bluuebunny/crossref_metadata_2025_split huggingface dataset:
Valeric Acid, a Gut Microbiota Product, Lowers Arterial Blood Pressure in Rats.
Authors: Ufnal Marcin , Onyszkiewicz Maksymilian , Gawrys-Kopczynska Marta , Aleksandrowicz Marta , Sawicka Aneta , Kozniewska Ewa , Samborowska Emilia 
Published: April 2020

Valeric acid (VA), a short‐chain fatty acid (SCFA), is produced by gut bacteria. The aim of the study was to evaluate if VA affect arterial blood pressure (BP) in rats. Furthermore, we investigated physiological concentrations of VA, and distribution of VA‐D9 to organs involved in the control of the circulatory system after the administration of VA‐D9 into the colon, a major site of microbiota metabolism. Hemodynamics were recorded in anesthetized, male, 14‐week‐old Wistar rats. A vehicle (0.9% NaCl), VA, or 3‐hydroxybutyrate, an antagonist of SCFA receptors GPR41/43 (ANT) were administered into the colon (IC) or intravenously (IV). Reactivity of mesenteric (MA) and gracilis muscle (GMA) arteries was tested in ex vivo conditions. SCFAs concentration was measured using ultra performance liquid chromatograph with mass spectrometer.Physiological concentration of VA in the colon content was ≈ 650μM and ≈ 0.4μM in systemic blood. VA‐D9 was detected in the liver, the heart, the brain and kidneys 5 min after the administration into the colon. The vehicle did not affect BP and heart rate (HR). VA acid produced a dose‐dependent decrease in BP, and at higher doses decrease in HR. The hypotensive effect of VA was inhibited by the ANT. In vitro, VA dilated GMA and MA. In MA the dilatory effect was reduced by the ANT.In conclusion, VA rapidly penetrates from the colon to systemic tissues exerting a hypotensive effect. The latter is mediated by vasodilation. The hypotensive effect of colon‐derived VA may have a therapeutic potential.Support or Funding InformationThis work was supported by the National Science Centre, Poland grant no. UMO‐2016/21/B/NZ5/02544.
Mechanisms of Phosphorylation‐Mediated Regulation of the RNAi Effector Protein Argonaute 2
Authors: Bibel Brianna , Elkayam Elad , Joshua-Tor Leemor 
Published: April 2020

Argonaute (Ago) proteins play a central role in gene regulation through a process called RNA interference (RNAi). Ago binds to small RNAs (sRNAs) such as small interfering RNA (siRNA) and microRNA (miRNA) to form the functional core of the RNA Induced Silencing Complex (RISC), which targets mRNAs with complementary sequences, ultimately leading to down‐regulation of the corresponding genes. It was previously shown that phosphorylation of a cluster of residues (824–834) in the “eukaryotic insertion” (EI) of Ago by CK1α can alleviate this repression in a variety of cell types. However, the mechanisms of such regulation were unknown. Using in‐vitro phosphorylation assays, we show that binding of miRNA‐loaded Ago2 to target RNA primes the EI for phosphorylation. Furthermore, we show that this phosphorylation reduces the affinity of the Ago‐miRNA complex to its target mRNA. While the EI of Ago2 contains 5 potential phosphorylation sites, we propose that specific residues in the EI are more susceptible to phosphorylation and dictate the pattern and sequence of the phosphorylation events. We also examined the structural requirements for the target to induce phosphorylation. These findings shed light on the manner in which phosphorylation of the EI acts to regulate the RNAi pathway central to gene regulation. The high conservation of potential phosphosites in the EI (both among Ago homologs and throughout eukaryotes) suggests that such a regulatory strategy may be a shared mechanism with broader impact.Support or Funding InformationThis work was supported by the NSF Graduate Research Fellowship Program and Cold Spring Harbor Laboratory’s Watson School of Biological Sciences. L.J. is an Investigator of the Howard Hughes Medical Institute.
Mechanisms of Myosteatosis in Obesity and the Effects of Muscle Hypertrophy
Authors: Corley Zachary L. , Padgett Caleb A. , Mintz James D. , Fulton David J. , Stepp David W. 
Published: April 2020

