Osteogenic differentiation of hESCs after culturing on fibrillar type I collagen coatings

This paper reports a new method to induce osteogenic differentiation of human embryonic stem cells (hESCs). Singly dissociated hESCs treated with ROCK inhibitor Y27632 were plated on type I fibrillar collagen (FC) coatings and achieved a high rate of survival. After culturing in osteogenic medium for 10 d, several cell colonies developed on the coatings. Cells forming the colonies showed homogenous, fibroblastic-like morphology. Subcultures of these cells on FC coatings in osteogenic medium developed an osteoblastic phenotype evidenced by strong staining for deposited minerals and expression of the ostoblastic related genes Runx2 and Col1a1. However, gene expression of osteocalcin (OC) was not detected, implying that while mineral deposition was mediated by the cells they did not mature during the culture period. FACS analysis showed the surface markers were 82.9% CD73⁺, 81.0% CD105⁺ and 99.8% CD45^-, consistent with the pattern of MSCs´. Additionally, in subsequent second subcultures these cells displayed a 5-fold increase in OC gene expression. We conclude that FC either exerted a unique cell-matrix interaction which promoted osteogenic differentiation or it selectively supported the survival and growth of cells committed to the osteoblastic lineage. Further characterization of the cells and the cell-collagen coating interactions are in progress.