bluuebunny/crossref_metadata_2025_split huggingface dataset:
The melanocortin receptor system in mediating anti‐inflammatory protection following cerebral ischemia reperfusion injury
Authors: holloway paul M , getting stephen j , gavins felicity 
Published: June 2021

The melanocortin receptor system (MRS) provides an attractive target for the treatment of stroke, due to their ability to initiate anti‐inflammatory and neuroprotective pathways. However, the relative contribution of the receptors that mediate these effects (MC1 MC3 and MC4) have yet to be determined. This current work aims to assess the therapeutic potential of targeting the MRS for treatment of stroke.The bilateral common carotid artery occlusion (BCCAo) mouse model of global stroke was utilised in conjunction with intravital microscopy, to quantify the neuro‐inflammatory response through real time visualisation of leukocyte recruitment in the cerebral microcirculation.BCCAo induced significant increases in leukocyte rolling and adhesion in the cerebral microcirculation. Treatment with the pan receptor agonist α‐MSH significantly reduced the ischemia‐reperfusion induced leukocyte recruitment. Antagonism of MC3 and MC4 using SHU9119 failed to abrogate the anti‐inflammatory effects of α‐MSH early in reperfusion (40 min), but was able to blunt the effects of α‐MSH by 2h.These results suggest that in the early stages of reperfusion, MC1 activation may be of particular importance in mediating protection within the cerebral microcirculation. However as the inflammatory reaction progresses the role of MC3 may become more prominent. This work was funded by the British Heart Foundation
Role of p47phox and Nox2 in the Promotion and Impairment of Collateral Growth
Authors: Distasi Matthew , Ingram David , Miller Steven , Unthank Joseph 
Published: June 2021

Anti‐oxidant therapy implicates a role for reactive oxygen species (ROS) in both successful collateral growth (CG) and its impairment in models of chronically elevated oxidative stress, including obese mice. NAD(P)H oxidase (Nox) is a major source of vascular ROS. This study investigates the roles of the Nox subunits p47phox and Nox2 in CG (assessed as % increase in collateral diameter compared to same animal control artery) in lean and obese mice. C57BL/6, p47phox−/−, and Nox2−/− mice were fed either standard chow (Lean) or a high‐fat (60% kcal from lard) diet to induce obesity. In lean mice after femoral artery ligation, significant CG occurred in wild‐type (WT) and Nox2−/− mice (171±43 and 112±21%, respectively, p<0.05) but not in p47phox−/− mice (52±31%). In obese mice, CG was greatly suppressed in WT mice (25±13%); however, whole body ablation of both Nox2 and p47phox attenuated the impairment (144±68 and 107±7% increase in collateral diameter, respectively, P<0.05). RT‐PCR analysis demonstrated an increase in p47phox (1.9X, P<0.02) and Nox2 (1.6X, P<0.07) subunit expression in obese vs. lean arteries. Transplantation of WT bone marrow into lean p47phox−/− did not prevent the suppression of CG. In conclusion, these data suggest that tissue resident p47phox is required for CG in lean mice and that obesity‐induced increases in p47phox and Nox2 expression mediate the impairment of CG in obese mice.Grant Funding Source: NIH T32 HL007910‐26
Fetal Endothelial Colony Forming Cells from pregnancies complicated by intrauterine growth restriction have reduced vasculogenic capacity
Authors: Bourque Stephane Louis , Sipos Peter , Sibley Colin , Baker Philip , Davidge Sandra , Crocker Ian 
Published: June 2021

