Msto 211h cells

HEK293T cells, a non-cancerous human embryonic kidney epithelial cell line with a stable expression of the SV40 large T antigen, were obtained from the RIKEN BioResource Center (Tsukuba, Japan). Human pancreatic cancer PANC-1, AsPC-1, and MIA PaCa-2 cells, human breast cancer MDA-MB-231 and MCF-7 cells, human prostate cancer PC-3 and LNCaP cells, human squamous cancer A431 cells, human cervical cancer HeLa cells, human melanoma MeWo cells, human mesothelioma MSTO-211H cells, and human lung cancer NCI-H2170 cells were obtained from the American Type Culture Collection (Rockville, MD). All cell lines were cultivated in DMEM/F12 (D/F) medium (Thermo Fisher Scientific, Waltham, MA) supplemented with 10% fetal bovine serum (FBS).
Human neutrophils were isolated from the blood of a healthy donor by the conventional method. In brief, the polymorphonuclear cell fraction was first collected from the whole blood by centrifugation at 2000 rpm for 30 min using the PolymorphPrep™ solution (Serumwerk Bernburg, Bernburg, Germany). The collected cells were then incubated with CD16 microbeads (Miltenyi Biotec, Bergisch Gladbach, Germany), and the unbound flow through the fraction was collected as the neutrophil-enriched fraction.

MSTO-211H cells were purchased from the American Type Culture Collection (ATCC), Manassas, VA, USA, and A549 human lung cancer cells were obtained from the Korean Cell Line Bank (KCLB), Seoul, Korea. These cell lines were grown in RPMI 1640 containing 100 units/mL penicillin, 0.1 mg/mL streptomycin, and 10% FBS. All cell lines used in the study were authenticated by the ATCC and KCLB using STR-PCR analysis. The cells were incubated in a humidified atmosphere of 5% CO₂ in air at 37°C and maintained in log phase growth. Cell viability was determined by spectrophotometrically measuring the mitochondrial conversion of MTT to formazan. After cells were treated with the specified drugs, MTT was added to the cell suspension for 4 h. After three washes with phosphate-buffered saline (PBS; pH 7.4), the insoluble formazan product was dissolved in dimethyl sulfoxide (DMSO). The optical density (OD) of each well was measured using a microplate reader (Titertek Multiskan; Flow Laboratories, North Ryde, New South Wales, Australia) at 590 nm. The OD resulting from formazan production in control cells was considered as 100% cell viability. All other measurements were expressed as a percentage of the control cell value.