bluuebunny/crossref_metadata_2025_split huggingface dataset:
Endothelial‐derived microparticles activates TLR4/JAK3/STAT3 pathway to induce acute lung injury
Authors: Ou Zhijun , Yuan Hao-Xiang , Li Yu-Quan , Wang Yan-Sheng , Li Yan , Ma Jian , Xu Jun , Ou Jing-song 
Published: April 2020

We previously reported that endothelial‐derived microparticles (EMP) could induce acute lung injury (ALI) and the mechanism is still not fully understood. Toll‐like receptor 4(TLR4) cascade has been considered to be involved in innate immune inflammation. As its downstream, janus kinases/signal transducers and activators of transcription (JAK/STAT), acts as a key signal pathway for immune responses. However, it remains unclear whether systemic circulating EMP could trigger TLR4‐pomoted JAK/STAT signaling contributing to ALI. EMP were isolated from human umbilical vein endothelial cells (HUVECs) stimulated with plasminogen activated inhibitor‐1. HUVECs were treated with/without different concentration of EMP. EMP induced inflammatory cytokines in cultured HUVECs. EMP could activated TLR4 and the downstream JAK3/STAT3 pathway. Silencing TLR4 inhibited the EMP‐induced JAK3/STAT3 pathway. EMP‐induced inflammatory response in plasma and bronchoalveolar lavage fluid, inflammatory cells infiltration. edema and JAK3/STAT3 pathway in the lung tissue, and ALI in C57BL6 mice. However, These effect induced by EMP were depressed in TLR4−/− mice. Our data demonstrated that EMP induce ALI by activating TLR4/JAK3/STAT3 pathway to promote inflammatory response. Our findings present the novel mechanisms by which EMP induce ALI.Support or Funding InformationThis research was financially supported by the National Natural Science Foundation of China (Grants 81670392, 81600382, 81770241, 81830013, 81970363 and Distinguished Young Scholar Grant 81325001), 973 project (2009CB522104) and International Cooperation project (2015DFA31070) from the Ministry of Science and Technology of China, Guangdong Natural Science Fund Committee (Grant 2015A030312009), the Changjiang Scholars Program from the Ministry of Education of China, the Sun Yat‐sen University Clinical Research 5010 Program.
Large Hypertrophy but Unmodified Specific Tension of Single Fibers of Body Builders
Authors: Monti Elena , Toniolo Luana , Marcucci Lorenzo , Martellato Ivan , Bondì Michela , Franchi Martino , Narici Marco , Reggiani Carlo 
Published: April 2020

IntroductionMuscle hypertrophy is the main outcome of training for body building. Previous studies showed a discrepancy between the functional (force/tension) and structural (cross‐sectional area, CSA) proprieties of leg extensor muscles measured in vivo and the corresponding parameters measured in single muscle fibers
[1]
[2]. In particular, a puzzling finding from these studies was that of a lower specific tension of skinned fibers of body builders (BB) compared to a control (C) population.In this study we aimed to investigate the possible reasons behind the alleged lower specific tension in single fibers of a strictly diet‐and‐training‐controlled BB population.MethodsFive male body builders (26±4.7 yr; 87.2±9.7 kg; 178.4±7.5 cm) with a training history of at least 5 years were recruited for this study. Five age‐matched recreationally active males (24.6±3.6 yr; 84.4±14.9 kg; 186.4±4.4 cm) were recruited as controls. After ethical approval, biopsies from the vastus lateralis (VL) muscle were collected together with in‐vivo quadriceps maximum isometric voluntary contraction (MVC) and architecture and CSA. Data analysis included (i) single skinned fiber mechanics (n=200 fibers; maximum force, CSA, specific tension and calcium diffusion), (ii) single fiber myosin quantification (n=100 fibers), (iii) slow and fast fiber CSA via histological section analysis. In addition, swelling ratio (skinned fibers CSA/histological CSA) was calculated for each fiber type and for each subject. A general linear mixed model was used to assess statistically significant differences.ResultsQuadriceps MVC and CSA were 64% and 33% higher in BB than C (p<0.05), VL pennation angle (PA) and thickness (MT) were 16% and 15% higher in BB than C (p<0.05). Quadriceps specific force was 25% higher in BB than C (n.s.). Single skinned type 2 CSA and force were 48% and 28% higher in BB than C (p<0.05), while specific tension was 19% lower (p<0.05). Type 1 fibers specific tension was 32% higher in BB than C (p<0.05). Myosin content and calcium diffusion were similar among populations. CSA determined in histological sections was 10% and 27% higher in BB than C (p<0.05) for type1 and 2 fibers, respectively. When type 1 and type 2 fiber tensions were corrected for individual swelling ratio, the difference between BB and C disappeared.ConclusionsAlthough at first sight these findings seem to confirm that in type 2 skinned fibers of BB specific tension is lower than in C, once accounting for fiber swelling, this difference disappears. This is further confirmed by the lack of difference in myosin concentration and calcium diffusion kinetics.In contrast, the results point to the existence of a differential swelling response to the skinning process by BB and C fibers, possibly due to adaptations of the fiber cytoskeleton proteins or specific permeability of the membrane of slow and fast fibers.
SPARC, a Muscle‐Secreted Protein, Improves Glucose Tolerance via AMP‐activated Protein Kinase in Skeletal Muscle
Authors: Aoi Wataru , Sakuma Kunihiro , Krook Anna 
Published: April 2020

