A Method for In Vitro Fabrication of Hybrid Bone/Cartilage Tissue Using Mouse Induced Pluripotent Stem Cells

In the developing embryo, bone and cartilage share the same progenitors. However, osteo-chondrogenic induction of mouse induced pluripotent stem cells (iPSCs) remains difficult. Here we describe a protocol to guide iPSCs to differentiate into osteochondral cells that form hybrid bone/cartilage constructs in vitro. Single mouse iPSCs are first reaggregated in ultra-low-attachment micro-space culture plates. At day 12, iPSC spheres are subjected to shaking culture and maintained in an osteogenic induction medium for 31 days (Os induction). In another condition, the osteogenic induction medium is replaced by chondrogenic induction medium at day 22 and maintained for a further 21 days (Os-Chon induction). Os induction produced robust mineralization and some cartilage-like tissue, whereas Os-Chon induction resulted in partial mineralization and a large area of cartilage tissue.