Preparation and functional evaluation of cell aggregates incorporating gelatin microspheres with different degradabilities.

The objective of this study was to investigate the viability and biological functions of cells in their aggregates incorporating gelatin microspheres with different degradabilities. After being prepared by a water-in-oil emulsion procedure, the gelatin microspheres were dehydrothermally crosslinked at 140°C for various time periods. In vitro degradation tests showed that the gelatin microspheres were slowly degraded slowly with an increase in the crosslinking time. When MC3T3-E1 cells were cultured with the gelatin hydrogel microspheres in the round U-bottom wells of 96-well microplates which had been coated with poly(vinyl alcohol), cell aggregates with homogeneously distributed gelatin microspheres were formed. A large amount of slowly degraded gelatin microspheres remained in the cell aggregates for long time periods, while a higher proliferation of MC3T3-E1 cells was observed. When evaluated as a measure of aerobic glycolysis, the ratio of l-lactic acid production:glucose consumption of MC3T3-E1 cells was lower for MC3T3-E1 cells in the cell aggregates incorporating slowly degraded gelatin microspheres than for aggregates incorporating rapidly degraded ones. The alkaline phosphatase activity and calcium content of MC3T3-E1 cells were higher for cell aggregates incorporating slowly degraded gelatin microspheres. It is possible that the incorporation of gelatin hydrogel microspheres with slow degradability enabled the permeation of oxygen and nutrients into the cell aggregates for longer time periods, resulting in better culture conditions for the survival, proliferation and differentiation of the cells.