bluuebunny/crossref_metadata_2025_split huggingface dataset:
PVN‐projecting NTS Neurons is Involved in Sympathetic Long‐Term Facilitation After Acute Intermittent Optogenetic Stimulation
Authors: Paundralingga Obed , Shimoura Caroline Gusson , Cunningham Joseph Thomas 
Published: April 2020

Caudal Nucleus Tractus Solitarius (NTS) neurons receive inputs from cardiorespiratory visceral afferents and send projections to the paraventricular nucleus of the hypothalamus (PVN). Acute intermittent optogenetic (AIO) stimulation of caudal NTS induces sympathetic long‐term facilitation (LTF), a progressive increase of sympathetic nerve activity. The contribution of PVN‐projecting NTS neurons to sympathetic LTF is unknown. This study used an intersectional viral approach to test the effects of acute intermittent optogenetic (AIO) stimulation of PVN‐projecting NTS neurons in sympathetic LTF. Adult male Sprague‐Dawley rats were anesthetized with isoflurane and injected rAAV2/Ef1a‐DIO‐hchR2(H134R)‐mCherry in the NTS and a retrograde AAV9.hSyn.HI.eGFP‐cre.WPRE.SV40 in the PVN four week prior to LTF experiments. This method was used to induce the CRE dependent expression of channel rhodopsin in PVN‐projecting NTS neurons. To measure LTF, we recorded splanchnic sympathetic nerve activity (SSNA), arterial pressure, and heart rate (HR) in anesthetized (urethane (800mg/kg) and α‐chloralose (80 mg/kg) cocktail delivered by intraperitoneal injection), paralyzed with gallamine triethiodide (20 mg/ml, 0.25 ml/h, iv) and mechanically ventilated adult rats. The caudal NTS was stimulated to induce AIO with a train of 10 stimuli each consisting of pulse of light of 10‐ms duration delivered at 20 Hz for 1 min every 6 min. Twenty minutes after AIO, SSNA amplitude increased by 25.36 ± 24.16% (n=3), and mean arterial pressure (MAP) increased by 7.43 ± 12.88 mmHg (n=3). During AIO induction, the acute SSNA and MAP varied with some rats demonstrating increases (n=2) or decreases (n=1) of SSNA, MAP, and HR. In other animals (n=3) where AIO decreases SSNA (‐58.96 ± 36.18%) and MAP (‐14.37 ± 8.57 mmHg), we tested the possible contribution of the baroreceptors in these decreases by bilateral muscimol (1–10 pmol/100nL) injections in the Caudal Ventrolateral Medulla (CVLM). AIO stimulation after inhibition of CVLM caused an increase in SSNA (1.293 ± 17.65%) and smaller decrease in MAP (−2.46± 6.2 mmHg). These results suggest that the splanchnic sympathetic LTF induced by acute intermittent stimulation of NTS neurons might be mediated by the PVN‐projecting neurons, although the acute responses may be excitatory or inhibitory. Further study is needed to clarify the mechanisms responsible for the differences in the cardiovascular and SSNA responses to acute, single optogenetic stimulation of PVN‐projecting NTS neurons. Despite the observed differences in the acute responses, the results indicate that sympathetic LTF can be induced by the optogenetic activation of NTS neurons that project to PVN.Support or Funding InformationNIH Grant P01 HL088052
Graduate Education Trends in the Anatomical Sciences: Are anatomy PhDs nearing extinction or adapting to change?
Authors: Kaza Niroop , Wilson Adam B. , Singpurwala Darius J. , Brooks William S. 
Published: April 2020