BackgroundDiminished numbers of ECFCs may be a risk factor for cardiovascular disease (CVD). Intrauterine growth restriction (IUGR) is a pregnancy complication associated with neonatal morbidity and mortality and long‐term sequelae, including CVD. We hypothesized that fetal ECFCs from human IUGR pregnancies would be fewer in number and exhibit altered vasculogenic potential.MethodsCord blood from uncomplicated and IUGR pregnancies were collected at delivery. ECFCs were counted by flow cytometry using discriminators: 7AAD‐/CD31bright/CD45‐/KDR+/CD34+. Vasculogenic potential was investigated by implanting collagen matrices populated with ECFCs into adult immuno‐deficient mice.ResultsECFC numbers were 67% lower in IUGR cord blood vs. controls (P<0.001), and exhibited a 71% longer doubling time (P=0.01). Blood vessel formation was reduced in collagen implants populated with IUGR‐derived ECFCs. In vitro, hypoxia (1% O2, 72h) induced‐ECFC senescence, and IUGR cells were more susceptible (P<0.05). Finally, IUGR‐derived ECFCs showed a 61% reduction in MMP‐2 release under hypoxic conditions (P<0.001).ConclusionECFCs from IUGR cord blood have intrinsic functional alterations, resulting in diminished vasculogenic potential and increased susceptibility to hypoxia. Supported by grants from the Canadian Institutes of Health Research and the Wellcome Trust.
Hydrogen sulfide mitigates diabetic nephropathy through NMDA receptor mediated renal remodeling
Authors: Kundu Sourav , Tyagi Aaron , Coley Denise , Sathnur Pushpakumar , Sen Utpal 
Published: June 2021

Diabetic nephropathy is a leading cause of vascular morbidity and mortality. Recently, the involvement of N‐methyl‐D‐aspartate receptor (NMDAR) and hydrogen sulfide (H2S) in diabetes associated complications has been implicated. Our aim in this study was to determine whether H2S mitigates diabetic nephropathy by modulating gap junction and matrix proteins by antagonizing NMDAR. We used a diabetic model (Akita, C57BL/6J‐Ins1Akita), matrix metalloproteinase‐9 knockout (KO) [(MMP‐9−/−)] and double KO of Akita/MMP‐9−/− mice and in vitro cell culture.ResultsDiabetic kidneys showed decreased levels of H2S and its enzymes CBS and CSE in both mRNA and protein level and increased expression of NMDA‐R1, connexin‐40, ‐43 (Cx‐40,‐43) and MMP‐9. Treatment with H2S reversed these effects. In double KO mice, NMDA‐R1 was high but Cx‐40, ‐43 was normal. Additionally, treatment with NMDA‐R1 blocker dizicilpine (MK801) failed to disrupt connexin. Glomerular endothelial cells treated with high glucose indicated dysregulated Cx‐40, ‐43. Inhibition of NMDA‐R1 by MK801, silencing MMP‐9 by siRNA or H2S treatment preserved gap junction proteins. We conclude that in diabetic nephropathy, remodeling is mediated by NMDA‐R1 associated MMP‐9 activation and H2S plays a significant role in mitigating nephropathy.
HSP27 regulation in acute unilateral obstructed kidney, along with RMIC and collecting duct cells subjected to mechanical, oxidative, and inflammatory stress
Authors: Carlsen Inge Gram , Nilsson Line , Frøkiær Jørgen , Nørregaard Rikke 
Published: June 2021

In vivo, renal medullary interstitial cells (RMIC) and collecting duct (CD) cells are subjected to mechanical, oxidative and inflammatory stress as a result of ureteral obstruction. Heat shock protein 27 (HSP27) is expressed in RMIC and CD cells in vivo and in vitro, and plays a role in stabilization of actin filaments. Both HSP27 and the phosphorylated‐form (pHSP27) have anti‐apoptotic functions.In vivo studies using rats subjected to acute unilateral ureteral obstruction (UUO) for 6h and 12h showed unchanged HSP27 protein level in kidney inner medulla (IM). pHSP27 protein level was increased in obstructed kidney IM compared to both sham and contralateral kidney. Further examination of HSP27 protein abundance in response to mechanical, oxidative, and inflammatory stress in vitro, we subjected RMIC and CD cell to pressure, H2O2 and interleukin 1â (IL‐1â) stimulation for 2–24h. Applying oxidative stress to RMIC and CD cells increased HSP27. Inflammatory stress with IL‐1â increased HSP27 and pHSP27. Pressure of 60 mmHg was explored directly in RMIC which increased HSP27.These data indicates that in response to acute UUO different forms of cellular stress increase expression of HSP27 and pHSP27, which may play a role in the cytoprotection of RMIC and CD cells.
NADPH oxidase 4 induces cardiac arrhythmia in zebrafish through ROS
Authors: Zhang Yixuan , Shimizu Hirohito , Siu Kin L , Chen Jau-Nian , Cai Hua Linda 
Published: June 2021

