bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of whey protein or carbohydrate supplementation on markers of muscle damage, muscular dysfunction and leg soreness
Authors: Witard Oliver Charles , Jackman Sarah R , Jeukendrup Asker E , Tipton Kevin D 
Published: June 2021

This study aimed to determine the impact of whey protein (WP) or carbohydrate (CHO) supplementation on recovery from exercise‐induced muscle damage. In a parallel, placebo‐controlled, single‐blind experimental trial, untrained males were randomly assigned to a WP (n=12), CHO (n=12) or placebo (n=12) supplementary group. Muscle damage was elicited by implementing an eccentric unilateral leg extensor protocol (15×10 repetitions at 170% peak isokinetic torque) using an isokinetic dynamometer. Diet was strictly controlled throughout the trial. Measures of muscle function, muscle damage, leg soreness and inflammation were taken pre‐ and post‐ (1, 8, 24, 48 and 72h) exercise. Maximal isometric strength (MIS) decreased by ~32% at 48h post‐ (662.3±32.2N) compared to pre‐exercise (451.2±38.9N) (P<0.01). Ratings of leg soreness peaked 24h following exercise. No effect of supplement was observed on post‐exercise plasma creatine kinase (CK) or myoglobin responses, MIS, visual analogue scale ratings or leg volume (P>0.05). Measures of muscle function and CK showed no correlation (r=−0.211) (P=0.402). These data suggest the chosen experimental model successfully increased muscular dysfunction, leg soreness and putative markers of muscle damage. However, supplementation with 40g/d WP or CHO failed to ameliorate these symptoms of muscle damage under conditions of dietary control.Protarmor 907 LSI whey protein was supplied by Armor Proteines, Saint‐Brice‐en‐Cogles‐FRANCE
Endothelial cilia are mechanosensory organelles
Authors: Nauli Surya M , Kawanabe Yoshifumi , Kaminski John J , Pearce William J , Ingber Donald E , Zhou Jing 
Published: June 2021

BACKGROUNDWhen challenged with extracellular fluid‐shear stress, vascular endothelial cells are known to release nitric oxide (NO), an important vasodilator. We here show that this mechanosensing signal transduction depends on apical membrane organelles, termed cilia.METHODS AND RESULTSCells with Pkd1null/null or Tg737orpk/orpk mutation encoded for polycystin‐1 or polaris, respectively, are unable to transmit extracellular shear stress into intracellular calcium signaling and biochemical NO synthesis. Cytosolic calcium and NO recordings further show that fluid‐shear sensing is a cilia‐specific mechanism, as other mechanical or pharmacological stimulation does not abolish calcium and NO signaling in polycystin‐1 and polaris mutant endothelial cells. Polycystin‐1 localized in the basal body of Tg737orpk/orpk endothelial cells is insufficient for a fluid‐shear stress response.CONCLUSIONWe demonstrate for the first time that the ability of endothelial cells to sense a low range of fluid shear depends on cilia and that cilia are compartments required for proper localization and function of the mechanosensitive polycystin‐1 molecule.
Omeprazole abrogates cell protection in an in vitro model of infectious gastritis
Authors: Blass Amy L. , Kohler Jonathan E. , Liu Jingjing , Mathew Jeff , Tai Kaniza , Kelly Edward , Soybe David I. 
Published: June 2021

