bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mechanistic Insights Into RNA Splicing As A Trigger For Arrhythmogenic Right Ventricular Cardiomyopathy
Authors: Bradford William H. , Liang Yan , Mataraarachchi Nirosh , Gu Yusu , Dalton Nancy D. , Peterson Kirk L. , Sheikh Farah 
Published: April 2020

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a genetic cardiac disease that causes sudden death in young adults and athletes. ARVC is termed a “disease of the desmosome” as 40% of mutations in ARVC patients are found in the cardiac desmosomal components, with plakophilin‐2 (PKP2) being the most frequently mutated desmosomal gene in ARVC. Evidence from humans suggests that altered RNA splicing may be a critical mechanism through which PKP2 patient genetics drive ARVC. Studies have suggested that approximately one third of all human disease‐causing mutations are a result of defects in RNA splicing; highlighting the importance of testing the relevance of this mechanism in human ARVC. However, there are no models and limited mechanistic insights into how mutations in RNA splicing impact ARVC disease pathogenesis. We hypothesize that RNA splicing mutations in PKP2 are sufficient to recapitulate classic disease features associated with human ARVC and can provide valuable mechanistic insight into how altered RNA splicing impacts disease. Through CRISPR‐Cas9 we generated a novel knock‐in mouse model harboring a human equivalent PKP2 mutation (IVS10‐1 G>C) that impacts RNA splicing and is sufficient to recapitulate all classic ARVC disease features. PKP2 homozygous mutant (PKP2 Hom) mice are viable at birth yet display adult hallmarks of ARVC including ventricular arrhythmias, right and left ventricular dysfunction, and fibro‐fatty replacement of myocardium leading to sudden death. RNA and sequencing analyses of exons spanning the PKP2 mutation site reveals a larger PKP2 transcript that retains a 54 base pair portion of the intronic sequence in PKP2 Hom hearts. However, RNA analysis of exons outside the PKP2 mutation site reveals PKP2 RNA transcripts at similar levels to wild type PKP2, suggesting that total RNA levels are not impacted by the mutation. Instead, we show that PKP2 Hom hearts express a higher molecular weight mutant PKP2 protein in the absence of endogenous PKP2. Analysis of PKP2 Hom neonatal hearts and cardiomyocytes reveal early desmosomal protein loss coupled with a susceptibility to baseline arrhythmias in the absence of overt cardiac disease features at this stage (e.g., no changes in heart weight/body weight ratios, no upregulation of cardiac stress and fibrotic markers). We provide a novel mouse model that highlights the sufficiency and molecular consequences of a PKP2 RNA splicing mutation that triggers all of the classic early and adult onset disease features associated with human ARVC.Support or Funding InformationNational Science Foundation (Graduate Research Fellowship Program), National Institutes of Health (NIH R01, HL142251‐A1)
Neuromodulation of afferent fibers for Postural Orthostatic Trachycardia Syndrome (POTS)
Authors: Rhoads Marie , Harper Rebecca K. , Snodgrass Danielle , Yan-Go Frisca , Sauerland Eberhardt K. , Harper Ronald 
Published: April 2020

