Human primary osteoblasts were isolated from the trabecular bone of adult femoral heads, provided by the Orthopaedic and Traumatology Unit at the hospital “Maggiore della Carità”, Novara, Italy. The samples represent surgical discharge materials and, therefore, their use does not need ethics committee approval. Written informed consent, specifying that residual material destined to be disposed of could be used for research, was signed by each participant before the biological materials were removed, in agreement with Rec(2006)4 of the Committee of Ministers Council of Europe on research on biological materials of human origin.
After being minced and washed in PBS, bone fragments were digested with collagenase/elastase as described in Bosetti et al. [86 (link)] and plated with Iscove’s modified Dulbecco’s medium (IMDM, Euroclone, Milan, Italy) supplemented with 10% fetal bovine serum (FBS, Hyclone GE Healthcare, Logan, UT, USA), 2 mM L-glutamine (L-Glu, Sigma-Aldrich, Milan, Italy), and PS at 37 °C in 5% CO2. Cells appeared within 10 days and were used for all the experiments up to the seventh passage after being characterized by alkaline phosphatase expression [87 (link)].
After being minced and washed in PBS, bone fragments were digested with collagenase/elastase as described in Bosetti et al. [86 (link)] and plated with Iscove’s modified Dulbecco’s medium (IMDM, Euroclone, Milan, Italy) supplemented with 10% fetal bovine serum (FBS, Hyclone GE Healthcare, Logan, UT, USA), 2 mM L-glutamine (L-Glu, Sigma-Aldrich, Milan, Italy), and PS at 37 °C in 5% CO2. Cells appeared within 10 days and were used for all the experiments up to the seventh passage after being characterized by alkaline phosphatase expression [87 (link)].