IL-23 production from DCs and macrophages stimulated by HBsAg is dependent upon endocytosis.

<p>(<b>A</b>) Monocytes were isolated from PBMCs of healthy blood donors and mDCs were induced by culturing in the presence of GM-CSF (50 ng/mL) and recombinant human IL-4 (5 ng/mL) for five days. Then, the mDCs were stimulated by HBsAg with or without ammonium chloride (10 µM), chloroquine (10 mM), Dynasore (80 µM), or HBsAg antibody (50 µg/mL) for 40 hours. ELISA was used to detect the concentration of IL-23 in the supernatants. The data represent one of three independent experiments with similar results. Error bars indicate SD. ***<i>P</i><0.001 vs. all other groups. DCs (<b>B</b>) and macrophages (<b>C</b>) were obtained from PBMCs for different stimulations and stainings. (<b>a</b>) Cells were stimulated with HBsAg (2 µg/mL) for 45 min and stained by isotype IgG (FITC). (<b>b</b>) Cells were directly stained with anti-MR mAb (FITC) without any stimulations. (<b>c</b>) Cells were stimulated with HBsAg (2 µg/mL) for 45 min and stained by anti-HBsAg mAb (FITC). (<b>d</b>) Cells were pretreated with MR-blocking antibody (10 µg/mL) for 30 min before the cells were stimulated by HBsAg (2 µg/mL) for 45 min and stained with anti-HBsAg (FITC). (<b>e</b>) Cells were pretreated with chloroquine (10 mM) for 10 min before stimulation by HBsAg (2 µg/mL) for 45 min and staining with anti-HBsAg (FITC). mDCs were stimulated by HBsAg (2 µg/mL) for 45 minutes, and the respective co-localizations of HBsAg with MR (<b>D</b>) or endosomal marker LysoTracker (<b>E</b>) were detected by confocal microscopy. (magnification 400×).