ObjectiveTo determine the mechanism of ectopic lipid deposition in skeletal muscle in obesity and if any impact is conferred by increasing muscle mass.MethodsSkeletal muscles from db/db mice, a well characterized model of human obesity, were isolated and myosteatosis assessed for lipid content via NMR, chemical assay and transmission electron microscopy. Genes involved in lipogenesis, lipid storage and oxidation were assessed by qPCR. To determine the effects of hypermuscularity, db/db mice were crossed with mice lacking myostatin, a myokine that inhibits muscle growth, to generate lean, obese, and hypermuscular obese mice.ResultsObese mice exhibited a 3‐fold increase in muscle triglyceride content, which appeared on electron microscopy as diffuse and heterogeneous lipid droplet accumulation in individual muscle fibers. Lipogenic pathways, assessed by expression of sterol regulatory element binding protein (SREBP) and its downstream targets stearyl CoA desaturase‐1 (SCD1), acetyl‐CoA carboxylase (ACC) and fatty acid synthase (FAS), were increased in obesity as was expression of perilipins. Muscle from obese mice was characterized by accumulation of mitochondria and increased expression of carnitine palmitoyltransferase I (CPT1). Deletion of myostatin produced a 40% increase in skeletal muscle mass and reversed the observed change in lipid content and gene expression.ConclusionObesity increases both lipogenesis and lipid oxidation in the skeletal musculature of obese mice. The balance of these processes favors lipid accumulation as evidenced by droplet formation and enhanced expression of droplet‐related proteins. Deletion of myostatin produced muscle hypertrophy and clearance of ectopic lipids. Specific links between muscle hypertrophy and excess muscle lipids remain to be defined.Support or Funding InformationTitle: Circadian Origins of Vascular Disease in Obesity | Investigator(s): Stepp, David W. and Fulton, David J., Augusta University, Augusta, GA | Agency: National Institute of Health (NIH) | Institute: National Heart, Lung and Blood Institute (NHLBI) | Type: Research Project (RO1) | Project #: 1RO1HL147159‐01
The Effects of TNF‐α on TTX Resistant NaV Current in Muscle Dorsal Root Ganglion Neurons after Femoral Artery Occlusion
Authors: Li Qin , Qin Lu , Li Jianhua 
Published: April 2020

Background/AimFemoral artery occlusion in rats has been used to study human peripheral artery disease (PAD). Using this animal model, a recent study suggests that increases of TNF‐α and its receptor lead to the exaggerated responses of sympathetic nervous activity and arterial blood pressure as metabolically sensitive muscle afferents are activated. Note that NaV1.8 channels are predominately presented in chemically sensitive thin fiber sensory nerves. The purpose of this study was to examine the role played by TNF‐α in regulating activity of NaV1.8 currents in muscle dorsal root ganglion (DRG) neurons of rats with PAD induced by femoral artery occlusion.MethodsDRG neurons from control and occluded limbs of rats were labelled by injecting the fluorescence tracer DiI into the hindlimb muscles five days prior to the experiments. A voltage patch clamp mode was used to examine TTX‐resistance (TTX‐R) NaV currents.ResultsSeventy‐two hours of femoral artery occlusion increased peak amplitude of TTX‐R and NaV1.8 currents in muscle DRG neurons. Likewise, after application of A803467 blocking NaV1.8 component of TTX‐R, TTX‐R currents were attenuated to a greater degree in DRG neurons of occluded limbs than in DRG neurons of control limbs. The inhibitory effects of A803467 on TTX‐R currents in muscle DRG neurons were 59 ± 2% in control and 75 ± 5% after occlusion (P<0.05 between two groups; n=6 in each group). TNF‐α exposure amplified TTX‐R and NaV1.8 currents in DRG neurons of control limbs and occluded limbs in a dose‐dependent manner. Notably, amplification of TTX‐R and NaV1.8 currents induced by TNF‐α was attenuated in DRG neurons with pre‐incubation of respective inhibitors of intracellular signal pathways p38‐MAPK, JNK and ERK.ConclusionOur data suggest that NaV1.8 is engaged in the role of TNF‐α in amplifying muscle afferent inputs as the hindlimb muscles are ischemic; and p38‐MAPK, JNK and ERK pathways are likely necessary to mediate the effects of TNF‐α.Support or Funding InformationThis study was supported by NIH P01 HL134609 & R01 HL141198
One‐Time Acute Elevation of Muscle Temperature Is Effective for Attenuation of Exaggerated Exercise Pressor Reflex in Rats with Peripheral Artery Disease
Authors: Qin Lu , Li Qin , Li Jianhua 
Published: April 2020

