Transepithelial Ion Transport Research Articles

In vitro platform to model the function of ionocytes in the human airway epithelium

BackgroundPulmonary ionocytes have been identified in the airway epithelium as a small population of ion transporting cells expressing high levels of CFTR (cystic fibrosis transmembrane conductance regulator), the gene mutated in cystic fibrosis. By providing an infinite source of airway epithelial cells (AECs), the use of human induced pluripotent stem cells (hiPSCs) could overcome some challenges of studying ionocytes. However, the production of AEC epithelia containing ionocytes from hiPSCs has proven difficult. Here, we present a platform to produce hiPSC-derived AECs (hiPSC-AECs) including ionocytes and investigate their role in the airway epithelium.MethodshiPSCs were differentiated into lung progenitors, which were expanded as 3D organoids and matured by air-liquid interface culture as polarised hiPSC-AEC epithelia. Using CRISPR/Cas9 technology, we generated a hiPSCs knockout (KO) for FOXI1, a transcription factor that is essential for ionocyte specification. Differences between FOXI1 KO hiPSC-AECs and their wild-type (WT) isogenic controls were investigated by assessing gene and protein expression, epithelial composition, cilia coverage and motility, pH and transepithelial barrier properties.ResultsMature hiPSC-AEC epithelia contained basal cells, secretory cells, ciliated cells with motile cilia, pulmonary neuroendocrine cells (PNECs) and ionocytes. There was no difference between FOXI1 WT and KO hiPSCs in terms of their capacity to differentiate into airway progenitors. However, FOXI1 KO led to mature hiPSC-AEC epithelia without ionocytes with reduced capacity to produce ciliated cells.ConclusionOur results suggest that ionocytes could have role beyond transepithelial ion transport by regulating epithelial properties and homeostasis in the airway epithelium.

Function of KvLQT1 potassium channels in a mouse model of bleomycin-induced acute lung injury.

Acute respiratory distress syndrome (ARDS) is characterized by an exacerbated inflammatory response, severe damage to the alveolar-capillary barrier and a secondary infiltration of protein-rich fluid into the airspaces, ultimately leading to respiratory failure. Resolution of ARDS depends on the ability of the alveolar epithelium to reabsorb lung fluid through active transepithelial ion transport, to control the inflammatory response, and to restore a cohesive and functional epithelium through effective repair processes. Interestingly, several lines of evidence have demonstrated the important role of potassium (K+) channels in the regulation of epithelial repair processes. Furthermore, these channels have previously been shown to be involved in sodium/fluid absorption across alveolar epithelial cells, and we have recently demonstrated the contribution of KvLQT1 channels to the resolution of thiourea-induced pulmonary edema in vivo. The aim of our study was to investigate the role of the KCNQ1 pore-forming subunit of KvLQT1 channels in the outcome of ARDS parameters in a model of acute lung injury (ALI). We used a molecular approach with KvLQT1-KO mice challenged with bleomycin, a well-established ALI model that mimics the key features of the exudative phase of ARDS on day 7. Our data showed that KvLQT1 deletion exacerbated the negative outcome of bleomycin on lung function (resistance, elastance and compliance). An alteration in the profile of infiltrating immune cells was also observed in KvLQT1-KO mice while histological analysis showed less interstitial and/or alveolar inflammatory response induced by bleomycin in KvLQT1-KO mice. Finally, a reduced repair rate of KvLQT1-KO alveolar cells after injury was observed. This work highlights the complex contribution of KvLQT1 in the development and resolution of ARDS parameters in a model of ALI.

The Eleutherococcus senticosus fruits' intractum affects changes in the transepithelial electric potential in the distal section of the rabbit's large intestine and inhibits hyaluronidase

