bluuebunny/crossref_metadata_2025_split huggingface dataset:
Acute IP Furosemide Increases Medullary PO2 in The Diabetic Rat Kidney
Authors: o neill julie , Fasching Angelica , Patinha Daniela , Pihl Liselotte , Franzen Stephanie , Palm Fredrik 
Published: June 2021

We recently observed medullary hypoxia during SGLT (sodium glucose linked transport) inhibition in the diabetic rat kidney. Underlying mechanisms are unclear but may reflect compensatory increases in sodium reabsorption in distal nephron segments, such as the medullary thick ascending limb (mTAL), where sodium transport is less efficient. To test this hypothesis, we investigated the impact of NKCC (sodium/potassium/chloride) co‐transporter inhibition on renal oxygen metabolism in the diabetic rat kidney.Diabetes was induced by intraperitoneal (IP) STZ (50mg/kg) injection in Sprague Dawley rats (n=11), 2 weeks prior to experimentation. Animals were surgically prepared for intravenous infusion of [3H]Inulin and [14C]PAH, the IP delivery of furosemide (NKCC inhibitor; 3mg/Kg) and the measurement of mean arterial pressure (MAP), left kidney function and medullary (M) PO2 (Clark electrode). Data± SEM was analyzed using a two‐way RM ANOVA. Baseline MAP, transported sodium (TNa), total kidney oxygen consumption (QO2), and MPO2 was 107±4 mmHg, 189±23 µmol/min, 23±3 µmol/min and 26±2 mmHg, respectively. IP furosemide, had no effect on MAP, reduced TNa and QO2 to 58±7 µmol/min and 8±2 µmol/min respectively and increased MPO2 to 36±2 mmHg (all P<0.05).Sodium transport via NKCC is clearly a major consumer of oxygen in the diabetic rat kidney. Moreover, the furosemide‐induced increase in MPO2 is in vast contrast to the reduction in medullary PO2, previously observed during SGLT inhibition. These data provide evidence that medullary hypoxia during SGLT inhibition in the diabetic rat kidney results from the diversion of TNa to the mTAL where sodium transport is less efficient.
Identification of Phosphate Regulated Transcripts in Ileum and Renal Proximal Tubules
Authors: Knöpfel Thomas , Kampik Nicole , Hernando Nati , Rubio Aliaga Isabel , Biber Jürg , Wagner Carsten 
Published: June 2021

Phosphate (Pi) is a vital mineral for every cell in order to maintain energy metabolism, cell signaling and structure. The kidney and intestine are two major organs in maintaining Pi homeostasis by their role in absorbing Pi from the diet and by the reabsorption of filtered Pi. Apical uptake of Pi into intestinal and proximal tubular epithelial cells is well characterized, whereas basolateral efflux mechanisms are poorly understood. The amount of Na‐Pi cotransporters (members of the Slc34 family) expressed at the apical membrane is regulated by changes in dietary Pi intake through mechanisms, which are not fully understood. The aim of this project is to identify transcripts in ileum and renal proximal tubules regulated by dietary Pi. Identified genes may be involved in the transcellular transport of Pi like the unknown basolateral transport pathway or contribute to adaption of apical Pi uptake and metabolism. C57BL/6 male mice (8 weeks old) were fed a high (1.2%) or low (0.1%) Pi diet for 3 days. Epithelial scrapings were prepared from ileum and micro‐dissected renal proximal tubules were collected. Renal and intestinal adaption was confirmed by qPCR for known Pi regulated transcripts. RNA was then analysed using the RNA sequencing service provided by the Functional Genomics Center Zurich. Comparison of the transcriptome of low and high dietary Pi conditions revealed 2029 genes to be differently (at least 2 fold changes) expressed in ileum and 327 in renal proximal tubules. Regulation of selected transcripts was confirmed by qPCR. Pathway analysis is currently being performed and selected candidates for basolateral Pi transporters will be expressed in Xenopus laevis oocytes.Funded by the Swiss National Science Foundation.
Renal Localization and Characterization of Monocarboxylate Transporter Family Member Slc16a14
Authors: Knöpfel Thomas , Atanassoff Alexander , Kampik Nicole , Hernando Nati , Wagner Carsten , Biber Jürg 
Published: June 2021

