7 days of a social stressor, the resident‐intruder paradigm. Social stress had divergent effects on behavior in EA and adult rats, promoting active coping behaviors in EA rats and decreasing active coping behaviors in adult rats as determined by the defensive burying test and response to swim stress. Mid‐adolescent rats exposed to social stress showed no changes in the behavioral endpoints. Effects of social stress in EA rats was not mimicked by chronic restraint stress. Because the locus coeruleus (LC)‐norepinephrine system has been implicated in certain active coping behaviors, LC neuronal activity was recorded in EA rats exposed to social stress and matched controls. LC spontaneous firing rates were higher in EA rats exposed to social stress. Moreover, intra‐LC infusion of the CRF antagonist, DPheCRF12‐41, inhibited LC neurons of stressed rats but not controls. These data suggest that exposure to social stress in early adolescence promotes tonic secretion of CRF into the LC to activate this system. This CRF‐induced activation of the LC‐norepinephrine system may underlie the promotion of active‐coping behaviors noted in socially stressed EA rats.MH058250
Effect of beta‐adrenergic stimulation on the rapid delayed rectifier K current in canine ventricular cardiomyocytes
Authors: Nanasi Peter 
Published: June 2021

Activation the PKA pathway has been reported to suppress the cardiac rapid delayed rectifier K current (IKr) in expression systems, however, the phosphorylation‐dependent regulation of IKr is still controversial. In the present study, therefore, the effects of activators and inhibitors of the PKA and PKC pathways were studied on IKr in canine ventricular myocytes using the whole cell patch clamp technique. Superfusion with isoproterenol (ISO), forskolin, or intracellular perfusion with PKA activator cAMP analogues (cAMP, 8‐Br‐cAMP, 6‐Bnz‐cAMP) increased IKr significantly by 30–50 %. Inhibition of PKA (by H‐89) and PKC (by GF‐109203X or chelerythine) decreased IKr significantly by 70–90 %. The IKr stimulator effect of ISO was fully eliminated by H‐89, chelerythine, and PKA activator cAMP analogues – but not by the EPAC activator 8‐pCPT‐2′‐O‐Me‐cAMP. The results indicate that beta‐adrenergic stimulation increases IKr in canine ventricular cells, and this effect involves the activation of both PKA and PKC.Financial support was provided by the Hungarian Research Fund (OTKA‐K68457).
Molecular Expression and Distribution of Calcium‐Activated Chloride Channel TMEM16A in Mouse Heart
Authors: Zhang Mi , Yoon Carol , Kuhn Andrew R. , Dai Yanping , Ye Linda , Singer Cherie A. , Burkin Dean J. , Duan Dayue Darrel 
Published: June 2021

BackgroundCalcium‐activated Cl− channels (CaCCs) are important in many cellular and physiological functions and in certain cardiac diseases. But the molecular identity of CaCCs in the heart is still elusive. Recent evidence suggests TMEM16A as the gene encoding the pore subunit of CaCCs. In this study, the molecular expression and tissue distribution of TMEM16A in mouse heart were quantitatively analyzed.MethodsTotal RNA was isolated from atria and ventricles of mouse heart (C57/BL6) and subjected to a quantitative real‐time PCR and Western blot analysis. TMEM16A mRNA and protein expression was calculated relative to 18S rRNA and GAPDH expression, respectively, within the same tissues.ResultsThe transcriptional level of TMEM16A as estimated by mRNA ratio of TMEM16A/18s rRNA was highest in the left atrium (2.13±1.5, n=3), followed by right atrium (1.31±0.38), right ventricle (0.11±0.01), and left ventricle (0.051±0.004). Immunoblots of atrial and ventricular wall protein extracts demonstrated TMEM16A immunoreactivity at 113 kDa. The protein expression levels of TMEM16A were consistent with the same tissue‐distribution pattern as the transcriptional levels.ConclusionThese results suggest that TMEM16A is heterogeneously expressed in mouse cardiac tissues, which may be responsible for the different functional activity of CaCCs in different tissues of the heart. (Supported by NCRR P‐20 RR‐15581)
Renal Microvascular Disease and the Responses to Renal Revascularization
Authors: Chade Alejandro R , Kelsen Silvia , Fails Fredrick , Bailey James 
Published: June 2021