Ethnopharmacological relevanceEleutherococcus senticosus (Rupr. et Maxim.) Maxim. has been used in traditional Russian medicine due to its recognized immunostimulant and anti-inflammatory activities. Compounds present in the fruits have demonstrated the capability to modulate the activity of enzymes such as hyaluronidase, suggesting their potential value in the development of effective therapies for various conditions where anti-inflammatory properties are beneficial, such as gastrointestinal diseases and tumor growth. Aim of the studyIn order to support the use of the fruits in folk medicine, this study is aimed to evaluate, post-mortem, the impact of E. senticosus fruits intractum (40 % extract made from fresh fruits) on the transepithelial electrogenic transport of sodium ions in the colon. The objective of this study was also to examine the impact of the intractum on proinflammatory serum hyaluronidase in children diagnosed with acute leukemia. MethodsThe study employed the Ussing technique to examine electrophysiological characteristics of isolated epithelial tissue, using the distal colon wall isolated from 10 New Zealand white male rabbits. The effect of the intractum on the inhibition of human serum hyaluronidase was examined with turbidimetric screening methods, using the blood samples collected from patients diagnosed with acute leukemia. ResultsFor the first time, we discovered that the intractum used in the stimulation fluid, caused hyperpolarization reactions in colon tissue. Statistical analysis showed that these reactions were significantly different in relation to the control. The intractum significantly inhibited hyaluronidase activity with the mean value by group of 60 %, and 40 % for aescin used as a control. ConclusionThe results support the traditional use of the fruits in inflammatory-related diseases. The use of intractum of E. senticosus on the distal colon wall demonstrates its protective effect on the wall integrity and in a relation to hyaluronidase inhibition may additionally indicate its anti-inflammatory property. Thus, the results mean that the intractum may be used in colon-related diseases.

Gasotransmitters do not prevent changes in transepithelial ion transport induced by hypoxia followed by reoxygenation.

How gaseous signalling molecules affect ion transport processes contributing to the physiological functions of the gastrointestinal tract under hypoxic conditions still needs to be clarified. The objective of the present study was to characterize the impact of gaseous signalling molecules on parameters of colonic ion transport during a hypoxia/reoxygenation cycle and the remaining secretory capacity of the epithelium after such a cycle. Short-circuit current (Isc) and tissue conductance (Gt) recordings in Ussing chamber experiments were performed on rat colon samples using CORM-2 (putative CO donor; 35 and 350 µM), sodium nitroprusside (NO donor; 100 µM), NaHS (fast H2S donor; 10- 1,000 µM), GYY 4137 (slowH2S donor; 50 µM) and Angeli's salt (HNO donor; 100 µM) as donors for gasotransmitters. Inhibition of endogenous synthesis of H2S was operated by inhibitors of cystathionin-γ-lyase, i.e.dl-propargylglycine (1 mM) or β-cyano-l-alanine (5 mM), and the inhibitor of cystathionine-β-synthase, amino-oxyacetate (5 mM). The fast gasotransmitter donors NaHS, sodium nitroprusside and Angeli's salt, administered 5 min before the onset of hypoxia, induced an increase in Isc. The response to the subsequently applied hypoxia was characterized by a decrease in Isc, which tended to be reduced only in the presence of the lowest concentration of NaHS (10 µM) tested. Reoxygenation resulted in a slow increase in Isc, which was unaffected by all donors or inhibitors tested. The stable acetylcholine derivative carbachol (50 µM) was administered at the end of each hypoxia/reoxygenation cycle to test the secretory capacity of the epithelium. Pretreatment of thetissue with the putative CO donor CORM-2 suppressed thesecretory response induced by carbachol. The same wasobserved when cystathionin-γ-lyase and cystathionin-γ-synthase were inhibited simultaneously. Under both conditions, Gt drastically increased suggesting an impaired tissue integrity. The present results demonstrate that none of the exogenous gasotransmitter releasing drugs significantly ameliorated the changes in epithelial ion transport during the hypoxia/reoxygenation cycle exvivo. In contrast, the putative CO donor CORM-2 exerted a toxic effect on the epithelium. The endogenous production of H2S, however, seems to have a protective effect on the mucosal integrity and the epithelial transport functions, which- when inhibited- leads to a loss of the secretory ability of the mucosa. This observation together with the trend for improvement observed with a low concentration of the H2S donor NaHS suggests a moderate protective role of low concentrations of H2S under hypoxic conditions.

Characterization of Commercially Available Human Primary Alveolar Epithelial Cells.