IntroductionHuman anatomy is the cornerstone of health sciences education, yet literature over the past 50 years has demonstrated a severe and continued shortage of anatomy educators. Many have postulated that the factors responsible for the dwindling numbers of anatomists are multiple.AimThis study (1) assessed trends in doctorates awarded in anatomy and related fields within the U.S. over the past 48 years and (2) evaluated the modern graduate education‐training paradigm in the anatomical sciences.MethodsRetrospective data relative to the anatomical sciences were compiled from the National Science Foundation (NSF) Survey of Earned Doctorates (SED), an annual census of research doctorates within the U.S. The number of doctorates in anatomy and related fields and the total number of doctorates by sex and race/ethnicity were plotted for trend analysis. Additionally, the number of currently active PhD anatomy training programs within U.S. allopathic & osteopathic medical schools were assessed. Curricula and major characteristics of each active program where evaluated through website searches and phone interviews with program directors. The IRB at UAB did not qualify this study as human subjects research.ResultsSince 1969, the number of PhDs awarded in Anatomy has declined, on average, by 3.3 graduates per year (Max = 163, 1982; Min = 9, 2017). Conversely, doctorates in Cell Biology, Developmental Biology, and Neuroscience have observed marked growth over the past 40 years at rates of 8.0, 4.3, and 31.3 graduates per year, respectively. Males have historically dominated the anatomical fields, but more females received doctorates than males from 2009–2016. The proportion of PhDs in the anatomical sciences awarded to underrepresented minorities (URMs) has steadily increased with an all‐time high of 11.1% in 2015, but still lags behind Science & Engineering (14.1%). The proportion of graduates entering employment directly after graduation declined from a high of 55% in 1969 to the current rate of 21% and is inversely proportional to the number of graduates entering additional training positions, which is 78% currently. As of August 2019, 21 active doctoral programs in anatomy were identified from among 187 institutions affiliated with a medical school. PhD programs were categorized as either Anatomy Education (N=8), Classic Anatomy (N=8), or Anthropology/Evolutionary Anatomy (N=5) programs based on coursework in anatomical science disciplines and education/teaching‐based curricula.Discussion & ConclusionDeclining numbers of anatomy PhD graduates and misalignment between training expectations and job market needs has contributed significantly to the shortage of anatomists. A concerted effort by all stakeholders to increase the diversity and number of anatomy educators is necessary to ensure the future health of the profession.
Biophysical Folding Sensors for the Detection of Misfolded Proteins by Hsp70
Authors: Karunnayake Chamithi , Page Richard C. 
Published: April 2020

Chaperoned ubiquitination complexes play a defining role in responding to chemical and pathological stresses and an understanding of how these complexes are regulated on a molecular level offers the opportunity to identify new avenues of treatment for a wide range of diseases. Hsp70 participates in chaperoned ubiquitination by recruiting and attempting to refold misfolded protein “clients” in an ATP‐dependent process. Misfolded clients interact with a binding cleft within the substrate binding domain (SBD) formed by a beta‐sheet subdomain (SBD‐beta) and helical lid subdomain. Understanding how Hsp70 recruits clients and positions clients relative to the E3 ligase CHIP is critical for understanding the core interactions between CHIP and Hsp70 that dictate the outcome of chaperoned ubiquitination. Herein we describe a set of folding sensors representing a spectrum of folded and unfolded states. When utilized in ubiquitination reactions this system provides a tunable folded/unfolded client with an Hsp70‐SBD binding site that provides control over the distance from the model unfolded client to the E2~Ub active site bound to the E3 ligase CHIP, thereby providing a measure of how folding state dictates degree of ubiquitination.Support or Funding InformationNIGMS R35 GM128595
Mitochondrial Protein Hyperacetylation in Rodents with Doxorubicin‐Induced Cardiac Dysfunction
Authors: Tomczyk Mateusz M. , Cheung Kyle G. , Xiang Bo , Agarwal Prasoon , Kereliuk Stephanie M. , Wilkins John A. , Dolinsky Vernon W. 
Published: April 2020

