bluuebunny/crossref_metadata_2025_split huggingface dataset:
CRISPR‐mediated deletion of a regulatory element downstream of the EDN1 gene increases expression of a long non‐coding RNA, EDN1‐AS
Authors: Douma Lauren G , Masten Sarah , Solocinski Kristen , Barral Dominique , Wingo Charles S , Cain Brian D , Gumz Michelle L 
Published: June 2021

Endothelin‐1 (ET‐1) is a peptide hormone that functions as a vasoconstrictor in the vasculature whereas in the collecting duct of the kidney it exerts blood pressure‐lowering effects via natriuretic actions. Aberrant ET‐1 signaling is associated with several pathological states including hypertension and chronic kidney disease. ET‐1 expression is regulated largely through transcriptional control of the gene that encodes ET‐1, Edn1. We recently discovered a long, non‐coding RNA (lncRNA) that appears to be antisense to the Edn1 gene, called EDN1‐AS. Because EDN1‐AS represents a potential novel mechanism to regulate ET‐1 expression, we examined the molecular regulation of EDN1‐AS expression and action. Using the ENCODE database and the UCSC genome browser, we located a putative glucocorticoid receptor response (GR) element upstream of the predicted transcription start site for EDN1‐AS. Preliminary chromatin immunoprecipitation results indicated that both GR and the circadian clock protein PER1 interact with this region of DNA in human renal proximal tubule HK‐2 cells. Using CRISPR, we deleted the region containing this potential regulatory element. Two homozygous deletion clones were identified following selection with puromycin (A11 and H3). An HK‐2 clone that went through the same selection procedure but was negative for the deletion served as the control cell cline. This deletion resulted in a significant increase in the expression of EDN1‐AS (2 fold, n=3–4, p<0.05). This increase in EDN1‐AS expression was associated with increased secretion of ET‐1 peptide from HK‐2 cells (KO H3=1.68 pg/mg vs. control line F1=1.53pg/mg, n=4, p= 0.004). Phenotypic characterization of these CRISPR clones revealed a difference in cell growth rates. Using a standard growth assay, we determined that the A11 clone exhibited a 3 fold increase in growth over 8 days compared to control cells (n=4, P<0.0001) and the H3 clone exhibited a 2 fold increase (n=4, p<0.001). These results support a role for EDN1‐AS as a novel regulatory mechanism of ET‐1 expression and cellular proliferation.Support or Funding InformationNIH T32DK104721 (LGD), NIH R21AG052861 (MLG), The Gatorade Trust through the UF Department of MedicineThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Modeling the 20‐HETE Binding Site on GPR75
Authors: Garcia Victor G , Gilani Ankit , Agostinucci Kevin , Adebesin Adeniyi , Falck John R , Schwartzman Michal L 
Published: June 2021

Recently, the cytochrome P450‐derived eicosanoid 20‐Hydroxyeicosatetraenoic acid (20‐HETE) was shown to activate the orphan receptor GPR75. The pairing of 20‐HETE and GPR75 activates a signaling pathway that elicits increases in endothelial angiotensin converting enzyme (ACE) and promotes the development of hypertension. Little is known about the critical binding sites/pockets across GPR75 involved in interacting with 20‐HETE that ultimately leads to receptor activation. Computational docking analysis of GPR75 uncovered a putative ligand binding site located between transmembranes five and six on GPR75 (SiteScore: 1.182 and Dscore: 1.303). Independent analyses of ligand‐receptor docking using 20‐HETE placed the lipid in close proximity to the putative binding pocket (Docking Score: −3.229, Glide Emodel: −29.045). Additional modeling of GPR75 alongside another proposed ligand, the chemokine CCL5 (RANTES), revealed a potential protein‐protein interaction model occurring at the top (extracellular face) between the first and seventh transmembranes of GPR75. This distant site suggests the possibility of distinct ligand‐receptor interactions that promote unique signaling cascades. In order to establish a functional assay to further investigate the significance of the proposed 20‐HETE binding site/pocket, we have adapted the PRESTO‐Tango system. This system allows for the quantification of ligand‐dependent β‐arresstin2 recruitment, a common hallmark of G‐protein coupled receptor activation, biased agonism and receptor inactivation/internalization. Preliminary studies using the GPR75‐Tango construct demonstrated a robust signal wherein 20‐HETE stimulates the recruitment of β‐arresstin2 after GPR75 activation (EC50=3μM). Further studies will be conducted to assess the significance of residues located within the putative binding pocket with respect to 20‐HETE‐mediated activation of GPR75.Support or Funding InformationThis study was supported by National Institute of Health grants HL39793(M.L. Schwartzman), HL39793‐01S1 (V. Garcia) and the Robert A. Welch Foundation (I‐0011, J.R. Falck).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A human‐origin probiotics cocktail exhibit cardio‐protective effects independent of GLP‐1 receptor signaling
Authors: Nagpal Ravinder , Mishra Sidharth P , Wang Shaohua , Tallant Ann , Varagic Jasmina , Kitzman Dalane , Yadav Hariom 
Published: June 2021