In vitro lung research requires appropriate cell culture models that adequately mimic invivo structure and function. Previously, researchers extensively used commercially available and easily expandable A549 and NCI-H441 cells, which replicate some but not all features of alveolar epithelial cells. Specifically, these cells are often restricted by terminally altered expression while lacking important alveolar epithelial characteristics. Of late, human primary alveolar epithelial cells (hPAEpCs) have become commercially available but are so far poorly specified. Here, we applied a comprehensive set of technologies to characterize their morphology, surface marker expression, transcriptomic profile, and functional properties. At optimized seeding numbers of 7,500 cells per square centimeter and growth at a gas-liquid interface, hPAEpCs formed regular monolayers with tight junctions and amiloride-sensitive transepithelial ion transport. Electron microscopy revealed lamellar body and microvilli formation characteristic for alveolar type II cells. Protein and single-cell transcriptomic analyses revealed expression of alveolar type I and type II cell markers; yet, transcriptomic data failed to detect NKX2-1, an important transcriptional regulator of alveolar cell differentiation. With increasing passage number, hPAEpCs transdifferentiated toward alveolar-basal intermediates characterized as SFTPC-, KRT8high, and KRT5- cells. In spite of marked changes in the transcriptome as a function of passaging, Uniform Manifold Approximation and Projection plots did not reveal major shifts in cell clusters, and epithelial permeability was unaffected. The present work delineates optimized culture conditions, cellular characteristics, and functional properties of commercially available hPAEpCs. hPAEpCs may provide a useful model system for studies on drug delivery, barrier function, and transepithelial ion transport invitro.

Tmprss2 maintains epithelial barrier integrity and transepithelial sodium transport.

The mouse cortical collecting duct cell line presents a tight epithelium with regulated ion and water transport. The epithelial sodium channel (ENaC) is localized in the apical membrane and constitutes the rate-limiting step for sodium entry, thereby enabling transepithelial transport of sodium ions. The membrane-bound serine protease Tmprss2 is co-expressed with the alpha subunit of ENaC. αENaC gene expression followed the Tmprss2 expression, and the absence of Tmprss2 resulted not only in down-regulation of αENaC gene and protein expression but also in abolished transepithelial sodium transport. In addition, RNA-sequencing analyses unveiled drastic down-regulation of the membrane-bound protease CAP3/St14, the epithelial adhesion molecule EpCAM, and the tight junction proteins claudin-7 and claudin-3 as also confirmed by immunohistochemistry. In summary, our data clearly demonstrate a dual role of Tmprss2 in maintaining not only ENaC-mediated transepithelial but also EpCAM/claudin-7-mediated paracellular barrier; the tight epithelium of the mouse renal mCCD cells becomes leaky. Our working model proposes that Tmprss2 acts via CAP3/St14 on EpCAM/claudin-7 tight junction complexes and through regulating transcription of αENaC on ENaC-mediated sodium transport.

Decaying kidney function during cirrhosis correlates with remodeling of distal colon aldosterone target gene expression.

Liver cirrhosis is associated to circulatory abnormalities leading to hypovolemia and stimulation of the renin-angiotensin-aldosterone system (RAAS). Advanced stages of the disease cause renal failure, impairing K+ and Na+ homeostasis. It has been proposed that the distal colon undergoes functional remodeling during renal failure, in particular by aldosterone-driven increased K+ excretion. In this study, we compared the transcriptional response of aldosterone target genes in the rat distal colon under two models of increased circulating aldosterone (one with concomitant RAAS activation) and in a model of secondary hyperaldosteronism induced by cirrhosis. The expression of a subset of these genes was also tested in distal colon biopsies from control subjects or patients with cirrhosis with varying levels of disease progression and treated or not with mineralocorticoid receptor inhibitor spironolactone. We examined known aldosterone-regulated transcripts involved in corticosteroid signaling and transepithelial ion transport. In addition, we included aldosterone-regulated genes related to cell proliferation. Our comparison revealed multiple aldosterone target genes upregulated in the rat distal colon during decompensated cirrhosis. Epithelial Na+ channel β and γ subunit expression correlated positively with plasma aldosterone concentration and negatively with glomerular filtration rate. Patients with cirrhosis showed increased expression of 11-β-hydroxysteroid-dehydrogenase 2 (11βHSD2), which was reverted by spironolactone treatment, suggesting a sensitization of the distal colon to aldosterone action. In summary, our data show that decaying kidney function during cirrhosis progression toward a decompensated state with hypovolemia correlates with remodeling of distal colon ion transporter expression, supporting a role for aldosterone in the process.NEW & NOTEWORTHY Liver cirrhosis progression significantly alters ion transporter subunit expression in the rat distal colon, a change that correlated well with declining kidney function and the severity of the disease. Our data suggest that the steroid hormone aldosterone participates in this homeostatic response to maintain electrolyte balance.

A Technique of Measurement of Gastrointestinal Luminal Nutrient Sensing and These Absorptions: Ussing Chamber (Short-Circuit Current) Technique.