BackgroundThe blood pressure (BP) response is augmented during static exercise in peripheral artery disease (PAD). Heating treatment was found to be effective for the attenuation of the BP response in PAD patients. However the underlying mechanism for the muscle temperature (Tm) on BP responses in PAD is still unclear.AimsWe first determined the effect of one‐time acute Tm elevation on the exaggerated exercise pressor reflex in PAD rats with 72 hours of femoral artery occlusion. We further determined if P2X receptor was engaged during this process because a lower Tm in occluded muscle can alter expression and function of P2X.MethodsThe BP response to static muscle contraction was recorded to examine the exercise pressor reflex. Also, αβ‐methylene ATP (αβ‐me ATP) was injected into the arterial blood supply of the hindlimb muscles to stimulate P2X receptors in the muscle afferent nerves. To elevate the Tm, a heating pad was placed locally on the hindlimb and the Tm was increased by ~1.5 °C. Then the Tm was stabilized and maintained for 5 min.ResultsThe BP response to static muscle contraction was examined 20 minutes before (pre‐heat), immediately following (acute heat) and 20 minutes after the end of elevation of Tm (recovery). Compared with control rats, a greater mean arterial pressure (MAP) response was observed in PAD rats in the pre‐heat session (27.50±2.07 mmHg in PAD rats/n=12 vs. 18.00±1.67 mmHg in control rats/n=9; P<0.05). However, following elevation of the Tm, the amplification of MAP response was similar in PAD rats and in control rats (21.25±3.00 mmHg in PAD rats/n=12 vs. 18.00±2.56 mmHg in control rats/n=8; P>0.05). During the recovery, a greater MAP response remained in PAD rats (27.54±0.90 mmHg in PAD rats/n=11 vs. 20.14±1.43 mmHg in control rats/n=7; P<0.05). Note that no significant difference in developed muscle tension during contraction was seen in control rats and PAD rats (P>0.05). When αβ‐me ATP (0.125mM) was given, the MAP responses were higher in PAD rats (n=7) than in control rats (n=7) (56.03±4.70 mmHg in PAD vs. 39.12±4.59 mmHg in control for pre‐heat, 59.63±3.55 mmHg in PAD vs. 39.92±5.20 mmHg in control after heating, and 63.33±2.51 mmHg in PAD vs. 35.57±4.38 mmHg in control for recovery; P<0.05 for all sessions between two groups).ConclusionOne‐time acute elevation of the Tm inhibits the exaggeration of the exercise pressor reflex in PAD rats. The effects of one‐time heat may not be through alternating muscle afferent nerves’ P2X receptors.Support or Funding InformationThis study was supported by NIH P01 HL134609 & R01 HL141198
Morphine blocks spinal respiratory motor plasticity via mechanisms that require toll‐like receptor 4 signaling
Authors: Tadjalli Arash , Novak Jonathan , Bolser Donald C. , Levitt Erica S. , Mitchell Gordon S. 
Published: April 2020

Opioid therapy is a common strategy for pain management; however, unfavorable side effects limit their clinical efficacy. Pro‐inflammatory cytokines oppose acute and chronic opioid‐induced analgesia and the ensuing inflammatory cascades that signal through toll‐like receptor 4 (TLR4)—pattern recognition innate immune receptors—are a significant contributor to some of the negative consequences of opioid therapy. While opioid‐induced inflammatory processes contribute to unwanted side‐effects (i.e. tolerance, dependence), it is unknown whether opioids, through pro‐inflammatory signaling, can affect the expression of plasticity within respiratory motor circuits. Adaptive plasticity within respiratory motor circuits is an important contributor to the preservation and/or restoration of respiratory function. For example, respiratory long‐term facilitation (LTF), a well‐studied model of respiratory motor plasticity, promotes breathing stabilization by increasing inspiratory drive to major breathing muscles. LTF is characterized by a long‐lasting compensatory increase in ventilation or respiratory motor nerve output triggered by exposures to acute intermittent hypoxia. Respiratory LTF is abolished by even a mild inflammation triggered by TLR4 receptor activation (e.g. via systemic lipopolysaccharides). Since opioid therapy activates TLR4 and induces neuro‐inflammation, we hypothesized that opioid delivery will abolish LTF through mechanisms that require TLR4 signaling. In adult Sprague Dawley rats, we show that acute systemic delivery of morphine, a prototypical opioid, blocks the expression of LTF in the phrenic motor system—a major motor pool that drives contractions of the diaphragm muscle. Furthermore, we show that morphine blocked phrenic LTF via mechanisms that require TLR4 signaling because systemic pre‐treatment with (+)‐naloxone—an opioid‐inactive TLR4 signaling inhibitor—restored phrenic LTF in morphine‐treated rats. Our findings suggest that loss of phrenic LTF following morphine treatment may remove an important stabilizing factor for breathing, potentially contributing to respiratory failure. Thus, attenuation of inflammatory signaling by opioid‐inactive compounds may help restore endogenous mechanisms of spontaneous plasticity within respiratory circuits, thereby minimizing opioid‐induced respiratory depression.Support or Funding InformationNIH SPARC Initiative (OT2OD023854)
Role of AMPK in Adiponectin‐Mediated Restoration of Nitric Oxide‐Dependent Flow Induced Dilation in the Human Microvasculature
Authors: Cohen Katie E. , Schulz Mary E. , Gutterman David D. , Freed Julie K. 
Published: April 2020

