Balb 3t3

All reagents and solvents were purchased from Merck (Darmstadt, Germany) and Acros Organics (New Jersey, US) and used without further purification. Flash chromatography was performed using silica gel with 200–300 mesh produced by Merck. All reactions and processes of flash chromatography were monitored by the TLC method using silica gel plates with fluorescence F254 and iodine visualization. The melting points were determined with Stuart melting point apparatus SMP30 and uncorrected. Fourier-transformed infrared absorption spectra of both solid and liquid samples were analysed with NICOLET 6700 FT-IR using diamond with ATR.
Microanalyses were performed on Flash Elemental Analyzer 110 series. The ¹H NMR and ¹³C NMR spectra were recorded on a Joel- 400 spectrometer at 400 MHz at 125 MHz using CDCl₃ and DMSO-d₆ as solvents and TMS as internal standard. Coupling constants (J) are expressed in hertz (Hz). Chemical shifts (δ) are given in parts per million (ppm). All target compounds have purity over 95%.
All ATCC bacterial strains (BAA-1556, BAA-1688, ATCC 6358, ATCC 35556, ATCC 25923 and ATCC 33591) and the three mammalian cell lines (WRL-68, Vero CCL-81 and BALB/3T3) were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). The additional 37 S. aureus clinical isolates were obtained from three local Hospitals in Peninsula Malaysia.

The adopted cells (breast cancer MCF-7 and MDA-MB-231 and human mammary epithelial MCF-10A cells) were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA) and cultured as indicated [18 (link)]. The mouse embryonic fibroblast BALB/3T3 and neuroblastoma SH-SY5Y were also obtained from American Type Culture Collection (ATCC, Manassas, VA, USA) and cultured in DMEM high glucose supplemented with 100 U mL⁻¹ penicillin/streptomycin and 10% bovine calf serum (BCS) or 10% fetal bovine serum (FBS), respectively.

Biophenotypic B myelomonocytic leukaemia MV4-11, human lung carcinoma A549, human breast carcinoma MCF-7, human colon carcinoma LoVo, its drug resistant line LoVo/DX, liver carcinoma HepG2, and normal mouse fibroblast BALB/3T3 cells were obtained from the American Type Culture Collection (Rockville, Maryland, USA). All lines are being maintained at the Institute of Immunology and Experimental Therapy, Wrocław, Poland. All cell lines were grown in a humid atmosphere at 37 °C with 5% CO₂ and were cultured in media according to the method described before.¹⁸

The mouse normal fibroblasts cell line Balb/3T3 (American Type Culture Collection ATCC^®, Old Town Manassas, VA, USA), the normal human dermal fibroblasts line (NHDF) (Lonza, Basel, Switzerland), the human epidermal keratinocyte line (HaCaT) (DKFZ, Heidelberg, Germany) [48 (link)] and the human neuroblastoma cell line (SH-SY5Y) (ATCC^®) were used in the study. The Balb/3T3 and HaCaT lines were cultured with DMEM medium (Lonza) with 10% foetal bovine serum (FBS) and 1% L-glutamine with penicillin and streptomycin solution (Sigma-Aldrich^®, St. Louis, MO, USA). NHDF was grown in FGM^TM Fibroblast Growth Medium BulletKit^TM (Lonza). SH-SY5Y was cultured with F-12 medium (Lonza) supplemented with 10% FBS (Sigma-Aldrich^®) and 1% L-glutamine with penicillin and streptomycin solution (Sigma-Aldrich^®). Cell culture was maintained in 5% CO₂ at 37 °C and 95% humidity. The cells were assessed twice a week, and the fresh medium was changed or passaged if confluence was approximately 70%.

BALB/3T3 (normal murine fibroblasts) and MCF 10A (human, non-tumorigenic mammary gland epithelial cells) were obtained from the American Type Culture Collection (ATCC; Rockville, Maryland, USA). The BALB/3T3 cell line was cultured in high glucose DMEM (Thermo Fisher Scientific) supplemented with 10% (v/v) fetal bovine serum (FBS) and 2 mM L-glutamine (Sigma-Aldrich). The MCF 10A cell line was cultured in Ham’s F12 medium with glutamine (Corning Costar) supplemented with 5% (v/v) FBS, 5% (v/v) horse serum, 10 µg/mL insulin, 0.05 µg/mL cholera toxin, 0.5 µg/mL hydrocortisone, and 20 ng/mL hEGF (all from Sigma-Aldrich). All culture media contained 100 µg/mL streptomycin (Sigma-Aldrich) and 100 U/mL penicillin (Polfa Tarchomin SA, Warszawa, Poland). The cells were grown at 37 °C in a humid atmosphere saturated with 5% CO₂.

Biphenotypic B myelomonocytic leukemia MV4-11, human lung carcinoma A549, human breast carcinoma MCF-7, and normal mouse fibroblast BALB/3T3 cells were obtained from American Type Culture Collection (Rockville, Maryland, USA). All the cell lines are being maintained at the Institute of Immunology and Experimental Therapy, Wroclaw, Poland. MV4-11 cells were cultured in RPMI 1640 medium (Gibco, UK) with 2 mM L-glutamine adjusted to contain 1.0 mM sodium pyruvate, and 10% fetal bovine serum (FBS) (all from Sigma-Aldrich, Germany). A549 cells were cultured in RPMI 1640+ Opti-MEM (1:1) (both from Gibco, UK), MCF-7 cells in Eagle medium (IIET, Wroclaw, Poland), BALB/3T3 in Dulbecco medium (IIET, Poland) supplemented with 2 mM l-glutamine and 1.0 mM sodium pyruvate, 10% fetal bovine serum (all from Sigma-Aldrich, Germany). The MCF-7 cell culture was supplemented with 0.8 mg/l of insulin (Sigma-Aldrich, Germany). All culture media were supplemented with 100 units/ml penicillin, and 100 µg/ml streptomycin (both from Polfa Tarchomin S.A., Poland). All cell lines were grown at 37 °C with 5% CO₂ humidified atmosphere (Łączkowski et al. 2014 (link), 2016a (link)).