The gastrointestinal (GI) tract is a series of hollow organs that play roles in food digestion and nutrient absorption. To perform these functions, they should recognize the luminal environment and elicit adequate physiological responses, including digestive juice secretion, peristaltic movements, etc. The Ussing chamber technique is an electrophysiological method for measuring transepithelial ion transport and permeability as short-circuit current (Isc) and transepithelial electrical tissue conductance (Gt) or resistance (TEER), respectively, in vitro. This technique can be applied for the measurement of luminal nutrient sensing and absorption. This article introduces practical methods for measuring luminal nutrient sensing and absorption using intestinal mucosa specimens isolated from humans and experimental animals.

Luteolin Enhances Transepithelial Sodium Transport in the Lung Alveolar Model: Integrating Network Pharmacology and Mechanism Study.

Luteolin (Lut), a natural flavonoid compound existing in Perilla frutescens (L.) Britton, has been proven to play a protective role in the following biological aspects: inflammatory, viral, oxidant, and tumor-related. Lut can alleviate acute lung injury (ALI), manifested mainly by preventing the accumulation of inflammation-rich edematous fluid, while the protective actions of Lut on transepithelial ion transport in ALI were seldom researched. We found that Lut could improve the lung appearance/pathological structure in lipopolysaccharide (LPS)-induced mouse ALI models and reduce the wet/dry weight ratio, bronchoalveolar protein, and inflammatory cytokines. Meanwhile, Lut upregulated the expression level of the epithelial sodium channel (ENaC) in both the primary alveolar epithelial type 2 (AT2) cells and three-dimensional (3D) alveolar epithelial organoid model that recapitulated essential structural and functional aspects of the lung. Finally, by analyzing the 84 interaction genes between Lut and ALI/acute respiratory distress syndrome using GO and KEGG enrichment of network pharmacology, we found that the JAK/STAT signaling pathway might be involved in the network. Experimental data by knocking down STAT3 proved that Lut could reduce the phosphorylation of JAK/STAT and enhance the level of SOCS3, which abrogated the inhibition of ENaC expression induced by LPS accordingly. The evidence supported that Lut could attenuate inflammation-related ALI by enhancing transepithelial sodium transport, at least partially, via the JAK/STAT pathway, which may offer a promising therapeutic strategy for edematous lung diseases.

Pharmacological inhibitors of the cystic fibrosis transmembrane conductance regulator exert off-target effects on epithelial cation channels

The cystic fibrosis transmembrane conductance regulator (CFTR) anion channel and the epithelial Na+ channel (ENaC) play essential roles in transepithelial ion and fluid transport in numerous epithelial tissues. Inhibitors of both channels have been important tools for defining their physiological role in vitro. However, two commonly used CFTR inhibitors, CFTRinh-172 and GlyH-101, also inhibit non-CFTR anion channels, indicating they are not CFTR specific. However, the potential off-target effects of these inhibitors on epithelial cation channels has to date not been addressed. Here, we show that both CFTR blockers, at concentrations routinely employed by many researchers, caused a significant inhibition of store-operated calcium entry (SOCE) that was time-dependent, poorly reversible and independent of CFTR. Patch clamp experiments showed that both CFTRinh-172 and GlyH-101 caused a significant block of Orai1-mediated whole cell currents, establishing that they likely reduce SOCE via modulation of this Ca2+ release-activated Ca2+ (CRAC) channel. In addition to off-target effects on calcium channels, both inhibitors significantly reduced human αβγ-ENaC-mediated currents after heterologous expression in Xenopus oocytes, but had differential effects on δβγ-ENaC function. Molecular docking identified two putative binding sites in the extracellular domain of ENaC for both CFTR blockers. Together, our results indicate that caution is needed when using these two CFTR inhibitors to dissect the role of CFTR, and potentially ENaC, in physiological processes.

