bluuebunny/crossref_metadata_2025_split huggingface dataset:
Virus replication and cytokine production in primary alveolar macrophages and airway epithelial cells induced by emerging canine H3N8 influenza virus (LB510)
Authors: Castleman William , Hancock Calvin 
Published: June 2021

Canine H3N8 influenza virus was first recognized as a serious infectious pulmonary disease in racing greyhound dogs in 2004. Disease resulted from direct transmission of commonly circulating equine H3N8 virus to dogs.We hypothesized that transmission of equine influenza virus to dogs was characterized by mutations in one or more genes that allowed the virus to replicate more efficiently in dog airway epithelial cells and/or alveolar macrophages and induce higher levels of proinflammatory cytokines. Primary alveolar macrophages and airway epithelial cells were isolated from dogs and inoculated with emergent and endemic canine H3N8 influenza viruses A/canine/Florida /43/2004 (canine/FL/04), canine/FL/05, canine/PA/09 as well as early and more recent equine viruses equine/KY/91, equine/NY/99 and equine/OH/03.Cytokines responses were measured by multiplex protein assay and/or quantitative PCR. Emergent and endemic canine viruses replicated to higher titers in dog macrophages than did the equine viruses from 1999 and 2003. Only endemic canine viruses from 2005 and 2009 replicated to a higher titer in isolated dog airway epithelial cells than did equine viruses from 1999 and 2005. Induction of TNF‐alpha in macrophages by equine and canine viruses was highly variable among macrophage isolates from different dogs. No significant differences were measured in epithelial cell production of IL‐8 in response to canine or equine H3N8 virus inoculation. Although small differences in productive virus replication were detected for canine influenza virus isolates compared to equine influenza virus isolates in macrophages and epithelial cells, differences in cytokine production were not detected.Grant Funding Source: Supported by NIH AI101625
Expedited RPE‐derivation from human embryonic stem cells by partial PARP‐1 inhibition (LB55)
Authors: Pennington Britney , Clegg Dennis 
Published: June 2021

The death of the retinal pigmented epithelium (RPE) induces the leading cause of blindness in the elderly, a disease called age‐related macular degeneration (AMD). Human embryonic stem cells (hESCs) have the potential to generate a limitless source of RPE for cellular therapies, but spontaneous differentiation methods require several months to produce mature cells. In 2013, a novel directed‐differentiation protocol described efficient derivation of RPE in 14 days using nicotinamide as a key‐inducing agent in the chemical cocktail. This study investigates the hypothesis that nicotinamide expedites RPE‐derivation by inhibiting poly(ADP‐ribose) polymerase‐1 (PARP‐1) during the neuralization component of differentiation. 3‐aminobenzamide (3‐ABA), a non‐hydrolyzable nicotinamide analogue and PARP‐1 inhibitor, was tested in place of nicotinamide. Gene expression analysis by qPCR, morphological assessment, and pigment quantification demonstrate that 3‐ABA affects differentiation during the first 4 days of differentiation, ultimately yielding cells with similar, yet attenuated, levels of RPE‐marker gene expression compared to controls. This suggests that nicotinamide contributes to RPE differentiation from hESCs by partially inhibiting PARP‐1. Since other mechanisms are likely involved, future investigation aims to identify additional pathways contributing to this efficient differentiation of RPE cells, which will be important for generating cells for therapeutic applications.Grant Funding Source: Supported by Sigma Xi, Fight For Sight, and the California Institute for Regenerative Medicine
Sex differences in the expression of MOR in areas critical for fear conditioning and extinction learning (LB557)
Authors: Morales Silva Roberto , Perez‐Torres Emily , Santini Edwin , Torres‐Reveron Annelyn 
Published: June 2021

