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S M Wilson

Publications and source records attributed to S M Wilson.

At least 19 recordsLinked to original sources
PubMed2006

Adenosine-evoked Na+ transport in human airway epithelial cells.

BACKGROUND AND PURPOSE: Absorptive epithelia express apical receptors that allow nucleotides to inhibit Na(+) transport but ATP unexpectedly stimulated this process in an absorptive cell line derived from human bronchiolar epithelium (H441 cells) whilst UTP consistently caused inhibition. We have therefore examined the pharmacological basis of this anomalous effect of ATP. EXPERIMENTAL APPROACH: H441 cells were grown on membranes and the short circuit current (I(SC)) measured in Ussing chambers. In some experiments, [Ca(2+)](i) was measured fluorimetrically using Fura -2. mRNAs for adenosine receptors were determined by the polymerase chain reaction (PCR). KEY RESULTS: Cross desensitization experiments showed that the inhibitory response to UTP was abolished by prior exposure to ATP whilst the stimulatory response to ATP persisted in UTP-pre-stimulated cells. Apical adenosine evoked an increase in I(SC) and this response resembled the stimulatory component of the response to ATP, and could be mimicked by adenosine receptor agonists. Pre-stimulation with adenosine abolished the stimulatory component of the response to ATP. mRNA encoding A(1), A(2A) and A(2B) receptor subtypes, but not the A(3) subtype, was detected in H441 cells and adenosine receptor antagonists could abolish the ATP-evoked stimulation of Na(+) absorption. CONCLUSIONS AND IMPLICATIONS: The ATP-induced stimulation of Na(+) absorption seems to be mediated via A(2A/B) receptors activated by adenosine produced from the extracellular hydrolysis of ATP. The present data thus provide the first description of adenosine-evoked Na(+) transport in airway epithelial cells and reveal a previously undocumented aspect of the control of this physiologically important ion transport process.

Adenosine↗
PubMed2006

Expression of intermediate-conductance, Ca2+-activated K+ channel (KCNN4) in H441 human distal airway epithelial cells.

Electrophysiological studies of H441 human distal airway epithelial cells showed that thapsigargin caused a Ca(2+)-dependent increase in membrane conductance (G(Tot)) and hyperpolarization of membrane potential (V(m)). These effects reflected a rapid rise in cellular K(+) conductance (G(K)) and a slow fall in amiloride-sensitive Na(+) conductance (G(Na)). The increase in G(Tot) was antagonized by Ba(2+), a nonselective K(+) channel blocker, and abolished by clotrimazole, a KCNN4 inhibitor, but unaffected by other selective K(+) channel blockers. Moreover, 1-ethyl-2-benzimidazolinone (1-EBIO), which is known to activate KCNN4, increased G(K) with no effect on G(Na). RT-PCR-based analyses confirmed expression of mRNA encoding KCNN4 and suggested that two related K(+) channels (KCNN1 and KCNMA1) were absent. Subsequent studies showed that 1-EBIO stimulates Na(+) transport in polarized monolayers without affecting intracellular Ca(2+) concentration ([Ca(2+)](i)), suggesting that the activity of KCNN4 might influence the rate of Na(+) absorption by contributing to G(K). Transient expression of KCNN4 cloned from H441 cells conferred a Ca(2+)- and 1-EBIO-sensitive K(+) conductance on Chinese hamster ovary cells, but this channel was inactive when [Ca(2+)](i) was <0.2 microM. Subsequent studies of amiloride-treated H441 cells showed that clotrimazole had no effect on V(m) despite clear depolarizations in response to increased extracellular K(+) concentration ([K(+)](o)). These findings thus indicate that KCNN4 does not contribute to V(m) in unstimulated cells. The present data thus establish that H441 cells express KCNN4 and highlight the importance of G(K) to the control of Na(+) absorption, but, because KCNN4 is quiescent in resting cells, this channel cannot contribute to resting G(K) or influence basal Na(+) absorption.

Animals↗
PubMed2005

Extensor tendon impingement in a gymnast.

Wrist injuries in the gymnast are due to the transformation of the upper extremity into a weight bearing entity. Gymnast wrist pain presents a difficult diagnostic and therapeutic challenge. Here, we present a new case of extensor tendon impingement in an elite gymnast. To our knowledge, there is no similar report in the literature.

Adult↗
PubMed2005

Closed rupture of the thumb flexor tendon pulleys.