Flow‐induced dilation (FID), the ability of an arteriole to dilate in response to increased flow, is primarily mediated by endothelial‐derived nitric oxide (NO) in healthy individuals, however is replaced by hydrogen peroxide (H2O2) in patients with coronary artery disease (CAD). Our laboratory has shown that adiponectin restores NO‐mediated FID in microvessels from patients with CAD. Adiponectin activates multiple signaling pathways that promote NO formation, including activation of neutral ceramidase to hydrolyze ceramide, a sphingolipid that in elevated amounts promotes FID mediated by H2O2. Our prior work has shown that both NO and H2O2 contribute as mediators of FID in diseased vessels treated with adiponectin and ceranib‐1, an inhibitor of neutral ceramidase. As adiponectin is also capable of activating AMP kinase (AMPK) to increase cellular NO levels, we tested the hypothesis that AMPK activation is required to restore NO‐mediated FID in diseased arterioles. Human microvessels (80–200μm) were dissected from adipose tissue collected from patients diagnosed with CAD. Following cannulation and equilibration, vessels were constricted with endothelin‐1. Using videomicroscopy, vessel diameters were measured in response to graded increases in intraluminal flow. To investigate whether activation of AMPK is a key mechanistic step in adiponectin‐induced restoration of NO‐mediated signaling, vessels from patients diagnosed with CAD were first treated with the AMPK inhibitor Compound C (1μM, 4hrs) followed by adiponectin treatment (2μg/mL, 16–20hrs). Dilation to flow was maintained during exposure to the nitric oxide synthase (NOS) inhibitor L‐NAME (100μM) (69.8% max dilation±2.7, n=2) as well as in the presence of the PEG‐Catalase (500U) (64.5%±21.2, n=4) compared to compound C and adiponectin alone (61.0%±9.4, n=4). To determine whether ceramide hydrolysis via neutral ceramidase contributes to the maintained dilation despite NOS inhibition, FID was also assessed in diseased arterioles treated with both compound C and ceranib‐1 prior to treatment with adiponectin. Both NO and H2O2 contributed to dilation under both AMPK and neutral ceramidase inhibition as dilation was observed in the presence of L‐NAME (33.6%±15.9, n=3) or PEG‐Catalase (51.7%±23.9, n=3). However, FID was decreased during exposure to both L‐NAME and PEG‐Catalase (11.6%±12.4, n=2). These preliminary data suggest that adiponectin promotes NO formation through multiple pathways and contributes to the redundancy of endothelial‐derived mediators in order to maintain FID.Support or Funding InformationThis research was supported by National Institute of Health (NHLBI) K08HL141562 (J.K.F.) and National Institute of Health 5R38HL143561‐02 (M.E.W.).
Activation of VTA NtsR1 Neurons Promotes Weight Loss Behaviors
Authors: Perez-Bonilla Patricia , Santiago-Colon Krystal , Matasovsky Jillian , Leinninger Gina M. 
Published: April 2020

Dopamine (DA) neurons in the ventral tegmental area (VTA) modulate motivation to engage in physical activity and food intake, dual behaviors that are disrupted in obesity. However, the functional heterogeneity of VTA DA neurons, and lack of molecular markers to distinguish them, has prevented determination of which DA neurons might be leveraged to support weight loss behaviors. We identified a distinct subset of VTA DA neurons defined by their expression of neurotensin receptor‐1 (NtsR1) and we hypothesized that these “VTA NtsR1 neurons” modulate feeding restraint and physical activity that could support weight loss. To test this hypothesis, we expressed Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) in the VTA NtsR1 neurons of normal weight and diet‐induced obese mice, permitting in vivo activation of VTA NtsR1 neurons by treatment with the DREADD‐ligand clozapine‐N‐oxide (CNO). Acute activation of VTA NtsR1 neurons (24hr) in normal weight and diet‐induced obese mice significantly decreased body weight by reducing food intake and increasing physical activity. Moreover, the weight loss of diet‐induced obese mice was maintained with daily activation of VTA NtsR1 over 7d. Intriguingly, activation of VTA NtsR1 neurons also suppressed how much mice would work to obtain sucrose rewards even when there was high motivation to consume (e.g. in fasted mice or diet‐induced obese mice.) These data suggest that VTA NtsR1 neurons may be useful to support weight loss behavior. Yet, the translational potential of NtsR1 was previously dismissed due to the broad expression of NtsR1 throughout the brain and periphery, and data that systemic NtsR1 agonists cause adverse hypothermia and vasodepressor responses. However, we found that specifically activating VTA NtsR1 neurons does not alter blood pressure, core body temperature or anxiety behavior. Thus, activation of VTA NtsR1 neurons during states of high‐appetitive drive such as fasting and obesity promotes dual behaviors that support weight loss without causing adverse cardiorespiratory effects or stress responses. In the future, strategies to selectively modulating VTA NtsR1 neurons could provide a safe, effective avenue for weight loss.Support or Funding InformationSupported by funding from the National Institutes of Health to Perez‐Bonilla (T32GM092715) and Leinninger (R01‐DK103808)
Effect of hyperglycaemia on the growth of lung pathogens in Artificial Sputum Media
Authors: Vasiljevs Stanislavs , Gupta Arya , Baines Deborah 
Published: April 2020

