Pulmonary Tissue Analysis of ASD-Induced Inflammation

After sampling BALF, pulmonary tissues were fixed, and then a paraffin block was prepared. The paraffin block was sectioned (4 µm) and then stained with hematoxylin and eosin (Sigma-Aldrich, St. Louis, MO, USA). To explore the effects of LTE on the expression of HO-1, NF-κB, Nrf2, and 8-OHdG in ASD-exposed pulmonary tissue, immunohistochemistry was conducted using a commercial kit (Vector Laboratories, Burlingame, CA, USA) as previously described [15 (link)]. The following primary antibodies were used: anti-rabbit p-p65 (dilution 1:200, Thermo Fisher Scientific), anti-rabbit HO-1 (dilution 1:200, Abcam), anti-mouse Nrf2 (dilution 1:200, Santa Cruz, CA, USA), and anti-mouse 8-OHdG (dilution 1:200, Santa Cruz). Quantitative analysis of the pulmonary inflammation and density of protein band was conducted using IMT i-Solution (IMT i-Solution Inc., Vancouver, BC, Canada).