Formation of Osteochondral Organoids from Murine Induced Pluripotent Stem Cells

Impact statement The regeneration of integrated articular cartilage and bone tissues from a single cell source has been a challenge in the field of osteochondral tissue engineering and osteoarthritis disease modeling. The goal of this study was to develop an osteochondral organoid system using a single murine induced pluripotent stem cell (iPSC) source in a scaffold-free system and to determine whether differentiated iPSCs retain the potential to undergo reinduction of pluripotency. Our findings indicate that sequential differentiation into chondrogenic and osteogenic lineages can be used to develop osteochondral organoids, and encapsulation within a cartilaginous matrix prevents the reinduction of pluripotency in differentiated iPSCs. Osteoarthritis is a debilitating joint disease that is characterized by pathologic changes in both cartilage and bone, potentially involving cross talk between these tissues that is complicated by extraneous factors that are difficult to study in vivo. To create a model system of these cartilage-bone interactions, we developed an osteochondral organoid from murine induced pluripotent stem cells (iPSCs). Using this approach, we grew organoids from a single cell type through time-dependent sequential exposure of growth factors, namely transforming growth factor beta-3 and bone morphogenic protein 2, to mirror bone development through endochondral ossification. The result is a cartilaginous region and a calcified bony region comprising an organoid with the potential for joint disease drug screening and investigation of genetic risk in a patient or disease-specific manner. Furthermore, we also investigated the possibility of the differentiated cells within the organoid to revert to a pluripotent state. It was found that while the cells themselves maintain the capacity for reinduction of pluripotency, encapsulation in the newly formed 3D matrix prevents this process from occurring, which could have implications for future clinical use of iPSCs.