Cytochrome c cyt c

Sodium hydrogen sulfide (NaHS), H₂O₂, PD98059 (an inhibitor of ERK1/2), LY294002 (an inhibitor of PI3K) and AG1478 (an inhibitor of EGFR) were purchased from Sigma Chemical Co. (St. Louis, MO, USA). The primary antibodies for anti-cleaved caspase-3 and -9, Bcl-2, cytochrome c (Cyt-c), c-myc, c-jun, c-fos and GAPDH were from Proteintech (Wuhan, China). The anti-ERK1/2, p-ERK1/2, PI3K, p-PI3K, GSK-3β, p-GSK-3β, Akt, p-Akt, HB-EGF, EGFR and p-EGFR antibodies were obtained from Cell Signaling Technology (Denver, CO, USA). Assay kits for lactate dehydrogenase (LDH) and creatine kinase (CK) were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). The Cell Counting Kit-8 (CCK-8) was obtained from Boster Bio-engineering Limited Company (Wuhan, China). The HB-EGF ELISA kit was purchased from Elabscience Biotechnology (Wuhan, China). All other chemicals were from Sigma or Santa Cruz.

After euthanasia, the liver and pancreas were processed, including 10% neutral formaldehyde fixing, paraffin embedding, 5 µm thickness sectioning, and H&E staining [26 (link)]. The pathological changes were observed using a stereomicroscope (SMZ25, Nikon, Tokyo, Japan). For immunohistochemistry, the sections of the liver were antigen retrieval, blocked with goat serum (37 °C for 20 min), and then incubated with primary antibodies. The antibodies included AMPK (1:100, Beyotime, Shanghai, China), Phosphorylated protein kinase RNA-like endoplasmic reticulum kinase (p-PERK, 1:200, Bioss, Beijing, China), Dynamic-related protein 1 (DRP1, 1:100, Beyotime), Cytochrome c (CytC, 1:100, Proteintech, Chicago, IL, USA), caspase 12 (1:100, Bioss), Inositol-requiring enzyme-1 (IRE1, 1:100, Proteintech) and Mitofusion 1/2 (MFN1/2, 1:100, Proteintech). Subsequently, the slides were visualized using DAB and then observed under a stereomicroscope (SMZ25, Nikon).

Sodium hydrogen sulfide (NaHS), N-acetyl-cysteine (NAC, an inhibitor of reactive oxygen species, ROS), Ammonium pyrrolidinedithiocarbamate (PDTC, a NF-κB inhibitor), AG 490 (a JNK2 inhibitor) were purchased from Sigma Chemical Co. (St. Louis, MO, USA). The anti-cleaved caspase-3 and -9, Bcl-2, cytochrome c (Cyt c), and GAPDH were from proteintech (Wuhan, China). Hoechst 33342 were from Santa Cruz (Bergheimer, Germany). The anti-IκBα-NF-κB and JNK2-STAT3 antibodies were obtained from Cell Signaling Technology (Danvers, USA). Assay kits for malondialdehyde (MDA), superoxide dismutase (SOD), Glutathione peroxidase (GSH-PX), catalase (CAT), lactate dehydrogenase (LDH), creatine kinase (CK) and ROS were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). All other chemicals were from Sigma or Santa Cruz.