Closed flexor tendon pulley ruptures are relatively rare injuries. All previously reported cases have been in the long finger pulleys. To our knowledge, there has not been a case of closed thumb flexor tendon pulley rupture reported in the literature. This paper presents two cases of this pathology and discusses appropriate treatment of it.

Adult↗
PubMed2005

Cystic fibrosis and airway submucosal glands.

The chronic pulmonary infections and inflammation associated with cystic fibrosis (CF) are responsible for almost all the morbidity and mortality of this disease. Our understanding of the mechanisms that underlie the very early stages of CF lung disease, that result directly from mutations in the CF gene, is relatively poor. However, the demonstration that the predominant sites of expression of the CF gene in normal lungs are the submucosal glands, together with the histological observations showing that hyperplasia of these glands and mucin occlusion of the gland ducts are the earliest signs of disease in the CF lung, suggest that malfunction of the submucosal glands may be an important factor contributing to the early pathophysiology of CF lung disease. This review describes the function of submucosal glands in normal lungs, and the way in which their function may be disrupted in CF and may thus contribute to the early stages of CF lung disease.

Cystic Fibrosis↗
PubMed2004

Thyroid hormone potentiates glucocorticoid-evoked airway Na+ transport without affecting alpha-ENaC transcription.

Glucocorticoid and thyroid hormones (T3) are important for the development of the lungs' Na+ absorbing phenotype, which is essential for the integrated functioning of the respiratory tract. Electrometric studies of H441 airway epithelial cells confirmed that dexamethasone increases apical Na+ conductance (GNa) and demonstrated that T3 facilitates this control over GNa. Assays of transcriptional activity showed that dexamethasone caused concentration-dependent activation of the human alpha-ENaC promoter (EC50 approximately 5 nM) but, despite its clear effect on GNa, T2 had no effect upon the transcriptional response to dexamethasone. The facilitation of Na+ transport may thus reflect control over events downstream to transcription.

Base Sequence↗
PubMed2004

A glucocorticoid-induced Na+ conductance in human airway epithelial cells identified by perforated patch recording.

The perforated patch recording technique was used to investigate the effects of dexamethasone (0.2 microm, 24-30 h), a synthetic glucocorticoid, on membrane conductance in the human airway epithelial cell line H441. Under zero current clamp conditions this hormone induced amiloride-sensitive depolarization of the membrane potential (V(m)). Lowering external Na(+) to 10 mm by replacing Na(+) with N-methyl-d-glucammonium (NMDG(+)) also hyperpolarized the dexamethasome-treated cells, whilst replacing Na(+) with Li(+) caused a small depolarization. Although V(m) was insensitive to amiloride in control cells, NMDG(+) substitution caused a small hyperpolarization and so an amiloride-insensitive cation conductance is present. Replacing Na(+) with Li(+) had no effect on V(m) in such cells. Voltage clamp studies of dexamethasone-treated cells showed that the amiloride-sensitive component of the membrane current reversed at a potential close to the Na(+) equilibrium potential (E(Na)), and replacing Na(+) with K(+) caused a leftward shift in reversal potential (V(Rev)) that correlated with the corresponding shift in E(Na). Lowering [Na(+)](o) to 10 mm, the concentration in the pipette solution, by substitution with NMDG(+) shifted V(Rev) to 0 mV, whilst replacing Na(+) with Li(+) caused a rightward shift. Exposing dexamethasone-treated cells to a cocktail of cAMP-activating compounds (20 min) caused a approximately 2-fold increase in amiloride-sensitive conductance that was associated with no discernible change in ionic selectivity and an 18 mV depolarization. Dexamethasone thus induces the expression of a selective Na(+) conductance with a substantial permeability to Li(+) that is subject to acute regulation via cAMP. These data thus suggest that selective Na(+) channels underlie cAMP-regulated Na(+) transport in airway epithelia.

Amiloride↗
PubMed2004

A regulated apical Na(+) conductance in dexamethasone-treated H441 airway epithelial cells.

