Cellular responses to deproteinized bovine bone mineral biofunctionalized with bone-conditioned medium.

OBJECTIVES The aim of the study was to investigate whether the osteoinductive properties of bone-conditioned medium (BCM) harvested from cortical bone chips within a clinically relevant short-term period can enhance the biologic characteristics of deproteinized bovine bone mineral (DBBM) in vitro. MATERIALS AND METHODS To assess the biofunctionalization of DBBM, the adhesive, proliferative, and differentiation properties of mesenchymal stromal ST2, pre-osteoblastic MC3T3-E1, and primary bone-derived cells grown on BCM-coated DBBM were examined by crystal violet staining of adherent cells, BrdU ELISA, and qRT-PCR, respectively. RESULTS BCM extracted within 20 min or 24 h in either Ringer's solution (BCM-RS) or RS mixed with autologous serum (BCM-RS + S) increased the adhesive properties of all three cell types seeded on DBBM. The 20-min BCM-RS preparation appeared more potent than the 24-h preparation. BCM-RS made within 20 min or 24 h had strong pro-proliferative effects on all cell types grown on DBBM. RS + S alone exhibited a considerable pro-proliferative effect, suggesting an impact of the serum on cellular growth. DBBM coated with BCM-RS or BCM-RS + S, made within 20 min or 24 h each, caused a significant induction of osteogenic differentiation marker expression with a higher potency of the BCM-RS + S. Finally, a strong additive effect of fresh bone chips combined with BCM-coated DBBM on the osteogenic differentiation of the three cell types was observed. CONCLUSIONS Altogether, the data strongly support the biofunctionalization of DBBM with BCM extracted within a clinically relevant time window of 20 min. CLINICAL RELEVANCE Pre-activation of non-osteoinductive biomaterials with BCM, prepared from autologous bone chips during a guided bone regeneration (GBR) procedure, bears the potential of an optimal treatment modality for bone defects in daily practice.