Abstract
Introduction: Alendronate is a well known inhibitor of osteoclastic activity and is used for the treatment of various metabolic bone diseases. Recent studies indicate that alendronate may have additional effect on osteoblastic activity. In this study, we evaluated the effect of alendronate on the osteogenic differentiation of mouse mesenchymal stem cells.
Materials and Methods: D1 cells, known as multipotent mouse mesenchymal stem cells, were cultured in the presence of osteogenic differentiation medium (ODM) for 7 days, then treated with alendronates, and 2 days later the cells were used for testing. The cell proliferation was analyzed using an MTT assay. Alizarin red staining was done for mineralization. Alkaline phosphatase activity was measured using a commercial ELISA kit. Calcification was analyzed using energy dispersive X-ray spectrophometric analysis (EDX). Osteogenic gene expression was analyzed using RT-PCR. The change of CD 44 expression was observed using confocal microscopy and FACS analysis.
Results: The D1 cells, in the presence of ODM, differentiated into osteoblasts. The evidence of osteogenic differentiation was confirmed by the positive stain by Alizarin red S, the increased activity of ALP, the increased mRNA expression of osteocalcin, a calcium peak in the EDX analysis, and by the positive immunofluorescence staining against CD 44 (an antigen detected on osteoblasts). With the addition of alendronate in the ODM medium, the osteogenic differentiation was enhanced. The enhancing effect was confirmed by the increased intensity of the Alizarin red S staining, higher activity of ALP, the higher mRNA expression of osteocalcin, a higher calcium peak in the EDX analysis, and by the increased immunofluorescence staining against CD 44 in the FACS analysis.
Conclusion: We have demonstrated that alendronate enhances osteogenic differentiation when given to mouse mesenchymal stem cells in association with the ODM. Alendronate may increase the bone density not only by inhibiting the osteoclastic activity but also by increasing the osteogenic differentiation of the mesenchymal stem cells.
The abstracts were prepared by Lynne C. Jones, PhD. and Michael A. Mont, MD. Correspondence should be addressed to Lynne C. Jones, PhD., at Suite 201 Good Samaritan Hospital POB, Loch Raven Blvd., Baltimore, MD 21239 USA. Email: ljones3@jhmi.edu