ObjectiveDoxorubicin (DOX) is an effective chemotherapeutic but has dose‐dependent cardiotoxic effects that limits its use in pediatric patients. Previous studies in our lab showed that DOX decreases expression of the mitochondrial lysine deacetylase SIRT3 and mitochondrial phospholipid cardiolipin (CL) in the mouse heart. We hypothesize that DOX impairs cardiac function as a consequence of reduced SIRT3 expression resulting in increased acetylation of mitochondrial proteins involved in cardiac energy and oxidative stress homeostasis.MethodsC57BL6 mice were given DOX (8.0mg/kg body weight) or saline control injections for 4 weeks. Transthoracic echocardiography was performed on all mice (n=10 per group) and parameters of cardiac structure, systolic and diastolic function were measured. Cardiac mitochondria were isolated from saline and DOX mice and an anti‐acetylated lysine antibody was used to enrich for tryptic digested peptides containing acetylated lysines followed by mass spectroscopy analysis (n=6).ResultsDOX treated mice exhibit decreased left ventricular posterior wall thickness (P<0.05), increased intraventricular relaxation time and reduced ejection fraction, compared to controls (P<0.05). Quantitative PCR of cardiolipin biosynthesis genes revealed Ptpmt1 and Crls1 gene expression was reduced by half with DOX compared to controls (p<0.01). In DOX treated mice, we observed an enrichment of cardiac mitochondrial acetylated peptides of proteins involved in metabolic, CL remodelling processes and oxidative stress resistance (eg. ATP5F1A, TFEα, SOD2, P<0.05, >2‐fold increase). MitoSOX staining of human induced pluripotent stem cell derived cardiomyocytes revealed an increase (2.5‐fold, p<0.001) in reactive oxygen species production with DOX treatment which was attenuated with adenoviral SIRT3 overexpression.ConclusionAlterations to the mitochondrial acetylome may be responsible for DOX‐induced cardiac dysfunction in mice. SIRT3 overexpression attenuates production of reactive oxygen species in human derived cardiomyocytes and SIRT3 may prevent cardiotoxic effects of DOX.
Self‐powered bioluminescence in a marine tube worm
Authors: De Meulenaere Evelien , Puzzanghera Christina , Deheyn Dimitri D. 
Published: April 2020

An increasing number of luminous organisms are being found in the ocean, presenting opportunities to discover new biochemical reactions associated with light production, and thus new bioreporter systems for biotechnology and biomedicine. The marine parchment tube worm Chaetopterus sp. is of particular interest because it secretes a luminous mucus that glows for a long time (hours, possibly days), although in seawater and isolated from any continuous metabolic support. This contrasts with the short flashes, or the bacterial, metabolically driven glow that are more typical for naturally observed bioluminescence. We have recently found that the light production in this mucus is activated by Fe2+ and that the long‐lasting glow may therefore be supported by the proven presence of the iron storage protein ferritin.We have also observed a temporary increase in the bioluminescence after exposure of the mucus to blue light specifically, which is capable of triggering Fe2+ release from ferritin through a photo‐reduction process. Since the mucus bioluminesces in the blue region of visible light, we are investigating whether it could be powering its own light production.Support or Funding InformationThis research is supported by the Air Force Office of Scientific Research (grant # FA955017 0189 to D. D. Deheyn).
Investigation of Cytochrome P450 2J2 Ligand Binding Modes
Authors: Motl Nicole , Scott Emily E 
Published: April 2020

Human cytochrome P450 2J2 (CYP2J2) plays a multifaceted role in cardioprotection and xenobiotic metabolism. CYP2J2 oxidizes endogenous polyunsaturated fatty acids P450s, including arachidonic acid and linoleic acid, to various epoxyeicosatrienoic acids (EETs). These EETs are implicated in vasodilation and protection against hypoxia‐reperfusion, but also promote tumor growth and metastasis, where inhibition may be a clinical advantage. Recently, it has been reported that CYP2J2 is inhibited by anti‐hypertensive drugs including manidipine, azelnidipine, and telmisartan. In its drug metabolism role, CYP2J2 metabolizes numerous xenobiotics including anti‐histamine and anti‐cancer agents. While kinetic and metabolic parameters for many of these endogenous and xenobiotic compounds have been reported, it remains to be elucidated how such a wide variety of ligands are accommodated by the CYP2J2 active site. This study determined the binding modes of a catalog of CYP2J2 ligands, including substrates, inhibitors, xenobiotics, and a broad panel of azole containing compounds for the purpose of better understanding their interactions with CYP2J2. The binding modes and affinities were determined by monitoring the spectral shift of the P450 heme Soret absorbance, which is indicative of perturbation of the heme environment. Such spectral shift indicated varying responses: displacement of the heme‐coordinated water (a type‐I binding mode), compound coordination to the heme iron (a type‐II binding mode), or no spectral shift. As the latter result can be consistent with no binding or a binding in the active site in a way that does not perturb the heme center, these latter compounds were investigated via a luminescence‐based inhibition assay to verify their interaction with CYP2J2. Overall, this study aides in improving our understating of different ligand scaffolds that are compatible with CYP2J2 binding to support the development of therapeutics and inhibitors of CYP2J2.Support or Funding InformationNational Institutes of Health Grant R37 GM076343
Acute Intermittent Hypoxia Results in Baroreflex Resetting to Higher Blood Pressures in Young Men but Not Young Women
Authors: Limberg Jacqueline K. , Jacob Dain W. , Baker Sarah E. , Ott Elizabeth P. , Ivie Clayton L. , Harper Jennifer L. , Acevedo Camila Manrique 
Published: April 2020