The mu opioid receptor (MOR) has a significant role in fear conditioning and extinction learning. Communication between amygdala, periaqueductal gray (PAG) and medial prefrontal cortex (mPFC) regulates fear conditioning and extinction. Fear conditioned female rats during the metaestrus (M, low hormones) stage of the estrous cycle, showed impaired extinction while rats during the proestrus (P, high hormones) stage showed normal extinction learning. We aim to elucidate how opioid modulators alter MOR expression during fear conditioning and extinction in both sexes. Female rats on M or P stages and males rats underwent auditory fear conditioning and immediately received MOR agonist morphine, or antagonist naloxone or saline. The next day rats received extinction trials. Western blots were used to measure MOR expression. In the PAG, female rats showed no change with either treatment. In male rats morphine produced an increase in MOR expression while naloxone had an opposite effect. In the PFC, morphine during P decreased MOR expression while naloxone had no effect. Increased MOR expression was observed in the amygdala of male rats after either morphine or naloxone treatments, while in females an increase in MOR was only observed during P. Our data portraits sex differences in MOR expression in the fear circuitry, which might be responsible for the persistence of fear responses during low ovarian hormone stages.Grant Funding Source: Supported by Molecular and Genomics core RR003050, Behavioral Core Facilities MD007579 , HPD335698 Nova Southeastern University to Edwin Santini and Dinah Ramos‐Ortolaza
Screening for and Identification of Novel Glucarpidase Producing Bacteria: Cloning and molecular characterisation of novel enzymes involved in ADEPT for cancer treatment (LB581)
Authors: Alqahtani Alanood , Alyafei Afrah , Abdallah Fatma , Latiff Aishah , Groves Mathew , Domling Alex , Goda Sayed 
Published: June 2021

Antibody Directed Enzyme Prodrug Therapy (ADEPT) is a novel therapy which has already been implemented in cancer therapy to solve the problem of drug resistance and lack of tumor selectivity. Repeated cycles of (ADEPT) and the use of glucarpidase in detoxification of cytotoxic methotrexate (MTX) are desirable but are hindered by the induced human antibody response to the glucarpidase.Novel glucarpidases with unrecognized epitopes may increase the prospect of repeated cycles of ADEPT for effective cancer therapy and could also be used to remove excess MXT.The aim of this study was to isolate and characterize new glucarpidase (former name: carboxypeptidase G2) producing bacteria in order to clone and overexpress the cognate genes for Antibody Directed Enzyme Prodrug Therapy (ADEPT), and for detoxification of methotrexate (MXT) used for cancer treatment.Over five hundred soil samples were collected from different fields in Egypt and Qatar where folate rich fruit and vegetables are routinely cultivated. Screening of these samples was performed using folate as the sole carbon and/or nitrogen source. Several molecular techniques including 16S rRNA were used to characterize the isolated strains.The work led to the isolation and characterization of four new glucarpidase producing strains, The novel native enzymes have been assayed using Methotrexate as a substrate, and characterised.We successfully cloned and molecular characterized a novel glucarpidase from one of the isolated strains which could be used in the ADEPT techniques for cancer treatment that could overcome the immune response problems.Grant Funding Source: Qatar National research fund, NPRP 6 ‐ 724 ‐ 3 ‐ 183
Cyclic nucleotides down regulate PAF receptor mediated effects in newborn ovine pulmonary arterial smooth muscle: significance in oxygen therapy of persistent pulmonary hypertension of the newborn (LB605)
Authors: Hanouni Mona , Cruz Belen , Opene Michael , Narvaez Vincent , Ibe Basil 
Published: June 2021

Platelet activating factor (PAF) is implicated in the pathogenesis of persistent pulmonary hypertension of the newborn (PPHN). Baseline PAFr activity is higher in fetal ovine pulmonary artery smooth muscle cells (PASMC) and is augmented by hypoxia. In fetal PASMC we have shown evidence of crosstalk between PAFr‐mediated cell signaling and activity of cyclic nucleotides (Cnucs) via respective receptors PKG and PKA. Since these mechanisms are not well described in the newborn counterpart, we studied effects of Cnucs on PAFr protein expression in newborn PASMC. Because long term effects of oxygen therapy on newborn lung are not well understood, we included various oxygen tension study conditions. We hypothesize that PPHN results from failure of newborn lamb pulmonary system to down regulate PAFr activity or to up regulate vasodilatory Cnuc activity.PASMC harvested from ovine newborns 6‐12d old were incubated for 24h with culture media or media containing 10μM cGMP or cAMP analogs or 0.1μM of PAF in normoxia (pO2~100torr), hypoxia (pO2 <40 torr) and hyperoxia (5% CO2, O2). Prostacyclin and TxA2 release were measured and compared to release by fetal PASMC. Proliferation studies were performed under the three conditions and PAFr, PKA and PKG expression were measured.Newborn PA baseline prostacyclin level was 9‐fold greater than in fetal PA, whereas baseline production of TxA2 by newborn PA was 7‐fold less. Overall, 24h hyperoxia attenuated cell proliferation across all treatments. Cnuc treatment attenuated PAF stimulation of proliferation and decreased PAFr expression.Elevated prostacyclin production over TxA2 suggests greater cyclooxygenase activity in newborn PA postnatally. Down regulation of PAFr effects by Cnucs indicates that normal newborn PA physiology favors vasodilatory pathways to minimize PAF‐induced hypertrophy or hyperplasia. Thus failure of newborn lung to anchor down regulation of vasoconstrictors with up regulation of vasodilators leads to pathogenesis of PPHN.
Cocaine self‐administration‐induced sensitization of the dopamine system (LB621)
Authors: Calipari Erin , Jones Sara 
Published: June 2021