Aim:Monochloramine (NH2Cl) is a thiol oxidant produced in H. pylori gastritis. Omeprazole (OME) is a proton pump inhibitor, and a thiol oxidant, used to treat gastritis. Our aim was to determine if OME influences cell injury by NH2Cl, and to test if effects of these thiol oxidants are mediated by changes in [Ca2+]i and [Zn2+]i.Methods:Isolated rabbit gastric glands were incubated with doses of NH2Cl for 3 hours, and cell viability was measured with Calcein‐AM. Chelators (BAPTA or TPEN) and a thiol reducing agent (DTT) were added, with or without OME. Results were normalized to Ringers controls.Results:NH2Cl caused cell death at all concentrations. DTT protected cells from NH2Cl‐induced damage. The promiscuous chelator BAPTA gave no protection. However, the zinc‐specific chelator TPEN protected completely. When cells were co‐incubated with OME, cell death occurred even in the absence of NH2Cl. Addition of TPEN to these cells was not protective despite high [Zn2+]i, but addition of DTT again prevented cell death. Conclusion: Chelation of intracellular Zn2+, but not Ca2+, protects cells exposed to NH2Cl. However, OME, commonly used to treat H. pylori gastritis, appears to cause zinc‐independent cell death. This can be reversed by the thiol‐reducing agent DTT, suggesting that OME acts downstream of Zn2+‐mediated injury. Funding: NIH R01DK069929‐03
Novel small molecules reverse myosin light chain kinase (MLCK) isoform 1 translocation and barrier dysfunction induced by tumor necrosis factor (TNF)
Authors: Graham W. Vallen , Marchiando Amanda M. , Wang Yingmin , Ostrov David A. , Turner Jerrold R. 
Published: June 2021

MLCK is central to TNF‐induced intestinal epithelial barrier dysfunction in vitro and in vivo. MLCK is expressed as two splice‐variants, MLCK1 and MLCK2, distinguished by a short insertion in MLCK1. MLCK1 knockdown increases epithelial barrier function in vitro. We therefore hypothesized that MLCK1 might specifically regulate the perijunctional actomyosin ring (PAMR) during TNF‐induced barrier dysfunction.TNF‐treated Caco‐2 monolayers demonstrated a progressive increase in PAMR‐associated MLCK1 within 30 min, 35.7±0.1% (P<0.05), that peaked at 4 hr, 118.8±0.1% (P<0.001), and correlated directly with TNF‐induced barrier loss (r=0.9). Structural modeling showed that the MLCK1 insertion created a binding pocket. In silico screening identified candidate small molecules targeting this pocket, of which, one molecule, SMKI2, bound the pocket with a predicted ΔG of −92 kcal/mole. 59.9±0.2% (P<0.001) of MLCK1 was displaced from the PAMR by 250 μM SMKI2. In contrast, enzymatic MLCK inhibition caused no MLCK1 displacement. However, like enzymatic inhibition, SMKI2 corrected TNF‐induced barrier loss in a dose‐dependent fashion.These data provide insight into the mechanism of TNF‐induced MLCK activation and PAMR myosin phosphorylation. Inhibition of MLCK1 targeting may be an important means of correcting barrier dysfunction without the potential toxicities of enzymatic MLCK inhibition.Supported by NIH
Acute Treatment of Isolated Colon with The Hydrogen Peroxide Scavenger, Catalase, Alleviates Ion Transport Dysfunction in Murine DSS Colitis
Authors: McCole Declan F , Barrett Kim E 
Published: June 2021

Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) contribute to epithelial dysfunction in inflammatory bowel disease (IBD). This study investigated the role of H2O2 in colonic ion transport defects in the murine DSS colitis model.Colitis was induced by administering DSS (4%) in drinking water for 5 days followed by 3 days on normal H2O. Distal colon from control and DSS mice was stripped of underlying smooth muscle and mounted in Ussing chambers. Tissues were bilaterally treated with the H2O2 scavenger, catalase (50, 100 or 300 U/ml), for 30 min prior to measuring short‐circuit current (Isc) responses to the cAMP agonist, forskolin (20 μM), or the Ca2+‐dependent muscarinic agonist, carbachol (CCh, 300 μM). Catalase (100 U/ml) treatment of tissues from DSS mice increased baseline Isc above that of untreated tissues (p<0.01; n=7) without affecting baseline Isc in control mouse colon. Catalase significantly restored the blunted Isc responses to forskolin (p<0.05; n=7) and CCh (p<0.05; n=7) in DSS mouse colon but had no effect in control mouse colon (n=4). Incubation of inflamed colon with catalase also abolished H2O2 release into bathing media, as measured by a colorimetric H2O2 assay (n=3).In summary, these data indicate that ongoing production of H2O2 in colitis contributes to the reduced capacity for active ion transport in inflamed colon.Supported by the Crohn's and Colitis Foundation of America and NIH.
Regulation of platelet‐activating factor (PAF) receptor by p85α subunit of phosphatidylinositol 3‐kinase (PI3K) in enterocytes
Authors: Lu Jing , Caplan Michael S , Jilling Tamas 
Published: June 2021