BackgroundRepetitive hypoxic apneas, similar to what is observed in sleep apnea, result in resetting of the sympathetic baroreflex to higher blood pressures (BP) in humans. This resetting of the baroreflex is associated with increased BP in preclinical models of sleep apnea (chronic intermittent hypoxia, CIH); however, the majority of our understanding comes from men. There are data to suggest female rats exposed to CIH do not develop high BP. With this, we sought to examine potential sex‐related differences in the effect of intermittent hypoxia (IH) on the sympathetic baroreflex. We hypothesized young men would exhibit resetting of the baroreflex to higher BP following IH, with no change in young women.MethodsBP and muscle sympathetic nerve activity (MSNA) were assessed in 17 men (30±1 yrs) and 10 women (28±1 yrs) during quiet normoxic rest prior to and following 30‐min of experimental IH [30‐s hypoxic exposures (0.05 FiO2) alternating with 90‐s room air breathing] resulting in 15 hypoxic events (91.7±0.5 % SpO2). Sympathetic baroreflex sensitivity was evaluated using the modified Oxford technique before and after 30‐min of IH.ResultsDiastolic BP increased in men following IH (71±2 to 74±2 mmHg; p<0.01) with no change in women (75±3 to 75±3 mmHg; p=0.97). Sympathetic baroreflex sensitivity did not change following IH in either group (Men: −1.8±0.2 to −1.9±0.3; Women: −1.6±0.3 to −2.1± 0.5 bursts/100 heart beats/mmHg; Main effect of IH, p=0.14; Interaction of group and IH, p=0.28).ConclusionAcute IH resets the sympathetic baroreflex to higher BP in young men, with no change in young women. Baroreflex sensitivity slopes were unchanged in either group following IH. Our results support sex differences in the effect of IH on sympathetic control of BP and show that young women are protected from the BP‐raising effects of IH. These data enhance our understanding of sex‐specific mechanisms that may contribute to the development of hypertension in sleep apnea.Support or Funding InformationNIH HL130339, Mayo Clinic Center for Biomedical Discovery
Protein splicing and tryptophan fluorescence of salt‐dependent inteins from Haloquadratum walsbyi
Authors: Yalala Vaishnavi , Chavez Angel , Lynch Abigeal K. , Mills Kenneth V. 
Published: April 2020

Protein splicing is a self‐catalyzed reaction by which an intein, an intervening polypeptide, catalyzes its own removal from exteins, the flanking polypeptides, as well as ligation of the exteins. We are studying the inteins that interrupt a cell division control protein in the halophilic archaea Haloquadratum walsbyi. Halophiles like H. walsbyi are able to survive in water with high concentrations of sodium chloride. H. walsbyi has four inteins that interrupt the cdc21 gene, and we have studied the two inteins (Hwa cdc21A and Hwa cdc21D) that lack an intervening homing endonuclease domain. We are studying the splicing activity under various salt concentrations for the inteins both separately and when expressed as part of a single fusion protein. Hwa cdc21A splices optimally at 2.5 M NaCl, while Hwa cdc21D splices optimally at 1.5 M NaCl. We also are using native Trp fluorescence to study the influence of salt concentration on the proper folding of the inteins.Support or Funding InformationThis work was supported by NSF grant MCB‐1517138, a Henry Dreyfus Teacher‐Scholar Award, and NIH Grant 1R15GM132817‐01.
Misoprostol attenuates hypoxia‐induced neonatal cardiomyocyte proliferation through Bnip3 and perinuclear calcium signaling
Authors: Gordon Joseph W. , Martens Matthew D. , Field Jared T. , Chapman Donald , Day Chelsea , Keijzer Richard , West Adrian R. , Ivanco Tammy L. 
Published: April 2020