The emerging role of the gut microbiome in cardiovascular diseases is posited to be mediated by the microbial metabolites, some of which yield beneficial effects while others may accelerate the heart diseases. The beneficial metabolites like short‐chain fatty acids, particularly butyrate, exhibit several beneficial effects on metabolic diseases including cardiovascular functions. We have previously demonstrated that butyrate promotes the production of glucagon‐like protein‐1 (GLP‐1). GLP‐1 and its therapeutic agonists are known to promote cardiovascular functions. Herein, we investigated the role of systemic deletion of GLP‐1 receptor (using GLP1R KO) on cardiac functions in 16 weeks high‐fat diet (HFD)‐fed obese mice and assessed the impact of feeding a butyrate‐producing probiotic cocktail (consisting 5 Lactobacillus and 5 Enterococcus strains isolated from infant's gut) on cardiac functions, using the non‐invasive high‐frequency ultrasound echocardiography system (VEVO, VisualSonics, Toronto, Ontario, Canada). GLP1R‐KO mice show reduced cardiac output and stroke volume, and higher ejection fraction and fractional shortening of the left ventricle compared GLP1R‐WT mice. In addition, the GLP1R‐KO mice also exhibited significantly reduced systolic and diastolic diameter and volume, along with higher ratio of aortic acceleration and ejection time. Interestingly, feeding of probiotic cocktail protected from such dysfunctions in GLP‐KO mice, and also all these measures remain unchanged in GLP‐WT mice with and without probiotics treatments. No significant differences in body weight and diet intake was observed, however, glucose and insulin tolerance tests were improved in both probiotic‐fed groups. Results indicate that GLP1‐receptor depletion causes cardiovascular dysfunctions and feeding of the human‐origin probiotic cocktail ameliorates such dysfunctions, and probiotics effects are independent of GLP‐1 receptor signaling. Further analyses to better understand the mechanisms of actions of probiotics cocktail contributing these effects, are directed to establish a new cardioprotective probiotic biotherapy in obesity and type 2 diabetes.Support or Funding InformationWe thank the support provide by Center of Cardiovascular Sciences Center, Wake Forest School of Medicine, and the Department of Defense funding PR170446, as well funds and services provided from Center for Diabetes, Obesity and Metabolism, Wake Forest Baptist Medical Center and National Center for Advancing Translational Sciences (NCATS), National Institutes of Health funded Wake Forest Clinical and Translational Science Institute (WF CTSI) through Grant Award Number UL1TR001420.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Left and right superior cervical ganglionectomy affect differently the cardiorespiratory patterning in the in situ arterially perfused brainstem preparation of the rat
Authors: Kola Gjinovefa , Coffee Gregory A , Getsy Paulina M , Hsieh Yee‐Hsee , Dick Thomas E , Dutschmann Mathias , Lewis Stephen J 
Published: June 2021

