Modified methods for efficiently differentiating human embryonic stem cells into chondrocyte-like cells.

Introduction: Human articular cartilage has a poor regenerative capacity. This often results in the serious joint disease- osteoarthritis (OA) that is characterized by cartilage degradation. An inability to self-repair provided extensive studies on AC regeneration. The cell-based cartilage tissue engineering is a promising approach for cartilage regeneration. So far, numerous cell types have been reported to show chondrogenic potential, among others human embryonic stem cells (hESCs).

Materials and methods: However, the currently used methods for directed differentiation of human ESCs into chondrocyte-like cells via embryoid body (EB) formation, micromass culture (MC) and pellet culture (PC) are not highly efficient and require further improvement. In the present study, these three methods for hESCs differentiation into chondrocyte-like cells in the presence of chondrogenic medium supplemented with diverse combination of growth factors (GFs) were evaluated and modified.

Results: The protocols established here allow highly efficient, simple and inexpensive production of a large number of chondrocyte-like cells suitable for transplantation into the sites of cartilage injury. The most crucial issue is the selection of appropriate GFs in defined concentration. The obtained stem-derived cells reveal the presence of chondrogenic markers such as type II collagen, Sox6 and Sox9 as well as the lack or significantly lower level of pluripotency markers including Nanog and Oct3/4.

Discussion: The most efficient method is the differentiation throughout embryoid bodies. In turn, chondrogenic differentiation via pellet culture is the most promising method for implementation on clinical scale. The most useful GFs are TGF-β1, -3 and BMP-2 that possess the most chondrogenic potential. These methods can also be used to obtain chondrocyte-like cells from differentiating induced pluripotent stem cells (iPSCs).