Fig. 3
Exosomes derived from SiO2-exposed macrophages promote inflammatory activation in monocytes/macrophages (A) The purified exosomes were labelled with PKH26 dye and then added to RAW264.7 macrophages and THP-1 macrophages. The uptake of PKH26-labelled exosomes was observed by fluorescence microscopy. The scale bar represents 20 μm. (B) Representative image of the morphological changes in RAW264.7 macrophages and THP-1 monocytes treated with PBS, NC-Exo, SiO2-Exo or SiO2 + GW4869-Exo. The scale bar represents 50 μm. (C-D). RT–PCR analysis of the expression of IL-1β, IL-6, TNF-α and IL-10 in RAW264.7 macrophages and THP-1 monocytes treated with PBS, NC-Exo, SiO2-Exo or SiO2 + GW4869-Exo. (E). ELISA analysis of the levels of proinflammatory factors in the supernatant of RAW264.7 macrophages treated with PBS, SiO2-Exo or SiO2 + GW4869-Exo. n = 3 each group. (F-G). Flow cytometric analysis of the proportions of iNOS+ cells among RAW264.7 macrophages and THP-1 monocytes after PBS, NC-Exo, SiO2-Exo or SiO2 + GW4869-Exo treatment. (H-I). Immunofluorescence analysis of iNOS expression in RAW264.7 macrophages (H) and THP-1 monocytes (I) after PBS, NC-Exo, SiO2-Exo or SiO2 + GW4869-Exo treatment. The scale bar represents 50 μm. The data are representative of three individual experiments and expressed as the mean ± SEM. The data were analysed by two-way ANOVA. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, ns = not significant. Abbreviations SiO2 = silica dust; NC-Exo = exosomes derived from cells without SiO2 exposure; SiO2-Exo = exosomes derived from SiO2-exposed macrophages; SiO2 + GW4869-Exo = exosomes derived from SiO2-exposed macrophages treated with GW4869 (10 µM)