Three-dimensional culture of osteoblastic cells using a radial-flow bioreactor

Objectives: In the field of regenerative medicine, a technique in which cells are cultured in 3 dimensions in vitro and transplanted to treat an extensive tissue defect has been performed in recent years. Various bioreactors have been developed to perform 3-dimensional culture for tissue regeneration. To regenerate the jawbone, we used a radial-flow bioreactor and ascertained whether osteoblasts could evenly proliferate over a scaffold with a diameter of 18 mm and a height of 10 mm.

Methods: To a type-1 collagen scaffold with 70-110 µm pore size, 80-95% porosity and 3-mm thickness, 5x10⁵ mouse osteoblast-like cells (MC3T3-E1) were seeded. Cells were incubated outside the reactor for 6 h to allow adhesion to the scaffold. These 6 pre-cultured scaffolds were placed so that the reactor was filled with scaffolds. Cells were then incubated for 1 week under conditions of 37°C, pH 7.4, DO 6.86 ppm, culture medium exchange 200 mL/day, and culture medium circulation 3 mL/min. For controls, cells were incubated for 1 week in the same manner without circulating culture medium. Histological analysis was performed to assess cell distributions inside the scaffold. Cell proliferation was assessed based on glucose consumption and DNA-based cell counts.

Results: Using the radial-flow bioreactor, culture medium was circulated sufficiently into a comparatively wide, thick scaffold using in this study. After 1 week of culture in the radial-flow bioreactor, glucose consumption increased, and cells were distributed throughout the scaffold. In addition, DNA-based cell counts increased after 1 week. Cells did not proliferate when culture medium was not circulated.

Conclusion: The present study suggests that a radial-flow bioreactor is useful for 3-dimensional culture of osteoblasts.