The superior cervical ganglia project to the nucleus tractus solitarii, modulate vasomotor tone and buffer cardio‐respiratory responses to chemoreceptor activation. Here, we report the differential effect of left versus right superior cervical ganglionectomy (SCGx) on cardiorespiratory activity in the in situ arterially perfused brainstem preparation. For these studies, we used male Sprague‐Dawley rat pups (P25). Four days after removal of either the left or right superior cervical ganglia (n=20, 10 in each side) or sham surgery (n=10), we recorded left and right phrenic (PNA) and cervical vagal (VNA) nerve activities, and monitored perfusion pressure. ‘Fictive’ cardio‐respiratory patterns were recorded for 10 min (baseline) and then in response to peripheral chemoreceptor activation (intra‐arterial injection of sodium cyanide, 100 μL of a 0.03% solution). At baseline and during the response to chemoreceptor stimulation, the left and right PNA and VNA were similar for all groups. At baseline, PNA burst frequency and amplitude were similar in the left‐, right‐ SCGx and sham groups but VNA was greatest for the left SGX group and the perfusion pressure had to be increased to maintain ‘fictive’ respiration for right‐SCGx group. During the chemoreceptor stimulation, the fR response was greatest for the sham group and greater for the right‐ than the left‐SCGx group. In contrast, the PNA amplitude response was greatest for right‐SCGx group and comparable for the left‐SCGx and sham groups. Despite the increase in baseline VNA, the increase in VNA response to chemo‐stimulation was similar for the left‐, right‐SCGx and sham groups. Finally, the left‐SCGx group had smallest increase in PP during chemoreceptor stimulation. These data indicate that the left and right superior cervical ganglia modulate vasomotor tone respiratory patterning differentially at rest and during chemoreceptor reflexes.Support or Funding InformationNIH SPARC 1OT2OD023860‐02 SJ LewisThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
WormBeat: A Strategy to Evaluate Pharyngeal Pumping Variability in the Nematode C. elegans
Authors: Harris Michael B , Shilleh Ahmad , Phillips Derek , Taylor Barbara E , Berlemont Renaud 
Published: June 2021

In C. elegans, the feeding behavior of pharyngeal pumping occurs spontaneously in proportion to metabolism and in response to stimuli. Age‐ and health‐related changes in tissue morphology and function correlate with declines in the frequency of pumping. As such, pumping frequency is an established index of C. elegans “health”, and pumping changes illustrate and quantify functional decline. Timing of pharyngeal pumping is controlled by two pairs of pharyngeal motor neurons (MC and M3). Each pump cycle corresponds to the propagation of a single pharyngeal muscle action potential, initiated by MC and terminated by M3. We currently model this system with MC acting as an oscillator and synaptically linked to the pharyngeal muscle. Pumping can change by influences on the pace and regularity of the oscillator and/or of the fidelity of pharyngeal muscle response to neuronal inputs.Traditionally, this phenomenon has been reported as an occurrence frequency, describing the number of pumping events observed per unit time over a period of observation. Variation in pumping frequency within and between subjects, and the relative coarseness of this measure may interfere with the resolution of subtle treatment effects and ignores both the mechanisms for resulting effects as well as the potential significance of pumping variability.We use the recently developed ScreenChip system (NemaMetrix) to detect individual pump events, and have developed an algorithm (WormBeat) to normalize variation between subjects and distinguish variability associated with MC neuron pace and neuromuscular fidelity of the pharynx. We present the validity of our algorithm and demonstrate enhanced resolution in detecting differences in pumping behavior using simulations representing ranges and distinct sources of biologically relevant variability. This presentation describes the theory and practice of the algorithm. The use of this analysis strategy on an experimental dataset is described in a companion presentation.Support or Funding InformationWork reported in this publication was supported by the National Heart Lung and Blood Institute and National Institute of General Medical Sciences of the National Institutes of Health under Award Numbers 1R15HL126105, 1SC2GM112570, UL1GM118979, TL4GM118980, and RL5GM118978. The work is solely the responsibility of the authors and does not necessarily represent the official view of the National Institutes of Health or any other funding body.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hypoxia Induces Emergence of Adult‐like Respiratory Motor Patterns in Pre‐metamorphic Bullfrog Brainstem Preparations
Authors: Janes Tara , Kinkead Richard 
Published: June 2021