Secreted protein acidic and rich in cysteine (SPARC) is an exercise‐responsive myokine. Although SPARC has been implicated in nutrient metabolism, the effect of this myokine on muscle metabolism is unclear. Here we investigated the effect of SPARC on glucose metabolism in skeletal muscle.SPARC‐knockout mice showed impaired systemic glucose metabolism. Chronic SPARC treatment improved systemic glucose tolerance and AMP‐activated protein kinase (AMPK) signaling in skeletal muscle of high‐fat diet‐induced obese mice. A protein interaction assay showed that SPARC strongly interacted with voltage dependent calcium channel. In cell culture experiments, SPARC increased glucose metabolism concomitant with activation of AMPK, and this effect was suppressed by inhibiting the calcium‐dependent signaling.Our findings suggest that SPARC treatment mimics the effects of exercise on glucose tolerance by enhancing AMPK‐dependent glucose uptake in skeletal muscle.Support or Funding InformationThis study was partly supported by the Japan Society for the Promotion of Science (JSPS) KAKENHI: Grant‐in‐Aid for Scientific Research (B) (grant numbers 25282199 and 17H02176).
MEF2C Mediates Chronic Pain in Rodent Neuropathy Models
Authors: Serafini Randal Alexander , Sakloth Farhana , Pryce Kerri , Bertherat Feodora , Akbarian Schahram , Zachariou Venetia 
Published: April 2020

Peripheral nerve injury triggers a number of maladaptive changes in both peripheral and central sites, leading to debilitating hypersensitivity, as well as cognitive, motivational, and affective deficits. Understanding the epigenetic and transcriptional events mediating pathological pain and synaptic reorganization will provide essential information for novel approaches towards the management of neuropathic pain. Our lab has previously demonstrated that prolonged peripheral nerve injury leads to an upregulation of histone deacetylase 5 (HDAC5) in the mouse Nucleus Accumbens (NAc), and revealed a dynamic role of HDAC5 in the antiallodynic efficacy of monoamine‐targeting antidepressants. HDAC5 negatively regulates the expression of Myocyte Enhancer Factor 2C (MEF2C), a transcription factor critical for synaptic plasticity in adulthood. Aberrant MEF2C expression contributes to clinical neuropsychiatric abnormalities such as autism and schizophrenia. By utilizing qPCR and RNAscope in situ hybridization, we found that long‐term peripheral nerve injury dynamically affects MEF2C expression in distinct cellular populations of chronic pain circuits. We also used adeno‐associated vectors to overexpress MEF2C in the NAc of adult mice in order to determine the impact of this transcription factor in the trajectory of sensory hypersensitivity associated with peripheral nerve injury. We found that while MEF2C overexpression did not affect the induction of sensory hypersensitivity, it did disrupt the maintenance of sensory hypersensitivity symptoms. Specifically, MEF2C over‐expressors recover from mechanical allodynia and show restored motivational drive. In conclusion, our findings suggest that manipulation of the plasticity‐mediating transcription factor, MEF2C, in the brain reward system holds promise for interrupting the chronification of pain, as well as its affective components. We are further investigating the mechanisms underlying neuropathy‐related changes in MEF2C expression and/or activity via phosphorylation‐specific protein analysis, in vivo DNA methyltransferase identification (DamID), and gene silencing tools for downstream molecular targets.Support or Funding InformationNS086444 & NS111351 NINDS,MSTP T32 NIH T32 GM007280,Irene & Eric Simon Brain Research Foundation Fellowship
The Influence of Selenium on Arsenic Hepatobiliary Transport
Authors: Zhou Janet R. , Kaur Gurnit , Arutyunov Denis , Leslie Elaine M. 
Published: April 2020