Renal angioplasty and stenting (PTRA) is the most frequent therapeutic approach for renal artery stenosis (RAS). However, renal function recovers in only 30% of the cases. The causes of these poor outcomes are still unknown. We hypothesize that preserving the renal microcirculation in the stenotic kidney will improve the responses to PTRA.RAS was induced in 12 pigs. In 6 pigs, VEGF‐165 was single‐infused intra‐renally (RAS+VEGF, 0.05 ug/kg). Additional animals were used as normal controls (n=6), and all were followed for 10 weeks. Single‐kidney function was assessed in vivo using ultra‐fast CT after 6 weeks of RAS. Then, RAS and RAS+VEGF underwent PTRA, VEGF repeated, and CT studies performed 4 weeks post‐PTRA. Pigs were then euthanized, the stenotic kidney removed, renal microvascular (MV) architecture reconstructed ex‐vivo using 3D micro‐CT, and renal fibrosis quantified.Degree of RAS and hypertension were similar in RAS and RAS+VEGF. Renal function and MV density were decreased in RAS but improved in RAS+VEGF.PTRA largely resolved RAS. Improvements in blood pressure and renal function were greater in RAS+VEGF+PTRA than in RAS+PTRA, accompanied by a 29% increase in MV density and decreased fibrosis.Preservation of the MV architecture and function in the stenotic kidney improved the responses to PTRA, indicating that renal MV rarefaction plays a role in determining the responses to PTRA. This study indicates that the damage and early loss of renal MV is an important determinant of the progression of renal injury in RAS and instigates often irreversible damage.Support: AHA‐SDG‐0830100N
Adenosine is involved in the arteriolar vasodilatory responses to a single skeletal muscle contraction
Authors: Ross Graeme A , Armstrong Marika L , Murrant Coral L 
Published: June 2021

To investigate the role of Adenosine (Ado) in response to a single contraction we stimulated 4–5 skeletal muscle fibres in a hamster cremaster preparation and measured the diameter of arterioles at the site of overlap with the stimulated muscle fibres before and after a single contraction (250ms train duration) at 4, 20 and 60Hz. Muscle fibres were stimulated prior to and following 30min incubation with Ado receptor antagonists 1,3‐dipropyl‐8‐sulphophenylxanthine (DPSPX, 10−5M) or xanthine amine congener (XAC,10−6M), or ecto‐5′‐nucleotidase inhibitor adenosine 5′‐[α,β‐methylene]diphosphate (AMPCP,10−5M). A dilation within 10s was observed at 4, 20 and 60Hz (1.0±0.2; 1.5±0.2; 2.0±0.2μm respectively) which was significantly attenuated by DPSPX (4Hz −0.1±0.2; 20Hz −0.2±0.3; 60Hz 0.04±0.2μm), XAC (4Hz 0.5±0.2; 20Hz 1.0±0.2; 60Hz 0.3±0.2μm), and AMPCP (4Hz 0.03±0.1; 20Hz −0.3±0.3; 60Hz −0.3±0.2μm). A secondary dilation at 20s at 20Hz (1.4±0.3μm) and 60Hz (1.1±0.1μm) was significantly attenuated by DPSPX (20Hz −0.1±0.1; 60Hz −0.1±0.3μm), XAC (20Hz 0.3±0.2; 60Hz 0.5±0.2) and AMPCP (20Hz 0.2±0.2; 60Hz 0.02±0.1μm). These findings implicate Ado in the vasodilations in response to a single muscle contraction.NSERC
Effect of arginase on NO‐mediated protein modifications in endothelial cells
Authors: Krotova K. , Zharikov S. , Block E. R. , Patel J. M. 
Published: June 2021