Vertebrate respiratory development involves the gradual transformation of primal respiratory behaviours into mature forms with specific, measurable characteristics. Yet, the impacts of environmental factors on respiratory maturation remain enigmatic. In pre‐metamorphic tadpoles of Lithobates catesbeianus (American bullfrog), gill ventilation predominates, while lung breathing is rarely observed. Following metamorphosis, lung breathing emerges as the dominant respiratory behaviour. Since accelerated metamorphosis (including respiratory development) constitutes an adaptive response to deterioration of the milieu, we hypothesized that hypoxia is sufficient to accelerate maturation of the motor command for lung breathing. Specifically, we predicted that hypoxia would elicit an “adult‐like” motor pattern in pre‐metamorphic tadpoles, including high amplitude lung bursts occurring episodically with a “ramping up” of burst amplitude within episodes. Using early‐stage tadpoles (Taylor‐Kollros: VI–IX), we recorded respiratory motor activity from cranial nerves V, VII and X in isolated brainstems before, during, and 2h after exposure to 15 min of moderate hypoxia (PwO2 range: 45–76 Torr). To test for long‐term effects brainstems were then incubated overnight in oxygenated artificial cerebral spinal fluid at 17°C and motor activity recorded 24h post‐hypoxia. In line with our predictions, lung motor activity in brainstems exposed to hypoxia exhibited characteristics of adult breathing while that of time‐matched controls resembled typical tadpoles. Lung burst amplitude was increased relative to baseline (+90%) at 2h post‐hypoxia. Additionally, hypoxia increased the incidence of lung burst episodes more than 300%, with a concurrent increase in the number of breaths per episode from an average of 2.2 at baseline to 3.3 at 2h post‐hypoxia (controls: 1.9 at 2h). The “ramping up” of lung burst amplitude during episodes was variable over time but gradually increased in prevalence above control levels by 24h post‐hypoxia. These data demonstrate that central hypoxia induces the emergence of “adult‐like” lung motor patterns in early pre‐metamorphic tadpoles.Support or Funding InformationThis work was supported by a Fonds de Recherche du Québec ‐ Santé (FRQS) Collaborative Scholarship awarded to T. Janes and an NSERC Discovery Grant awarded to R. Kinkead.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Energetic cost of intestinal ion transport pathways in marine teleosts
Authors: Grosell Martin 
Published: June 2021

A number of studies have attempted to determine the metabolic cost of osmoregulation in teleost fish by measuring standard or resting metabolic rates with the anticipation that metabolic demands would be lowest at isosmotic salinities. However, there is no consensus among these studies and estimates of osmoregulatory costs based on whole animal metabolic rates range from a 2–30% of resting metabolic rates. Both interspecies differences and difficulties in determining true standard metabolic rates not influenced by spontaneous activity/anxiety/stress contributes to the lack of consensus in field. Isolated osmoregulatory tissues have rarely been employed even though they avoid problems of organismal responses. The few studies using isolated tissues have focused on the gills and estimate cost in the range of 2–4%, a range that matches theoretical considerations. Such low metabolic demands are challenging to capture by measurements of intact animals. To date, no studies have attempted to quantify the metabolic demand of osmoregulatory processes by the marine teleost intestine despite its central role in osmoregulation. Multiple parallel Cl− uptake pathways with different energetic efficiency are present in the marine teleost intestinal epithelium and suggests osmoregulatory metabolic demands comparable to that of the gill tissue. The least expensive Cl− uptake pathway is via NKCC2 consuming 0.17 ATP/Cl− while anion exchange via SLC26a6 is consuming ≥ 1 ATP/Clyet, marine teleost fish employ the SLC26a6 pathway extensively. The theoretical intestinal contribution to marine teleost osmoregulation and the seemingly energetically expensive strategy of employing anion exchange will be presented.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Chemical and Physical Properties of Gill Plasma Membranes Are Altered in Response to Thermal Acclimation in the Antarctic Notothenioid Fish Notothenia coriiceps
Authors: Biederman Amanda M , Crockett Elizabeth L 
Published: June 2021