Monocarboxylate transporter 14 (MCT14) is an orphan member of the solute carrier family 16 for monocarboxylates, thyroid hormone and aromatic amino acids. The kidney shows the highest abundance of MCT14 mRNA, however, the renal localization, function and regulation are poorly understood. The aim was to localize MCT14 in murine kidney and address the question of its possible function. To this end an antibody was raised against a peptide located at the extracellular side of MCT14 and its specifity was confirmed by peptide protection and heterologous expression of MCT14 in Xenopus laevis oocytes.Colocalization experiments with markers for different nephron segments indicate that MCT14 is localized to the apical membrane of the thick ascending limb of the loop of Henle. Members of the Slc16 family showed changes in protein and mRNA abundance in kidney due to metabolic acidosis. MCT14 itself showed an upregulation under acidotic conditions in both mRNA and protein levels, but the expression pattern in the kidney was not altered.Uptake experiments in Xenopus laevis oocytes expressing MCT14 suggest that MCT14 does not transport lactate or pyruvate unlike the classical monocarboxylate transporters of the Slc16 family. To identify the substrate of MCT14 we used a mass spectrometry based approach in order to cover many substrates. Therefore Xenopus laevis oocytes expressing MCT14 and corresponding negative controls were incubated in mouse serum. Intracellular metabolites were then analyzed by mass spectrometry in the Functional Genomics Center Zurich. Major substrate candidates identified by mass spectrometry are currently being examined to confirm whether they are substrates of MCT14.Funded by the Swiss National Science Foundation.
CFTR and SLC5A8 Associate in Thyroid Epithelial Cells
Authors: Fong Peying , Li Yonghai , Ganta Suhasini , Chen Grace 
Published: June 2021

The cystic fibrosis transmembrane conductance regulator, CFTR, is expressed at the apical membranes of many epithelial cells, where it mediates cAMP‐stimulated anion secretion. In thyroid, CFTR may indirectly and/or directly furnish iodide for hormone synthesis. CFTR functionally interacts with many membrane transporters via PSD‐Discs large homologous‐ZO‐1 (PDZ) domain‐containing scaffolding proteins, thereby forming macromolecular transport complexes. SLC5A8 is an electrogenic, sodium‐coupled monocarboxylate transporter that also is expressed at thyrocyte apical membranes. CFTR and SLC5A8 contain the identical PDZ domain interaction motif; preliminary studies utilizing overexpression systems indicate a role for PDZ adaptor proteins in linking CFTR and SLC5A8. The present studies use the Fisher rat thyroid cell line, FRTL‐5, to test the hypothesis that PDZ proteins facilitate CFTR/SLC5A8 interactions in an endogenously expressing system. Expression of CFTR (~150 kDa), SLC5A8 (~68 kDa) and NHERF‐1 (~50 kDa) in total FRTL‐5 lysates was confirmed by immunoblot analysis using anti‐CFTR (M3A7 and Cystic Fibrosis Foundation Therapeutics (CFFT) 660), anti‐SLC5A8 (Abcam) and anti‐NHERF‐1 (Alomone) antibodies. Anti‐SLC5A8‐immunoprecipitated (IPd) fractions of FRTL‐5 lysates reacted with both CFFT 660 and anti‐NHERF‐1. Fractions IPd using anti‐CFTR (M3A7) were probed with anti‐SLC5A8 and anti‐NHERF‐1. M3A7‐IPd fractions showed positive reactivity with anti‐SLC5A8, but NHERF‐1 was not detected. These data suggest that although CFTR and SLC5A8 associate in FRTL‐5 cells, the participation of NHERF‐1 is not required. Support: NIH/NIGMS Award 1R15GM101674‐01A1 (to PF); Office of the Director, NIH Award Number T35OD010979.
Gastric iodide secretion; possible role in antimicrobial oxidant hypoiodous acid production
Authors: Akiba Yasutada , Kaji Izumi , Said Hyder , Narimatsu Kazuyuki , Kaunitz Jonathan 
Published: June 2021