Treating H441 cells with dexamethasone raised the abundance of mRNA encoding the epithelial Na(+) channel alpha- and beta-subunits and increased transepithelial ion transport (measured as short-circuit current, I(sc)) from <4 microA.cm(-2) to 10-20 microA.cm(-2). This dexamethasone-stimulated ion transport was blocked by amiloride analogs with a rank order of potency of benzamil >or= amiloride > EIPA and can thus be attributed to active Na(+) absorption. Studies of apically permeabilized cells showed that this increased transport activity did not reflect a rise in Na(+) pump capacity, whereas studies of basolateral permeabilized cells demonstrated that dexamethasone increased apical Na(+) conductance (G(Na)) from a negligible value to 100-200 microS.cm(-2). Experiments that explored the ionic selectivity of this dexamethasone-induced conductance showed that it was equally permeable to Na(+) and Li(+) and that the permeability to these cations was approximately fourfold greater than to K(+). There was also a small permeability to N-methyl-d-glucammonium, a nominally impermeant cation. Forskolin, an agent that increases cellular cAMP content, caused an approximately 60% increase in I(sc), and measurements made after these cells had been basolaterally permeabilized demonstrated that this response was associated with a rise in G(Na). This cAMP-dependent control over G(Na) was disrupted by brefeldin A, an inhibitor of vesicular trafficking. Dexamethasone thus stimulates Na(+) transport in H441 cells by evoking expression of an amiloride-sensitive apical conductance that displays moderate ionic selectivity and is subject to acute control via a cAMP-dependent pathway.

Cell Membrane↗
PubMed2003

Secretion of acid and base equivalents by intact distal airways.

Secretion of HCO(3)(-) by airway submucosal glands is essential for normal liquid and mucus secretion. Because the liquid bathing the airway surface (ASL) is acidic, it has been proposed that the surface epithelium may acidify HCO(3)(-)-rich glandular fluid. The aim of this study was to investigate the mechanisms by which intact distal bronchi, which contain both surface and glandular epithelium, modify pH of luminal fluid. Distal bronchi were isolated from pig lungs, cannulated in a bath containing HCO(3)(-)-buffered solution, and perfused continually with an aliquot of similar, lightly buffered solution (LBS) in which NaCl replaced NaHCO(3)(-) (pH 7 with NaOH). The pH of this circulating LBS initially acidified (by 0.053 +/- 0.0053 pH units) and transepithelial potential difference (PD) depolarized. The magnitude of acidification was increased when pH(LBS) was higher. This acidification was unaffected by luminal dimethylamiloride (DMA, 100 microM) but was inhibited by 100 nM bafilomycin A(1) (by 76 +/- 13%), suggesting involvement of vacuolar-H(+) ATPase. Addition of ACh (10 microM) evoked alkalinization of luminal LBS and hyperpolarization of transepithelial PD. The alkalinization was inhibited in HCO(3)(-)-free solutions containing acetazolamide (1 mM) and by DMA and was enhanced by bumetanide (100 microM), an inhibitor of Cl(-) secretion. The hyperpolarization was unaffected by these maneuvers. The anion channel blocker 5-nitro-2-(3-phenylpropylamino)benzoate (300 microM) and combined treatment with DMA and bumetanide blocked both the alkalinization and hyperpolarization responses to ACh. These results are consistent with earlier studies showing that ACh evokes glandular secretion of HCO(3)(-) and Cl(-). Isolated distal airways thus secrete both acid and base equivalents.

Acetylcholine↗
PubMed2003

Use of a phage-based assay for phenotypic detection of mycobacteria directly from sputum.

The phage amplified biologically assay is a new method for rapid and low-cost phenotypic determination of the drug sensitivities of Mycobacterium tuberculosis isolates and the detection of viable organisms in patient specimens. Infection of slowly growing mycobacteria with phage (phage D29) was followed by chemical virucide destruction of extracellular phage. Infected mycobacteria were mixed in culture with rapidly growing sensor cells, which the phage can also infect; i.e., lytic amplification of phage occurs. The aims of the present study were to optimize the speed and sensitivity of the assay and reduce its cost for developing countries by using an M. tuberculosis-spiked sputum model with (i). identification of inhibitory components of sputum and optimization of decontamination methods; (ii). simplification of the washing and development steps; (iii). reduction of the use of high-cost components, e.g., oleate-albumin-dextrose-catalase (OADC) supplement; and (iv). optimization of virucide treatment. The following results were obtained. (i). An inhibitory factor in sputum which could be removed by treatment of the sample with sodium dodecyl sulfate or NaOH decontamination was identified. (ii). A microcentrifuge-based approach with thixotropic silica as a bedding and resuspension agent was developed as an alternative to conventional centrifugation medium exchange. The yield was increased 228-fold, with increased speed and reduced cost. (iii). At present, after extracellular inactivation of phage, the ferrous ammonium sulfate (FAS) virucide is sequestered by dilution with an expensive supplement, OADC. Sodium citrate with calcium chloride was found to be a cost-effective after treatment with the FAS protectant and offered greater protection than OADC. Kinetic-lysis experiments indicated that an infection time of 1 to 3 h prior to FAS addition was optimal. (iv). Amplification of the signal (which corresponded to the burst size) was shown by allowing lysis prior to plating in a spiked medium model (up to 20-fold) and a spiked sputum model (up to 10-fold). A liquid culture detection method capable of detecting approximately 60 viable M. tuberculosis organisms in 1 ml of sputum was developed. Taken together, these improvements support the routine application of the assay to sputum specimens.

