Fig 1: Inhibition of DPEP1 alleviates BLM-induced pulmonary fibrosis in mice, and its effect is related to the regulation of the TGF-β1/Smad3 pathway and arachidonic acid metabolism. (A) Representative microphotographs of HE and Masson’s trichrome-stained lung sections from the control, BLM, and BLM + cilastatin groups (n=4). Scale bar: 100 μm. (B) Quantification of Masson’s trichrome-positive fibrotic areas (n=4). (C) Hydroxyproline content measured within the lung tissues across the three cohorts (n=3). (D) Relative mRNA expressions of TNF-α, IL-1β, and IL-6 in murine lung tissues analyzed via qPCR (n=5). (E-H) Immunoblotting analysis evaluating intra-pulmonary inflammatory markers (COX-2, TGF-β1, and Smad3), with GAPDH serving as the loading control (n=3). (I-K) Tissue concentrations of arachidonic acid, PGE2, and cAMP within the lung (n=4). The uncropped Western blot images corresponding to (E) are provided in Figure S8. *, P<0.05; **, P<0.01; ***, P<0.001. BLM, bleomycin; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; HE, hematoxylin and eosin; IL, interleukin; NC, negative control; qPCR, quantitative polymerase chain reaction; TGF-β1, transforming growth factor beta 1.
Fig 2: Expression of DPEP1 mRNA of lung tissue and blood in patients with IPF. AE, acute exacerbation; IPF, idiopathic pulmonary fibrosis.
Fig 3: DPEP1 expression is upregulated and arachidonic acid content is decreased in a bleomycin-induced mouse model of pulmonary fibrosis. (A,B) Immunoblotting and densitometric analysis of fibronectin, collagen I, and DPEP1 protein expression in lung tissues harvested from control and BLM-treated mice, with GAPDH serving as the loading control (n=3). (C) Relative DPEP1 mRNA transcripts quantified via qPCR within the lungs of control and BLM cohorts (n=3). (D-F) Representative IHC microphotographs of DPEP1 in murine lungs (D), alongside corresponding statistical assessments of the IHC-positive area (E) and signal intensity (F) (n=3; scale bar: 100 μm). (G) Confocal images illustrating dual immunofluorescence staining of DPEP1 (green), CD31 (red), and DAPI (blue) in the respective lung tissues (scale bar: 50 μm). (H) Determination of endogenous arachidonic acid concentration within the lungs of both groups (n=4). The uncropped Western blot images corresponding to (A) are provided in Figure S7. *, P<0.05; **, P<0.01; ***, P<0.001. BLM, bleomycin; DAPI, 4',6-diamidino-2-phenylindole; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; IHC, immunohistochemistry; NC, negative control; qPCR, quantitative polymerase chain reaction.
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