Systemic hypoxia resulting from preterm birth, altered lung development, and cyanotic congenital heart disease is known to impede the regulatory and developmental pathways in the neonatal heart. Although the molecular mechanisms are still unknown, metabolic stressors resulting from neonatal hypoxia drive aberrant glycolysis and cardiomyocyte proliferation, which may be initially adaptive, but ultimately can program the heart for early life pathological remodeling. Recent evidence suggests the prostaglandin E1 analogue misoprostol is cytoprotective in the hypoxia‐exposed neonatal heart by promoting the alternative splicing of Bnip3, resulting in the generation of an isoform lacking the third exon (Bnip3ΔExon3) known as small Nip (sNip). Using a rodent model of neonatal hypoxia, in combination with rat primary ventricular neonatal cardiomyocytes and H9c2 cells, we sought to determine if misoprostol can prevent cardiomyocyte proliferation and what the key molecular mechanisms might be in this pathway. In cultured myocytes, molecular markers and fluorescent indicators of proliferation, such as cylcin‐D1 and PHH3, were significantly elevated with hypoxia exposure and were attenuated with misoprostol treatment (p<0.01). Moreover, hypoxia reduced mitochondrial respiration and increased cytosolic production of NADH, which were restored by misoprostol treatment. In addition, both sNip and misoprostol promoted cardiomyocyte maturation by increasing nuclear calcium accumulation, and drove the nuclear localization of NFATc3 (p<0.01). Gene expression studies identified that misoprostol and sNip reduced the expression of the proliferative MEF2C‐myocardin‐BMP10 pathway, while favoring expression of the cardiac maturation factors BMP2 and MEF2A (p<0.01). Furthermore, an siRNA targeting sNip restored hypoxia‐induced proliferation in misoprostol‐treated cells. Finally, we observed a significant increase in left ventricular nuclei number and PHH3 staining in hypoxia‐exposed rat pups, which were also attenuated with misoprostol treatment (p<0.05). These findings suggest a novel mechanism by which misoprostol promotes Bnip3 alternative splicing and myocyte maturation, thereby blocking hypoxia‐induced neonatal cardiomyocyte proliferation.Support or Funding InformationThis work is supported by the Natural Science and Engineering Research Council of Canada, the Heart and Stroke Foundation of Canada, and the Children’s Hospital Research Institute of Manitoba.
Structural studies of human Fis1 reveals a dynamic region important for Drp1 recruitment and mitochondrial fission
Authors: Hill Blake , Enger John , Harwig Megan , Ihenacho Ugochukwu , Widlansky Michael 
Published: April 2020

Fission protein 1 (Fis1) and dynamin‐related protein 1 (Drp1) were initially described as being evolutionarily conserved for mitochondrial fission, yet the role of Fis1 in human fission is unclear and disputed by many. In budding yeast where Fis1 helps to recruit Drp1 from the cytoplasm to mitochondria for fission, an N‐terminal “arm” of Fis1 is required for function. The yeast Fis1 arm interacts intramolecularly with a conserved surface that governs in vitro interactions with yeast Drp1. In human Fis1, NMR and x‐ray structures show a different arm conformation and its importance for human Drp1 recruitment is unknown. Here, we use MD simulations and comparisons to experimental NMR chemical shifts to show the human Fis1 arm can adopt an intramolecular conformation akin to that observed with the yeast molecule. This finding is further supported through intrinsic tryptophan fluorescence and NMR experiments on human Fis1 with and without the arm. Using NMR, the human Fis1 arm is also observed to be sensitive to environmental changes. The importance of these findings are revealed in cellular studies where the removal of the hFis1 arm reduces Drp1 recruitment and mitochondrial fission suggesting an important role for Fis1 in human mitochondrial fission.Support or Funding InformationThis project was supported by the following National Institutes of Health grants: R01GM067180 (to RBH), and R01HL128240 (to MEW).