Postural Orthostatic Trachycardia Syndrome, or POTS, is characterized by low arterial blood pressure, inappropriate cardiovascular and breathing responses to tilt, dizziness and syncope to sudden standing, and a range of other autonomic signs, including thermoregulatory issues in the periphery and frequent manifestations of migraine pain. Interventions are few, and typically include volume repletion by saline and enhanced fluid intake. In the course of evaluating neuromodulatory procedures for migraine pain (n=62), several subjects showed classic POTS symptoms, many of which spontaneously resolved with the intervention for migraine. Non‐invasive neuromodulation of two different areas, one being the auditory canal containing mandibular nerve V3, cranial nerves VII, IX, and X and cervical nerves C2 and C3; and the other being regional stimulation of V1 and V2, together with C2 and C3 sensory areas, caused significant declines in dizziness, hand cooling, and other POTS symptoms. The neuromodulatory device stimulating the auditory canal consisted of bilateral silicon impressions of the auditory canals, each containing a mechanical vibratory device which reached a range of cranial nerves, and especially the IXth (glossopharyngeal) nerve to the carotid baroreceptors. The other neuromodulatory device applied vibrating signals to the cutaneous surface over areas innervated by V1 and V2. The trials consisted of a 10‐min control period, followed by 25‐min stimulation, and finished with a 10‐min control period. Continuous thoracic wall movement, pulse oximetry, and ECG were collected, and beat‐by‐beat blood pressure values were inferred from the pulse transit time of the cardiac R wave and oximetry signal. All procedures were approved by the UCLA Institutional Review Board. Subjects who began the trial with low blood pressure increased systolic and diastolic pressures. In two subjects, repeated trials benefitted critical signs (hand warming, dizziness, syncope to sudden movement). These findings suggest that repeated activation of afferent input to baroreceptors can elicit both short‐ and long‐term effects on blood pressure control. The means by which this “learning" or adaptation of cardiovascular regulatory systems remain unknown, but raise the possibility that a low‐cost, non‐invasive intervention can provide another means to restore effective blood pressure control in a syndrome with few options for treatment approaches.Support or Funding InformationThis research was supported in part by the Fidelity Charitable Nancy Adams and Scott Schoen Fund and the Kraig and Linda Kupiec Family Trust.
Assessment of CYP2A6 and CYP2C19 Genetic Variation on Cotinine‐N‐Oxide Formation and their Potential Effect on the Nicotine Metabolic Ratio (NMR)
Authors: Perez Paramo Yadira X. , Watson Christy J.W. , Chen Gang , Natesan Senthil , Lazarus Philip 
Published: April 2020

SignificanceThe nicotine metabolic ratio (NMR) of 3‐hydroxycotinine (3HC)/cotinine in smokers is a determinant of CYP2A6 activity and is widely used in clinical practice to help select smoking cessation pharmacotherapy. The major mode of metabolism of nicotine is via the formation of cotinine and subsequently to 3HC by the enzyme CYP2A6. Cotinine is also metabolized to cotinine‐N‐glucuronide and to cotinine‐N‐oxide (COX), accounting for ~7 and 5% of urinary nicotine metabolites, respectively, in smokers. The goal of the present study was to investigate the enzymes that catalyze COX formation and determine whether genetic variation in these enzymes may affect this pathway.MethodsWe used specific inhibitors of the major hepatic cytochrome P450 (CYP) enzymes including CYPs 1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, and 3A4 in cotinine‐N‐oxidation reactions using pooled human liver microsomes (HLM). COX formation was monitored by UPLC‐MS/MS, and enzyme kinetic analysis was performed using microsomal fractions from CYP‐overexpressing HEK293 cell lines. Additionally, genotype‐phenotype analysis was performed for a panel of 113 HLM specimens.ResultsInhibition of COX formation was only observed when using inhibitors of CYPs 2A6, 2B6, 2C19, 2E1, and 3A4. CYPs 2A6 and 2C19 exhibited similar oxidation