Bacteriophages↗
PubMed2002

Hormonal modulation of Na(+) transport in rat fetal distal lung epithelial cells.

Isolated rat fetal distal lung epithelial (FDLE) cells were cultured (approximately 48 h) on permeable supports in medium devoid of hormones and growth factors whilst P(O2) was maintained at the level found in either the fetal (23 mmHg) or the postnatal (100 mmHg) alveolar regions. The cells became incorporated into epithelial layers that generated a basal short-circuit current (I(SC)) attributable to spontaneous Na(+) absorption. Cells at neonatal P(O2) generated larger currents than did cells at fetal P(O2), indicating that this Na(+) transport process is oxygen sensitive. Irrespective of P(O2), isoprenaline failed to elicit a discernible change in I(SC), demonstrating that beta-adrenoceptor agonists do not stimulate Na(+) transport under these conditions. However, isoprenaline did elicit cAMP accumulation in these cells, indicating that functionally coupled beta-adrenoceptors are present. Further experiments showed that isoprenaline did increase I(SC) in cells treated (24 h) with a combination of tri-iodothyronine (T(3), 10 nM) and dexamethasone (200 nM). Studies of basolaterally permeabilised cells showed that these hormones are essential for the isoprenaline-evoked increase in the apical membrane's Na(+) conductance (G(Na)), whereas isoprenaline-evoked changes in apical Cl(-) conductance (G(Cl)) can occur in both control and hormone-treated cells. Irrespective of their hormonal status, FDLE cells thus express beta-adrenoceptors that are functionally coupled to adenylate cyclase, and allow beta-adrenoceptor agonists to modulate the apical membrane's anion conductance. However, T(3) and dexamethasone are needed if these receptors are to exert control over G(Na). These hormones may thus play an important role in the functional maturation of the lung by allowing beta-adrenoceptor-mediated control over epithelial Na(+) channels in the apical plasma membrane.

1-Methyl-3-isobutylxanthine↗
PubMed2002

Regulation of intracellular pH in Calu-3 human airway cells.

The Calu-3 human cell line exhibits features of submucosal gland serous cells and secretes HCO(3)(-). The aim of this study was to identify the HCO(3)(-) transporters present in these cells by studying their role in the regulation of intracellular pH (pH(i)). Calu-3 cells were grown on coverslips, loaded with the pH-sensitive fluorescent dye BCECF, and their fluorescence intensity monitored as an indication of pH(i). Cells were acidified with NH(4)Cl (25 mM, 1 min) and pH(i) recovery recorded. In the absence of HCO(3)(-), initial recovery was 0.208 +/- 0.016 pH units min(-1) (n = 37). This was almost abolished by removal of extracellular Na(+) and by amiloride (1 mM), consistent with the activity of a Na(+)-H(+) exchanger (NHE). In the presence of HCO(3)(-) and CO(2), recovery (0.156 +/- 0.018 pH units min(-1)) was abolished (reduced by 91.8 +/- 6.7 %, n = 7) by removal of Na(+) but only attenuated (by 63.3 +/- 5.8 %, n = 9) by amiloride. 4,4-Dinitrostilbene-2,2-disulfonic acid (DNDS) inhibited recovery by 45.8 +/- 5.0 % (n = 7). The amiloride-insensitive recovery was insensitive to changes in membrane potential, as confirmed by direct microelectrode measurements, brought about by changing extracellular [K(+)] in the presence of either valinomycin or the K(+) channel opener 1-EBIO. In addition, forskolin (10 microM), which activates the cystic fibrosis transmembrane conductance regulator Cl(-) conductance in these cells and depolarises the cell membrane, had no effect on recovery. Removal of extracellular Cl(-) trebled pH(i) recovery rates, suggesting that an electroneutral, DNDS-sensitive, Cl(-)-HCO(3)(-) exchanger together with a NHE may be involved in pH(i) regulation and HCO(3)(-) secretion in these cells. RT-PCR detected the expression of the electrogenic Na(+)-HCO(3)(-) cotransporter NBC1 and the Cl(-)-HCO(3)(-) exchanger (AE2) but not the electroneutral Na(+)-HCO(3)(-) cotransporter NBCn1.

