Fig. 8

Inhibiting Bach1-SAT1 Pathway Alleviates FMP-Induced Pulmonary Fibrosis in vivo. Embryonic day 17 (ED17) chicken embryos were exposed to HPPE via air cell injection, and then 24 h-old chickens were exposed to 0.6 mg/kg FMP via aerosol inhalation. Bach1 and SAT1 expressions in chicken lung tissues were assessed with qRT-PCR and immunohistochemistry. Extent of pulmonary fibrosis were determined with H&E, Masson and Sirius red staining. Protein expression levels of ALOX15, SAT1, GPX4 and TGF-β were assessed with western blotting. Data were expressed as mean ± SD, N = 5 per group; * indicates significant difference from the blank control group (p < 0.05); ** indicates significant difference from the blank control group (p < 0.01); # indicates significant difference from 0.6 mg/kg FMP exposure group (p < 0.05); ## indicates significant difference from 0.6 mg/kg FMP exposure group (p < 0.01). (A) Schematic of animal treatments and qRT-PCR analysis of Bach1 and downstream HOMX1 expression in chicken lung tissues following air cell-injection of HPPE (4 mg/kg) at ED17 (N = 3). (B) Representative IHC images showing Bach1 and SAT1 protein expression in chicken lung tissues exposed to 0.6 mg/kg FMP, with or without pretreatment of 4 mg/kg HPPE, along with semi-quantifications (N = 4). Each image was captured under a 400× optical microscope, with the scale bar in the lower-right corner representing 100 μm. The lower-left corner of the image indicates a 200× magnification of the field of view, with the scale bar representing 100 μm. (C) Representative pathological pictures of chicken lung tissue sections from animals exposed to 0.6 mg/kg FMP, with or without pretreatment of 4 mg/kg HPPE, Yellow arrows represent positive areas. (D) Quantifications of Ashcroft score and fibrotic lesion ratio from C(N = 5). (E) Representative western blotting images for ALOX15, SAT1, GPX4 and TGF-β in the lung tissues samples from animals exposed to 0.6 mg/kg FMP, with or without pretreatment of 4 mg/kg HPPE, along with semi-quantifications (N = 4)