Ingested iodide is absorbed in the small intestine, followed by uptake into the thyroid follicles, whereas iodide in the systemic circulation is secreted via the salivary glands and the stomach in an entero‐gastric loop. Since the gastric mucosa generates luminal H2O2 via dual oxidase, and hypoiodous acid (HOI) is an antimicrobial oxidant with strong antifungal and antiviral activity, we hypothesized that gastric iodide secretion (GIS) might be related to the production of luminal HOI in the stomach.We measured GIS in the perfused stomach in vivo using flow‐through iodide sensitive electrode. Sodium iodide (NaI) was given IV or intraduodenally (Id). Transmucosal iodide movement was measured in an Ussing chamber.Immunostaining localized the Na+/I‐ symporter (NIS) to the basolateral membranes in fundic surface cells and pyloric gland cells. Mucosal NIS mRNA expression was highest in the fundus and antrum to levels similar to that in the thyroid. Bolus iv injection of NaI dose‐dependently increased luminal iodide output in the stomach, reduced by pretreatment with the NIS inhibitor ITB5. Id NaI gradually increased GIS, paralleled by an increase of HOI output into the gastric lumen. Antral > fundic iodide secretion rates were present, enhanced by the presence of luminal amiloride or serosal forskolin, suggesting that amiloride 'unmasks' electrogenic epithelial Na+ uptake via the basolateral NIS, and apical iodide secretion is related to cAMP‐mediated anion secretion.These results suggest that GIS may affect host defenses via luminal HOI production, complementing antimicrobial gastric acid secretion.Supported by VA Merit Review, NIH R01 DK54221
OCTN1‐Mediated Ergothioneine Uptake Protects Lung Epithelial Cells From Tobacco Smoke‐Induced Damage
Authors: Ehrhardt Carsten , Nickel Sabrina , Selo Mohammed Ali , Clerkin Caoimhe , Salomon Johanna , Talbot Brian , Walsh John 
Published: June 2021

Smoking is a major contributing factor for chronic obstructive pulmonary disease (COPD). Tobacco smoke exposure causes oxidative stress which results in alveolar wall destruction, mucus hyper secretion, inflammation and defective tissue repair. Ergothioneine (ET), an amino acid transported by OCTN1, has been shown to exhibit antioxidant and cytoprotective capacities in vitro. Hence, the objective of this study is to understand the role of OCTN1 and the effect of ET on human alveolar cells when subjected to tobacco smoke‐induced oxidative stress.Confocal microscopy and Western blot was used to determine the presence of OCTN1 in A549 cells and freshly isolated human alveolar epithelial type II (ATII) and type I‐like (ATI‐like) cells in primary culture. A549 cells were cultured in medium containing 1 mM of ET and its intracellular accumulation was measured by LC‐MS/MS over a 3‐week period. A549 cells were also was exposed to various concentrations of cigarette smoke extract (CSE). Prior to the experiment, cells were either cultured for 1 week in the presence or absence of 1 mM ET. An MTT assay was used to assess cell viability for the difference exposures to CSE.Immunoblot and CLSM showed OCTN1 expression in A549, ATII and ATI‐like cells. The uptake of ET by A549 epithelial cells was saturated after approximately 7 days of culture. Western blot data supported the CLSM results and also indicated that OCTN1 levels were increased in the presence of ET. The MTT assay showed that cells pre‐treated with ET were less affected by the cytotoxic effects of CSE.These preliminary data indicate that OCTN1 might play a role in the pathogenesis of tobacco smoke‐induced COPD by regulating ergothioneine transport.
Osteopontin‐Stimulated Apoptosis in Cardiac Myocytes Involves Reactive Oxygen Species and Mitochondrial Pathway
Authors: Dalal Suman , Singh Mahipal , Singh Krishna 
Published: June 2021

