Modulation of epithelial sodium channel activity by lipopolysaccharide in alveolar type II cells: involvement of purinergic signaling
ABSTRACT Pseudomonas aeruginosa is a gram-negative bacterium that causes chronic infection in cystic fibrosis patients. We reported recently that P. aeruginosa modulates epithelial Na(+) channel (ENaC) expression in experimental chronic pneumonia models. For this reason, we tested whether LPS from P. aeruginosa alters ENaC expression and activity in alveolar epithelial cells. We found that LPS induces a approximately 60% decrease of ENaC apical current without significant changes in intracellular ENaC or surface protein expression. Because a growing body of evidence reports a key role for extracellular nucleotides in regulation of ion channels, we evaluated the possibility that modulation of ENaC activity by LPS involves extracellular ATP signaling. We found that alveolar epithelial cells release ATP upon LPS stimulation and that pretreatment with suramin, a P2Y(2) purinergic receptor antagonist, inhibited the effect of LPS on ENaC. Furthermore, ET-18-OCH3, a PLC inhibitor, and Go-6976, a PKC inhibitor, were able to partially prevent ENaC inhibition by LPS, suggesting that the actions of LPS on ENaC current were mediated, in part, by the PKC and PLC pathways. Together, these findings demonstrate an important role of extracellular ATP signaling in the response of epithelial cells to LPS.
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ABSTRACT: BACKGROUND: Pseudomonas aeruginosa airway infection is associated with a high mortality rate in cystic fibrosis. Lipopolysaccharide (LPS), a main constituent of the outer membrane of P. aeruginosa, is responsible for activation of innate immune response but its role on airway epithelium ion transport, is not well known. The aim of this study was to determine the role for P. aeruginosa LPS in modulating chloride secretion and intracellular calcium in the human bronchial epithelial cell line, 16HBE14o-. METHODS: We used intracellular calcium imaging and short-circuit current measurement upon exposure of cells to P. aeruginosa LPS. RESULTS: Apical LPS stimulated intracellular calcium release and calcium entry and enhanced chloride secretion. This latter effect was significantly inhibited by CFTR(inh)-172 and BAPTA-AM (intracellular Ca(2+) chelator). CONCLUSIONS: Our data provides evidence for a new role of P. aeruginosa LPS in stimulating calcium entry and release and a subsequent chloride secretion via CFTR in human bronchial epithelium.Journal of cystic fibrosis: official journal of the European Cystic Fibrosis Society 07/2012; 12(1). DOI:10.1016/j.jcf.2012.06.007 · 3.82 Impact Factor
Conference Paper: Test and debug features of the RTO7 chip[Show abstract] [Hide abstract]
ABSTRACT: The Philips RTO7 chip consists of a complete receive chain from RF up to and including digital demodulation for Bluetooth-like radio communication. This paper describes both the implementation and verification of the test and debugs hardware for the digital core of the RTO7. The core-based DfT and DfD flow of the RTO7 is presented. The experimental results show that the RTO7 is both a fully testable and debuggable chip. State dump analysis results are also presented, showing that the state dumps obtained in the application are 100% stable, and match the state dumps made in simulation, and on the digital test system.Test Conference, 2005. Proceedings. ITC 2005. IEEE International; 12/2005
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ABSTRACT: A balance sheet describing the integrated homeostasis of secretion, absorption, and surface movement of liquids on pulmonary surfaces has remained elusive. It remains unclear whether the alveolus exhibits an intra-alveolar ion/liquid transport physiology or whether it secretes ions/liquid that may communicate with airway surfaces. Studies employing isolated human alveolar type II (AT2) cells were utilized to investigate this question. Human AT2 cells exhibited both epithelial Na(+) channel-mediated Na(+) absorption and cystic fibrosis transmembrane conductance regulator-mediated Cl(-) secretion, both significantly regulated by extracellular nucleotides. In addition, we observed in normal AT2 cells an absence of cystic fibrosis transmembrane conductance regulator regulation of epithelial Na(+) channel activity and an absence of expression/activity of reported calcium-activated chloride channels (TMEM16A, Bestrophin-1, ClC2, and SLC26A9), both features strikingly different from normal airway epithelial cells. Measurements of alveolar surface liquid volume revealed that normal AT2 cells: 1) achieved an extracellular nucleotide concentration-dependent steady state alveolar surface liquid height of ∼4 μm in vitro; 2) absorbed liquid when the lumen was flooded; and 3) secreted liquid when treated with UTP or forskolin or subjected to cyclic compressive stresses mimicking tidal breathing. Collectively, our studies suggest that human AT2 cells in vitro have the capacity to absorb or secrete liquid in response to local alveolar conditions.Journal of Biological Chemistry 11/2010; 285(45):34939-49. DOI:10.1074/jbc.M110.162933 · 4.60 Impact Factor