Indirect evidences implicate that oxidative stress might play an important role in cardiac arrhythmogenesis. Recently, we identified an intriguing link between NADPH oxidase 4 (NOX4) and atrial fibrillation in humans. The present study utilizes cardiac specific GFP transgenic zebrafish to examine effects of NOX4 injected RNA on cardiac rhythm. Full‐length human NOX4 was cloned, transcribed, and RNA injected to one‐cell stage fish embryos. Remarkably, 51.0% of the fish developed arrhythmia (irregular heart beat) after 24–30 hrs. To quantify the intervals among heart rates of each embryo, we recorded consecutive digital images (30 frames/second) of each embryo and analyzed by custom‐made software. The variation of intervals in NOX4 embryos was significantly bigger than that of control embryos (0.038±0.0020 vs. 0.027±0.0026 second/beat, n=11–17, all data as Mean±SEM, p=0.0028). The NOX4 embryos also had increased superoxide production determined by electron spin resonance (5.3±0.7 vs. 10.9±1.5 μM/min/mg, n=9–12, p<0.01). Co‐injection of an ATG morpholino led to reduced arrhythmogenesis (to 20.2%) and blunted superoxide production. We also generated catalytically inactive NOX4 RNA (proline 437 substituted with histidine in the NADPH binding domain), which failed to induce arrhythmia and overproduction of superoxide (n=5, p=0.67). In conclusion, our data have identified a novel phenotype of cardiac arrhythmia in NOX4 overexpressed zebrafish, providing new insights into mechanistic studies of arrhythmogenesis.This study has been supported by National Heart, Lung and Blood Institute (NHLBI), and American Heart Association.
Searching for the Na1 site of hSGLT1
Authors: Loo Donald D.F. , Hirayama Bruce A. , Jiang Xuan , Wright Ernest M. 
Published: June 2021

The crystal structure of vSGLT has provided the location of a Na+ binding site that is conserved in the LeuT structural family, the Na2 site. In human SGLT1 there is strong functional evidence for two Na+ binding sites, one of which (Na2) is conserved in homology models. The location of the Na1 site is unknown in hSGLT1 but in the LeuT family it is colocalized with the substrate binding site. To search for the Na1 site in hSGLT1, we measured the intrinsic Na+ affinity of sugar binding site mutants and the gating and ligand coordinating residues (H83C, Y290C, W291C and Q457C) from the [Na+] dependence of the SGLT1 capacitive currents (charge movement) in the absence of glucose. For wild‐type and Q457C hSGLT1 the Na+ sensitivity of the charge movement was ≈−100 mV per 10‐fold decrease in external [Na+]. This was dramatically reduced for H83C, Y290C and W291C hSGLT1, ≈−50 mV per 10‐fold decrease in external [Na+]. In addition, the Na+/glucose coupling coefficient for hSGLT1 decreased from 2 to 1 for mutants Y290F and W291F. The loss of Na+ affinity for mutants H83C, Y290C and W291C, and the reduction in coupling indicates that these resides may be involved in Na+ binding. We hypothesize that the Na1 site in hSGLT1 is close to the substrate binding site as in LeuT and BetP.Supported by NIH grant DK19567.
Discovery of an inward rectifying potassium channel inhibitor with preference for Kir2.3, Kir3.X and Kir7.1
Authors: Raphemot Rene , Lonergan Daniel , Nguyen Thuy T. , Utley Thomas J. , Gogliotti Rocco , Hopkins Corey R. , Lewis Michelle , Lindsley Craig W. , Weaver C. David , Denton Jerod S. 
Published: June 2021

