2.7 Cell culture and seeding using MG63

Osteoblast-like osteosarcoma MG-63 cells we cultured in Dulbecco’s modified Eagle’s medium (DMEM, Hyclone, Logan, UT, USA) containing 10% fetal bovin serum (FBS, Gibco, Rockville, MD, USA) and 1% penicillin/streptomycin (PS, Sigma, St. Louis, MO, USA) in an atmosphere of 5% CO₂ at 37°C for 4 days, and the culture medium was replaced every day. 5×10⁴ cells/scaffold was seeded onto each scaffold. Cell proliferation was assessed by cell counting kit-8 (CCK-8, Dojindo Molecular technologies, Japan) assay. The culture media and CCK-8 solution were mixed at a ratio of 10:1. The optical density (OD) level of extracted suspension was determined by measuring at 450 nm using a micro reader (UVM 340, Elisa, USA).