PAF and intestinal epithelial apoptosis play important roles in the development of necrotizing enterocolitis. We revealed that activation of the PAF receptor (PAFR), a member of the G‐protein coupled receptor (GPCR) family, induces apoptosis through a PI3K/Akt‐dependent pathway in enterocytes. Since direct binding of p85a subunit of PI3K to certain GPCRs was shown to disrupt GPCR recycling and signaling, we sought to study the interaction between PAFR and the p85a subunit of PI3K in intestinal epithelial apoptosis. We revealed that intestinal epithelial cells (IEC‐6) over‐expressing p85a subunit had significantly lower basal levels of caspase 3 activity compared to controls and failed to respond to PAF. Over‐expression of p85a in IEC‐6 cells resulted in sustained activation of Akt and prevented PAF‐induced Akt dephosphorylation. In live cells, over‐expressed p85a in IEC/p85a cells induced the formation of large perinuclear vesicles that were decorated with p85a. When it was transfected into p85a‐expressing cells, PAFR was recruited to these vesicles and there was a co‐localization between p85a and PAFR. This formation of a complex was confirmed by immunoprecipitation. In summary, p85a and PAFR interaction could be a mechanism to regulate PAFR endocytosis and PAFR‐activation‐induced apoptosis.This may provide insights to design therapeutic interventions for NEC. Supported by NIH HD37581 and DK062960.
Rho Kinase inhibitors impair the response of chicken ductus arteriosus to oxygen and other vasoconstrictors
Authors: Villamor Eduardo , van der Sterren Saskia , Agren Pia , Zoer Bea , Blanco Carlos E , Cogolludo Angel L , Perez‐Vizcaino Francisco 
Published: June 2021

Accumulated evidence demonstrates that RhoA/Rho kinase‐mediated increases in calcium sensitivity of the contractile apparatus contributes to ductus arteriosus (DA) contraction. We hypothesized that the contribution of Rho kinase to DA tone is developmentally regulated. In the 19‐day DA (total incubation 21 days), the contractions induced by oxygen, KCl, the inhibitor of KV channels 4‐aminopyridine (4‐AP), the α1‐adrenoceptor agonist phenylephrine, and endothelin‐1 were markedly impaired by the presence of the Rho kinase inhibitors Y‐27632 (10μM) and hydroxyfasudil (10μM). The 15‐d DA did not respond to oxygen or phenylephrine and the response to KCl and 4‐AP was also impaired by Y‐27632 and hydroxyfasudil. During contractions induced by KCl (125 mM), Y‐27632 and hydroxyfasudil (1nM‐10 μM) produced a concentration‐dependent relaxation with a respective ‐log EC50 of 5.7±0.2 and 5.58±0.2 in the 19‐d DA; and 5.0±0.2 and 4.98±0.2 in the 15‐d DA (P<0.05 vs. 19‐d). In conclusion, our data indicate that activation of Rho kinase is a key signalling pathway in oxygen‐, as well as membrane depolarization‐ and receptor stimulation‐induced contraction of chicken embryo DA and that a developmental increase in this pathway occurs during prenatal life.Supported by a grant from “Fundación de Investigación Médica Mutua Madrileña”
Pentoxifylline attenuates fMLP‐induced oxidative burst in human neutrophils through PKA dependent and independent mechanisms: Effect on Ras, Raf, and MAPK activation
Authors: Deree Jessica , Costantini Todd , Woon Tiffany , Loomis William H , Coimbra Raul 
Published: June 2021

