The temporal transcriptomic signature of cartilage formation

Chondrogenesis is a multistep process whereby cartilage progenitor cells generate a tissue with distinct structural and functional properties. Although several approaches to cartilage regeneration rely on the differentiation of implanted progenitor cells, the temporal transcriptomic landscape of in vitro chondrogenesis in different models has not been reported. Using RNA sequencing, we examined the differences between gene expression patterns during early cartilage formation in micromass cultures of embryonic limb bud-derived progenitors. Principal component analysis indicated a progressively different and distinct transcriptome during chondrogenesis. Differentially expressed genes (DEGs) based on pairwise comparisons of samples from consecutive days were classified into clusters and analysed. We confirmed the involvement of the top DEGs in chondrogenic differentiation with pathway analysis. We discovered several chondrogenesis-associated transcription factors and new collagen subtypes not previously linked to cartilage formation. These results provide, for the first time, a detailed insight into the molecular mechanism of in vitro chondrogenesis and provide a new gold standard for the temporal transcriptomic landscape of chondrogenic differentiation that may serve as a platform for new approaches in cartilage tissue engineering. Highlights Chondroprogenitors derived from chicken embryonic limb buds were differentiated into chondrocytes. RNA-sequencing revealed expression levels of 24,145 transcripts. We examined the global expression patterns during chondrogenesis and defined 6 clusters. Using WGCNA analysis, we created a module of genes that had similar patterns to the chondrogenic transcription factor SOX9. We created a list of transcription factors with a hitherto unexplored role in chondrogenesis.