West China School of Stomatology, Sichuan University, Sichuan, China.
J Endod. 2010 Feb;36(2):238-43. doi: 10.1016/j.joen.2009.09.007. Epub 2009 Oct 23.
The Wnt signaling pathway plays an important role in tissue development by acting on proliferation, differentiation, and cell fate decisions. Because the role of Wnt6 in tooth development was still unknown, the purpose of this study was to investigate the role of Wnt6 in tooth morphogenesis and dental tissue mineralization by elucidating its effect on human dental papilla cells (hDPCs) in vitro.
Human dental papilla cells were enzymatically separated from tooth germs. Recombinant adenovirus encoding full-length Wnt6 cDNA was constructed to overexpress Wnt6, and the biologic effects of Wnt6 on hDPCs were investigated. Wnt6-transduced changes in hDPC proliferation were examined by means of a 5-bromodeoxyuridine (BrdU) incorporation assay and cell cycle analysis. Wnt6-transduced changes in hDPC differentiation were investigated by evaluating alkaline phosphatase (ALPase) activity, by a mineralization assay, and analysis of mineralization-related gene expression including ALP, type I collagen (Col I), osteonectin (ON), osteopontin (OPN), bone sialoprotein (BSP), and dentin matrix protein-1 (DMP-1).
Wnt6 overexpression had no significant effect on the proliferation of hDPCs by BrdU incorporation assay and flow cytometric analysis. Wnt6 enhanced differentiation of hDPCs into functional odontoblast-like cells with up-regulated activity of ALPase and the expression of mineralization-related genes such as ALP, Col I, ON, OPN, BSP, and DMP-1. Wnt6 overexpression also promoted the mineralization of hDPCs.
Our findings verified that Wnt6 plays an important role in tooth development by promoting hDPC differentiation, without significant effects on hDPC proliferation.
Wnt 信号通路通过作用于增殖、分化和细胞命运决定，在组织发育中发挥重要作用。由于 Wnt6 在牙齿发育中的作用尚不清楚，本研究旨在通过阐明其对人牙髓细胞（hDPC）的体外作用，研究 Wnt6 在牙齿形态发生和牙组织矿化中的作用。
通过酶消化从牙胚中分离出人牙髓细胞。构建了编码全长 Wnt6 cDNA 的重组腺病毒以过表达 Wnt6，并研究了 Wnt6 对 hDPC 的生物学影响。通过 5-溴脱氧尿苷（BrdU）掺入测定和细胞周期分析检测 Wnt6 转导对 hDPC 增殖的影响。通过评估碱性磷酸酶（ALPase）活性、矿化测定以及分析矿化相关基因表达（包括 ALP、I 型胶原（Col I）、骨连接蛋白（ON）、骨桥蛋白（OPN）、骨唾液蛋白（BSP）和牙本质基质蛋白-1（DMP-1））来研究 Wnt6 转导对 hDPC 分化的影响。
通过 BrdU 掺入测定和流式细胞术分析，Wnt6 过表达对 hDPC 的增殖没有显著影响。Wnt6 增强了 hDPC 向功能性成牙本质细胞样细胞的分化，ALPase 活性和矿化相关基因如 ALP、Col I、ON、OPN、BSP 和 DMP-1 的表达上调。Wnt6 过表达还促进了 hDPC 的矿化。
我们的研究结果证实，Wnt6 通过促进 hDPC 分化在牙齿发育中发挥重要作用，而对 hDPC 增殖没有显著影响。
