Collagen I scaffolds cross-linked with beta-glycerol phosphate induce osteogenic differentiation of embryonic stem cells in vitro and regulate their tumorigenic potential in vivo.

Embryonic stem cells (ESCs) have the potential to differentiate into all tissues of the adult organism. This, along with the ability for unlimited self-renewal, positions these cells for regenerative medicine approaches based on tissue engineering strategies. With the objective of developing a treatment regime for skeletal injuries and diseases, this study presents a novel protocol that effectively induces ESC differentiation into osteogenic and chondrogenic lineages while concurrently eliminating observed tumorigenicity during the period of observation after transplantation in vivo. Exposure to a collagen I matrix polymerized with beta-glycerol phosphate (BGP) induced the osteogenic differentiation of the ESCs with an efficiency of >80% without purification and/or lineage-specific cell selection. Furthermore, when the collagen I matrix was supplemented with chondroitin sulfate, chondrogenesis was promoted instead of osteogenesis. Interestingly, without purification of the differentiated cells from the collagen I matrix, these constructs did not lead to the formation of teratomas or tumors when implanted subcutaneously in a severe combined immunodeficiency (SCID). Furthermore, if undifferentiated ESCs were mixed with collagen I and then injected immediately (i.e., without previous in vitro differentiation), again, no teratomas or tumors were observed, whereas undifferentiated ESCs without collagen scaffolds all produced teratomas in this bioassay system. These results suggest that collagen I scaffolds not only induce osteogenic differentiation of ESCs, but also prevent ESCs from producing unwanted tumors when injected in vivo.