Fig. 4
Characteristics of the alveolar epithelial cells in the Fibrotic pulmonary dust foci (fPDF) in rat lungs after inhalation exposure to TiO2 NP (8 mg/m3). A: Double staining for the AT2 TF marker TTF1 (green in the nucleus) and the AT2 differentiation marker LPCAT1 (brown in the cytoplasm) in the fPDFs. Staining of severe (B) and mild (C) lesions indicate the existence of TTF1-negative AT2 cells. Red arrows indicate TTF1-negative and LPCAT1-positive cells. Bar: 200 μm in A, 25 μm in B and C. D: Double staining for the AT1 TF marker Hop (green in the nucleus) and the AT2 differentiation marker LPCAT1 (brown in the cytoplasm) in the fPDFs. Staining of severe (E) and mild (F) lesions indicate the existence of Hop-positive AT2 cells (Red arrows). Bar: 200 μm in D, 25 μm in E and F. G: Triple staining for Sox9 (green in the nucleus), the AT1 differentiation marker Pdpn (brown in the cell membrane), and the AT2 differentiation marker LPCAT1 (red in the cytoplasm) in the fPDFs. H: High magnification of Fig. 4G indicates the presence of cells that were positive for Sox9 and negative for the lineage markers LPCAT1 and PDPN in the dysplastic gland within the fPDF (Red arrows). I: Triple staining for bronchial epithelial lineage marker Sox2 (green in the nucleus), the AT1 differentiation marker Pdpn (brown in the cell membrane), and the AT2 differentiation marker LPCAT1 (red in the cytoplasm) in the fPDFs. J: High magnification of Fig. 4I indicates the presence of Sox2-positive AT2 cells in the bronchiolar-alveolar duct junction. Bar: 200 μm in G, I. K: Summary of immunostaining results. Lin: Abbreviation for lineage, referring to AT1 or AT2 cells. pos: positive, neg: negative, AEC: alveolar epithelial cell