ЦИСТАМИН И ЦИСТИН ПОДАВЛЯЮТ ТРАНСПОРТ Na+ В ЭПИТЕЛИИ КОЖИ ЛЯГУШКИ

Amphibian skin and other isolated epithelial systems are classical model objects for studying the mechanisms of transepithelial ion transport. Na+ transport in osmoregulatory epithelium is a complex, multicomponent system, which involves Na+ transporting proteins and signaling cascades localized in various cell membranes. The protein components of this system may be a target for oxidative stress. At the same time, the effect of oxidizing and reducing agents on Na+ transport in native epithelial systems, such as frog skin epithelium, practically has not been studied. Using voltage-clamp technique the effect of disulfide-containing oxidizing agents cystine and cystamine on Na+ transport in frog skin was investigated. It has been shown for the first time that the treatment of the skin of the Rana temporaria frog with cystamine or cystine at a concentration of 10 μg/ml suppresses the transepithelial Na+ transport. It was also found that the preincubation with dithiothreitol, which reduces disulfide bonds in proteins, prevents the inhibitory effect of cystamine and cystine. The obtained results indicate that Na+ transport in the frog skin is sensitive to oxidative stress and is modulated by disulfide-containing oxidizing agents, such as cystamine or cystine, and also that the effect of cystamine and cystine on Na+ transport in the frog skin epithelium is mediated by their interaction with functionally important cysteine residues of Na+-transporting proteins.

Mucins and CFTR: Their Close Relationship.

Mucociliary clearance is a critical defense mechanism for the lungs governed by regionally coordinated epithelial cellular activities, including mucin secretion, cilia beating, and transepithelial ion transport. Cystic fibrosis (CF), an autosomal genetic disorder caused by the dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) channel, is characterized by failed mucociliary clearance due to abnormal mucus biophysical properties. In recent years, with the development of highly effective modulator therapies, the quality of life of a significant number of people living with CF has greatly improved; however, further understanding the cellular biology relevant to CFTR and airway mucus biochemical interactions are necessary to develop novel therapies aimed at restoring CFTR gene expression in the lungs. In this article, we discuss recent advances of transcriptome analysis at single-cell levels that revealed a heretofore unanticipated close relationship between secretory MUC5AC and MUC5B mucins and CFTR in the lungs. In addition, we review recent findings on airway mucus biochemical and biophysical properties, focusing on how mucin secretion and CFTR-mediated ion transport are integrated to maintain airway mucus homeostasis in health and how CFTR dysfunction and restoration of function affect mucus properties.

Taste Receptor Activation in Tracheal Brush Cells by Denatonium Modulates ENaC Channels via Ca2+, cAMP and ACh.

Mucociliary clearance is a primary defence mechanism of the airways consisting of two components, ciliary beating and transepithelial ion transport (ISC). Specialised chemosensory cholinergic epithelial cells, named brush cells (BC), are involved in regulating various physiological and immunological processes. However, it remains unclear if BC influence ISC. In murine tracheae, denatonium, a taste receptor agonist, reduced basal ISC in a concentration-dependent manner (EC50 397 µM). The inhibition of bitter taste signalling components with gallein (Gβγ subunits), U73122 (phospholipase C), 2-APB (IP3-receptors) or with TPPO (Trpm5, transient receptor potential-melastatin 5 channel) reduced the denatonium effect. Supportively, the ISC was also diminished in Trpm5−/− mice. Mecamylamine (nicotinic acetylcholine receptor, nAChR, inhibitor) and amiloride (epithelial sodium channel, ENaC, antagonist) decreased the denatonium effect. Additionally, the inhibition of Gα subunits (pertussis toxin) reduced the denatonium effect, while an inhibition of phosphodiesterase (IBMX) increased and of adenylate cyclase (forskolin) reversed the denatonium effect. The cystic fibrosis transmembrane conductance regulator (CFTR) inhibitor CFTRinh172 and the KCNQ1 potassium channel antagonist chromanol 293B both reduced the denatonium effect. Thus, denatonium reduces ISC via the canonical bitter taste signalling cascade leading to the Trpm5-dependent nAChR-mediated inhibition of ENaC as well as Gα signalling leading to a reduction in cAMP-dependent ISC. Therefore, BC activation contributes to the regulation of fluid homeostasis.

Colonic Fluid and Electrolyte Transport 2022: An Update.

