Effect of Dexamethasone and 1,25$(OH)_2D_3$on Proliferation and Osteogenic Differentiation of Cultured Human Bone Marrow Stromal Cells

Background: It is crucial, in the case of regenerating bone by tissue-engineering technique, that osteoblast progenitors are proliferated and induced to differentiate to osteoblasts sequentially at the proper time. Osteoblasts can be obtained from bone itself or from osteoblast progenitors in bone marrow, even though the amount of human marrow stromal cells in marrow aspirate is usually scanty. These cells, however, have been known demonstrate the potential to easily proliferate and differentiate in osteoblasts, chondroblasts or adipocytes according to different microenvironmental factors. We evaluated the effect of dexamethasone and 1,25 on the proliferation, differentiation, and mineralization of human marrow stromal cells in vitro. Methods: We used twelve bone marrow aspirates obtained from different healthy bone marrow donors. Culture plates were randomly divided into the following four experimental groups; group 1 was cultured with control medium only, group 2 with control medium containing 1,25, group 3 with control medium containing dexamethasone, and group 4 with control medium containing both 1,25 and dexamethasone. -thymidine uptake, protein content of cell lysates, alkaline phosphatase activities and alkaline phosphatase histochemistries were measured. Alizarin Red-S staining and quantification of dissolved dye were also performed. Results: Combined stimulation of marrow stromal cells with both 1,25 and dexamethasone was found to be effective to maintain stable long-term culture of the cells and to increased differentiation and mineralization of the cells. Synthesis and mineralization of matrix were highest when the cells were stimulated with 1,25 alone during the early culture phase. However, 1,25 shortened the lifespan of the cells. Interestingly, mineralization was higher in female donor cells than in male donor cells when stimulated with dexamethasone alone or with both dexamethasone and 1,25. Neither 1,25 nor dexamethasone affected cell proliferation. Conclusion: Our results suggest that the synergistic effect of dexamethasone and 1,25 is important in maintaining long-term culture and differentiation of human marrow stromal cells. It is preferable to administer 1,25 after the attachment of cultured osteoblasts to biomaterials has been established, since it could shorten cell survival despite the great increase of mineralization at the early culture phase.