Elevated glucose concentration in the airway surface liquid due to high blood glucose (hyperglycaemia) has been associated with the increased risk of developing pulmonary infection. It has recently been shown that lungs are colonisedby resident bacteria and are not truly sterile. Several chronic lung conditions, such as cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD), have been shown to induce changes in this bacterial population.Here we sought to develop an in vitro model using artificial sputum media (ASM) to investigate the effect of hyperglycaemia on Staphylococcus aureus (SA) and Pseudomonas aeruginosa (PA), two of the most common pathogens in CF patients.SA (ATCC29213) and PA (H174) were grown in the artificial sputum media (ASM) at varying glucose concentrations. Approximately 3×105 bacteria were added to 15 mL ASM and grown at 37°C with constant shaking. Two‐way ANOVA with Geisser‐Greenhouse correction and Pearsons correlation coefficients were used for statistical analysis.ASM supplemented with 4 mM glucose had a significant effect on the growth rate of both S. aureus and P. aeruginosa when compared with ASM with no glucose (p = 0.0028, n=5 for SA and p < 0.0001, n=5 for PA). Compared with growth in Mueller Hinton broth, only P. aeruginosa exhibited increased growth rate in the presence of 4 mM glucose (p = 0.0082, n = 3). Growth of both, S. aureus and P. aeruginosa, was correlated with the depletion of glucose in the media (p = 0.0002, n=5 for PA and 0.0038, n=5 for SA). The production of lactate by S. aureus was not related to the depletion of glucose as concentrations for both reached zero at the sixth hour of growth.As ASM mimics the nutrients available in the lungs, it facilitates the study of lung pathogens in the presence of glucose. The established in vitro assay in this study can be further utilised to examine the competition and/or co‐existence between multiple pathogens that are known to reside within the lungs.Support or Funding InformationResearch funded by MRC‐DTP
Development of New Methods for Enhanced Conformational Sampling of GPCRs
Authors: Serrano Erik , Abrol Ravinder 
Published: April 2020

G protein coupled receptors (GPCRs) are integral membrane proteins that enable a cell to respond to extracellular stimuli like light, small molecules, peptides, and proteins. These receptors have evolved as highly flexible proteins that possess multiple functionally important conformational states, which enable these receptors to activate pleiotropic signaling events inside the cells. Recent progress in membrane protein structural biology techniques and cryo‐electron microscopy is providing structural evidence of two or more distinct receptor conformations for these receptors. Computational structural modeling approaches can complement and supplement these studies by predicting all functionally important states of a receptor, however, they run into the classical protein conformational sampling problem. We are developing a computational biophysical method called Enhanced Conformational Markov‐state Sampling in Membrane BiLayer Environment (EnCoMSeMBLE) to solve this sampling problem for GPCRs. We are utilizing Markov‐State‐Models to combine the Boltzmann sampling of receptor conformations from molecular dynamics (MD) with the brute‐force sampling of receptor conformations from our previously developed ActiveGEnSeMBLE method. The method was applied to the A2A receptor starting from the active state by only using traditional MD simulations. This is not surprising because the pre‐activce states is a downhill direction on the energy landscape. However, applying both MD and ActivateGEnSeMBLE methods was able to predict pre‐active states from the inactive states which, classical MD cannot achieve. Our results indicated that our methods provide an unbiased method to sample the conformational landscape. This can protentional sample unexplored regions of the conformational landscape and find novel conformations that can elucidate the mechanistic properties that GPCRs possess.Support or Funding InformationThis project was funded through a grant from the National Institutes of Health (NIH) Building Infrastructure Leading to Diversity (BUILD) #5TL4GM118977 or # 5RL5GM118975.