activity against cotinine with Vmax/KM values of 4.4, and 4.2 nL•min−1•mg−1, respectively; the Vmax/KM value for the 2B6 isoform was of 2.2 nL•min−1•mg−1. The activity observed for both CYPs 2E1 and 3A4 were low, with KM’s greater than 10 mM. We measured the levels of COX formation in 113 HLM specimens and genotyped variants in CYPs 2A6 (*2, *9, and *14), 2B6 (*2, *5, *9, rs4803419 and rs8109525) and 2C19 (*2 and *17) using qPCR. An association between decreased levels of COX formation and CYP variants in the 2C19 (up to 35% reduction) and 2A6 (up to 38% reduction) isoforms was observed. No statistically significant correlation was observed with variants in the 2B6 isoform.ConclusionsThese results demonstrate that genetic variants in CYPs 2A6 and 2C19 are associated with decreased COX formation, potentially affecting the levels of free cotinine and consequently the calculated NMR in smokers and clinical practices surrounding smoking cessation therapies.Support or Funding InformationThis work is supported by grants from NIH, National Institutes of Environmental Health Sciences (grant R01‐ES025460; to P. Lazarus), the Fulbright‐Garcia Robles Program and CONACyT I2T2 (to Y. Perez‐Paramo), Health Sciences and Services Authority of Spokane, Washington (grant WSU002292 to College of Pharmacy and Pharmaceutical Sciences, Washington State University)
Hyperoxic Lung Injury in Newborn Mice is Abrogated by Neutralizing mAb 2A11 to Gastrin‐Releasing Peptide (GRP)
Authors: Sunday Mary E. , Cuttitta Frank , Cambey Colette 
Published: April 2020

RationaleGastrin‐releasing peptide (GRP) promotes fetal lung development in mice and baboons. Postnatally, GRP levels in lung normally drop, but rise in newborns who later develop bronchopulmonary dysplasia (BPD), chronic lung disease of newborns, with interstital fibrosis and arrested alveolarization. In premature baboons, blocking anti‐GRP mAb2A11 abrogated BPD. Rodent BPD models could help to clarify mechanisms by which GRP mediates BPD. Mice are born at the canalicular stage of lung development, ~24–28 wks human gestation, when BPD peaks.MethodsNewborn (NB) timed‐pregnant Swiss Webster mice were treated with continuous 75% O2 and twice weekly injections of mAb MOPC21 (negative control IgG1) or mAb2A11. On day 18, right lungs were inflation‐fixed and paraffin embedded. Myofibroblasts were immunostained with alkaline‐phosphatase‐conjugated mAb1A4 (Sigma‐Aldrich), developed with Vector Red. Eight‐ten random lung 20x images were captured per mouse. Using ImageJ, the area‐percent of SMA+ alveolar tissue was determined as: (area of SMA+ alveolae)/(alveolar tissue) ×100%. Mean linear intercept (MLI) was used to determine alveolar size with a fixed grid with lines totaling 1‐mm: 1/(#air‐tissue interfaces)×1000 = mean distance between two air‐tissue interfaces in microns. Data are shown as mean ±SEM, with significance at p < 0.05 (one‐way t‐test).ResultsNormoxic (NO) mice ± MOPC or 2A11, had area‐percent SMA+ alveoli unchanged (5.1–6.3 ±0.5–1.4). Hyperoxia+MOPC led to 3‐fold increased SMA (15.6±2%). Hyperoxia+2A11 reduced SMA to 10.3±1.2%; P<0.01 comparing hyperoxia+2A11 vs. hyperoxia+MOPC. 2A11 had no effect on MLI with either normoxia or hyperoxia, possibly due to insufficient time. We performed Affymetrix cDNA analysis of lung RNA from hyperoxic mice +2A11 or MOPC for 18d. HO+2A11 vs. HO+MOPC altered expression of >1000 genes (FC>10, P<0.05), many potentially protective vs. lung injury/fibrosis.
Increased gene expression included: nucleoredoxin, an antioxidant related to thioredoxin (FC x78); surfactant protein A (FC x140); Tom20, mitochondrial transport protein that reduces lung injury (x74), and multiple other mitochondrial genes (>20x).
