Optimization of bioartificial bone graft construction in vitro

Objective To investigate the influencing factors and mechanisms of human fetal osteoblasts seeding and culture in porous β-tricalcium phosphate (β-TCP) particles,and to optimize the bioartifical bone graft construction in vitro.Methods A newly designed perfusion bioreactor was employed for human fetal osteoblasts seeding and culture in porous β-TCP particles.Static seeding was served as the negative control.The effects of different conditions,including the time of seeding,density of cell suspension,flow rate of cell seeding and culture,on cell adhesion and growth were evaluated by the cell viability assay (MTT) and measurement of the cell seeding efficiency,histological and histomorphometric study.Results Perfusion seeding technique was superior to static seeding.Perfusion seeding and culture of fetal cells in β-TCP scaffolds resulted in significantly higher cell viability and growth rates than those by static seeding followed by perfusion culture.The time of seeding,density of cell suspension and flow rate of seeding and culture were determinants of the cell seeding and growth.The optimal conditions for constructing bioartificial bone included the time of seeding between 12 and 24 hours,the density of cell suspension between 2× 105/ml and 5× 105/ml,the flow rate of seeding 1 ml/min and the perfusion flow rate for incubation between 0.5 ml/min and 2 ml/min within the first 24 hours and 2 ml/min thereafter.Conclusions The in vitro perfusion technique is effective for seeding and culture of human fetal osteoblasts in porous β-TCP particles.The time of seeding,cell density and flow rate markedly impact on the outcomes of bioartificial bone construction in vitro.The optimization of conditions for bioartificial bone construction may facilitate clinical translation and promotion. Key words: Bone substitutes ;  Bioreactors;  Cytoskeleton