Jc 1 t 3168

About 5 × 10⁵ cells CaSki or SiHa cells were seeded onto a 60 mm dish containing 5 mL of culture medium. ABT-737 (2.5 or 5.0 µM) or As₂O₃ (2.0 µM) was added 48 hr. At 30 min prior to harvesting, these cells were stained at 37 °C using a 2.5-µM final concentration of 5,5,6,6′-tetrachloro-1,1,3,3′-tetraethylbenzimi-dazolylcarbocy-anine iodide dye (JC-1, T3168, Invitrogen, Carlsbad, CA, USA; Thermo Fisher Scientific, Inc., San Jose, CA, USA) to detect the MMP by fluorescence microscopy and flow cytometry using CellQuest 5.1 software (BD Biosciences, San Jose, CA, USA).

Dulbecco’s modified eagle’s medium (DMEM), trypsin-EDTA, penicillin-streptomycin and dimethyl sulfoxide (DMSO), MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) (M2128), Hoechst 33258 (B1155), Crystal Violet (C0755), and DCF (D6883) were purchased from Sigma-Aldrich (St. Louis, MO, USA). All tissue culture-ware was from Nunc (Roskilde, Denmark). Biotin Blocking System, peroxidase substrate (DAB), and peroxidase buffer were from DAKO (Glostrup, Denmark). Proteinase inhibitors were from Roche (Madrid, Spain). ECL Western blotting substrate was from Pierce (Thermo Fisher Scientific, Rockford, IL, USA). BCL-2 siRNA (h) (sc-29214), BCL-xL siRNA (h) (sc-43630), and scrambled controls were purchased from Santa Cruz Biotechnology (Dallas, TX, USA), while a second siRNA for BCL-2 (ID#s1915) and for BCL-xL (ID#s1920) were obtained from Ambion Life technologies (Carlsbad, CA, USA). Lipofectamine2000 (11668-027), Novex Sharp Pre-stained Protein Standard (LC5800), and JC-1 (T-3168) were from Invitrogen Life Technologies (Carlsbad, CA, USA). Sorafenib (BAY 43-9006, Nexavar) and Regorafenib (BAY 73-4506, Stivarga) are manufactured by Bayer. A-1331852 and ABT-199 (Venetoclax) were purchased from Selleckchem (Houston, TX, USA).

Dulbecco's Modified Eagle's Medium (DMEM), trypsin-EDTA, penicillin-streptomycin and dimethyl sulfoxide (DMSO), MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) (M2128), Hoechst 33258 (B1155), Crystal Violet (C0755), and DCF (D6883) were purchased from Sigma-Aldrich (St. Louis, MO). All tissue culture-ware was from Nunc (Roskilde, Denmark). Biotin Blocking System, peroxidase substrate (DAB) and peroxidase buffer were from DAKO (Glostrup, Denmark). Aquatex was from Merck (Darmstadt, Germany). The ABC kit was from Vecstain (Burlingame, CA). Proteinase inhibitors were from Roche (Madrid, Spain). ECL western blotting substrate was from Pierce (Thermo Fisher Scientific, Rockford, IL). BCL-2 siRNA (h) (sc-29214), BCL-xL siRNA (h) (sc-43630), MCL-1 siRNA (h) (sc-35877) and scrambled controls were purchased from Santa Cruz Biotechnology (Dallas, TX). Lipofectamine2000 (11668-027), Novex Sharp Pre-stained Protein Standard (LC5800) and JC-1 (T-3168) were from Invitrogen Life Technologies (Carlsbad, CA). Sorafenib (BAY 43-9006, Nexavar) and Regorafenib (BAY 73-4506, Stivarga) are manufactured by Bayer. A-1155463, ABT-737, AT101, HA-14, ABT-263 (Navitoclax) and ABT-199 (Venetoclax) were purchased from Selleckchem (Houston, TX).