Colonic epithelial cells are responsible for maintaining a delicate balance between luminal secretion and the absorption of fluids and ions. This review aims to discuss and update the model of colonic electrolyte secretion and absorption via the cystic fibrosis transmembrane regulator (CFTR), epithelial sodium channel (ENaC), Na-K-Cl cotransporters (NKCC1 and 2), Na-H exchangers (NHE1–4), colonic H,KATPase, and several other key components involved in multi-level transepithelial ion transport. Developments in our understanding of the activity, regulation, localization, and relationships of these ion transporters and their interactions have helped forge a more robust understanding of colonic ion movement that accounts for the colonic epithelium’s role in mucosal pH modulation, the setting of osmotic gradients pivotal for fluid retention and secretion, and cell death regulation. Deviations from homeostatic ion transport cause diarrhea, constipation, and epithelial cell death and contribute to cystic fibrosis, irritable bowel syndrome (IBS), ulcerative colitis, and cancer pathologies. Signal transduction pathways that regulate electrolyte movement and the regulatory relationships between various sensors and transporters (CFTR as a target of CaSR regulation and as a regulator of ENaC and DRA, for example) are imperative aspects of a dynamic and comprehensive model of colonic ion homeostasis.

Mucus concentration-dependent biophysical abnormalities unify submucosal gland and superficial airway dysfunction in cystic fibrosis.

Cystic fibrosis (CF) is characterized by abnormal transepithelial ion transport. However, a description of CF lung disease pathophysiology unifying superficial epithelial and submucosal gland (SMG) dysfunctions has remained elusive. We hypothesized that biophysical abnormalities associated with CF mucus hyperconcentration provide a unifying mechanism. Studies of the anion secretion-inhibited pig airway model of CF revealed elevated SMG mucus concentrations, osmotic pressures, and SMG mucus accumulation. Human airway studies revealed hyperconcentrated CF SMG mucus with raised osmotic pressures and cohesive forces predicted to limit SMG mucus secretion/release. Using proline-rich protein 4 (PRR4) as a biomarker of SMG secretion, CF sputum proteomics analyses revealed markedly lower PRR4 levels compared to healthy and bronchiectasis controls, consistent with a failure of CF SMGs to secrete mucus onto airway surfaces. Raised mucus osmotic/cohesive forces, reflecting mucus hyperconcentration, provide a unifying mechanism that describes disease-initiating mucus accumulation on airway surfaces and in SMGs of the CF lung.

First Author Spotlight

AnnouncementFirst Author SpotlightPublished Online:18 Jan 2022https://doi.org/10.1152/ajprenal.2022.322.1.AUMoreSectionsPDF (347 KB)Download PDFDownload PDFPlus ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInEmail NICOLAS MONTALBETTIRenal-Electrolyte Division, Department of Medicine, University of Pittsburgh, Pittsburgh, PennsylvaniaFigure 1.Nicolas MontalbettiDownload figureDownload PowerPointNicolas Montalbetti is a Research Instructor in the Renal-Electrolyte Division, Department of Medicine, at the University of Pittsburgh. He obtained his PhD in physiology at the University of Buenos Aires, where he studied transepithelial ion transport. His research interest focuses on basic neurophysiological aspects of the urinary tract system in health and disease [Photo: Fig. 1].https://doi.org/10.1152/ajprenal.00167.2021JAMES VOTAVADepartment of Nutritional Sciences, University of Wisconsin-Madison, Madison, WisconsinFigure 2.James VotavaDownload figureDownload PowerPointJames Votava is a PhD student in the Department of Nutritional Sciences at the University of Wisconsin-Madison, currently mentored by Dr. Brian Parks. He earned his BS in Biological Sciences from the University of Wisconsin-Madison before beginning his graduate studies in the laboratory of Dr. Rick Eisenstein. His research interest focuses on identifying new genetic modifiers of human disease. Outside of the laboratory, James enjoys playing recreational sports and spending time with his dog [Photo: Fig. 2].https://doi.org/10.1152/ajprenal.00537.2020ZHEN WANGDepartment of Physiology and Biophysics, University of Mississippi Medical Center, Jackson, MississippiFigure 3.Zhen WangDownload figureDownload PowerPointZhen Wang is an assistant professor of the Department of Physiology and Biophysics at University of Mississippi Medical Center. He received his PhD from the Pharmacology Department of the University of Arkansas for Medical Sciences and completed his postdoctoral training in the Department of Physiology and Biophysics, University of Mississippi Medical Center, under the mentorship of Dr. John Hall. His major research interest is to understand the pathophysiology of diabetic cardiovascular diseases and to determine the molecular mechanisms of target organ injury induced by diabetes, hypertension, and obesity [Photo: Fig. 3].https://doi.org/10.1152/ajprenal.00296.2021DAN-DAN ZHANGGuangdong-Hong-Kong Joint Laboratory on Immunological and Genetic Kidney Diseases, Guangdong Academy of Medical Sciences, Guangdong Provincial People’s Hospital, Guangzhou, ChinaFigure 4.Dan-Dan ZhangDownload figureDownload PowerPointDan-Dan Zhang has received her PhD at the Department of Physiology of Harbin Medical University in 2018, and she has continued her postdoctoral training in Dr. Wang and Dr. Lin’s laboratories in the Department of Pharmacology of New York Medical College. Her research has focused on studying the regulation of renal potassium and sodium channels in the aldosterone-sensitive distal nephron. Since 2021, she has worked in Guangdong Provincial People’s Hospital in China [Photo: Fig. 4].https://doi.org/10.1152/ajprenal.00306.2021This article has no references to display. Back to Top Next FiguresReferencesRelatedInformationRelated articlesBladder infection with uropathogenic Escherichia coli increases the excitability of afferent neurons 15 Dec 2021American Journal of Physiology-Renal PhysiologyDysregulation of the sensory and regulatory pathways controlling cellular iron metabolism in unilateral obstructive nephropathy 05 Jan 2022American Journal of Physiology-Renal PhysiologyTransient receptor potential cation channel 6 contributes to kidney injury induced by diabetes and hypertension 05 Jan 2022American Journal of Physiology-Renal PhysiologyROMK channels are inhibited in the aldosterone-sensitive distal nephron of renal tubule Nedd4-2-deficient mice 23 Dec 2021American Journal of Physiology-Renal Physiology More from this issue > Volume 322Issue 1January 2022Pages i-ii Crossmark Copyright & PermissionsCopyright © 2022 the American Physiological Society.https://doi.org/10.1152/ajprenal.2022.322.1.AUHistory Received 3 August 2021 Accepted 24 November 2021 Published online 18 January 2022 Published in print 1 January 2022 PDF download Metrics Downloaded 184 times