Inward rectifying potassium (Kir) channels play key physiological roles in diverse cell types and may represent novel drug targets for disease. In cardiac myocytes, Kir channels contribute to the resting membrane potential and the late repolarization phase of the action potential. Homomeric Kir2.3 and heteromeric Kir3.1/3.4 channels are expressed almost exclusively in the atria and are postulated drug targets for atrial fibrillation, a common arrhythmia that predisposes patients to stroke. Efforts to assess the utility of Kir2.3 and Kir3.1/3.4 channels as therapeutic targets have been hampered by the lack of drug‐like compounds targeting these channels. We therefore performed a fluorescence‐based high‐throughput screen for small‐molecule inhibitors of Kir3.X channel function. Several novel inhibitors were discovered. One compound, termed VU573, inhibits Kir3.X channels with a half‐maximal inhibition concentration of 1.9 μM. Subsequent electrophysiology‐based counterscreens revealed that VU573 inhibits Kir2.3 and Kir7.1 with similar potency, but is less active toward Kir1.1, Kir2.1, and Kir4.1. VU573 is the first selective inhibitor for Kir2.3 and most potent Kir7.1 inhibitor known to date. We are currently using medicinal chemistry to improve the potency and selectivity of VU573 for Kir2.3 and Kir3.X and understand the structural requirements for inhibition of these channels.NIH R01 award to Jerod S. Denton DK082884
Regulation of ion transport and ASL height by the anti‐inflammatory mediator, lipoxin A4 in normal and cystic fibrosis bronchial epithelium
Authors: Urbach Valerie , Higgins Gerard , Al-Alawai Mazen , Costello Richard W , McNally Paul , Verriere Valia , Harvey Brian J 
Published: June 2021

Mutations of CFTR results in defective Cl− secretion and Na+ hyperabsorption which contributes to a reduction of the airway surface liquid layer height (ASLh) thus promoting bacterial colonization and inflammation. Lipoxin A4 (LXA4) which plays a central role in resolving inflammation is decreased in cystic fibrosis (CF). We have investigated the role of LXA4 in modulating ion transport and ASLh in CF and non‐CF bronchial epithelial cell lines (CuFi‐1 and Nuli‐1) and primary cultures grown under an air‐liquid interface. ASLh were measured using confocal microscopy and ion transport using electrophysiological techniques.LXA4 (1nM) significantly increased ASLh in CF and non‐CF epithelia. This effect was inhibited by bumetanide, amiloride, Boc‐2 (FPR2 antagonist), extracellular hexokinase, reactive blue 2 and NF340 (P2Y and P2Y11 receptor antagonist). LXA4 stimulated a carbenoxolone (Panexin‐1 inhibitor)‐sensitive apical ATP release, intracellular Ca2+ mobilization and Cl− secretion and inhibited Na+ absorption. These effects of LXA4 involving apical ATP release, inhibition of Na+ absorption and stimulation of Cl− secretion to enhance ASL dynamics open up a new therapeutic avenue to promote mucociliary clearance in CF airways.
Perivagal capsaicin alters the response of vagal motoneurons to Thyrotropin Releasing Hormone (TRH)
Authors: Holmes Gregory M. , Swartz Emily M , Travagli R. Alberto 
Published: June 2021

A supposedly selective degeneration of vagal afferent C‐fibers by perivagal capsaicin (CAP) has often been used to identify the mechanism of action of GI‐ peptides. The actions of many of these peptides have been attributed to activation of vagal afferent fibers based on the dramatic reduction of their effects following perivagal CAP pretreatment. We have reported recently that, similar to surgical vagotomy, perivagal CAP induces dendritic degeneration, reduction of ChAT‐immunoreactivity and, possibly, death of a subpopulation of vagal preganglionic (DMV) neurons.We tested the hypothesis that perivagal CAP treatment induces functional impairment of rat DMV neurons.Monolateral perivagal CAP treatment (0.1 and 1%) was performed 7–10 days prior to monitoring gastric motility response induced by DMV microinjection of TRH (100pmoles), which increases motility via actions exclusively at DMV.Microinjection of TRH in DMV reduced gastric motility by 42±16 and 76±5% in rats pretreated with 0.1 or 1% CAP vs control (N=5 for all). Conversely, gastric motility in response to microinjection of TRH in the contralateral DMV was not altered (N=3–4).Our data show that perivagal CAP treatment induces functional impairment of DMV neurons and indicates that perivagal CAP treatment should not be used as an exclusive means to investigate the site of action of gastrointestinal peptides.Funds: NIDDK 55530
The opioid‐induced suppression of augmented (‘sigh’) breaths is dose‐dependent and prevented by co‐administration of naloxone
Authors: Bell Harold James , Pankuch Glenn 
Published: June 2021

