Abstract
Insulin is an inducer of chondrocyte hypertrophy and growth plate chondrogenesis, although the specific molecular mechanisms behind these effects are mostly unknown. Our aim was to investigate whether insulin-induced chondrocyte hypertrophy occurs through a modification in the amount of O-linked N-acetylglucosamine (O-GlcNAc)-modified proteins and in the expression of the key enzymes of this pathway, O-GlcNAc transferase and O-GlcNAcase (OGA). We also studied if O-GlcNAc accumulation per se, induced by an OGA inhibitor, was able to induce pre-hypertrophic chondrocyte differentiation both in vitro and in vivo. Insulin-induced differentiation of ATDC5 pre-chondrocytes occurred alongside a gradual increase in the accumulation of O-GlcNac-modified proteins (O-GlcNAcylated proteins), as well as an increase in the expression of O-GlcNAc transferase and OGA. In the absence of insulin, O-GlcNAc accumulation induced by thiamet-G, a specific OGA inhibitor, was able to increase the gene expression of differentiation markers, as well as the activity of MMP-2 and -9. Thiamet-G also activated pERK, p-JNK, and p-p38 and the O-GlcNAcylation of Akt. Thiamet-G administration to C57/bl mice induced a significant expansion in the growth plate height and in the hypertrophic zone height. Therefore, our results show that O-GlcNAc glycosylation has chondromodulating activity.