Three-dimensional collagen gel culture promotes osteoblastic phenotype in bone marrow derived cells.

When calvarial cells or osteogenic cell lines were cultured in type I collagen gel, calcification was observed early and diffusely compared to monolayer culture. Bone marrow derived cells were cultured in three-dimensional type I collagen gel to investigate whether the cells can differentiate into osteogenic cells. In terms of efficient induction of osteogenic differentiation, we compared collagen gel culture to type I collagen coated dish culture and collagen-free plastic dish culture by morphology, alkaline phosphatase activity, and mRNA expression for type I collagen and osteopontin. Bone marrow derived primary cells formed colonies consisting of fibroblastic cells positively expressing alkaline phosphatase activity. Mineral deposition was observed in both primary and the 3rd passaged cells cultured in collagen gel, whereas the 3rd passaged cells on plastic dishes failed to be mineralized. Cells in collagen gel showed higher alkaline phosphatase activity than those in the other two methods suggesting that three-dimensional collagen network stimulated osteoblastic differentiation effectively. The expression level for type I collagen mRNA of the cells in collagen gel was three times higher in the 3rd passaged cells, and was slightly decreased in primary cells compared to the other two methods. The osteopontin mRNA expression of the cells in collagen gel was four times higher in the 3rd passaged cell culture but lower in primary cell cultures. These results suggested that collagen gel culture might be a useful environment for osteogenic induction of passaged cells derived from bone marrow.