Over 200 million people worldwide are exposed to the proven human carcinogen arsenic at levels exceeding the World Health Organization guideline of 10 μg/L. In animal models arsenic and selenium are mutually protective via the formation and biliary excretion of the seleno‐bis (S‐glutathionyl) arsinium ion [SeAs(GS)2]−, which allows for the fecal elimination of both compounds. Consistent with this, selenium deficiency in humans living in arsenic endemic regions is associated with an increased risk of arsenic‐induced disease. These observations have led to the initiation of human selenium supplementation trials. Despite these ongoing trials in arsenic endemic regions, the influence of selenium on human hepatic handling of arsenic is not adequately understood. Furthermore, supplementation trials have utilized different chemical forms of selenium with unknown influence on efficacy. In the liver, multidrug resistance protein 2 (MRP2/ABCC2) transports arsenic metabolites, including [SeAs(GS)2]− into bile, and the related MRP4 (ABCC4) transports other arsenic metabolites into sinusoids. We hypothesized that selenium will increase the biliary excretion of arsenic from HepaRG cells, an immortalized cell line used as a surrogate for primary human hepatocytes. To test this hypothesis, we studied the influence of selenite (SeIV), selenide (SeII), methylselenocysteine (MSC) and selenomethionine (SeMet) on arsenic efflux from HepaRG cells. The expression of genes involved in hepatic arsenic methylation (As3MT) and export (ABCC2 and ABCC4) were assessed. Crude membrane preparations subjected to immunoblots were used to evaluate the presence of MRP2 and MRP4 proteins. The influence of arsenic on the levels of these proteins was also assessed similarly. Fluorescence microscopy after treatment with 5(6)‐carboxy,2’,7’‐dichlorofluorescein (CDF) diacetate was performed to visualize the canalicular networks and assess MRP2 function. Transport across sinusoidal and canalicular membranes was measured after treatment of HepaRG cells with 1 μM 73AsIII ± selenium (in the forms SeIV, SeII, MSC or SeMet) using B‐CLEAR® technology. Biliary excretion indices (BEIs) were calculated to quantify the extent of arsenic export into bile. ABCC2, ABCC4, and As3MT are expressed by HepaRG Cells. MRP2 and MRP4 proteins were detected, and their expression was increased by the presence of AsIII. CDF accumulation in canalicular networks suggested that MRP2 was functional. At a 5 minute time point, the BEI of 73AsIII alone was 17%. The addition of SeII increased biliary excretion of 73AsIII to 30%. Biliary excretion of 73AsIII was lost in the presence of SeIV, SeMet and MSC. Arsenic underwent biliary excretion in HepaRG cells and this was stimulated by SeII, and inhibited by SeIV, SeMet and MSC. These data have implications for the form of selenium utilized for selenium supplementation trials in arsenic endemic regions.Support or Funding InformationCanadian Institutes of Health Research (CIHR)
Engineering Temperature Sensitive Alleles in Saccharomyces cerevisiae to Study the Genetic Interactions Between Ran point mutants and its Accessory Proteins
Authors: Tsepal Tenzin 
Published: April 2020

Ran is an essential GTPase protein involved in bidirectional transportation between nucleus and cytoplasm. We engineered temperature sensitive (TS) mutant alleles of Ran accessory proteins in S. cerevisiae to study the genetic interactions. For essential proteins like Ran and its accessory proteins, TS mutants give insight into gene function without knocking out genes. By creating these TS mutant strains, we will be able to study the genetic interactions between Ran point mutants and TS mutant Ran accessory proteins to closely interrogate the biophysics and mechanism of the Ran GTPase cycle and it’s role nucleocytoplamsic transport.Support or Funding InformationThe 2019 NIH Summer Undergraduate Research Program at UMass Medical School. NIH Grant # R25 HL092610‐11
Sodium Caprate Enhances Transcellular LPS Transport via Clathrin‐Mediated Endocytosis in Rat Jejunum
Authors: Akiba Yasutada , Kaunitz Jonathan D. 
Published: April 2020

