Anti rabbit ppar γ

Liver tissues were stored at −70 °C. Tissue samples were homogenized in ice-cold RIPA lysis buffer (Millipore, Billerica, MA, USA) for protein extraction. Tissue debris was removed by centrifugation, and the resulting supernatants were collected and analyzed for protein concentration by the BCA protein assay kit (Thermo Scientific, Bartlesville, OK, USA). The protein was separated on a 10% SDS polyacrylamide gel and then transferred to nitrocellulose membranes (Hybond, GE Healthcare Life Sciences, Little Chalfont, UK). The membranes were incubated with specific primary antibodies overnight at 4 °C. The primary antibodies included anti-mouse β-actin, anti-mouse PPAR-α (Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti-rabbit TNF-α, anti-rabbit PPAR-γ, and anti-rabbit GLUT4 (Cell Signaling Technology, Inc., Beverly, MA, USA). After washing, the membranes were allowed to react with diluted horseradish peroxidase-conjugated secondary antibodies, including goat anti-rabbit IgG antibody and horse anti-mouse IgG (Cell Signaling Technology, Inc., Beverly, MA, USA) at room temperature for 2 h. An enhanced chemiluminescence system (SuperSignal, Thermo Scientific, Rockford, IL, USA) was used to visualize antibody–antigen complexes.

Specific primary and secondary antibodies were used for Western blotting. Antibodies against anti‐rabbit AMPK, anti‐rabbit phospho‐AMPK, anti‐rabbit phospho‐CaMKK, anti‐rabbit phospho‐LKB1, anti‐rabbit phospho‐protein kinase A (PKA), anti‐rabbit ACC, anti‐rabbit phospho‐ACC, anti‐rabbit Sirtuin1 (Sirt1), anti‐rabbit FAS, anti‐rabbit TGFβ, anti‐rabbit phospho‐Smad2 (Ser465/467)/Smad3 (Ser423/425) (Smad2/3), anti‐rabbit α‐smooth muscle actin (α‐SMA), anti‐rabbit HSL, anti‐rabbit phospho‐HSL, anti‐rabbit C/EBPα, anti‐rabbit C/EBPβ, anti‐rabbit PPARγ, anti‐rabbit IgG, and anti‐mouse IgG were purchased from Cell Signaling Technology (Beverly, MA). Anti‐mouse β‐actin was obtained from Sigma as an internal control.