Abstract
Cellular differentiation comprises a progressive, multistep program that drives cells to fabricate a tissue with specific and site distinctive structural and functional properties. Cartilage constitutes one of the potential differentiation lineages that mesenchymal stem cells (MSCs) can follow under the guidance of specific bioactive agents. Single agents such as transforming growth factor beta (TGF-β) and bone morphogenetic protein 2 in unchanging culture conditions have been historically used to induce
in vitro
chondrogenic differentiation of MSCs. Despite the expression of traditional chondrogenic biomarkers such as type II collagen and aggrecan, the resulting tissue represents a transient cartilage rather than an
in vivo
articular cartilage (AC), differing significantly in structure, chemical composition, cellular phenotypes, and mechanical properties. Moreover, there have been no comprehensive, multicomponent parameters to define high-quality and functional engineered hyaline AC. To address these issues, we have taken an innovative approach based on the molecular interrogation of human neonatal articular cartilage (hNAC), dissected from the knees of 1-month-old cadaveric specimens. Subsequently, we compared hNAC-specific transcriptional regulatory elements and differentially expressed genes with adult human bone marrow (hBM) MSC-derived three-dimensional cartilage structures formed
in vitro
. Using microarray analysis, the transcriptome of hNAC was found to be globally distinct from the transient, cartilage-like tissue formed by hBM-MSCs
in vitro
. Specifically, over 500 genes that are highly expressed in hNAC were not expressed at any time point during
in vitro
human MSC chondrogenesis. The analysis also showed that the differences were less variant during the initial stages (first 7 days) of the
in vitro
chondrogenic differentiation program. These observations suggest that the endochondral fate of hBM-MSC-derived cartilage may be rerouted at earlier stages of the TGF-β-stimulated chondrogenic differentiation program. Based on these analyses, several key molecular differences (transcription factors and coded cartilage-related proteins) were identified in hNAC that will be useful as molecular inductors and identifiers of the
in vivo
AC phenotype. Our findings provide a new gold standard of a molecularly defined AC phenotype that will serve as a platform to generate novel approaches for AC tissue engineering.
