P ikkα β

Antibodies from Affinity Biosciences: p-Smad1/5/8 (1:1000, #AF8313), Smad1/5/8 (1:1000, #AF0614), PPARγ (1:1000, #bs-4590R), UCP1 (1:1000, #72298). Antibodies from Beyotime: HA tag (1:1000, #AH158), Flag tag (1:1000, #AF519). Antibodies from Sangon Biotech: p-ACC (1:1000, #D155180), ACC (1:1000, #D155300). Antibodies from Abcam: BMP8A (1:1000, #154373). Antibodies from ZenBio: p-AMPK (1:1000, #R26252), AMPK (1:1000, #380431), PGC-1α (1:1000, #381615). Antibodies from CWBIO: goat anti-Rabbit IgG HRP secondary antibody (1:4000, #CW0103S), goat anti-Mouse IgG HRP Conjugated (1:4000, # CW0102). Antibodies from Bioss: p38 MAPK (1:1000, #bs-0637R), p-p38 MAPK (1:1000, #bs-2210R), β-actin (1:2000, #bs-0061R), p-JNK (1:1000, #bs-1640R), JNK (1:1000, #bs-2592R), C/EBPα (1:1000, #AF6333), p-Smad2/3 (1:1000, #AF3367), Smad2/3 (1:1000, #AF6367), p-ERK1/2 (1:1000, #AF1015), ERK1/2 (1:1000, #AF0155), p-p65 (1:1000, #AF2006), p65 (1:1000, #AF5006), p-IKKα/β (1:1000, #AF3013), IKKα/β (1:1000, # AF6014).

Proteins were extracted from injured abdominal arteries and equivalent amounts of protein from each sample were electrophoresed on a 5% discontinuous polyacrylamide minigel, and transferred onto nitrocellulose membranes. The membranes were blocked at 4 °C overnight with 5% non-fat dry milk, then incubated with antibodies for NF-κB (Thermo Fisher, USA), IκB-α, P-IκBα, IKK-α, and P-IKKα/β (all from BIOSS, China) at 4 °C overnight. The membranes were washed and incubated with secondary antibodies for 30 min at room temperature. The immune complexes were visualized using the enhanced chemiluminescence method. Protein levels were quantified by using PhotoShop (version 18.0, Adobe Corporation, USA).