Protein Expression Profile in IBS Rats

Protein extracts were obtained from colon tissue samples of control, IBS-Std, and IBS-KD rats using standard procedure [7 (link)]. Aliquots of 50 µg of total protein extracts from each sample were loaded into 4–15% pre-cast polyacrylamide gels (Bio-Rad, Milan, Italy) for western blot analysis. Anti-PrxIII (Ab FRONTIER, Seoul, Korea), anti-PGC-1α (Ab NOVUS, Centennial, CO, USA), anti-COX-2, anti-TLR-4, anti-PPAR-γ, anti-COX-4, anti-BECLin-1, anti-LC3, anti-VDAC1 (Abcam, Cambridge, UK), and anti-β-actin (Cell Signaling, Danvers, MA, USA) were used as primary antibodies. The proteins were detected by chemiluminescence (ECL, Thermo Scientific, Rockford, IL, USA), and the densitometric analysis of each protein-related signal was obtained using the Molecular Imager Chemidoc^TM (Bio-Rad, Milan, Italy) and normalized against β-actin expression.