The ability of cocaine to bind to, and inhibit the dopamine transporter (DAT) mediates the reinforcing effects of the compound and is essential for the maintenance of self‐administration behaviors. As such, it is important to determine the changes in cocaine potency following cocaine self‐administration as they have important implications for 1.) how the DAT changes in response to repeated psychostimulant administration as well as 2.) changes in the reinforcing potential for drugs such as cocaine. Although sensitization paradigms, which result in sensitization of cocaine‐induced locomotor behavior and dopamine (DA) overflow following repeated cocaine injections, are often used as a model to study drug addiction, similar effects have been difficult to demonstrate following cocaine self‐administration. We have recently shown that intermittent access (IntA) conditions can result in enhanced cocaine potency at the DAT in the nucleus accumbens immediately following self‐administration. However, traditional sensitization paradigms often show enhanced effects following withdrawal/abstinence periods. Here we aimed to determine a time‐course of IntA‐induced DA sensitization and how a seven‐day abstinence period influences these effects. We show that while three days of IntA is not sufficient to alter evoked DA release, a seven‐day abstinence period following IntA results in augmented release. Additionally, cocaine potency is enhanced following as little as 3‐day IntA and further augmented following an abstinence period. Further, IntA plus abstinence enhanced cocaine‐induced DA release as compared to all other groups, demonstrating that following abstinence, cocaine’s ability to increase DA release and inhibit uptake at the DAT, two separate mechanisms for increasing DA levels, is augmented. Thus, the enhanced cocaine potency and cocaine‐induced release following abstinence could enhance the reinforcing efficacy and facilitate drug‐seeking for cocaine following periods of abstinence/withdrawal.Grant Funding Source: Supported by NIH grants from NIDA [R01 DA024095, R01 DA03016, R01 DA14030, P50DA006634 (SRJ), T32 DA007246 and F31 DA031533 (ESC)]
Ovariectomy regulates the production of prostanoids and the MAPK pathway in rat mesenteric arteries (LB675)
Authors: Ferrer Mercedes , Andres Monica , Crabtree Matthew , del Campo Lara , del Campo Marta , Villalobo Antonio 
Published: June 2021

Sex hormones regulate vascular function through nitric oxide (NO), thromboxane A2 (TXA2), and prostaglandin E2 (PGE2) among other factors. Since NO, TXA2 and PGE2 are known to regulate the epidermal growth factor receptor (EGFR) and downstream signaling pathways, the purpose of this study was to investigate whether the loss of ovarian function alters the production of these effectors regulating the activity of the mitogen‐activated protein kinase/extracellular‐regulated kinases 1 and 2 (MAPK‐ERK1/2), an EGFR downstream pathway. For this purpose, mesenteric arteries from control and ovariectomized Sprague‐Dawley rats (6‐months old) were used to analyze: i) the release of NO, TXA2 and PGE2; ii) the vascular reactivity to acetylcholine (Ach), the NO donor sodium nitroprusside, the TXA2‐mimetic compound U‐46619, and exogenous PGE2; and iii) the activation status of MAPK(ERK1/2). Our results show that ovariectomy: i) does not change NO release while decreases its vasodilator action; ii) increases the release and vasoconstrictor action of TXA2 and PGE2; and iii) increases the basal activity of the MAPK(ERK1/2) pathway. We conclude that activation of the EGFR/MAPK axis by these effectors could contribute to increase the proliferation rate of vascular smooth muscle cells, leading to increase peripheral resistance and hypertension.Grant Funding Source: Supported by the Fondo de Investigaciones Sanitarias (PI0011406) to MF.
Sodium Nitrite Supplementation Alters the Human Metabolome in Healthy Middle‐Aged and Older Adults (LB703)
Authors: Johnson Lawrence , de Picciotto Natalie , Cruickshank‐Quinn Charmion , Reisdorph Nichole , Giordano Tony , DeVan Allison , Seals Douglas , Sindler Amy 
Published: June 2021

