Lox imvi

CMeC-1, CMeC-2, KMeC, LMeC melanoma cell lines [16 (link)] were provided by Professor Nobuo Sasaki (University of Tokyo); the DEN haemangiosarcoma cell line [17 (link)] by Professor Douglas Thamm (Colorado State University); the melanoma 12 cell line [18 (link)] by Professor Michael Kent (University of California, Davis). The ARCE mast cell tumour line was provided by Dr Richard Elders (formerly RVC, University of London, now at University of Edinburgh). The canine cell lines C2, DH82, A72, D17, CF33MG, CF35MG and MDCK and the human melanoma cell line LOXIMVI, were obtained from ATCC.

The human melanoma cell lines A375, MeWo, LOX IMVI, and WM793 were obtained from the American Type Culture Collection (ATCC), and WM 266.4 was provided by Dr. Michael Davies (MD Anderson). Cells were cultured in Dulbecco’s modified Eagle medium containing 5% fetal bovine serum at 37°C and in 5% CO2 and 95% air. For DETA NONOate treatment, cells were cultured to 75% confluency, and freshly prepared DETA NONOate stock was added to the final concentration as indicated in the text. After treatment times, cells were washed twice with cold phosphate-buffered saline and were harvested for analysis of pAKTS473 levels, AKT kinase activity, AKT substrate phosphorylation, and PTEN activity. An iNOS expressing construct was established as described by us previously (10 (link)). Cells were transfected with plasmids using Lipofectamine 2000 (Invitrogen), according to the manufacturer’s specifications.

The human melanoma cell lines M14, A375, A2058, and LOXIMVI were obtained from the American Type Culture Collection (ATCC; Manassas, VA, USA). The cells were cultured in RPMI-1640 supplemented with 10% fetal bovine serum (FBS) at 37 °C in a humidified incubator containing 5% CO₂. Normal human skin cells (Detroit 551) were obtained from ATCC and maintained in dulbecco modified eagle medium (DMEM) high glucose (Lonza, Basel, Switzerland) supplemented with 10% FBS. Frugoside was obtained from NPBANK (Gungsan, Korea). Antibodies against poly-(adenosine diphosphate-ribose) polymerase (PARP), caspase-3, phospho(p)-JNK (Thr183/Tyr185), p-p38 (Thr180/Tyr182), p-ERK (Thr202/Tyr204), JNK, p38, and ERK were purchased from Cell Signaling Technology (Danvers, MA, USA). Antibodies against Prx2, Prx3, Prx-SO2, and hemagglutinin epitope (HA) were purchased from Abfrontier (Seoul, Korea). Antibodies against Srx and tubulin were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Antibodies to cytochrome c were purchased from BD Pharmingen (San Jose, CA, USA). SB202190, N-acetyl-L-cysteine (NAC), diphenyleneiodonium chloride (DPI), U0126, and hydrogen peroxide solution were purchased from Sigma (St Louis, MO, USA).