Reduced gene expression included: Adam10, profibrotic; amphiregulin, elevated in BPD; amyloid beta A4, toxic fragment; laminin B1, which usually declines by birth; Tgfb1i4, profibrotic; and Stat1, profibrotic.ConclusionsAfter 18d exposure to 75% O2, newborn mice have >3‐fold more interstitial myofibroblasts that are significantly reduced by mAb2A11. Many pulmonary genes are up‐ or down‐regulated by mAb2A11 consistent with a multigenic downstream effect on reducing interstitial fibrosis via interacting signaling pathways. Mice can provide a useful model for carrying out preclinical investigations.Support or Funding InformationThis work was supported by NIH grants #R01‐HL44984 (M.E.S.), U01‐HL52638 (M.E.S.), and a Scientific Research Bequest from the Jenny Lillian Semans Koortbojian Trust Fund
Cerebro, the Virtual Lab ‐ a mobile neuroanatomy game that is both fun and educational
Authors: Marais Olivia , Tourigny Katherine , Lellis-Santos Camilo , Burke Kim , Pucci Niccolo , Riisager Oliver , Jeon Sean , Zhang Weiyi , Lastovikova Julia , Kopak Jeannette , Krebs Claudia 
Published: April 2020

Neuroanatomy remains one of the more complex disciplines in anatomical sciences education. It requires both a deep understanding of 3D anatomical relationships within the brain and a translation of that knowledge to 2D slices through the brain – an important skill, which enables the interpretation of clinical imaging studies of the brain. An additional challenge for learners is to understand the anatomy of the main ascending and descending tracts through the brainstem as well as the location of the crossing to the contralateral side for these pathways. Mastery of neuroanatomy requires both a conceptual understanding and practice in identifying structures with confidence on slices through the brainstem and the forebrain.Gamification of learning is a concept that aims to harness the fun and rewarding qualities of a game to make learning appear effortless.In this project we put together a multidisciplinary team of neuroanatomy content experts, game & software designers, and artists to develop a neuroanatomy game that is easy to use and yields learning results for students. The target audience was identified and characterized and the user experience and user interface were designed for this target audience. Iterative, agile design cycles with multiple sessions of user testing resulted in a final product that serves the needs of undergraduate students seeking a high yield return on their time investment for learning. A narrative arc was developed for the game and chapters with three levels from beginner to expert were created based on the learning objectives of typical neuroanatomy lab sessions. Principles of design thinking and iterative software design were applied in the creation of this game so that the final product would meet both the learning objectives of the course and the objective to keep the game fun and competitive.The game was designed to fit into the existing learning ecosystem for neuroanatomy, which includes interactive online resources, 3D experiences, and lab sessions that are integrated with clinical sciences.
The Role of Magnesium Biodegradable Material in Peripheral Nerve Repair
Authors: Abu-Niaaj Lubna F. , Harris Greg , Pixley Sarah 
Published: April 2020

Peripheral nerve injuries can greatly diminish a patient’s quality of life. The use of permanent metallic material implants, including stainless steel, introduce a toxicity risk, due to release of allergenic particles upon chronic use, The significant difference in hardness versus bodily tissues is also unsatisfactory for long‐time placement. Biodegradable biomaterial scaffolds are an innovative alternative therapeutic methodology to enhance tissue repair. Magnesium is a biodegradable metal which has received significant interest recently because it is both closer in hardness to tissues and it biodegrades safely, releasing Mg ions, playing many beneficial roles in the human body. Mg ions regulate ion channel opening, DNA replication, protein synthesis and are a crucial cofactor for >300 enzymes. Serum deficiencies of Mg result in general inflammation and are implicated in disorders such as Parkinson’s and Alzheimer’s diseases. Thus, Mg metal biodegradable implants have great promise as a new generation of tissue repair materials. In vivo studies