Lipopolysaccharides (LPS) entry from the intestinal lumen to the circulation is associated with systemic inflammation. We have found that LPS is transcellularly transported to portal vein during long‐chain fatty acid (LCFA) exposure via CD36‐ and lipid raft‐mediated pathways. We examined the effects of medium‐chain fatty acids (MCFA) on LPS transport in rat jejunum.FITC‐LPS was applied to the mucosal bath of Ussing chambered muscle‐stripped jejunal and distal colonic mucosa‐submucosa preparations, in which short‐circuit current (Isc) and tissue electrical resistance (TER) were measured. Serosal appearance of FITC‐LPS was measured with or without luminal application of sodium caprate (C10, 10 mM). FITC‐dextran 4kDa (FD4) m‐to‐s movement was also separately measured.Luminal application of caprate increased FITC‐LPS m‐to‐s transport without any changes in FD4 m‐to‐s movement and TER in the jejunum, whereas caprate remarkably increased FITC‐LPS transport with increased FD4 movement and decreased TER in the distal colon, suggesting that caprate enhances transcellular LPS transport in the jejunum, but increases paracellular permeability in the distal colon. In the jejunum, C10‐mediated FITC‐LPS m‐to‐s transport was inhibited by pretreatment with luminal Pitstop2 (30 μM), a selective inhibitor of clathrin‐mediated endocytosis (CME), whereas the CD36 inhibitor sulfosuccinimidyl oleate (SSO, 0.1 mM), the lipid raft inhibitor methyl‐β‐cyclodextrin (MβCD, 1 mM), or the dynamin inhibitor Dynasore (50 μM) had no effect. In contrast, luminal OA (30 mM) with taurocholate (TCA, 0.1 mM) also increased FITC‐LPS transport with SSO, MβCD and Dynasore‐sensitive, but Pitstop2‐insensitive manner. In vivo, jejunal luminal perfusion of C10 with FITC‐LPS increased portal venous FITC‐LPS appearance 15 min after perfusion, higher than by OA/TCA, which was then inhibited by co‐perfusion of Pitstop2.These results suggest that luminal MCFA and LCFA differentially induce transcellular LPS transport in the jejunum; MCFA via CME, but LCFA via the CD36/lipid raft/caveolin pathway. Dietary fatty acids may impact LPS‐associated diseases, including metabolic disease and multiple organ failure.Support or Funding InformationVA Merit Review and Shire Pharmaceuticals
A K‐RAS Inhibitor Abrogates Self‐Renewal of Pancreatic Cancer Stem Cells via K‐RAS – NF‐κB – SOX2 Axis
Authors: Stanslas Johnson , Teh Yuan Han , Ho Kok Lian , Ramasamy Rajesh , Sagineedu Sreenivasa Rao 
Published: April 2020