Functional significance and physiological regulation of essential trace metals in fish.

Trace metals such as iron, copper, zinc and manganese play essential roles in various biological processes in fish, including development, energy metabolism and immune response. At embryonic stages, fish obtain essential metals primarily from the yolk, whereas in later life stages (i.e. juvenile and adult), the gastrointestine and the gill are the major sites for the acquisition of trace metals. On a molecular level, the absorption of metals is thought to occur at least in part via specific metal ion transporters, including the divalent metal transporter-1 (DMT1), copper transporter-1 (CTR1), and Zrt- and Irt-like proteins (ZIP). A variety of other proteins are also involved in maintaining cellular and systemic metal homeostasis. Interestingly, the expression and function of these metal transport- and metabolism-related proteins can be influenced by a range of trace metals and major ions. Increasing evidence also demonstrates an interplay between the gastrointestine and the gill for the regulation of trace metal absorption. Therefore, there is a complex network of regulatory and compensatory mechanisms involved in maintaining trace metal balance. Yet, an array of factors is known to influence metal metabolism in fish, such as hormonal status and environmental changes. In this Review, we summarize the physiological significance of iron, copper, zinc and manganese, and discuss the current state of knowledge on the mechanisms underlying transepithelial metal ion transport, metal-metal interactions, and cellular and systemic handling of these metals in fish. Finally, we identify knowledge gaps in the regulation of metal homeostasis and discuss potential future research directions.

Impact of Campylobacter spp. on the Integrity of the Porcine Gut.

Simple SummaryCampylobacteriosis is recognised as a leading food-borne zoonotic enteric disease of humans, mainly caused by Campylobacter jejuni, and to a minor extent by C. coli. In general, pigs are symptomless carriers primarily of C. coli, but may also harbour C. jejuni. In a swine infection model, weaned piglets were intragastrically inoculated with C. coli (ST-5777; n = 8), with C. jejuni (ST-122; n = 10), or with both strains (n = 8) and 11 piglets served as control. The health status was monitored and the influence on the intestinal barrier was investigated using the Ussing chamber technique and histological examinations. After inoculation, no clinical manifestations were noted. No gross lesions were observed during dissection four weeks post inoculation, and no pathohistological changes were detected in the intestinal mucosal sections. On the other hand, in the caecum of C. jejuni mono-inoculated pigs, we recognized an impact on transepithelial transport processes. We observed an increased Cl− secretion by using the Ussing chamber technique.Campylobacter (C.) is the most common food-borne zoonosis in humans, which mainly manifests with watery to bloody diarrhoea. While C. jejuni is responsible for most cases of infection, C. coli is less frequently encountered. The object of the study was to prove the clinical impact of mono- and co-colonisation of C. coli and C. jejuni on weaned piglets in an infection model and to investigate the impact on transepithelial transport processes in the jejunum and caecum. At an age of eight weeks, eight pigs were infected with C. coli (ST-5777), 10 pigs with C. jejuni (ST-122), eight pigs with both strains, and 11 piglets served as control. During the four-week observation period, no clinical signs were observed. During dissection, both strains could be isolated from the jejunum and the caecum, but no alteration of the tissue could be determined histopathologically. Mono-infection with C. jejuni showed an impact on transepithelial ion transport processes of the caecum. An increase in the short circuit current (Isc) was observed in the Ussing chamber resulting from carbachol- and forskolin-mediated Cl− secretion. Therefore, we speculate that caecal colonisation of C. jejuni might affect the transport mechanisms of the intestinal mucosa without detectable inflammatory reaction.

