Figure 1.

Mechanics-induced YAP localization and CD44 expression in fibroblasts. (A-B) Western blot analysis of YAP in nuclear extracts of lung tissues after different treatments at day 56. Lamin B was used as a loading control. Data shown are representative of three independent experiments. Error bars indicate mean ± SD (**, P < 0.01). (C) Distribution of YAP was determined by immunohistochemical staining in the lungs after saline or crystalline silica treatment at day 56. Arrows point to cells in the lung with silicotic lesions with predominantly nuclear YAP staining. Scale bar, 50 μm. (D, F) NIH-3T3 fibroblasts cultured on soft (1 kappa) or stiff (60 kappa) gel-coated coverslips were immunostained with an antibody recognizing YAP (D) and CD44 (F). The percentage of cells with predominantly nuclear YAP staining was quantified at D. Nuclei were counter-stained with DAPI. Scale bar, 50 μm (n=3; ***, P < 0.001). (E) Bioinformatic analysis of protein-protein interaction networks in STRING v.10. A screenshot from STRING shows a network associated with CD44 and YAP. The red nodes show extracellular matrix proteins; the green nodes show the association between CD44 and YAP; the blue nodes show cell signaling proteins in cytoplasm associated with YAP. (G) Immunofluorescent analysis of α-SMA and CD44 in lung sections with silicotic lesions after saline or crystalline silica treatment at day 56. Arrows point to fibroblasts expressing CD44. Representative images of the staining are shown (n=3). Scale bar, 100 μm.