2 7 dichlorodihydrofluorescein diacetate dcfda fluorescence kit

Dulbecco’s Modified Eagle’s Medium (DMEM) with glucose and without glucose and all medium additives were obtained from Gibco BRL (Grand Island, NE, USA). Methylthiazolyldiphenyl-tetrazolium bromide (MTT), glucose, metformin, N-acetyl-l-cysteine (NAC), FH535, and 2-7-Dichlorodihydrofluorescein diacetate (DCFDA) fluorescence kit were purchased from Sigma Aldrich (St. Louis, MI, USA). PD98059 was purchased from Calbiochem (La Jolla, CA, USA). LDH, Caspase-3 activity assay kit and ATP measurement kits were purchased from BioVision (Milpitas, CA, USA). The antibodies specific for ERK, p-ERK, GSK3β, p-GSK3β, and PARP were purchased from Cell Signaling Technology (Danvers, MA, USA). The antibodies specific for Bax and Bcl-2 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA), and the antibody specific for GAPDH was purchased from Thermo Fisher (Rockford, IL, USA).

Cearoin was isolated from the ethanol extract of dried heartwoods of D. odorifera as described previously [27 (link)]. Cearoin stock solution was prepared in DMSO and stored at −20 °C, which was diluted using serum-free media during the treatment. Similarly, stock solution of anticancer drug cyclosphosphamide was prepared in DMSO and diluted in serum-free media during treatment. Dulbecco’s Modified Eagle’s Medium (DMEM) and all medium additives were obtained from Gibco BRL. Cyclophosphamide, Sodium nitrite, N-Acetyl-l-cysteine (NAC), NG-methyl-l-arginine acetate (L-NMMA), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), and 2-7-dichlorodihydrofluorescein diacetate (DCFDA) fluorescence kit were purchased from Sigma Aldrich (St Louis, MI, USA). PD98059 was purchased from Calbiochem (La Jolla, CA, USA). Caspase-3 activity kit was purchased from Biovison (Milpitas, CA, USA). The antibodies specific for α-spectrin, PARP, LC3B, p-ERK, ERK, p-JNK and JNK were purchased from cell signaling Technology (Danvers, MA, USA). The β-actin was purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA) and GAPDH was purchased from Thermo Fisher (Illinois, Rockford, AL, USA).