Penicillin streptomycin solution

Huh‐7, a human hepatocarcinoma cell line, and A549, an adenocarcinomic human alveolar basal epithelial cell line, were both cultured in 10 cm dishes (Nunclon; Thermo Fisher Scientific, Waltham, MA, USA) and maintained in high‐glucose Dulbecco's modified Eagle medium supplemented with 10% fetal bovine serum (Gibco, Gaithersburg, MD, USA) and 1% penicillin streptomycin solution (Corning, NY, USA). T24, a human bladder carcinoma cell line, was cultured in 10 cm dishes and maintained in McCoy's 5a medium (modified) with l‐glutamine (1.5 mm) adjusted to contain sodium bicarbonate (2.2 g L⁻¹) and supplemented with 10% fetal bovine serum (Gibco) and 1% penicillin streptomycin solution (Corning). The cells were kept at 37 °C in a 5% CO₂ humidified chamber. The media were monitored daily and replaced regularly every 2 to 3 days. The cells were subcultured using 0.5% trypsin‐EDTA (Corning) upon reaching 80–90% confluence.

The RAW264.7 cells are macrophage-like cells derived from mice. The KG-1 cell line is made up of macrophages isolated from the bone marrow of a human. Both cell lines were purchased from ATCC, USA. The Raw264.7 cells were cultured in High-glucose Dulbecco's Modified Eagle Medium (DMEM-HG; Life technology, NY, USA) consisting of 10% fetal bovine serum (Thermo, Logan, UT) and 1% penicillin–streptomycin solution (Corning, NY, USA). KG-1 cells were cultured in Iscove's Modified Dulbecco's Medium (IMDM; Life technology, NY, USA) consisting of 10% fetal bovine serum (Thermo, Logan, UT) and 1% penicillin–streptomycin solution (Corning, NY, USA). The cells were incubated at 37 °C and 5% CO₂. When cells reached 80% confluence density, cells were separated by Accutase ® (Innovative Cell Technologies, SD, USA) and reseeded at a ratio 1:6.

HCT116 cells were cultured in McCoy’s 5A media (Gibco #16600082) and 10% FBS (Avantor #97068–085) and 1X Penicillin-Streptomycin Solution (Corning #30002CI). HEK 293T cells were cultured in DMEM with 4.5 g / L glucose, L-glutamine & sodium pyruvate (Corning #10–0130CV) and 10% FBS (Avantor #97068–085) and 1X Penicillin-Streptomycin Solution (Corning #30002CI). Cells were cultured at 37 °C in 5% CO₂. Cell passage was conducted with trypsin (Corning #25–053-CI). Cells were washed with 1X PBS pH 7.4 (Gibco #70011044) before media change and passaging.

HCT116 (CCL-247) and HeLa S3 (CCL-2.2) cells were obtained from ATCC. HCT116 cells were grown in McCoy’s 5A medium with 10% fetal bovine serum (Gemini Bio Products) and penicillin/streptomycin solution (Corning). HeLa S3 cells were grown in MEM-alpha (Corning) with 10% fetal bovine serum, sodium pyruvate, and penicillin/streptomycin solution (Corning). Cell lines were maintained in a humidified atmosphere at 37°C, 5% CO₂ and were routinely screened for mycoplasma infection using MycoAlert (Lonza).
Aminoguanidine (AG; 1 mM) was used in all studies including the addition of exogenous polyamines to limit extracellular polyamine oxidation by bovine serum amine oxidase (21 (link)). DFMO was provided by Dr Patrick Woster at the Medical University of South Carolina. AG, N⁸-AcSpd, N¹-AcSpd, N-AcPut, Put, Spd, and Spm were purchased from Sigma Chemical Co. N¹,N⁸-diAcSpd was purchased from Cayman Chemical Co. The polyamine transport inhibitor Trimer44NMe was provided by Dr Otto Phanstiel (University of Central Florida).

The HaCaT cells were cultured in Dulbecco’s modified Eagle’s medium, supplemented with 10% fetal bovine serum (Gibco^®, Grand Island, NY, USA) and 1% penicillin–streptomycin solution (Corning^®, Manassas, VA, USA). The NIH3T3 fibroblast cells were cultured in Dulbecco’s modified Eagle’s medium supplemented with 10% bovine calf serum (Gibco^®, Penrose, AKL, NZ) and 1% penicillin–streptomycin solution (Corning^®, Manassas, VA, USA). Both cells were cultured in a humidified 5% CO₂ atmosphere at 37 °C and prepared for the cell proliferation assay and wound healing scratch assay.