Pentoxifylline (PTX) has been proven to be an inhibitor of fMLP‐induced neutrophil (PMN) oxidative burst and is thought to function by increasing cAMP and Protein kinase A (PKA). We hypothesized that PTX diminishes production of the respiratory burst by both PKA dependent and independent mechanisms.Human PMNs (1 x 106 cells/sample) were isolated and stimulated with fMLP (1μmol/L) and PTX. PMN activation was determined by the cytochrome C reduction method in the presence and absence of p38 MAPK (SB203580 10 mM), ERK 1/2 (PD98059 50 mM), and PKA inhibitors (H89 10 mM). Western blot analysis of Ras, c‐Raf, p38 MAPK, and ERK 1/2 was performed in PMNs exposed to fMLP and PTX.PTX produced a decrease in the reduction of cytochrome C that was diminished but not abrogated by H89 exposure. The reduction in Ras activation seen with PTX was not effected by the presence of H89. The attenuation in phosphorylation of c‐Raf p38 MAPK, and ERK 1/2 observed with PTX was only partially reversed with H89.PTX attenuates fMLP induction of oxidative burst through effects on Ras, c‐Raf, p38 MAPK, and ERK 1/2. The attenuation of early events by PTX in the cascade leading to production of the oxidative burst is PKA independent, while the effects of PTX on signaling distal to c‐Raf phosphorylation rely on both PKA‐dependent and independent mechanisms.Source of Support: Department of Surgery, University of California San Diego
Retinoid signaling alterations in amyotrophic lateral sclerosis
Authors: Kolarcik Christi Lynn , Bowser Robert 
Published: June 2021

Amyotrophic lateral sclerosis (ALS) is characterized by the progressive degeneration and death of motor neurons (MNs) in the brain and spinal cord. Previous reports demonstrated decreased transthyretin (TTR) levels in cerebrospinal fluid of ALS patients. As TTR impacts retinoid signaling, we examined this pathway using immunohistochemistry and immunoblot. Differences in cytoplasmic proteins including cellular retinol binding protein (CRBP) and cellular retinoic acid binding proteins (CRABP)‐I and II were observed. CRBP immunoreactivity was higher in ALS spinal cord MNs while CRABP‐I was decreased in ALS. CRABP–II was concentrated to the nucleus in ALS MNs while control MNs exhibited a diffuse cytoplasmic pattern. We also assessed the expression and distribution of retinoic acid nuclear receptors. RARβ exhibited increased immunoreactivity in MN nuclei of ALS spinal cord, and RXRβ was increased in MNs of ALS spinal cord. Preliminary studies utilizing a motor neuron‐enriched culture system have evaluated the functional significance of these alterations. Our results indicate that retinoic acid signaling is altered in ALS and components of this pathway may represent novel therapeutic targets.Sources of research support: NIH grant ES013469 (RB) and NIH T32 EB001026 (CK)
Nogo‐B limits intima‐media thickening during mouse vein graft adaptation
Authors: Muto Akihito , Yi Tai , Fitzgerald Tamara N , Fancher Tiffany , Magri Dania , Yu Jun , Sessa William C , Dardik Alan 
Published: June 2021

Successful vein graft adaptation to the arterial environment is critical to maintain long‐term graft patency. However, the mechanism of vein graft adaptation remains unclear. Since Nogo‐B protects against vascular injury, we determined whether Nogo‐B plays a mechanistic role, limiting intima‐media thickening, during vein graft adaptation.Intrathoracic inferior vena cava was harvested from wild‐type (WT) or Nogo‐B knockout (Nogo‐KO) mice and placed as an interposition vein graft into the infrarenal aorta of WT mice. Vein grafts were harvested and analyzed after 2 or 3 weeks.In WT mouse vein grafts Eph‐B4 immunoreactivity decreased without increased Ephrin‐B2 immunoreactivity (n=4).Nogo‐KO vein grafts had increased intima‐media area compared to WT vein grafts (.25 mm2 vs .16 mm2, p<.05). However, Nogo‐KO vein grafts also had diminished Eph‐B4 immunoreactivity, similar to the decrease in WT vein grafts.Mouse vein grafts exhibit decreased markers of venous identity, recapitulating changes in humans and rats. Nogo‐KO vein grafts have loss of venous identity, similar to WT vein grafts, but increased intima‐media thickening. This data suggests that Nogo‐B plays a critical role during venous adaptation to an arterial environment independently of mechanisms that determine venous identity, and is also consistent with a general protective role of Nogo‐B in response to injury of the vasculature.
How to attach the limb to the body axis
Authors: Streicher Johannes , Pomikal Christine 
Published: June 2021

