Chitosan enhances mineralization during osteoblast differentiation of human bone marrow-derived mesenchymal stem cells, by upregulating the associated genes

Objectives: Chitosan is widely used as a scaffold for bone tissue engineering. However, up-to-date, no previous detailed study has been conducted to elucidate any mechanism of osteogenesis by chitosan itself. Here, we have evaluated effects of chitosan-coated tissue culture plates on adhesion and osteoblast differentiation processes of human mesenchymal stem cells (hMSCs), isolated from adult bone marrow. Materials and methods: Tissue culture plates coated with chitosan at different coating densities were used to evaluate the effects on hMSC adhesion and osteoblast differentiation. hMSCs were induced to differentiate into osteoblasts on the chitosan-coated plates and were evaluated using established techniques: alkaline phosphatase assay, demonstration of presence of calcium and real time PCR. Results: The cells adhered to plates of lower coating density of chitosan, but formed viable cell aggregates at higher coating density (100 mu g/sq.cm). Coating density of 25 mu g/sq.cm, supporting cell adhesion was chosen for osteoblast differentiation experiments. Differentiating hMSCs showed higher mineral deposition and calcium content on chitosan-coated plates. Chitosan upregulated genes associated with calcium binding and mineralization such as collagen type 1 alpha 1, integrin-binding sialoprotein, osteopontin, osteonectin and osteocalcin, significantly. Conclusions: We demonstrate for the first time that chitosan enhanced mineralization by upregulating the associated genes. Thus, the study may help clinical situations promoting use of chitosan in bone mineralization, necessary for healing non-union fractures and more.