Study of Morphology and Biochemistry of Rat Bone Marrow Mesenchymal Stem Cells Before and After Osteogenic Differentiation: A Comparative Study

The aim of this study was to compare the morphological and biochemical character of mesenchymal stem cells and osteoblasts in vitro. After extraction of rat bone marrow mesenchymal stem cells and culturing them till the 3rd passage, these cells were cultured in media containing beta-glycerol phosphate, dexamethasone and ascorbic acid for 21 days. Then morphology of the mesenchymal cells and differentiated one were investigated using Hoechst and acridine orange. In addition the level of matrix deposition, level of calcium, activity of alkaline phosphatase in mesenchymal cells and differentiated on as well as metabolic activity of osteoblasts with the help of MTT was determined. Morphological study showed delocalization of nuclei and roundness of cytoplasm in differentiated cells as compared to mesenchymal stem cells. In addition mineralization of the matrix started from the 10th day and reached the maximum level at the 21st day. Also from the day 10 the level of calcium increased significantly (p<0.05) but activity of alkaline phosphatase decreased significantly (p<0.05) in differentiated cells. Where as during the differentiation process the metabolic activity of the cells increased significantly (p<0.05). In addition to morphological differences, calcium content, alkaline phosphatase activity and metabolic activity of the mesenchymal cells and osteoblasts also differs. Therefore in addition to alizarin red staining these factors also can be used to investigate the differentiation processes in vitro.