While the question “How to make a limb” is extensively addressed in terms of pattern formation, morphogenesis, and genetic background, the ontophyletic scenario of attaching the limb to the axial skeleton via girdles has been paid much less attention. In a comparative study we have investigated pelvis morphogenesis and axio‐appendicular linkage in species from different vertebrate classes. Series of developmental stages from frog tadpoles1, chick embryos and mouse embryos2 were serially sectioned, and 3D‐reconstructed3. The 3D‐models were aligned to 4D‐visualizations of developmental dynamics and further analyzed.Our data reveal, that in all three species ‐ at the morphological level ‐ the pelvic element originates from a single anlage in close relation to the femur. Shape and orientation, however, differ already at initial stages and are followed by divergent subsequent morphogenesis. The timing and patterns of ossification are significantly different. In frog tadpoles a connective tissue strand serves a guiding function for attachment to the sacral vertebra1.These results demonstrate substantial differences in the establishment of the axio‐appendicular linkage among vertebrates and prompt for clarification of their genetic and epigenetic background.
Retroconversion of docosahexaenoic acid (22:6) to eicosapentaenoic acid (20:5) is undetectable in serum of young women
Authors: Brenna J. Thomas , Zhang Ying , Vandal Milène , Cunnane Stephen C. 
Published: June 2021

Background. The polyunsaturated fatty acid (PUFA) 22:6 is widely accepted to be synthesized from 20:5 via these steps: 20:5 → 22:5 → 24:5 → 24:6 → 22:6. Many papers report apparent retroconversion of 22:6 to 20:5, presumed to be by reverse of this pathway, but this hypothesis has not been directly tested in human adults.Methods. Four young adult women were given a single oral dose of [U‐13C]‐22:6, and serum was collected several times on day 1 and weekly until 28 days post‐dose. FAME were prepared and analyzed by high precision GC‐combustion isotope ratio mass spectrometry, and data are expressed as nmol 13C‐FA per ml serum.Results. On average, 22:6* peaked at 24 h and dropped smoothly until day 28, when it remained well above baseline. In contrast, no other n‐3 PUFA showed an increase in labeling of more than a few percent above baseline, and there were no trends attributable to retroconversion. 16:0* peaked at 11% of the 22:6* peak on day 1, dropped to baseline, then gradually increased to a plateau at 4 days, while 18:0* peaked on day 2 at about 7% of the 22:6* peak. These changes were driven primarily by one subject.Conclusions. Increases in serum 20:5 often ascribed to increasing dietary 22:6 may not be due to retroconversion but to other mechanisms such as sparing of 20:5 oxidation. Alternatively, retroconversion may be driven by much higher levels of dietary 22:6. Funding: Canadian Institutes of Health Research.
Biotransformation of soy isoflavones and enhanced cholesterol lowering effect with an oligofructose‐enriched inulin in equol producers
Authors: Wong Julia MW , Kendall Cyril WC , de Souza Russell , Liu Zhen , Emam Azadeh , Jenkins David JA 
Published: June 2021

