Comparative investigation of viability, metabolism and osteogenic capability of tissue-engineered bone preserved in sealed osteogenic media at 37 0C and 4 0C

Preservation of tissue-engineered (TE) bone is one of the key problems needed to be solved for its clinic application and industrialization. Traditional cryopreservation has been restricted because of the damages caused by ice formation and solution. Hypothermic preservation at 4 0C has been widely used for the preservation of transplanted organ despite potential negative effects on viability of cells and tissue. 37 0C is the best temperature for maintaining cellular bioactivities. However, 37 0C also has a potential negative effect on preserved cells due to consumption of nutrients and accumulation of by-products. No studies have reported which temperature is more suitable for the preservation of TE bone constructs. The current study explored the feasibility of preservation of TE bone constructs in sealed osteogenic media at 37 0C and 4 0C. Human bone marrow stromal cells (hBMSCs) were seeded into partially demineralized bone matrix (pDBM) scaffolds and cultured for 7 days to form TE bone constructs. The constructs were preserved in sealed osteogenic media at either 37 0C or 4 0C for 5, 7, 9 and 11 days, respectively. Growth kinetics, viability, metabolism and osteogenic capability were evaluated to explore the feasibility of preservation at 37 0C and 4 0C. The constructs cultured in osteogenic media at humidified 37 0C/5%CO2 served as the positive control. The results demonstrated that all the constructs preserved at 4 0C showed negative osteogenic capability at all time points with a much lower level of growth kinetics, viability and metabolism compared to the positive control. However, the constructs preserved at 37 0C showed good osteogenic capability within 7 days with a certain level of growth kinetics, viability and metabolism, although an obvious decrease in osteogenic capability was observed in the constructs preserved at 37 0C over 9 days. These results indicate that the preservation of TE bone constructs is feasible at 37 0C within 7 days in sealed osteogenic media.