Hff 1 cells

Human skin fibroblast 1 (HFF1) cells were purchased from (ATCC, Manassas, VA, USA) and cultured in a basic growing medium of DMEM low glucose (Life Technologies, Carlsbad, CA, USA), supplemented with 10% fetal bovine serum (FBS Life Technologies, Carlsbad, California, USA), 2 mM L-glutamine (Euroclone, Milano, Italy) and 1% of penicillin/streptomycin (Euroclone, Milano, Italy). Cells at passage 5 were then exposed to freeze-dried Myrtus extracts, previously suspended in cultured medium at three different concentrations of 0.5, 0.75 and 1 mg/mL, for 12 and 24 h. After treatment with the extracts, cells were incubated with 100 µM H₂O₂ in basic growing medium for 1 h, to induce oxidative stress. Other cells were cultured in growing medium without extracts, then exposed to H₂O₂ treatment (H₂O₂), representing our control of oxidative stress. HFF1 cultured in the presence of 100 µg/mL ascorbic acid (AA)(Sigma-Aldrich, Germany), then exposed to H₂O_2, represent a positive control of antioxidant activity. Cells grown in basic medium alone, without any treatment, were considered the negative control (Ctrl).

All cells cultured in DMEM with 10% FBS (Invitrogen, Carlsbad CA) and 100 μ/mL Penicillin/Streptomycin (Invitrogen) unless otherwise noted. NIH 3T3 cells (ATC#CRL-1658) were obtained from the ATTC. HFF-1 cells (ATCC#SCRC-1041) were obtained from the ATTC. Mouse fibroblasts were generated as described (Gasser et al., 2005 (link)) and provided by Pr D Raulet. MCA-205 were received from Pr. L. Linear (UCSF). All cell lines tested negative for mycoplasma as described by Young et al. (2010) (link). Mithramycin A and Butyrate were purchased from Sigma (St. Louis, MO). RGFP966 was purchased from Seleckchem (Houston, TX). Antibodies recognizing Sp1 (product # 07–645), Acetylated Histone 3 (product # 06–599) were purchased from Millipore (Billerica, MA). Antibodies against histone 3 (clone D1H2), HDAC3 (Clone 7G685), and antibody recognizing CKII substrate (#8738) were purchased from Cell Signaling Technologies (Danvers MA). Antibody recognizing CK2β (Product PA5-27416) was purchased from Thermo Fisher (Waltham, MA). Antibody against Sp3 (D-20) was purchased from Santa Cruz Biotechnology. Plasmid encoding HSV-1 ICP0 was provided by the Knipe Lab (Harvard). Plasmid encoding HCMV IE1 was provided by the Weinberger Lab (UCSF).

Human dermal fibroblast HFF-1 cells (ATCC SCRC-1041) and U937-DC-SIGN cells (ATCC CRL-3253) were infected with all the Flavivirus isolates at an MOI of 0.1, and supernatants were harvested for titration every 24 h for 6 days.

A375 (human malignant melanoma) and MCF7 (human breast adenocarcinoma) cell lines were purchased from ECACC, Salisbury, UK. HFF-1 cells (human foreskin fibroblasts) were obtained from ATCC. A375 and MCF7 cells were cultivated in RPMI 1640 and HFF cells in DMEM medium (HyClone Laboratories, Inc., Logan, UT, USA). Growth media were supplemented with 2 mM L-glutamine (PAA Laboratories, Pasching, Austria), 10% fetal calf serum, 100 IU/ml penicillin, and 100 µg/ml streptomycin (HyClone Laboratories, Inc., Logan, UT, USA). Cells were incubated at 37 °C in a high-humidity atmosphere with 5% CO₂ and were passaged three times a week.