in the Pixley lab have shown that peripheral nerve axons regenerating across an injury gap, react positively to the presence of the Mg, including giving indications that they may be drawn towards the magnesium wire within the tissue. The long term goal is to understand cellular reactions to magnesium (Mg) metal, which will aid the development of a clinical application in peripheral nerve repair.ObjectivesToward investigating the underlying cellular mechanisms, this study sought to 1) design in vitro cell culture systems to determine the effects of selected types of Mg metal wires being tested for use in clinical practice. Then, 2) we sought to determine the in vitro responses to Mg metal by two cell types that are important in peripheral nerve repair: Schwann cells and the class of immune cells, macrophages, that are the first to attach to any implant material and that also play a large role in nerve repair.Materials and MethodsCellular interactions with the selected Mg wires were studied in vitro using immunostaining and phagocytosis assays. The cells analyzed were mouse Schwannoma cells (RT4‐D6P2T) and a mouse macrophage cell line (RAW264.7).ResultsSchwannoma cells showed aggregates around the edge of the immobilized Mg wires, though, no significant migration towards/away from Mg wires, but macrophages appear to react to the Mg wires. The research is continuing to determine the mechanism of cellular interactions with the Mg metal, and whether these interactions would influence the quality of the peripheral nerve repair.Support or Funding InformationFunding was provided by the NSF ERC RMB EEC‐0812348
Studies of Continuous Lorcaserin Plus Buprenorphine in Rat Fentanyl Self‐Administration
Authors: Merritt Christina , Stutz Sonja , Fox Robert , Anastasio Noelle , Moeller F. Gerard , Cunningham Kathryn 
Published: April 2020

AimsOpioid overdose deaths and the rise in problematic opioid use patterns that indicate the development of opioid use disorder (OUD) have reached crisis levels in the United States. Medication‐assisted treatment (MAT) with therapeutics such as the partial μ‐opioid receptor agonist buprenorphine mitigate withdrawal, reduce mortality, and reduce intake of abused opioids (e.g., fentanyl, heroin). Still, the opioid epidemic has crystalized the need to identify adjunctive therapeutics to substantially improve the odds of successful recovery. In the present study, we tested the hypothesis that treatment with the FDA‐approved, 5‐HT2CR agonist lorcaserin (Belviq®), previously shown to reduce opioid intake in the preclinical self‐administration (SA) model, may serve as an efficacious adjunct to suppress opioid intake and/or prevent escalation of opioid use after cessation of MAT. We employed osmotic minipumps to chronically administer buprenorphine and/or lorcaserin to achieve stable levels and mimic a clinical dosing regimen.MethodsMale Sprague‐Dawley rats (n=34) were implanted with osmotic minipumps [sham, lorcaserin (1 mg/kg/day), buprenorphine (3 mg/kg/day), or the combination] following stable acquisition of fentanyl self‐administration. Rats resumed fentanyl self‐administration for 10 days and were then monitored for an additional ten days after removal of osmotic pumps. During this phase, the efficacy of acute lorcaserin (1 mg/kg) to impact fentanyl intake was evaluated.ResultsChronic treatment with buprenorphine or the combination, but not lorcaserin alone, attenuated fentanyl intake relative to sham‐operated rats [F3,30=15.3, p<0.05]. Acute treatment with lorcaserin attenuated fentanyl intake relative to saline‐treated rats [F1,12=51.81, p<0.05].ConclusionsThe efficacy of acute lorcaserin to suppress opioid intake extended to the illicit opioid fentanyl. Further, chronic buprenorphine, with or without lorcaserin, resulted in sustained inhibition of fentanyl intake. Taken together, these studies provide support for further interrogation of the efficacy or lorcaserin as a viable adjunct pharmacotherapy for MAT.Support or Funding InformationSupported by the NIDA U54 DA038999
Cardiac TRPV1‐Afferent Signaling Promotes Arrhythmogenic Ventricular Remodeling after Myocardial Infarction
Authors: Yoshie Koji , Rajendran Pradeep A. , Massoud Louis , Mistry Janki , Swid M. Amer , Wu Xiaohui A. , Sallam Tamer A. , Zhang Rui , Goldhaber Joshua I. , Salavatian Siamak A. , Ajijola Olujimi A. 
