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.

P. aeruginosa LPS stimulates calcium signaling and chloride secretion via CFTR in human bronchial epithelial cells

“…The role of PLC in generating calcium release has been described in airway epithelial cells and LPS can stimulate PLC in alveolar epithelial cells [4]. It cannot be excluded that such mechanism occurs in 16HBE14o − cells upon LPS stimulation.…”

“…P. aeruginosa inhibits sodium absorption via ENaC in alveolar epithelial cells [4]. However, it has been reported that airway epithelial cell lines have a tendency to lose their amiloride-sensitivity in culture and that ENaC activity varies according culture conditions of the 16HBE14o − cells [25].…”

“…LPS from P. aeruginosa inhibits sodium absorption via ENaC in mammalian alveolar epithelial cells [4]. In contrast, LPS from Salmonella enterica stimulated sodium absorption by increasing Na/K ATPase expression in guinea-pig tracheal epithelium [5].…”

Purinergic Signaling in the Airways

“…Flagellin of P. aeruginosa inhibits ENaC expression in airway epithelium, reducing Na ϩ absorption to enhance local mucociliary clearance, a mechanism that is attenuated in CF (Kunzelmann et al, 2006). The action of LPS on ENaC was mediated in part by the protein kinase C and phospholipase C pathways and by purinergic signaling (Boncoeur et al, 2010). In intact distal bronchi of porcine lungs, UTP stimulated Cl Ϫ secretion by a Ca 2ϩ -independent mechanism and inhibited Na ϩ absorption by a Ca 2ϩ -dependent mechanism; both effects are likely to favor increased hydration of the airway surface and may therefore be beneficial in CF .…”

Human Alveolar Type II Cells Secrete and Absorb Liquid in Response to Local Nucleotide Signaling

“…Similarly, purinergic signaling may contribute to the integrated control of ion/liquid homeostasis within the alveolar space. Nucleotide receptors and adenosine receptors (12,22) have been identified in mouse and rat AT2 cells, and ATP or UTP has been reported to regulate Na ϩ and Cl Ϫ transport in cultured rat AT2 cells (23,24). Whether a role for nucleotide release in response to the mechanical stresses of tidal breathing plays a role in regulating human alveolar liquid homeostasis is as yet unknown.…”