Studying the Activity of Neuropeptides and Other Regulators of the Excretory System in the Adult Mosquito.

Studies of insect physiology, particularly in those species that are vectors of pathogens causing disease in humans and other vertebrates, provide the foundation to develop novel strategies for pest control. Here, a series of methods are described that are routinely utilized to determine the functional roles of neuropeptides and other neuronal factors (i.e., biogenic amines) on the excretory system of the mosquito, Aedes aegypti. The Malpighian tubules (MTs), responsible for primary urine formation, can continue functioning for hours when removed from the mosquito, allowing for fluid secretion measurements following hormone treatments. As such, the Ramsay assay is a useful technique to measure secretion rates from isolated MTs. Ion-selective microelectrodes (ISME) can sequentially be used to measure ion concentrations (i.e., Na+ and K+) in the secreted fluid. This assay allows for the measurement of several MTs at a given time, determining the effects of various hormones and drugs. The Scanning Ion-selective Electrode Technique uses ISME to measure voltage representative of ionic activity in the unstirred layer adjacent to the surface of ion transporting organs to determine transepithelial transport of ions in near real time. This method can be used to understand the role of hormones and other regulators on ion absorption or secretion across epithelia. Hindgut contraction assays are also a useful tool to characterize myoactive neuropeptides, that may enhance or reduce the ability of this organ to remove excess fluid and waste. Collectively, these methods provide insight into how the excretory system is regulated in adult mosquitoes. This is important because functional coordination of the excretory organs is crucial in overcoming challenges such as desiccation stress after eclosion and before finding a suitable vertebrate host to obtain a bloodmeal.

Extracellular phosphate enhances the function of F508del-CFTR rescued by CFTR correctors

Background: The clinical response to cystic fibrosis transmembrane conductance regulator (CFTR) modulators varies between people with cystic fibrosis (CF) of the same genotype, in part through the action of solute carriers encoded by modifier genes. Here, we investigate whether phosphate transport by SLC34A2 modulates the function of F508del-CFTR after its rescue by CFTR correctors.Methods: With Fischer rat thyroid (FRT) cells heterologously expressing wild-type and F508del-CFTR and fully-differentiated CF and non-CF human airway epithelial cells, we studied SLC34A2 expression and the effects of phosphate on CFTR-mediated transepithelial ion transport. F508del-CFTR was trafficked to the plasma membrane by incubation with different CFTR correctors (alone or in combination) or by low temperature.Results: Quantitative RT-PCR demonstrated that both FRT and primary airway epithelial cells express SLC34A2 mRNA and no differences were found between cells expressing wild-type and F508del-CFTR. For both heterologously expressed and native F508del-CFTR rescued by either VX-809 or C18, the magnitude of CFTR-mediated Cl− currents was dependent on the presence of extracellular phosphate. However, this effect of phosphate was not detected with wild-type and low temperature-rescued F508del-CFTR Cl− currents. Importantly, the modulatory effect of phosphate was observed in native CF airway cells exposed to VX-445, VX-661 and VX-770 (Trikafta) and was dependent on the presence of both sodium and phosphate.Conclusions: Extracellular phosphate modulates the magnitude of CFTR-mediated Cl− currents after F508del-CFTR rescue by clinically-approved CFTR correctors. This effect likely involves electrogenic phosphate transport by SLC34A2. It might contribute to inter-individual variability in the clinical response to CFTR correctors.