Published: April 2020

Chronic sympathoexcitation is implicated in ventricular arrhythmogenesis (VAs) following myocardial infarction (MI), but the critical neural pathways involved are not well understood. Cardiac adrenergic function is partly regulated by sympathetic afferent reflexes, transduced by spinal afferent fibers expressing the TRPV1 channel. The role of chronic TRPV1 afferent signaling in VAs is not known. We hypothesized that persistent TRPV1 afferent neurotransmission promotes VAs after MI. Using epicardial Resiniferatoxin (RTX) to deplete cardiac TRPV1‐expressing fibers, we dissected the role of this neural circuit in VAs after chronic MI in a porcine model. We examined the underlying mechanisms using molecular approaches, immunohistochemistry, in vitro and in vivo cardiac electrophysiology, and simultaneous cardio‐neural mapping. Epicardial RTX depleted cardiac TRPV1 afferent fibers and abolished functional responses to TRPV1 agonists. Ventricular tachycardia/fibrillation (VT/VF) was readily inducible in MI subjects by programmed electrical stimulation or cesium chloride administration, however, TRPV1 afferent depletion prevented VT/VF induced by either method. Mechanistically, TRPV1 afferent depletion neither altered cardiomyocyte action potentials and calcium transients; nor the expression of ion channels and calcium handling proteins. However, it attenuated fibrosis and mitigated electrical instability in the scar‐border zone. In vivo recordings of cardiovascular‐related stellate ganglion neurons (SGNs) revealed that MI enhances SGN function and disrupts integrated neural processing. Depleting TRPV1 afferents normalized these processes. Taken together, these data indicate that after MI, TRPV1 afferent‐induced adrenergic dysfunction promotes fibrosis, adverse cardiac remodeling, and worsens border zone electrical heterogeneity, resulting in electrically unstable ventricular myocardium. We propose targeting TRPV1‐expressing afferent to reduce VT/VF following MI.Support or Funding InformationNIH/NHLBI DP2 OD024323‐01
Amplified Protein Kinase C Signaling in Alzheimer's Disease
Authors: Lorden Gema , Wosniak Jacob M. , Dozier Lara E. , Patrick Gentry N. , Gonzalez David J. , Roberts Amanda J. , Newton Alexandra C. 
Published: April 2020

Protein kinase C (PKC) isozymes are tightly regulated kinases that transduce signals from receptor‐mediated hydrolysis of membrane phospholipids. Whereas loss‐of‐function mutations in PKC are associated with cancer, germline mutations that enhance the activity of one isozyme, PKCα, are associated with Alzheimer’s Disease (AD). Here we show that introducing a highly penetrant AD‐associated variant of PKCα (M489V; increases catalytic activity by 30%) into a mouse model accelerates brain pathogenesis and results in a cognitive defect compared to wild‐type mice. Specifically, behavioral studies reveal a cognitive impairment in mice harboring this mutation compared to WT mice at 12 months of age, as assessed by performance on a Barnes Maze test, which measures spatial learning and memory. Additionally, the presence of the PKCα mutation in an AD mouse model overexpressing the amyloid precursor protein with the Swedish mutation, significantly exacerbated the cognitive decline as early as 6 months of age. To better understand the molecular basis for the cognitive defect, we analyzed the phosphoproteome and synapse morphology of mice harboring the PKCα M489V mutation. Analysis of the brain phosphoproteome of PKCα M489V mice vs wild‐type mice identified a variety of substrates with altered phosphorylation including known substrates of PKC involved in synaptic depression. Phosphorylation of one such protein, MARCKS, was increased both in mice harboring the PKCα mutation as well as human brains from patients with AD. Analysis of spine density revealed a modest (approximately 10%) but highly significant reduction in the number of spines per micron in the homozygous M489V mice compared to the wild‐type mice. Reduced spine density is consistent with the enhanced phosphorylation of MARCKS observed in brains from the M489V mice. In summary, this work reveals that a single amino acid change in the catalytic domain of PKCα is sufficient to cause increased phosphorylation of PKC substrates, neurite degeneration, and cognitive decline. These data strongly support inhibition of PKCα as a potential therapeutic target in AD.Support or Funding InformationCure Alzheimer's Fund