Mitani K, Haruyama N, Hatakeyama J, Igarashi K
Department of Oral Dysfunction Science, Tohoku University Graduate School of Dentistry, Sendai, Japan.
Oral Dis. 2013 Mar;19(2):169-79. doi: 10.1111/j.1601-0825.2012.01967.x. Epub 2012 Aug 2.
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.
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.
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.
M180 and LRAP accelerate chondrogenic differentiation and maturation of ATDC5 cells.
釉原蛋白是牙釉质中含量最丰富的基质蛋白。在釉原蛋白异构体中，全长釉原蛋白（M180）和富含亮氨酸的釉原蛋白肽（LRAP）在多种组织中表达，并被认为是间充质细胞中的信号分子。在此，我们研究了M180和LRAP对软骨生成细胞系ATDC5的影响，以探讨釉原蛋白在软骨生成中的作用。
将含重组小鼠M180或LRAP蛋白的培养基或对照培养基与软骨生成刺激培养基混合，分析培养的ATDC5细胞的表型变化、基因表达水平和细胞增殖情况。
与对照组相比，添加釉原蛋白分别在培养14天和21天时增加了碱性磷酸酶活性和糖胺聚糖分泌。定量PCR（Q-PCR）分析显示，LRAP在分化7天时增加了Runx2、Col2a1和聚集蛋白聚糖的基因表达水平。此外，在培养28天时，M180和LRAP均显著增加了碱性磷酸酶、聚集蛋白聚糖、Col10a1和骨桥蛋白的基因表达水平。溴脱氧尿苷检测和Wnt信号通路的Q-PCR分析表明，M180和LRAP均降低了细胞增殖，但可能通过改变非经典Wnt信号通路诱导细胞分化。
M180和LRAP加速了ATDC5细胞的软骨生成分化和成熟。
