Mineralization of matrix vesicles isolated from a human osteosarcoma cell line in culture with water-soluble chitosan-containing medium

Chitosan is a natural bioactive material. Although it has been reported that chitosan promotes osteogenesis in bone lesions, little is known about how chitosan modulates this process. The present study was designed to investigate the effect of water-soluble chitosan relative to initiation of biologic mineralization, especially in the matrix-vesicles-(MVs) mediated process in vitro. A human osteoblastic cell line (NOS-1) was used. After 3 days of incubation, the number of cells and alkaline phosphatase (ALP) activity increased significantly in the chitosan group. RT-PCR analysis revealed that chitosan induced an increase in the expression of bone morphogenetic protein-2 mRNA after 7 days of incubation. MVs were isolated from NOS-1 cells using a collagenase digestion and ultracentrifugation method. ALP activity of MVs isolated from chitosan-supplemented cells was significantly higher than that of the control group. Furthermore, isolated MVs were incubated in medium supplemented with Na-β-glycerophosphate without fetal bovine serum. Needle-like crystals were observed in association with MVs after 24 h of incubation. These needlelike crystals were densely accumulated in the chitosan group. The present findings suggest that water-soluble chitosan would promote osteoblast proliferation and differentiation and may be useful for the acceleration of initial biologic mineralization.