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Figure 1 | Respiratory Research

Figure 1

From: Oxidative stress augments toll-like receptor 8 mediated neutrophilic responses in healthy subjects

Figure 1

Detection of toll-like receptor (TLR) 8 in human polymorphonuclear cells (PMNs), and the effects of TLR 7/8 ligand R848 on interleukin(IL)-8 release. (A) TLR8 in PMN was detected by immunocytochemistry. Left panel indicates isotype control. Right panel shows TLR8 immunoreactivity in PMN. (Original magnification: × 400, Scale bars = 10 μm). (B) TLR8 expression was analyzed by flow-cytometry. PMNs were stained by anti-human TLR8 (solid lines) or the isotype control (gray histograms) in the permeabilized (left panel) and unpermeabilized condition (right panel). Left panel indicates both intercellular and cell surface expression of TLR8. Right panel shows cell surface expression alone. (C-F) Effect of R848 on the release of IL-8, and effect of bafilomycin or dexamethasone on the R848-induced IL-8 release from PMN. (C) PMNs were treated with 10 μM R848. The media were harvested at various time points and assayed for IL-8 by ELISA. (D) PMNs were treated for 24 hrs with R837, a ligand of TLR7, or various concentrations of R848, a ligand of TLR 7/8. Media were assayed for IL-8 by ELISA. (E, F) PMNs were treated with 10 μM R848 or vehicle in the presence of various concentrations of bafilomycin, an inhibitor of endosomal acidification (E), or dexamethasone (F). Media were assayed for IL-8 by ELISA. All values are mean values ± SEM of three to four separate experiments. *p < 0.05, **p < 0.01, compared with the values of control; +p < 0.05, ++p < 0.01, compared with the values of the vehicle-pretreated and 10 μM R848-treated group.

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