Antarctic notothenioid fishes are highly stenothermal as a result of living under relatively stable conditions for millions of years. In a warming climate, preservation of membrane structure and function at elevated body temperatures will be critical to the survival of these species. Many ectothermic organisms can respond to thermal variation by altering membrane compositions to preserve membrane structure (i.e., homeoviscous adaptation). It is largely unknown to what extent Antarctic notothenioids possess the capacity to remodel their biological membranes in response to thermal change. We studied effects of thermal acclimation on physical (fluidity, permeability, oxygen solubility) and biochemical (lipid composition) properties of plasma membranes from the gills of an Antarctic notothenioid, Notothenia coriiceps. Animals were acclimated to 0 and 5°C for a minimum of 6 weeks. Plasma membranes were prepared from the gill, and membrane fluidity was measured from 0 to 30°C. Cholesterol contents and phospholipid compositions were analyzed in membranes, and permeability to water was measured in gills at both 0 and 4°C. Oxygen partition coefficients were determined in membrane samples to assess oxygen solubility. Membrane fluidity was reduced in the 5°C‐acclimated group, compared with animals held at 0°C (P<0.0001) and exhibited perfect (100%) homeoviscous efficacy. Membranes from 5°C‐acclimated fish also contained 1.2‐fold greater cholesterol (P<0.01) and 1.1‐fold greater long‐chain fatty acids (i.e., fatty acids with at least 20 carbon molecules per chain) (P<0.05). Water permeability was reduced 1.5‐fold with 5°C acclimation (P<0.05), exhibiting near‐perfect (96%) homeostatic efficacy. In contrast, oxygen solubility of membrane lipids was unchanged. The biochemical alterations with acclimation are in accord with changes in both fluidity and permeability, suggesting lipid restructuring occurred to conserve membrane structure at elevated temperature. Taken together, these results provide evidence of homeoviscous adaptation, and conservation of permeability, with temperature acclimation in gill plasma membranes from an Antarctic notothenioid.Support or Funding InformationSupported by NSF ANT 1341602 and the Ohio University Student Enhancement Award fund.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Ontogeny of renin gene expression and juxtaglomerular apparatus in birds
Authors: Hoy Jessica , Nishimura Hiroko , Yaoita Eishin , Gomez Robert Ariel , Sequiera‐Lopez Maria Luisa S 
Published: June 2021

The renin‐angiotensin system (RAS) plays a crucial role in the regulation of blood pressure, fluid electrolyte homeostasis, and vascular development. Renin or a renin‐like enzyme evolved in primitive vertebrates and is conserved throughout the vertebrate phylogeny. The ontogenic development of renin, however, has not been well understood. We aimed to determine expression patterns and relative abundance of renin mRNA in pre‐ and postnatal chickens (Gallus gallus, White Leghorn breed) using primers designed by the NCBI PrimerBlast tool. On embryonic day 13 (E13) embryos show renal tubules, undifferentiated mesenchymal structures, and a small number of developing glomeruli. Maturing glomeruli are seen in post‐hatch day 4 (D4) and day 30 (D30) kidneys, indicating that nephrogenic areas still exist in 4‐week‐old kidneys. In E13 embryos, renin mRNA measured by qPCR in the adrenal glands is equivalent to the expression in the kidneys, whereas in post‐hatch D4 and D30 maturing chicks, renal renin expressions became much higher (10–15‐hold) and relative renin expression became lower in the liver and adrenals. Also, renin expression becomes clearly visible by in situ hybridization in the juxtaglomerular (JG) area in D4 and D30, but not in E13 embryos. The results suggest that in chickens, renin evolved in both renal (presumably in arteries and arterioles) and extrarenal organs at an early stage of ontogeny and, with maturation, is more localized in the JG area of the kidney. Clear JG structures are not morphologically detectable in E13 embryos, but are visible in 4‐week‐old chicks, supporting this concept. The early appearance of the chicken angiotensin type 1 receptor agrees with renin ontogeny.Support or Funding InformationNIH DK091330 and DK096373 to M.L.S.S.L.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.