We tested whether the morphine (MOR) induced suppression of augmented breaths (ABs) can be prevented by the non‐specific, competitive opioid receptor antagonist naloxone (NX). Further we determined whether the suppression of ABs follows a dose‐response pattern in the low‐end recommended dose range. We studied breathing in 8 rats on 4 separate days, giving four dosages of MOR s.c. in randomized order: 0 mg/kg (saline control), 1, 3, and 5 mg/kg. In a second set of 8 animals, we co‐administered either {MOR (5mg/kg) + NX (5 mg/kg)}, or {MOR (5mg/kg) + saline control (1 ml/kg)} on two separate days and monitored animals for 2 hours. MOR suppressed ABs in a typical doseresponse fashion, from 6.5 ± 1.6 ABs/15mins (control) to 1.5 ± 1.5, and 0.6 ± 1.8 ABs/15mins in the 3 and 5 mg/kg dose conditions respectively (both p<0.001). 5 mg/kg MOR caused ABs to be completely eliminated in all but one of the 8 animals we studied. In the second experiment, MOR caused a rapid and potent suppression of ABs across a two‐hour monitoring interval that was prevented by NX co‐administration. There were significantly more ABs when NX was co‐administered with MOR, regardless of the time interval across the 2 hours at which they were assessed post‐injection (p<0.001). In summary, MOR suppresses ABs in a dose‐response manner in the low‐end of the recommended dosage range, and this effect occurs via typical opioid‐receptor mediated pathways.This study received financial support from the Department of Medicine, Penn State College of Medicine, in the form of laboratory start‐up funds.
Ventilatory long‐term facilitation is altered in tryptophan hydroxylase 2 knock out mice
Authors: Hickner Stephen James , Angoa-Perez Mariana , Kuhn Donald M , Mateika Jason H 
Published: June 2021

PurposeThis study was designed to examine the impact of serotonin (5HT) in the central nervous system (CNS) on ventilatory long‐term facilitation (vLTF) in spontaneously breathing mice.MethodsWe exposed six mice with a null mutation for tryptophan hydroxylase 2 (KO) and six wild type mice (WT) to 12 episodes of hypoxia (10 % oxygen) and measured ventilation via whole body plethysmography.Results

KO
WT

Baseline VT
0.21 ± 0.03*
0.17 ± 0.01

Baseline Bf
150.5 ± 7.1*
169.5 ± 6.0

Recovery VT
0.22 ± 0.02*
0.16 ± 0.01

Recovery Bf
152.9± 11.2*
186.2 ± 4.8§

VT – tidal volume (ml); Bf – breathing frequency (breaths/min);
– significantly different from WT mice (p < 0.05);
– significantly different from baseline (p < 0.05).

KO mice displayed an elevated VT and a decreased Bf under baseline conditions. LTF was evident in the WT (see Bf measures) but not the KO mice following exposure to hypoxia.ConclusionsLack of 5HT in the CNS is accompanied by the absence of LTF following acute exposure to intermittent hypoxia.
Brain 11β‐hydroxysteroid dehydrogenase activity: which enzyme?
Authors: Formenti Silmara , Gomez-Sanchez Celso E , Colombari Eduardo , Gomez-Sanchez Elise P 
Published: June 2021

