Osteogenic performance of MC3T3-E1 cells on granular hydroxyapatite scaffold
The initial seeding density plays a crucial role in determining the effectiveness of bone tissue regeneration utilising scaffold. One of the factors that influence cell osteoblast differentiation is cell seeding related factor. Therefore, this study aims to determine the effect of different initial seeding densities on the osteoblast performance of MC3T3-E1 cells in hydroxyapatite granular scaffold. MC3T3-E1 cells at different seeding density (5x105 and 1x106 cells/cm2) were cultured in the 2-dimensional flask and 3-dimensional hydroxyapatite scaffold. Cells in both cultures were analysed in term of morphology using CellB software while biochemical activity was assessed using Alkaline Phosphatase (ALP) assay. MC3T3-E1 showed a mononucleated, fibroblast-like shape cell with extended cytoplasmic projection. Cell seeding density of 1x106 cells/cm2 reached 90% confluency while 5x105 cells/cm2 reached 70% confluency on day six of culture. Variations in cell seeding density significantly influenced the cell osteodifferentiation. During 2-Dimensional osteoblast differentiation, lower initial seeding density resulted in higher ALP activity. However, cells were seeded on hydroxyapatite scaffold, by increasing the seeding density from 5x105 to 1x106 cells/cm2 resulted in decreased of ALP activity. This study has shown that the seeded cell population in the 3-dimensional hydroxyapatite scaffolds clearly affected the degree of osteoblast cell differentiation in which a higher seeding density was not necessarily better. The seeding density played a major role in influencing the corresponding cell differentiation.
Keywords: Seeding density; osteoblast; Granumas®; alkaline phosphatase