Increased osteopontin (OPN) expression associates with increased myocyte apoptosis and myocardial dysfunction. Recently, we have shown that OPN stimulates cardiac myocyte apoptosis via the involvement of endoplasmic reticulum stress and mitochondrial death pathways. The objective of this study was to evaluate the production and source of reactive oxygen species (ROS), and their role in OPN‐stimulated myocyte apoptosis. Treatment of adult rat ventricular myocytes (ARVMs) with OPN (20 nM) induced protein oxidation as analyzed by protein carbonylation assay. Direct monitoring of ARVMs using total ROS detection kit and DCFDA (2',7'‐dichlorofluorescin diacetate; a fluorogenic dye) staining showed increased number of ROS‐positive cells in response to OPN. OPN induced mitochondrial depolarization, and decreased the expression of mitochondrial antioxidant enzyme, manganese superoxide dismutase‐2 (SOD‐2). Transmission electron microscopy revealed that OPN induces mitochondrial abnormalities including swelling and reorganization of cristae. OPN‐mediated increase in the levels of cytosolic cytochrome c and apoptosis were significantly inhibited by N‐acetyl‐cysteine (antioxidant), apocynin (NADPH oxidase inhibitor), MnTBAP [Mn(III)tetrakis (4‐benzoic acid) porphyrin; SOD mimic] and glibenclamide (mitochondrial KATP channel blocker). These data suggest that OPN induces ROS production, and affects mitochondrial morphology and integrity. Mitochondria may serve as a source of ROS production through the opening of mKATP channels.Supported by Department of Veterans Affairs (BX002332) and NIH
Temporal Heterogeneity In Apoptosis Execution
Authors: Vorobjev Ivan , Barteneva Natasha 
Published: June 2021

Apoptotic process is highly heterogeneous, and a long‐standing question is what parameters define time and reversibility of the apoptotic response at a single cell level. Using multicolor fluorescence assay we characterized at a single‐cell and population levels sequence of apoptotic events induced by extrinsic (TRAIL) and intrinsic (staurosporine, actinomycin D and etoposide) apoptotic stimuli in Jurkat and HeLa cells. Regardless of the stimulus first visible event on a single cell level was rapid apoptotic volume decrease (AVD) and Na+ influx tightly coordinated with mitochondria outer membrane permeabilization (MOMP) and mitochondrial depolarization. These events happened largely asynchronous in the cell population. Activation of caspases 3/7 is slow process at a single cell level. It always starts after MOMP with significant delay and caspase activity linearly accumulates in a given cell for several hours.Cell‐to‐cell variability of the beginning of MOMP is well approximated by Gaussian distribution with CV about 35‐40%, while the γ‐distribution model describes cell‐to‐cell variability in the pre‐MOMP and MOMP‐to‐caspase activation stages. Sorting different subpopulations after induction of apoptosis we show that cells with intact mitochondria can recover after washout of apoptotic stimulus, cells after MOMP recover rarely and cells with caspase 3/7 activity can not recover at all. We propose a double‐stroke model for apoptosis execution/cell recovery.
Truncation of GSK‐3β in Cardiac Mitochondria is the Master Switch of the mPTP
Authors: Hurst Stephen , Gomez Ludovic , Jhun BongSook , O‐Uchi Jin , Sheu Shey‐Shing 
Published: June 2021