Objective: To assess whether the hypocholesterolemic effect of soy can be enhanced with an oligofructose‐enriched inulin (OF‐IN) by promoting the biotransformation of colonic soy isoflavones (i.e. equol production). Method: 23 hyperlipidemic adults (11M, 12F; 58±7y; LDL‐C, 4. 1±0.7mmol/L) completed three 4‐week phases: OF‐IN (10 g/d) and low‐fat dairy (control), and soy foods (isoflavones, 61 mg/d) with or without OF‐IN (10 g/d). Fasting blood lipids and anthropometrics were measured biweekly. Urine (24h) was collected at baseline and at the end of each phase. Results: A stepwise reduction in TChol:HDL‐C was observed for OF‐IN (0.25±0.13, P=0.08), soy (–0.07±0.08, P=0.41) and soy plus OF‐IN (–0.36±0.14, P=0.02), compared to baseline. Preliminary analysis suggested that this effect was seen most clearly in equol producers (n=7). TChol:HDL‐C for equol producers was 0.09±0.16 on OF‐IN, –0.23±0.14 on soy, and –0.72±0.42 on soy plus OF‐IN, compared to baseline. For non‐equol producers, TChol:HDL‐C was 0.41±0.17, 0.04±0.09, and –0.17±0.08, respectively. Conclusion: Intake of soy in conjunction with OF‐IN resulted in greater improvements in the lipid profile than their intake alone, suggesting a synergistic effect. This association was more prominent in equol producers possibly related to increased biotransformation of soy isoflavones.Funding: Heart and Stroke Foundation of Ontario, ORAFTI Group
Creation of long‐lasting blood vessels in vivo
Authors: Fukumura Dai , Au Patrick , Tam Joshua , Koike Naoto , Jain. Edwin L Rakesh K. 
Published: June 2021

A major challenge in tissue engineering and regenerative medicine is the creation of stable vasculature. One approach in vascularizing an engineered tissue is to add the cellular components of blood vessels (endothelial and perivascular cells) directly in the tissue‐engineered construct. We have shown the formation of long‐lasting vascular networks in vivo by co‐implanting human umbilical cord vein endothelial cells (HUVECs) and 10T1/2 mesenchymal precursor cells in a three‐dimensional fibronectin‐type I collagen scaffold. Because mesenchymal precursor cells differentiate into perivascular cells and fortify blood vessels in vivo, the resulting vessels are stable and functional for one year. In contrast, the HUVEC‐alone constructs showed minimal perfusion and disappeared by two months. These findings indicate that fostering endothelial‐perivascular cell interactions in vivo offers a promising approach for engineering blood vessels. Furthermore, the tissue‐engineered blood vessel model provides an ideal platform to study cellular and molecular mechanisms of vessel formation and maturation. The potential of different sources of vascular cells in the creation of engineered vessels in vivo can also be tested with this model.Supported in part by NIH grants (P01CA80124, R01CA96915).
Transcriptional pathways in heart development
Authors: Black Brian L , Kang Jione , Harris Ian S , Schachterle William 
Published: June 2021

The second heart field (SHF) is a cardiac progenitor population that resides in the splanchnic and pharyngeal mesoderm and is progressively added to the linear heart tube at the time of cardiac looping to gives rise to the outflow tract (OFT), right ventricle (RV), ventricular septum, and portions of the left ventricle. Although much has been learned regarding the contributions of the SHF to the developing heart, several crucial questions remain regarding the molecular pathways that control SHF development. The LIM‐homeodomain transcription factor ISL1 is required for SHF development and sits at or near the top of a transcriptional cascade for the development of this progenitor population. Likewise, the MADS domain transcription factor MEF2C is required for RV and OFT development. We identified Mef2c as a direct transcriptional target of ISL1 via a novel SHF‐specific enhancer, establishing a direct link between these two essential factors.We have recently identified a novel mesoderm‐specific enhancer from the Isl1 gene, which we are analyzing to define early steps in SHF specification. This enhancer directs expression to the SHF from early cardiac crescent stage through pharyngeal mesoderm development. We are defining the pathways upstream of this novel Isl1 enhancer, and we are continuing our analyses of the Mef2c SHF enhancer with the goal of defining the mechanism through which ISL1 results in sustained activation in the RV and OFT after Isl1 expression itself is extinguished. These data and a SHF transcriptional network involving ISL1, GATA4, Nkx2‐5, MEF2C and other factors will be presented.