It is known that arginase is able to modify Nitric Oxide (NO) production through competition with NO synthase for the common substrate L‐arginine. We hypothesize that by modifying NO production arginase may affect NO‐mediated protein modifications – S‐nitrosylation and nitration. We studied NO production and arginase activity in human endothelial cells (EC) exposed to hypoxia. We found that the exposure of EC to 1% O2 for 24h induces an increase in NO production and an elevation in the expression of arginase II (Arg II) isoform which localizes in mitochondria. To study the effect of Arg II on S‐nitrosylation and nitration we used the arginase‐specific inhibitor BEC. After exposure of EC to hypoxia with or without BEC, nitrated proteins were precipitated with nitro‐tyrosine antibody, and then nitration of individual proteins was analyzed by Western blotting. In another set of experiments S‐nitrosylated proteins in cell lysates were analyzed by biotin‐switch assay. We found that the treatment of EC with BEC induced remarkable nitration of the mitochondrial protein cytochrome c oxidase (COX) both in normoxia and hypoxia. For non‐mitochondrial protein GAPDH treatment with BEC upregulated both S‐nitrosylation and nitration only in hypoxic conditions. Hence, we report the ability of Arg II to modify NO‐mediated protein modifications in EC.This work was supported by Florida DOH (KK), VA and NIH (JMP).
Endothelial ephrin‐B2 is essential for acute arterial vasodilation and ischemic recovery in adult mice
Authors: Wang Rong A. , Lin Yuankai , Jiang Weiya , Ng Jennifer , Jina Asmita 
Published: June 2021

Arterial vasodilation is critical for acute increases in tissue perfusion, yet molecular mechanisms of arterial‐specific dilation are unknown. Here we show that the cell surface protein ephrin‐B2, expressed in arteries but not veins, is a critical regulator of arterial dilation. Mice lacking the ephrin‐B2 gene specifically in endothelial cells were defective in acute arterial vasodilation in response to increased flow and acetylcholine. The protein, not RNA, levels of endothelial nitric oxide (NO) synthase (eNOS) along with its product NO, a vasodilator, were diminished in mutant arteries, but not veins. Ischemic recovery after femoral artery occlusion was also impaired in the mutants. S‐nitrosoglutathione, an NO donor, alleviated both the vasodilatory and ischemic protection defects in mutant mice. Thus, our data suggest that ephrin‐B2, potentially acting through eNOS, is a specific regulator of arterial dilation that is necessary to protect against ischemia.This work was supported by NIH R01 HL075033 to R.A.W., NIH T32 CA009043 to Y.L., and by a generous grant from the Foundation for Accelerated Vascular Research.
Urinary Renin Excretion is augmented in Chronic Angiotensin II‐infused Sprague‐Dawley Hypertensive Rats
Authors: Liu Liu , McCormack Michael , Seth Dale M , Kobori Hiroyuki , Navar L. Gabriel , Prieto Minolfa C 
Published: June 2021

Renin is upregulated in principal cells of the collecting ducts of angiotensin II (AngII)‐dependent hypertensive rats; however it remains unclear if increased distal nephron‐derived renin is secreted into the urine during AngII‐dependent hypertension. Urinary renin (uRen), angiotensinogen (uAGT) and AngII (uAngII) excretion was measured in chronic AngII‐infused male Sprague‐Dawley rats [80 ng/min, SC minipumps for 14 d, n=5] and control, sham‐operated rats [n=5]. Systolic blood pressures increased in the AngII rats by Day 5 and continued to increase throughout the study (Day 13; AngII: 175±10 vs. sham: 116±2 mm Hg). Although plasma renin activity was suppressed in the AngII‐infused rats (AngII: 0.3±0 vs. sham: 5.5±2 ng Ang I/ml/h); renin content in renal medullary tissues did not change (AngII: 10,564±1,476 vs. sham: 9,438±1,588 ng Ang I/h/mg; p=ns). Excretion of uAGT and uAngII increased in the AngII rats compared to sham rats, respectively [uAGT (AngII:1107±106 vs. sham:60±26 ng/day); uAngII (AngII: 3813±431 vs. sham: 2080±361 fmol/day). Importantly, uRen excretion rate at Day 13 was markedly increased in the AngII rats (AngII: 9±3 vs. sham: 1±1×10−5 Enzyme Units/day; p<0.05). Augmented uRen and uAGT may contribute to increased intrarenal AngII formation in AngII‐dependent hypertensive rats.NIH [IdeA (P20‐RR‐017659‐06), Tulane‐BIRCWH (K12HD043451)]; and AHA (09PRE2209079 and 09BGIA2280440).
Habitual aerobic exercise reverses age‐associated increases in transforming growth factor beta 1 in carotid arteries of mice
Authors: Marshall Kurt D , Seals Douglas R , Geolfos Candace , Lesniewski Lisa A , Fleenor Bradley S 
Published: June 2021

