FIGURE 10.

Exosomes of skin mast cells induce features of AD in HaCaT and dermal fibroblast cells. (A) Skin mast cells were pretreated with GW4869 (20 μM) for 2 h, and treatment with DNCB (5 μM) for 1 h was followed. HaCaT cells and dermal fibroblast cells were treated with the culture medium of skin mast cells for 12 h. Immunoblot was performed. Representative blots of three independent experiments were shown. (B) Exosomes isolated from skin mast cells treated with or without DNCB (5 μM) for 24 h were visualized by negative staining electron microscopy. (C) Shows size distributions of exosomes employing nanoparticle tracking analysis (NTA). (D) Immunoblot shows the presence of CXCL13 in the exosomes of DNCB-treated skin mast cells. Representative blots of three independent experiments were shown. (E) Exosomes were isolated from skin mast cells treated with or without DNCB for 1 h. Exosomes (5 μg) were then added to HaCaT cells or skin dermal fibroblast cells for 24 h followed by immunoblot and invasion assays. *, p < 0.05; ***, p < 0.001. Average values of three independent experiments were shown. (F) Immuno-gold staining images using anti-TSG101, a known membrane marker for the exosomes, and anti-CXCL13 antibody. Twenty-five and 10 nm gold particles show the presence of TSG101 and CXCL13, respectively.