Glycogen Synthase Kinase‐3β (GSK‐3β) is proposed as one of the key molecules that regulate cell death during ischemia reperfusion (IR) injury in the heart, largely via modulating mitochondria permeability transition pore (mPTP) activity, where both Ca2+ and oxidative stress are at peak levels. However, the exact mechanism of how GSK‐3β controls cardiomyocyte survival and death has remained elusive. In non‐cardiac cells, μ‐Calpain activated by high levels of Ca2+ and ROS can truncate GSK‐3β at both the N and C termini to generate a constitutively‐active GSK‐3β by removing the inhibitory phosphorylation sites. Therefore, we hypothesize that truncation of GSK‐3β in cardiac mitochondria can modulate mPTP function via the phosphorylation and binding affinity of mPTP component proteins.Using mitochondrial fractionation and proteinase‐K digestion, we found that GSK‐3β is truncated by μ‐Calpain in the mitochondria matrix. Using co‐immunoprecipitation (IP) we determined that truncated GSK‐3β associates with and phosphorylates cyclophilin D (mCypD). In addition, P‐mCypD increases the binding affinity to ATP‐Synthase, a component of the mPTP pore. Overexpression of a mitochondrial targeted Calpain inhibitory peptide based on the Calpastatin inhibitory domain in H9c2 cardiac myoblasts significantly blocked ionomycin‐induced cell death, as measured by Caspase 3/7 activation, to a greater extent than pretreatment with cyclosporine A, a potent mPTP inhibitor. In conclusion, Calpain‐mediated truncation of GSK‐3β in the mitochondrial matrix leads to phosphorylation of mCypD, allowing P‐mCypD to bind to and open the mPTP pore complex in cardiac cells.Funding Source: NIH Grant 5T32AA007463‐28
PAH Exposure Impacts Cardiac Function in Xenopus laevis Embryos
Authors: Pinette Julia , Sestak Madison , Johnson Kirsten , Whittemore Susan 
Published: June 2021

Using Xenopus laevis, a well‐established model system for vertebrate heart development, we assessed the impact of three common polyaromatic hydrocarbons (PAHs): phenanthrene (PHE), pyrene (PYR), and fluoranthene (FLA) on heart rate (HR), interbeat variability (IBV), incidence of atrioventicular (AV) block, and stroke volume (SV) and compared effects with the well‐characterized PAH benzo(a)pyrene (BaP) and with DMSO (vehicle) controls.A 72‐hr exposure (during heart valve formation) was associated with tachycardia for all PAHs at all doses (0.25‐25 μM). Increased HRs ranged from 20‐35% (p<0.05‐0.001). With the exception of 25 μM PHE and FLA, PAH‐induced tachycardia persisted at 96 hr (during atrial partitioning), while bradycardia was observed in embryos with a mature heart phenotype after 120 hr exposure to 25 μM FLA and PHE (14% and 17% decrease in HR; p<0.05, 0.01, respectively).While PAH exposure did not alter IBV, an indicator of ventricular arrhythmia, 25 μM FLA and PHE treatment did interfere with normal cardiac conduction. The incidence of AV block with FLA (25 μM) was 40%, with similar, but less dramatic, dose‐dependent responses observed for PHE. In contrast, none of the DMSO or 0.5 μM BaP‐treated animals exhibited AV block, with 5% incidence for 5 μM BaP. SV was unaffected by 120 hr exposure to any of the PAHs at any dose.The embryonic heart generates the hemodynamic forces needed for development of the circulation. These results suggest that early exposure to these common PAHs compromises cardiac function and may impact circulatory development. All three PAHs have been detected in significant amounts in human milk and cord blood, underscoring the need to gain a better understanding of their potential for cardiotoxicity.
Depression‐like Symptoms are Associated with Dysbalance in Vascular Norepinephrine Uptake. This is Worsening by Antidepressants and Increases the Risk for Hypertension.
Authors: Matchkov Vladimir , Wiborg Ove , Bouzinova Elena , Aalkjaer Christian 
Published: June 2021