Amiloride↗
PubMed2002

Beta-adrenoceptor-mediated control of apical membrane conductive properties in fetal distal lung epithelia.

Distal lung epithelial cells isolated from fetal rats were cultured (48 h) on permeable supports so that transepithelial ion transport could be quantified electrometrically. Unstimulated cells generated a short-circuit current (I(sc)) that was inhibited (~80%) by apical amiloride. The current is thus due, predominantly, to the absorption of Na(+) from the apical solution. Isoprenaline increased the amiloride-sensitive I(sc) about twofold. Experiments in which apical membrane Na(+) currents were monitored in basolaterally permeabilized cells showed that this was accompanied by a rise in apical Na(+) conductance (G(Na(+))). Isoprenaline also increased apical Cl- conductance (G(Cl-)) by activating an anion channel species sensitive to glibenclamide but unaffected by 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS). The isoprenaline-evoked changes in G(Na(+)) and G(Cl(minus sign)) could account for the changes in I(sc) observed in intact cells. Glibenclamide had no effect upon the isoprenaline-evoked stimulation of I(sc) or G(Na(+)) demonstrating that the rise in G(Cl-) is not essential to the stimulation of Na(+) transport.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed2001

Hypoxic activation of an amiloride-sensitive cation conductance in alveolar epithelial cells.

Imposing hypoxia (P(O(2)) = 23 mmHg) upon A549 cells elicited increased G(amil) although previous work had predicted a fall in this parameter. G(amil) appeared to be dependent upon glucocorticoid-driven gene expression, a process inhibited by ERK, an enzyme activated by oxidative stress. However, hypoxia transiently activated this enzyme and the response was blocked by glucocorticoids, showing that the rise in G(amil) occurs only if ERK activation is suppressed. Fluorimetric assays showed that lowering P(O(2)) elicited H(2)O(2) formation indicating that this maneuver actually imposes oxidative stress, thus explaining how hypoxia can elicit responses normally associated with a rise in P(O(2)).

Amiloride↗
PubMed2001

Mutations in Cdh23 cause nonsyndromic hearing loss in waltzer mice.

Mutations at the waltzer (v) locus result in deafness and vestibular dysfunction due to degeneration of the neuroepithelium within the inner ear. Here, we use a positional cloning approach to show that waltzer encodes a novel cadherin (Cdh23), which is most closely related to the Drosophila Fat protein. A single nucleotide deletion in the v(J) allele and a single nucleotide insertion in the v allele are predicted to truncate each protein near the N-terminus and produce a functional null allele. In situ hybridization analysis showed that Cdh23 is expressed in the sensory hair cells of the inner ear, where it has been suggested to be a molecule critical for crosslinking of the stereocilia. In addition, Cdh23 is expressed in the urticulo-saccular foramen,the ductus reuniens, and Reissner's membrane, suggesting that Cdh23 may also be involved in maintaining the ionic composition of the endolymph. Finally, mutations in human CDH23 have recently been described for two loci, DFNB12 and USH1D, which cause nonsyndromic deafness, identifying waltzer as a mouse model for human hearing loss.

Alleles↗
PubMed2001

Oxygen-evoked Na+ transport in rat fetal distal lung epithelial cells.

Monolayer cultures of rat fetal distal lung epithelial (FDLE) cells generated larger spontaneous short circuit currents (ISC) when maintained (48 h) at neonatal alveolar PO2 (100 mmHg) than at fetal PO2 (23 mmHg). When cells were shifted between these atmospheres in order to impose a rise in PO2 equivalent to that seen at birth, no rise in ISC was seen after 6 h but the response was fully established by 24 h. Studies of basolaterally permeabilised cells revealed a small rise in apical Na+ conductance (GNa) 6 h after PO2 was raised but no further change had occurred by 24 h. A substantial rise was, however, seen after 48 h. Reporter gene assays showed that no activation of the -ENaC (epithelial Na+ channel -subunit) promoter was discernible 24 h after PO2 was raised but increased transcriptional activity was seen at 48 h. Studies of apically permeabilised cells showed that a small rise in Na+ pump capacity was evident 6 h after PO2 was raised and, in common with the rise in ISC, this effect was fully established by 24 h. The rise in ISC thus develops 6-24 h after PO2 is raised and is due, primarily, to increased Na+ pump capacity. The increase in GNa thus coincides with activation of the -ENaC promoter but these effects occur after the rise in ISC is fully established and so cannot underlie this physiological response. The increased transcription may be an adaptation to increased Na+ transport and not its cause.

Amiloride↗