We studied 11β‐HSD activity in rat kidney (positive control) and brain homogenates and subcellular fractions. Samples were incubated with 0.5 mM NAD+ or NADP+ and [1,2 3H] corticosterone for 1h at 37° C. Steroids were extracted, separated by TLC, and the radioactivity represented by corticosterone and 11‐dehydrocorticosterone read by a bioscanner. Conversion of corticosterone to dehdrocorticosterone by homogenates was significantly increased by NADP+, but not NAD+: Cerebel: Control‐24.1±0.4%, NADP+‐60.9±0.8%, NAD+‐21.9±0.7%; Hypothal: Control‐8.2±0.1%, NADP+‐23.6±0.8%, NAD+‐ 7.3±0.0%; Hippoc: Control‐24.2±0.9%, NADP+‐62.1±0.6%, NAD+‐21.9±0.2%, p<0.001 in all). Conversion was significantly greater isolated in microsomes compared to nuclei and mitochondria, and for NADP+ vs NAD+: Cerebel microsomes: NAD+‐5.4±1.7%, NADP+‐34.3±4.6%, p<0.001; Hypothal microsomes: NAD+‐0%; NADP+‐9.2±1.8%, p<0.001; and Hippoc microsomes: NAD+ 0.6±0.6% vs NADP+ 23.1±1.4%, p<0.001). Thiram inhibited NAD+‐dependent HSD activity in 11‐β HSD2‐ transfected CHO cells; it did not in the brain. These data suggest a novel NADP+‐dependent 11β‐HSD that may allow aldosterone to activate some brain mineralocorticoid receptors.Support: VA Medical Research Funds, NIH HL27255 & HL75321, CAPES & FAPESP
A novel signaling pathway for regulation of ATP‐binding cassette transporter A1 expression
Authors: Yang Hong , Chen Xinping , Zhang Hongfeng , Lin Xinghua , Okoro Emmanuel U , Zhou LiChun , Guo Zhongmao 
Published: June 2021

The objective of this study is to characterize the signaling pathway for regulation of ATP‐binding cassette transporter A1 (ABCA1) expression. Quantitative real‐time RT‐PCR assay and Western blot analysis revealed that treatment of macrophages with reelin, apolipoprotein E (ApoE) or ApoE‐containing lipoproteins increased ABCA1 mRNA and protein expression. In addition, these reagents significantly increased the level of phosphorylated disabled‐1 (Dab1), phosphatidylinositol 3‐kinase (PI3K), protein kinase C‐æ (PKC‐æ), and specificity protein 1 (Sp1), increased the amount of Sp1 bound to the ABCA1 promoter, and induced PKC‐æ binding with Sp1. Mutation of the Sp1 binding site in the ABCA1 promoter suppressed the ABCA1 promoter activity induced by reelin, ApoE or ApoE‐containing lipoproteins. The upregulation effect of these reagents on ABCA1 expression was suppressed by: 1) knockdown of Dab1, very low‐density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2) with siRNAs, 2) inhibition of PI3K and PKC with kinase inhibitors, 3) overexpression of kinase‐dead PKC‐æ, and 4) inhibition of Sp1 DNA‐binding with mithramycin A. These results, together with the known function of these signaling proteins, suggest that ABCA1 expression can be upregulated by activation of the VLDLR/ApoER2‐Dab1‐PI3K‐PKCæ‐Sp1 signaling cascade.Grant support: SC1HL101431 (HY) and R01HL089382 (ZMG).
PhenoMiner: an interactive tool for physiologists integrating phenotype data using multiple ontologies
Authors: Dwinell Melinda , Shimoyama Mary , Nigam Rajni , Liu Weisong , Tutaj Marek , De Pons Jeff , Wang Shur-Jen , Smith Jennifer , Lowry Timothy , Hayman G. Thomas , Laulederkind Stanley , Petri Victoria , Jayaraman Pushkala , Worthey Elizabeth , Munzenmaier Diane , Jacob Howard 
Published: June 2021

AimThe Rat Genome Database (RGD, http://rgd.mcw.edu) has been developing tools and resources to aid researchers in integration of genomic and phenomic data derived from rat research for the past 10 years. Following the development of the Phenotypes and Models Portal, a tool was created to integrate phenotype data sets from multiple sources. The new PhenoMiner tool delivers an interactive tool to query phenotype data by strain, phenotype, measurement method and experimental conditions.MethodsRGD was established primarily as a genomic resource but has gathered and curated physiological data for many years. The Phenotypes and Models Portal provides both new types of phenotype data, as well as different ways to access existing data in RGD. PhenoMiner was developed to integrate data collected from multiple laboratories, using a wide variety of techniques, and across many inbred, transgenic and knockout strains of rats. Data are integrated through the use of four ontologies (rat strain, clinical measurement, measurement method, experimental conditions).ResultsThe more than 12,000 records can be viewed graphically or in tabular format, allowing for easy comparison of phenotypes across strains.ConclusionPhenoMiner promises to be an effective and user‐friendly method to access, retrieve, and compare physiological data and identify ideal disease models for further studies.