Sh sy5y human neuroblastoma

The cell lines of SH‐SY5Y human neuroblastoma had been gained from the American Type Culture Collection (ATCC, USA). Cells were cultured in DMEM high-glucose culture fluid (Sigma, USA), including 100 U/mL double antibiotics as well as 10% fetal bovine serum (Sigma, USA).

HeLa human cervix adenocarcinoma cells, SH-SY5Y human neuroblastoma, and HDFa human normal fibroblast as control cell lines were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA). Cells were cultured in RPMI 1640 medium (Labtek Eurobio, Milan, Italy), supplemented with 10% FCS (Euroclone, Milan, Italy) and 2 mM L-glutamine (Sigma-Aldrich, Milan, Italy), at 37 °C, and a 5% CO₂ atmosphere. γ-T, α-T, p-cym, and myr were purchased from Sigma-Aldrich (Sigma-Aldrich, Milan, Italy). γ-T, α-T, p-cym and myr were percolated twice through activated basic alumina and once through silica to remove impurities and traces of hydroperoxides. The compounds were dissolved in DMSO in a 30 mM stock solution. In cell treatments, the final DMSO concentration never exceeded 0.1%.

Human HeLa cervical carcinoma and human SH-SY5Y neuroblastoma cells were obtained from the American Type Culture Collection (ATCC). Both cell lines were maintained in DMEM (Dulbecco’s Modified Eagle’s Medium), supplemented with 10% fetal bovine serum (Biowest, ref. S181B-500, Labclinics, Madrid, Spain) and 1% of a mixture of antibiotic-antimycotic (Gibco, USA) and were grown in a humidified atmosphere with 5% CO₂ at 37°C. HeLa and SH-SY5Y cells were subcultured in a 10 mL Petri dish (Nunc, Roskilde, Denmark). At confluence, cells were detached with trypsin (Tryple Express, Gibco, USA) and were resuspended in complete growth medium and plated in a 6- or 96-well dish (Nunc, Roskilde, Denmark) as necessary.

Human SH-SY5Y neuroblastoma and rat C6 glioblastoma cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, United States). SH-SY5Y cells were maintained in DMEM media (Fisher, Houston, TX, United States) supplemented with 15% FBS, penicillin-streptomycin (Sigma-Aldrich, St. Louis, MO, United States) and grown in an incubator at 37°C in the presence of 5% CO₂ (Hearst et al., 2011 (link)). C6 cells were maintained in DMEM media (Fisher, Houston, TX, United States) supplemented with 10% FBS, penicillin-streptomycin (Sigma), and grown in an incubator at 37°C in the presence of 5% CO₂. GFP control or dMMP plasmid DNA was transfected into cell lines using either Lipofectamine 2000 (Invitrogen, Carlsbad, CA, United States), or FuGene 6 (Roche, Indianapolis, IN, United States) according to the manufacturer’s instructions. SH-SY5Y stable cell lines were produced by transfecting with GFP using FuGene 6 transfection reagent and selected by G418 (G418 sulfate 600 μg/ml, Sigma) resistance as previously described (Hearst et al., 2011 (link)). Cell viability post transfection was assessed using the MTT assay using methods as previously described (Datki et al., 2003 (link)). All experiments were conducted with three independent biological repeats.