Fig. 1.

Coculture of osteoblasts and fibroblasts promoted osteogenic differentiation. (a) Schematic diagram of MC/NIH cocultures. MC3T3-E1 (osteoblasts) and NIH/3T3 (fibroblasts) were cocultured and characterized as time lime shown. (b) Morphology of coculture cells at different ratios was imaged by a phase contrast microscope on day 1 and 2. Green “M” indicated MC3T3-E1 cells; Red “N” indicated NIH/3T3 cells; yellow dotted line indicated the edge between MC3T3-E1 and NIH/3T3 cells. (c) Real-time qPCR analysis of osteogenic associated genes (RUNX2, ALP, BSP and OCN) at different time points as indicated in (a). MC: MC3T3-E1; NIH: NIH/3T3. Protein levels of ALP (blue) and OCN (red) was analyzed by ALP staining (d) and IF staining (f) respectively. The activity of ALP in different groups in 24-well plates was quantified by ImageJ software (e). Images and data are representative of n = 3 individual experiments, and bar heights and error bars were represented as means ± SD. Scale bars: 100 μm (b&e).