A375 human melanoma cell line

A375 human melanoma cell line was obtained from American Type Culture Collection (ATCC). A375 was maintained in the DMEM (Gibco) supplemented with 10% FBS and cultured at 37 °C in a 5% CO₂ incubator.

The A375 human melanoma cell line was purchased from the American Type Culture Collection (ATCC Manassas, Virginia). The WM1366 and MeWo cell lines were obtained from Dr. Saldano Ferrone (Massachusetts General Hospital, Boston, MA). Ixazomib (MLN2238) and bortezomib (Velcade, PS-341) were obtained from Millennium Pharmaceuticals, Inc. (Cambridge, MA). Recombinant human IFN-α was obtained from Schering-Plough, Inc. (Kenilworth, NJ).

The B16 mouse melanoma cells stably transfected with murine apelin (B16 Ap) or with the empty plasmid vector (B16 Mock) have been previously described in ref.^{63 (link)}. The A375 human melanoma cell line was obtained from the American Type Culture Collection (Manassas, VA). All cell lines were maintained in Dulbecco’s modified Eagle’s medium (DMEM; Sigma Aldrich Corp., St. Louis, MO, USA) supplemented with 10% fetal bovine serum (FBS; Sigma Aldrich Corp.) and 1% penicillin/streptomycin (Sigma Aldrich Corp.) at 37 °C in a humidified incubator with 5% CO₂.

The melanoma cell line A375-M6, isolated in our laboratory from a lung metastasis in a SCID bg/bg mouse i.v. injected with A375 human melanoma cell line [27 (link)], obtained from American Type Culture Collection (ATCC, Rockville, MD, USA); the melanoma cell line FO-1, the FO-1 cell lines transfected with pSV2neo alone (FO-1 neo) or with human wild type β2m gene and pSV2neo (FO-1β2), the human melanoma cell lines WM266-4 (from ATCC) and M21 (kindly provided by Antony Montgomery, The Scripps Research Institute, La Jolla, CA, USA) were cultured in Dulbecco’s Modified Eagle Medium high glucose (DMEM 4500, EuroClone, Figino, MI, Italy) supplemented with 10% fetal bovine serum (FBS, Boerhinger Mannheim, Mannheim, Germany) and 2mM L-glutamine (EuroClone, MI, Italy) at 37 °C in a humidified atmosphere containing 90% air and 10% CO₂.
Cells were harvested from subconfluent cultures by incubation with a trypsin-EDTA solution (EuroClone, MI, Italy), and propagated every three days. Viability of cells was determined by trypan blue exclusion test. Cultures were periodically monitored for mycoplasma contamination using Chen’s fluorochrome test [28 (link)].

A375 human melanoma cell line was purchased from American Type Culture Collection (Manassas, VA, USA). BLM human melanoma cell line was kindly provided by Dr. G.N. van Muijen from Radbound University Nijmegen Medical Center (Department of Pathology, Nijmegen, The Netherlands). Cells were stored in liquid nitrogen and, after resuscitation, they were kept in culture for 10–12 weeks. In particular, they were cultured in humidified atmosphere of 5% CO2/95% air at 37  °C, in DMEM medium supplemented with antibiotics, glutamine and 7.5% (A375 cells) or 10% (BLM cells) FBS. Cell line authenticity was assessed by the Short Tandem Repeat (STR) profile analysis, as explained by ATCC Standards Development Organization (SDO) in ANSI Standard (ASN-0002).

C4‐2 and PC3 human prostate cancer cell lines, A549 human lung cancer cell line, A375 human melanoma cell line, HCT116 human colorectal cancer cell line and HEK 293T human embryonic kidney cell line were obtained from the American Type Culture Collection. C4‐2, PC3, A549 and HCT116 cells were cultured in Roswell Park Memorial Institute (RPMI)−1640 medium (Gibco, cat. #11875119), while A375 cells and HEK 293T cells were cultured in Dulbecco's Modified Eagle Medium (DMEM, Gibco, cat. #10569044). Both media were supplemented with 10% foetal bovine serum (FBS, ExCell, cat. #FCS500) and 100 units/mL penicillin/streptomycin (Gibco, cat. #15140122). The cells were incubated in a 5% (vol/vol) CO₂ and 95% (vol/vol) air incubator (ThermoFisher Co.) at 37°C.