Aging increases carotid artery stiffness and adventitial collagen in mice. These events are reversed by voluntary wheel running (VR). Transforming growth factor β1 (TGF‐β1) modulates extracellular matrix proteins with aging, but the effect of habitual exercise on TGF‐β1 expression in large elastic arteries is unknown. Carotid artery stiffness was greater in old (O, 31 mo, n=4) vs. young (Y, 7 mo, n=4) adult cage‐control B6D2F1 mice (8.53 ± 2.00 vs. 3.015 ± 1.19 AU, p < 0.05), but not in O (n=4) given access to running wheels (mean distance = 1 km/d) for 12 weeks (3.61 ± 0.83 AU, p < 0.05). Immunohistochemical analysis of carotid arteries from Y (n=6) and O (n=8) cage control mice revealed increases in adventitial TGF‐β1 with aging (5.7E‐05 ± 5.9E‐06 vs. 3.6E‐05 ± 3.5E‐06 AU, P < 0.05), which were reversed by VR in O (n=4) (3.6E‐05 ± 4.2E‐06 AU). In a primary culture of adventitial fibroblasts obtained from Y adult (6 mo) Sprague‐Dawley rats, 24‐hr treatment with TGF‐β1 increased type I collagen (1.38 ± 0.07 vs. 1.0 ± 0.08, P < 0.05, western blot) and superoxide (O2−) production (4514 ± 317 vs. 2828 ± 185, P < 0.05, electron paramagnetic resonance). These preliminary data are consistent with the working hypothesis that habitual aerobic exercise may reverse large elastic artery stiffening with aging in part by reducing O2− production, which inhibits adventitial TGF‐β1‐induced adventitial collagen accumulation.AG013038 and HL007822
Role of 17[beta]‐estradiol and estrogen receptor [beta] in the genomic response to hypertrophy
Authors: Kararigas Georgios , Fliegner Daniela , Regitz‐Zagrosek Vera 
Published: June 2021

We and others have clearly indicated an important role for 17β‐estradiol (E2) and estrogen receptor β (ERβ) in attenuating pressure overload (PO)‐induced left ventricular hypertrophy (LVH). However, the underlying molecular mechanisms are poorly understood. Here, we investigated the role of E2/ERβ signaling in PO‐induced LVH at a genome‐wide level.Female wild‐type (WT) and ERβ knockout (ERβ−/−) cycling C57Bl6 mice were subjected to transverse aortic constriction (TAC) or sham surgery. At nine weeks post operation, left ventricles were harvested and RNA was hybridized to the Mouse Genome 430 2.0 Array. The microarray data were analyzed with R and Bioconductor.Assessment of single gene differential expression with a false discovery rate‐adjusted P < 0.05 identified 197 probe sets regulated in WT, 941 in ERβ−/− and 366 in both WT and ERβ−/− mice. Gene set enrichment analysis based on 1000 permutations revealed 26 pathways modulated in WT, while almost the double were modulated in ERβ−/− mice. Of these, 16 were modulated in both WT and ERβ−/− mice. Among others, the cytochrome P450 pathway was repressed in WT/TAC mice, the apoptosis pathway was induced in ERβ−/−/TAC mice, while the ECM‐receptor interaction pathway was induced in both WT and ERβ−/−/TAC mice.In conclusion, we provide evidence that the E2/ERβ axis plays an important role in the genomic response to LVH modifying the expression of several genes/pathways.