A chemically-defined protocol for generating chondrocytes from human embryonic stem cells.

Progress in generating robust differentiation protocols for efficient and scalable production of defined cell lineages from human embryonic stem cells (hESc) has been slow. Amongst the obstacles to be addressed are those inherent to standard hESc culture and differentiation practices including the use of feeder cells, serum and animal-derived matrices. These components are biologically complex, undefined and highly variable between batches, inhibiting the development of consistently reproducible protocols (1,2).
We have developed for hESc a new 3-Stage directed differentiation protocol (DDP) to generate chondrocytes, the specialized cells that form cartilage tissue (3). The protocol is segmented into stages that mimic the developmental processes that occur in cell lineage specification during embryogenesis. Pluripotent hESc, are established as feeder-free cultures (4) prior to DDP being carried out in monolayer culture on a sequence of matrix protein-coated surfaces with a chemically-defined medium sequentially supplemented with growth factors to direct differentiation through developmental intermediate cell populations of primitive streak/mesendoderm (Stage 1), mesoderm (Stage 2) and then chondrocytes (Stage 3) (Figure 1).
This unique protocol is highly efficient, scalable and completely chemically-defined, thus making it appropriate for translation towards clinical applications of chondrocytes, as well as providing a defined system for characterizing the molecular mechanisms regulating hESc differentiation.