Amelogenin splice isoforms stimulate chondrogenic differentiation of ATDC5 cells.

OBJECTIVE

Amelogenins are the most abundant matrix proteins in enamel. Among the amelogenin isoforms, full-length amelogenin (M180) and leucine-rich amelogenin peptide (LRAP) are expressed in various tissues and are implicated as signalling molecules in mesenchymal cells. Here, we examined the effects of M180 and LRAP on a chondrogenic cell line, ATDC5, to investigate the role of amelogenins in chondrogenesis.

MATERIALS AND METHODS

Recombinant mouse M180- or LRAP-protein-containing medium or control medium was mixed with a chondrogenesis-stimulating medium, and changes in the phenotype, gene expression levels and cell proliferation of cultured ATDC5 cells were analysed.

RESULTS

The addition of amelogenins increased alkaline phosphatase activity and glycosaminoglycan secretion at 14 and 21 days of culture, respectively, as compared with the control. Quantitative PCR (Q-PCR) analysis revealed that LRAP increased the gene expression levels of Runx2, Col2a1 and Aggrecan at 7 days of differentiation. Moreover, both M180 and LRAP significantly increased the gene expression levels of ALP, Aggrecan, Col10a1 and osteopontin at 28 days of culture. Bromodeoxyuridine assay and Q-PCR analysis for Wnt signalling indicated that both M180 and LRAP reduced proliferation, but induced the cell differentiation possibly through altered non-canonical Wnt signalling.

CONCLUSION

M180 and LRAP accelerate chondrogenic differentiation and maturation of ATDC5 cells.