Volume-sensitive release of organic osmolytes in the human lung epithelial cell line A549: role of the 5-lipoxygenase

Abstract

Pathophysiological conditions challenge cell volume homeostasis and perturb cell volume regulatory mechanisms leading to alterations of cell metabolism, active transepithelial transport, cell migration, and death. We report that inhibition of the 5-lipoxygenase (5-LO) with AA861 or ETH 615-139, the cysteinyl leukotriene 1 receptor (CysLT₁) with the antiasthmatic drug Zafirlukast, or the volume-sensitive organic anion channel (VSOAC) with DIDS blocks the release of organic osmolytes (taurine, meAIB) and the concomitant cell volume restoration following hypoosmotic swelling of human type II-like lung epithelial cells (A549). Reactive oxygen species (ROS) are produced in A549 cells upon hypotonic cell swelling by a diphenylene iodonium-sensitive NADPH oxidase. The swelling-induced taurine release is suppressed by ROS scavenging (butylated hydroxytoluene, N-acetyl cysteine) and potentiated by H₂O₂. Ca²⁺ mobilization with ionomycin or ATP stimulates the swelling-induced taurine release whereas calmodulin inhibition (W7) inhibits the release. Chelation of the extracellular Ca²⁺ (EGTA) had no effect on swelling-induced taurine release but prevented ATP-induced stimulation. H₂O₂, ATP, and ionomycin were unable to stimulate the taurine release in the presence of AA861 or Zafirlukast, placing 5-LO and CysLT₁ as essential elements in the swelling-induced activation of VSOAC with ROS and Ca²⁺ as potent modulators. Inhibition of tyrosine kinases (genistein, cucurbitacin) reduces volume-sensitive taurine release, adding tyrosine kinases (Janus kinase) as regulators of VSOAC activity. Caspase-3 activity during hypoxia is unaffected by inhibition of 5-LO/CysLT₁ but reduced when swelling-induced taurine loss via VSOAC is prevented by DIDS excess extracellular taurine, indicating a beneficial role of taurine under hypoxia.

Keywords: 5-lipoxygenase; cell volume regulation; cysteinyl leukotriene 1 receptor; hypoxia; ischemia; lung adenocarcinoma; reactive oxygen species; taurine homeostasis.