Reduced nitric oxide (NO) bioavailability contributes to physiological dysfunction with aging. Oral sodium nitrite supplementation (SN) increases circulating nitrite, a precursor of NO, and improves some physiological functions in middle‐aged and older (MA/O) adults. To gain insight into the underlying molecular mechanisms, the metabolome was assessed by liquid chromatography‐mass spectrometry in plasma samples from MA/O adults before and after SN (80 mg/d capsules, TheraVasc, Inc.) (n=8, 59±2 yrs) or placebo (P, n=7, 61±3 yrs) (randomized, double blind). Of ~4,000 metabolites detected, significant changes in 37 intermediary metabolites were observed in SN vs. P groups (minimum 1.5 fold‐change, p<0.05). Several metabolites associated with a pro‐aging phenotype changed with SN, including pro‐inflammatory mediators and molecules related to sphingolipid, glycan, glycerophospholipid, fatty acyl, polyketide, and choline metabolism. Importantly, these SN‐induced changes occurred in the absence of changes in clinical characteristics including body composition and blood lipid profiles (all p>0.05). These preliminary results suggest that SN influences human plasma metabolome pathways linked to aging. Further targeted analysis will allow the identification of novel biomarkers in metabolic pathways modulated by SN and offer insight into the mechanisms by which SN improves specific domains of physiological function in MA/O adults.Grant Funding Source: Supported by NIH HL107105, RR025780, AG000279
Sodium taste elicits ATP secretion in mouse taste buds (LB71)
Authors: Huang Anthony , Wu Sandy , Roper Stephen 
Published: June 2021

Taste buds contain two types of cells that directly participate in gustatory transduction‐‐ Receptor (Type II) and Presynaptic (Type III) cells. Receptor cells respond to sweet, bitter and umami taste stimulation. Presynaptic cells have been identified with confidence as sour (acid) taste‐sensing cells. Using confocal Ca2+ imaging in a lingual slice preparation, Tomchik et al. (2007) showed that some vallate taste cells responded to salt (Na+) taste. Recently, ion channels (ENaC) believed to underlie salt taste transduction were reported to be in taste cells that were neither Receptor (Type II) or Presynaptic (Type III) cells. Yet, mechanisms of Na+ taste transduction and the identity of the cells that respond to Na+ stimulation remain controversial. Using Ca2+ imaging on isolated mouse vallate taste cells loaded with Fura 2, we demonstrate here that a subset of cells, neither Receptor nor Presynaptic cells, show Ca2+ mobilization in response to Na+ stimulation. Removing extracellular Ca2+ had no effect on responses evoked by Na+ stimulation. In marked contrast, applying 1 μM thapsigargin, a SER Ca‐ATPase inhibitor, or 10 μM U73122, a PLC blocker, eliminated Na+ responses, suggesting that Na+ stimulation triggers PLC/IP3‐mediated release of Ca2+ from intracellular stores. Next, we used CHO cells expressing P2X receptors as biosensors to monitor ATP release from taste buds and isolated taste cells. Na+ stimulation elicited robust biosensor responses that were blocked by suramin, confirming that Na+ stimulation elicits ATP secretion from taste buds/cells. Carbenoxolone (5 μM) or probenecid (250 μM), blockers of pannexin1 hemichannels, reversibly blocked Na‐evoked ATP secretion. Our data indicate that salt taste stimulus triggers a dedicated population of taste cells to secrete ATP via pannexin1hemmichannels.Grant Funding Source: Supported by SIUSOM and NIH/NIDCD DC007630
TRAF6 is critical for Satellite cell function and skeletal muscle regeneration upon injury (LB815)
Authors: Hindi Sajedah , Kumar Ashok 
Published: June 2021

