Ap52269

5 μm thick HIR-injured lung sections embedded in paraffin and cells grown on sterile coverslips and then subjected to the HIR protocol were prepared for immunofluorescence assays. The samples were stained with anti-8-OHdG antibody (1 : 200; LS-C415095, LSBio) at 4°C overnight, followed by anti-rabbit or anti-mouse IgG (1 : 1000), according to the manufacturer's instructions. Cell nuclei were stained with DAPI. All images were captured using an EVOS FL fluorescence microscope (EVOS FL, Life Technology), and 10 randomly selected fields of each sample were semiquantified.
Immunohistochemical staining was performed to detect 4-hydroxynonenal (4-HNE) and Nrf2 expression in lung sections embedded in paraffin with anti-4-HNE antibody (1 : 100; ab46545, Abcam) and anti-Nrf2 antibody (1 : 100, AP52269, Abgent), respectively.

Western blotting was performed following standard procedures. Anti-Nrf2 antibody (1 : 100, AP52269, Abgent), anti-Brg1 antibody (1 : 100; ab110641, Abcam), anti-NQO1 antibody (1 : 100; ab28947, Abcam), anti-HO-1 antibody (1 : 100; sc-10,789, Santa Cruz), and secondary antibody (1 : 2000; Millipore) were used to detect protein expression. Anti-β-actin and anti-lamin B2 (Proteintech) were used at 1 : 5000. Images were acquired by a Tanon 5500 imaging system (Tanon, Shanghai). The images were scanned with the ImageJ scanning software, and the data were expressed as relative values to sham or control values.