Upright transmission fluorescence microscope

The left lung of the mice was fixed in formalin for 48 h, then dehydrated, embedded and sectioned. Then, lung sections (4 μm) were prepared and stained with hematoxylin–eosin (H&E) and Masson’s trichrome. Quantification of pulmonary fibrosis was performed as described previously [45 (link)]. In brief, images were photographed with an upright transmission fluorescence microscope (Olympus) and analyzed by Image-Pro Plus Version 6.0 (Media Cybernetics, Inc, American). The software selection tool can select the entire lung tissue area and automatically calculate the total pixels, P_w, of the region. It can then use the same method to calculate the total pixels, P_f, of the fibrosis region (fibrosis ratio = fibrosis area total pixel, P_f/total lung total pixel, P_w).

Skin samples were fixed in 10% formalin, dehydrated, embedded in paraffin, and cut into sections 5 µm thick. After deparaffinizing in xylene and rehydrating using an alcohol series, the sections were stained with hematoxylin and eosin (H&E), Masson’s trichrome, and Picrosirius red (Solarbio, Beijing, China). Images were randomly photographed with an upright transmission fluorescence microscope (Olympus, Tokyo, Japan) and analyzed by Image-Pro Plus Version 6.0.

Left lungs were fixed in 10% formalin for 24 h and embedded in paraffin. Then, lung sections (4 μm) were prepared and stained with hematoxylin–eosin. Quantification of pulmonary fibrosis was performed as described previously (Jiang et al., 2004 (link)). In brief, images were photographed with an upright transmission fluorescence microscope (Olympus) and analyzed by Image-Pro Plus Version 6.0 (Media Cybernetics, Inc. American). The software selection tool can select the entire lung tissue area and automatically calculate the total pixel Pw of the region. It can then use the same method to calculate the total pixel Pf of the fibrosis region (fibrosis ratio = fibrosis area total pixel Pf/total lung total pixel Pw).