Skeletal muscle satellite cells are a subpopulation of muscle‐stem cells responsible for carrying out muscle regenerative events. Paired box‐protein Pax7 has been shown to be indispensable for driving proper regeneration as well as supplementing the reservoir of satellite‐cell pool. However, signaling mechanisms that regulate the expression of Pax7 remain less understood. In the present study, we demonstrate that TNF receptor‐associated factor 6 (TRAF6) positively regulates the expression of Pax7 in vivo and in vitro. Deletion of TRAF6 under a tamoxifen‐induced Pax7 promoter led to profound muscle regeneration defects following BaCl2‐mediated injury. Regenerating muscle of tamoxifen‐administered mice failed to up‐regulate Pax7 and displayed a significant reduction in Myf5 transcript levels with no effect on MyoD, myogenin, Mhc3 and Mhc4 compared with vehicle‐administered mice. Similarly, cultured muscle progenitors lacking TRAF6 also displayed a significant reduction in Pax7 expression limiting their proliferative capacity. Deletion of TRAF6 had no effect on myotube formation when cells where plated at high density. Collectively, our study reveals an unprecedented requirement of TRAF6 in regulating Pax7 function in adult skeletal muscle.Grant Funding Source: Supported by NIH R01AR059810
Autophagy Induction in AC‐16 Human Cardiomyocytes: Effects of Resveratrol and Rapamycin (LB835)
Authors: Balagopal Gauthami , Kim Jae‐Sung , Leeuwenburgh Christiaan 
Published: June 2021

Mitochondrial dysfunction and increased oxidative stress contribute to the deterioration of cardiomyocyte and heart function with advancing age. Autophagy is the process by which damaged mitochondria are removed from cells. Increasing autophagy may help reverse some detrimental effects of aging on the heart. We tested the hypothesis that resveratrol, a plant polyphenol, and rapamycin, an immunosuppressant, will induce autophagy in the AC‐16 human cardiomyocytes (AC‐16 HC) independently and in combination will produce synergistic and/or additive effects.AC‐16 HC cells were incubated for 24 hours with resveratrol (50uM), rapamycin (1uM) and a 1:1 combination of the two treatments. Induction of autophagy was assessed by protein concentration and western blot analysis using the autophagy indicator LC3B antibody. ImageLab densitometry software was used to assess the band intensity.Treatment of the cells with the respective concentrations of resveratrol and rapamycin resulted in the up‐regulation of autophagy in the AC‐16 HC cells. The combination approach showed a modest upregualtion, but there was no significant synergistic effect.Taken together, resveratrol and rapamycin independently and in combination have up‐regulated autophagy in the AC‐16 HC cells. The dose response of resveratrol and rampamycin on the induction of autophagy and the optimum functional or physiological concentration required for in‐vivo changes remain to be determined. The optimum exposure time coursemust also be determined to optimize the duration of increased autophagy flux. Future studies are needed to optimize and fully understand the role of resveratrol and rampamycin through autophagy in human cardiomyocytes. Grant Funding Source: Supported by: HHMI‐Science for Life Program and NIH/NIA P30 AG028740
Vasopressin induces apical aquaporin‐2 trafficking via mal2‐mediated transcytosis in renal collecting duct cells (LB840)
Authors: Chen Pei‐Yu , Yu Ming‐Jiun 
Published: June 2021

In kidney collecting duct cells, vasopressin (AVP) triggers redistribution of aquaporin‐2 (AQP2) water channel protein from intracellular vesicles to the apical plasma membrane to regulate renal water excretion. In our previous proteomic study, we identified Mal2, a lipid‐raft membrane protein known to mediate apical membrane protein trafficking via transcytosis. This study examined roles of Mal2 in AVP‐induced apical AQP2 trafficking via transcytosis. Time course immunofluorescence showed staining of AQP2 in the apical as well as the lateral plasma membrane of the mpkCCD cells after AVP stimulation. The lateral AQP2 staining disappeared gradually with concomitant appearance of apical AQP2 staining, suggesting both direct and indirect mechanisms for apical AQP2 targeting upon AVP stimulation. When the lateral membrane was fixed with tannic acid, a portion of AQP2 accumulated in the lateral space of the cells in response to AVP or forskolin, consistent with an indirect apical AQP2 targeting mechanism via transcytosis upon AVP or forskolin stimulation. The AVP or forskolin‐induced lateral AQP2 staining in the presence of tannic acid was completely absent in the cells lacking Mal2. Our data are compatible with a role of Mal2 in AVP‐induced indirect apical AQP2 targeting via the transcytotic mechanism.Grant Funding Source: Supported by 101‐2320‐B‐002‐038‐MY3, NSC TAIWAN