Osteoinduction properties of graphite-like amorphous carbon films evaluated in-vitro

Graphite-like amorphous carbon (a-C) films were deposited by magnetron sputtering on medical grade stainless steel (SS) substrates. The purpose of the study was the evaluation of amorphous carbon as a coating for medical implants in contact with bone, in which the formation of bone on the surface is of special interest to improve the implant performance. The initial step consisted of the evaluation of the cytocompatibility of the a-C films. Cytocompatibility was evaluated by seeding human osteoblast cells on the surfaces and measuring their adhesion, proliferation and viability up to seven days. These data are also of interest for the osteoinduction process because the initial phase of the bone-forming activity of osteoblasts entails the attachment, proliferation, cell differentiation and extracellular matrix synthesis, followed by a second phase of bone matrix mineralization. The evaluation in-vitro of the osteoinduction properties of the surfaces consisted on the study of cell differentiation by measuring the alkaline phosphatase activity, the morphological examination of the mineralized extracellular matrix by electron microscopy, and the evaluation of a selection of proteins involved in the bone-formation process.The cytocompatibility (cell attachment, proliferation and viability) was determined in comparison with SS, titanium coatings and the plastic tissue control. We observed a superior cellular adhesion and proliferation in the a-C surfaces, while the viability was slightly better on the Ti coatings, but nevertheless, the a-C values were much higher than in the plastic control, indicating that the a-C films are no toxic and biocompatible. On the other hand, all the results of the osteoinduction; ALP specific activity and expression of bone-growth specific proteins indicated that the a-C surface significantly promotes the cell differentiation, leading to the formation of the extracellular matrix and its mineralization.