Antidepressants affecting norepinephrine (NE) reuptake are known to increase the risk for hypertension. Anhedonia ‐ a decreased ability to experience pleasures ‐ is a central symptom in depression and can be evoked in preclinical models exploring the role of the stressful environment in mediating affective signs and autonomic function.Some rats exposed to Chronic Mild Stress (CMS) developed depression‐like symptoms (i.e. anhedonia, measured by sucrose intake) while others were resilient to 8 wks of CMS. Normal blood pressure and decreased cardiac function suggested increased peripheral resistance in rats with depression‐like symptoms. Chronic treatment with escitalopram (SSRI) did not affect blood pressure but normalized behavior in ~50% of treated rats, e.g. responders.Resistance (small mesenteric and femoral) arteries from CMS and non‐stressed rats responded similarly to NE in vitro. Inhibition of neuronal reuptake with cocaine increased NE sensitivity more in rats with depression‐like symptoms than in resilient and non‐stressed groups. Corticosterone‐sensitive extra‐neuronal catecholamine uptake was diminished in rats exposed to CMS. These changes were associated with upregulation of the neuronal NE transporter and reduced expression of the extra‐neuronal transporter (OCT‐2) in arteries from 'depressed' rats. Escitalopram further upregulated neuronal NE reuptake in all groups.Our results indicate that depression‐like symptoms are associated with changes in NE uptake in the vascular wall and could explain pro‐hypertensive action of some antidepressants.
The Role of Nucleus Tractus Solitarii (nTS) Neurons in Cardiorespiratory Dysfunction Associated with Alzheimer's Disease
Authors: Ostrowski Tim , Kline David , Hasser Eileen , Ostrowski Daniela 
Published: June 2021

Alzheimer's disease (AD) is associated with dysfunction of cardiorespiratory control. The mechanisms behind these alterations are not understood. The nTS is critical in cardiorespiratory homeostasis and reflex function, and may play a crucial role in AD dysautonomia. Here we examined autonomic and respiratory regulation, and neuronal alterations in the nTS of an established rodent AD model (CNS Streptozotocin (STZ)‐induced). Results are compared to saline‐injected control rats. In vivo plethysmography revealed a strong trend for blunted hypoxia/chemoreflex‐induced increases in breathing (p=0.068). Furthermore, baroreflex‐mediated sympathoexcitation to hypotension was markedly blunted in AD rats (p蠄0.01), consistent with orthostatic hypotension observed in AD patients. To examine the neuronal alterations in the nTS, patch clamp recordings were made in in vitro slices. When compared to control, AD rats show significantly increased network activity (p蠄0.01), spontaneous spike discharge and enhanced discharge to current injection (~40% increase vs. control; p蠄0.001). Our results suggest increased basal excitation of the nTS neuronal network and blunted cardiorespiratory reflex responses, which may play an important role in the autonomic and respiratory dysfunction observed in AD patients.Supported by: RO1 HL098602, AHA 12POST11670002
Influence of oxygen enriched gases during decompression on bubble formation and endothelial function
Authors: Valic Zoran , Segrt Ivana , Bozic Josko , Glavas Duska , Obad Ante , Glavicic Igor , Valic Maja 
Published: June 2021

Breathing oxygen enriched gases during decompression procedure leads to decreased time spent at specific decompression stop and ultimately to shorter dives with same amount of time on the bottom. However, breathing oxygen can damage endothelial function with reduction in flow mediated vasodilation (FMD). Five female and five male experienced SCUBA divers, with a mean age of 39±3 years (mean±SEM), were included in this investigation. Each diver performed three dives to 45 meters of sea water and 20 minutes bottom time using air. Air, nitrox 50 or nitrox 99 were used during decompression stops in accordance to V‐planner software. Order of the dives was randomized, with one day of recovery between each dive. Brachial artery function (FMD) was determined before and after each dive using ultrasound. Same device was used to measure number of venous gas bubbles after diving. Total diving times using air, nitrox 50 or nitrox 99 were 65, 45 and 43 minutes, respectively. Bubble grades after corresponding dives were: 1.4±0.4, 1.9±0.4 and 0.9±0.2, and did not show statistical difference. FMDs after corresponding dives were: 7.6±3.9%, 1.0±8.6% and 7.4±3.7, and did not show statistical difference although function of the arteries was greatly affected if divers used nitrox 50 as gas during decompression. In conclusion, we would recommend protocol which employs breathing nitrox 99 during decompression since it leads to the shortest dive with no significant change in both bubble number and FMD comparing to dive with air.This study was supported by Croatian Ministry of Science, Education and Sports.