Nestin-GFP–/NG2-DsRed+cells, but not Nestin-GFP+/NG2-DsRed+cells, increase and participate in pulmonary fibrosis. (A) Representative photomicrographs of lung sections from Nestin-GFP/NG2-DsRed mice (control) showing blood vessels with CD31+ endothelial cells and pericytes (NG2-DsRed+). All panels show identical areas with CD31 staining, NG2-DsRed, Nestin-GFP+, Hoechst, and combined fluorescent images. Nestin-GFP– and Nestin-GFP+ pericytes (NG2-DsRed+) surround capillaries. Region in yellow box shows type-1 and type-2 pericytes close to CD31+ blood vessels, magnified in (C). (B) Immunohistochemical staining with an antibody against type I collagen (Col I) in lung sections from Nestin-GFP/NG2-DsRed double-transgenic mice showing matrix deposition 2 weeks after bleomycin treatment. All panels show the same lung area with Col I, NG2-DsRed, Nestin-GFP+, Hoechst, and merged fluorescent images. Region in yellow box shows area with dense collagen accumulation at the lung injury site, magnified in (D). (C) Type-1 and type-2 pericytes close to CD31+ blood vessels, magnified from (A). (D) Dense collagen accumulation at the lung injury site, magnified from (B). Note that some type-1 pericytes (Nestin-GFP–/NG2-DsRed+) produce collagen type I after lung injury, indicated by a white arrow. (E) Number of type-1 and type-2 pericytes before and after pulmonary injury (n = 3 mice; 10 lung sections from each). (F) Percentage of cells producing type I collagen in NG2-DsRed–/Nestin-GFP– cell populations, and type-1 and type-2 pericytes. Scale bars = 100 μm.