Mnsod

Cells were harvested and lysed for total protein extraction. Protein concentration was determined using Bio-Rad DC protein assay kit (Bio-Rad Laboratories) according to manufacturer's protocol. Aliquots of equal amounts of protein from the cell lysates were subjected to Western blot analysis. Antibodies used include those specific for MnSOD(BD Biosciences), catalase (proteintech), caspase3 (Cell Signaling Technology), caspase9 (Cell Signaling Technology), β-catenin (Cell Signaling Technology) and β-actin (Sigma, MO, USA).

Protein samples were prepared from liver homogenates in Laemmli sample buffer, run on SDS-polyacrylamide gels (4-15% TGX stain-free gel, Bio-Rad), and transferred to the polyvinylidene difluoride (PVDF) membrane. The membranes were blocked, incubated with primary antibodies overnight at 4 °C, followed by secondary antibodies, and developed using the chemiluminescence imaging system (Bio-Rad). Following primary antibodies were used: OPA1 (BD Biosciences, 612606; 1:1000); caspase-3 (Cell Signaling, 9662; 1:1000), PARP-1 (Cell Signaling, 9542; 1:1000), β-actin (Sigma, A1978; 1:40000), TOM20 (Proteintech, 11802-1-AP; 1:1000), cytochrome c (BD Biosciences, 556432; 1:5000), elF2α (Cell Signaling, 9722; 1:1000), phospho-elF2α (Cell Signaling, 9721; 1:1000), FGF21 (Proteintech, 26272-1-AP; 1:1000), LC3 A/B (Cell Signaling, 4108; 1:1000), PGC1α (Invitrogen, PA5-38022; 1:500), OMA1 (Santa Cruz, sc-515788; 1:100), and mitochondria total OXPHOS rodent WB cocktail (Abcam, ab110413; 1:1000). JNK (Cell Signaling, 9252; 1:1000), p-JNK-Thr183/Tyr185 (Cell Signaling, 9255; 1:500), MCU (Sigma, HPA016480; 1:1000), NCLX (Proteintech, 21430-1-AP; 1:1000), CypD (Proteintech, 18466-1-AP; 1:1000), MnSOD (BD Biosciences, 611580; 1:1000), GPx1/2 (Santa Cruz Biotechnology, sc-133160; 1:200), and CYP2E1 (Proteintech, 19937-1-AP; 1:300).