Fulton’s index [the weight of the right ventricle (RV) divided by the weight of the left ventricle (LV) + septum] was determined to assess RVH. Additionally, RV and LV + septum weights were normalized to body weight (22 (link)).
Plasma IL-1b was quantified using the Rat IL-1 beta Platinum ELISA kit (Thermo Fisher Scientific, Waltham, MA, United States) according to the manufacturer’s instructions.
Western blots were performed on lung tissue as previously described (12 (link)). Briefly, lung tissue was suspended in RIPA buffer containing protease and phosphatase inhibitors and sonicated on ice. Protein content was determined by the Bradford method and Western blotting performed using 1:500 dilution of mouse anti-IκB-α antibody (sc-1643, Santa Cruz Biotechnology, Dallas, TX, United States) at 4°C overnight followed by a 60 min incubation with an anti-mouse secondary antibody conjugated to horseradish peroxidase (Santa Cruz). Blots were then probed for β-actin (ab6276, 1:4000, Abcam, Cambridge, MA, United States) for 60 min at room temperature. Chemiluminescence generated by Super Signal West Femto substrate (Thermo Fisher Scientific) was detected and quantified using a Kodak Image Station and software. Signals were normalized to β-actin and expressed as fold change relative to OR animals.
Plasma IL-1b was quantified using the Rat IL-1 beta Platinum ELISA kit (Thermo Fisher Scientific, Waltham, MA, United States) according to the manufacturer’s instructions.
Western blots were performed on lung tissue as previously described (12 (link)). Briefly, lung tissue was suspended in RIPA buffer containing protease and phosphatase inhibitors and sonicated on ice. Protein content was determined by the Bradford method and Western blotting performed using 1:500 dilution of mouse anti-IκB-α antibody (sc-1643, Santa Cruz Biotechnology, Dallas, TX, United States) at 4°C overnight followed by a 60 min incubation with an anti-mouse secondary antibody conjugated to horseradish peroxidase (Santa Cruz). Blots were then probed for β-actin (ab6276, 1:4000, Abcam, Cambridge, MA, United States) for 60 min at room temperature. Chemiluminescence generated by Super Signal West Femto substrate (Thermo Fisher Scientific) was detected and quantified using a Kodak Image Station and software. Signals were normalized to β-actin and expressed as fold change relative to OR animals.