Tissues and cell lines

Human primary osteosarcoma tissues (n=20) and matched adjacent normal tissues (n=20) were obtained from 20 patients (13 males and seven females; 17.6±6.8 years of age) attending the Department of General Surgery, Tianjin Medical University General Hospital, from 2012 to 2014. All the patients gave informed consent to participate in the study. All patients had a histological diagnosis of primary osteosarcoma according to the clinicopathological criteria of the International Union for Cancer Control. Histological assessment of tissue samples confirmed that each osteosarcoma sample contained >70% tumor cells. Patient consents the use of tissue specimens in this research study, and the study protocols were approved by the Research Ethics Committee of Tianjin Medical University General Hospital.

Human osteosarcoma cell lines (SAOS-2, MG-63, U-2 osteosarcoma) and human osteoblasts cell line hFOB (OB3) were purchased from the American Type Culture Collection (ATCC) (Manassas, VA, USA). The osteosarcoma and osteoblast cells were maintained in culture according to the vendor’s instructions. In brief, SAOS-2 and U-2 osteosarcoma cells were cultured in McCoy’s 5a Medium (Modified) (ATCC). MG-63 cells were cultured in Eagle’s Minimum Essential Medium (EMEM) (ATCC), while hFOB (OB3) cells were cultured in a 1:1 mixture of Ham’s F12 Medium Dulbecco’s Modified Eagle’s Medium, with 2.5 mM l-glutamine (without phenol red). All cells were cultured in media containing 10% fetal bovine serum (FBS) and 1% penicillin–streptomycin (100 U/mL penicillin and 100 μg/mL streptomycin) and maintained in a humidified incubator under standard conditions.