BackgroundGrowing evidence indicate that pancreatic cancer stem cells (pCSCs) being the culprit of cancer recurrence and metastasis. These chemoresistant cells remain undifferentiated, while giving rise to their differentiated progenies in an asymmetrical cell division known as self‐renewal to sustain tumor maintenance and progression. Therefore, targeting the self‐renewal of pCSCs is no doubt a key strategy to eradicate them. Here, we are the first to describe 4,6‐dichloro‐2‐methyl‐3‐aminoethyl‐indole (DCAI), a known K‐RAS binder that inhibits K‐RAS signaling, as a novel anti‐pCSC agent; its mechanism of action in abrogating the self‐renewal of pCSCs was elucidated.MethodsCell viability assay was used to determine the chemoresistantance of pancreatic cancer cell lines with clinically relevant K‐RAS mutations: PANC‐1 (K‐RASG12D), Capan‐2 (K‐RASG12V), and MIA PaCa‐2 (K‐RASG12C). Tumorsphere assay, which is a hallmark test platform for evaluating potential self‐renewal inhibitors in vitro was utilized. We employed this assay to investigate the effect of DCAI on the self‐renewal of pCSCs. Briefly, tumorspheres were treated with test compounds, followed by counting intact tumorspheres (diameter ≥ 60 μm). The molecular players of stem cell maintenance, K‐RAS and NF‐κB pathways in tumorspheres, before and after treatments, were examined by Western blotting.ResultsCapan‐2 (GI25 > 100 μM) and PANC‐1 (GI25 = 90 ± 17 μM) were equally ranked as the most resistant cell line to gemcitabine, as compared with MIA PaCa‐2 (GI25 = 0.053 ± 0.009 μM) cells. This profound chemoresistance coincided with the tumorsphere‐forming ability, which was found solely in Capan‐2. The Capan‐2 tumourspheres were remarkably irresponsive to gemcitabine (IC50 > 100 μM). We were intrigued to learn that DCAI (IC50 = 5 μM) abrogated the formation of Capan‐2 tumorspheres more effectively than vismodegib (IC50 > 20 μM), an anti‐pCSC agent. Several studies collectively suggested a link between K‐RAS signaling, NF‐κB signaling, and SOX2 expression. In our study, Capan‐2 tumourspheres demonstrated hyperactivations of MAPK and AKT pathways, both of which are downstream of K‐RAS. Activation of P65 NF‐κB signaling molecule was observed in respond to the hyperactivation of K‐RAS signaling, which eventually led to the overexpression of SOX2. Upon treatment with DCAI, K‐RAS and NF‐κB signalings were significantly inhibited and SOX2 expression was greatly reduced in Capan‐2 tumourspheres.ConclusionOur present study revealed that DCAI is a promising agent pertaining to targeting the self‐renewal of pCSCs via K‐RAS – NF‐κB – SOX2 axis. It is suggested that K‐RAS is a valid therapeutic target in pCSCs for eradicating pancreatic cancer recurrence and metastasis. Targeting K‐RAS could form a solution to the problem that remains unsolved for many decades.Support or Funding InformationMalaysian Ministry of Education (Fundamental Research Grant Scheme, 04‐01‐17‐1905FR)
Stimulation of the β2 adrenergic receptor rapidly increases phosphorylation of the Na+‐Clcotransporter (NCC)
Authors: Poulsen Søren Brandt , Cheng Lei , Penton David , Murali Sathish Kumar , Kortenoeven Marleen , Loffing Johannes , Fenton Robert A. 
Published: April 2020

Adrenergic signaling is proposed to modulate activity of the thiazide‐sensitive Na+‐Clcotransporter (NCC) by direct effects on the renal distal convoluted tubule (DCT). However, there are conflicting data on whether, α1, β1 or β2‐adrenoceptors mediate these effects by activation of the with‐no‐lysine kinases (WNKs) and STE20/SPS1‐related proline alanine‐rich kinase (SPAK) kinase pathway or by other signaling pathways. Here, we determined using RT‐PCR that the β2‐adrenergic receptor was abundantly expressed in mpkDCT cells (a model of the DCT) and in DCT cells isolated from parvalbumin EGFP mice by fluorescence‐activated cell sorting. In mice administered salbutamol (β2 adrenergic receptor agonist used to treat asthma) for 30 min, phosphorylated NCC levels increased, indicating increased activity. However, mice developed hypokalemia, a condition that also increases NCC phosphorylation. To investigate direct effects of β2 adrenergic signaling on DCT cells independent of K+, the effects of salbutamol on mpkDCT cells and in kidney slices/tubule suspensions were examined. Salbutamol gradually increased NCC phosphorylation in the pharmacologically relevant concentration range of 0.01–10 μM, an effect observed after 15 min and maintained at 60 min. Changes in NCC phosphorylation were associated with increased cAMP levels in tubule suspensions, kidney slices and mpkDCT cells. A role for the NCC modulatory PKA/I‐1/PP1 pathway was suggested by attenuated effects of salbutamol on phosphorylated NCC during PKA/PP1 pathway inhibition or in slices from I‐1 KO mice. Inhibition of the WNK‐SPAK pathway with a WNK inhibitor also attenuated, but did not prevent, the salbutamol‐induced phosphorylation of NCC. In conclusion, β2 adrenergic receptor signaling modulates NCC phosphorylation in vivo and ex vivo via PKA and WNK‐dependent pathways. We speculate that this response may be a compensatory mechanism to limit electrogenic renal K+ secretion during β2 adrenergic receptor stimulation of muscle. Furthermore, frequent use of salbutamol in e.g. asthmatic patients may lead to renal NaCl retention and influence blood pressure.Support or Funding InformationThe Novo Nordisk Foundation, The Leducq Foundation and the Danish Medical Research Council.