Box plots corresponding to four peaks differentially displayed.

<p>Two sphingomyelin (m/z 489.3 and 725.5, upper panels) and two phosphatidylcholine (m/z 782.6 and 810.6, lower panels) species are represented. Relative intensity corresponds to the area of each peak of the spectrum related to the total ion count. Box plots are constructed from log-transformed relative intensity data.</p

Peaks differentially displayed in either healthy controls <i>vs</i> CF patients (*) or in CF mild <i>vs</i> CF severe patients (**) with p<0.05 after T-test analysis.

<p>All peaks are [M+H]⁺ except 489.27 and 725.53, which correspond to [M+Na]⁺. # Theoretical identity (not confirmed by MSⁿ).</p

Effect of TNF-α and DRM destabilization on AA release.

<p>Calu-3 cells were incubated overnight with either ³H-labelled AA, treated with 100 U/mL TNF-α for 10 min, 10 mM mβCD for 1 h, with or without preincubation with 15 µM pyrrolidine for 45 min, or with a combination of the different treatments. For combined treatments, TNF-α was added for the last 10 min of incubation. After incubation, supernatants were collected and radioactivity measured by a scintillation counter. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0007116#s3" target="_blank">Results</a> are expressed as percent increment with respect to control. Asterisks denote p<0.05 with respect to control unless indicated otherwise, n = 3.</p

Effect of TNF-α and DRM destabilization on eicosanoid production.

<p>Calu-3 cells were treated with either 100 U/mL TNF-α for 10 min, 10 mM mβCD for 1 h, with or without preincubation with 15 µM pyrrolidine for 45 min, or with a combination of the different treatments. For combined treatments, TNF-α was added for the last 10 min of incubation. After incubation the supernatant was collected, either immediately or after 3 h of incubation in fresh medium, and subjected to ELISA for LTB4 and PGE2 determination. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0007116#s3" target="_blank">Results</a> are expressed as percent of control values. Asterisks denote p<0.05 with respect to control, n≥3.</p

Effect of CFTR inhibition on eicosanoid, AA and IL-8 release.

<p>A: Iodide efflux (CFTR activity) measurements on Calu-3 cells. Effect of 10 min incubation with 10 µM Inh172 in Calu-3 cells (left). Effect of forskolin activation (right). <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0007116#s3" target="_blank">Results</a> are representative of three experiments. B: Calu-3 cells were treated with either 20 µM Inh172 or Gly-101 for 20 min. In a separate experiment, HeLa cells were treated with 20 µM Inh172 for 20 min. The supernatant was collected right after treatment and subjected to ELISA for LTB4 and PGE2 determination. C: Calu-3 cells were incubated with or without 20 µM Inh172 for 20 min. After incubation, supernatants were removed and fresh DMEM medium containing fetal calf serum was added. After 3 h supernatants were harvested for IL-8 determination. All results are expressed as percent of control values.</p

Impact of TNF-α and DRM destabilization on IL-8 release.

<p>IL-8 production by Calu-3 cells after proinflammatory stimulation, functional inhibition of CFTR and DRM disruption. Calu-3 cells were incubated with either 100 U/mL TNF-α for 10 min, 10 mM mβCD for 1 h, or with a combination of both treatments. After incubation, supernatants were removed and fresh DMEM medium containing fetal calf serum was added. After 3 h of incubation, supernatants were harvested for IL-8 determination. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0007116#s3" target="_blank">Results</a> are expressed as percent of control values. Asterisks denote p<0.05 with respect to control, n≥3.</p