Nerve Growth Factor Promotes Differentiation of Odontoblast-Like Cells Szilvia Arany, * Souichi Koyota, and Toshihiro Sugiyama Department of Biochemistry, Akita University School of Medicine, Akita, Japan ABSTRACT Contemporary strategies in tooth repair markedly rely on the newest findings on the cellular and biological components of dental development. Among several identified bioactive molecules, neurotrophins were recently proposed to affect tooth germ cell proliferation, differentiation, and cell–extracellular matrix interactions. The present study attempted to explore the effect of nerve growth factor (NGF) on a spontaneously immortalized dental papilla mesenchymal cell line. NGF induced differentiation of odontoblast-lineage cells with subsequent biomineralization in vitro. Here we showed that normalized transcript levels of tissue-specific markers such as DSPP and DMP1 were elevated significantly, indicating cell differentiation and maturation processes. We performed innovative gene expression analysis of TM14, a matricellular protein and novel member of the fibulin family. TM14 expression followed a pattern similar to that of DMP1, which suggests its important role in cell–matrix and intercellular interactions during dentin calcification. Alkaline phosphatase enzyme assay confirmed the extracellular matrix calcifications in NGF-supplemented groups. Thus, NGF was characterized as a potent promoter of mineralization during dentin formation. For the first time, we included TM14 in odontoblast genotype analysis and proved that NGF also promotes in vitro odontoblast differentiation. Collectively, these results highlight the importance of NGF during tooth morphogenesis, as well as urge the elaboration of complex epithelial–mesenchymal tissue cultures, where further elucidation of the signaling factor network could be completed. J. Cell. Biochem. 106: 539–545, 2009. ß 2009 Wiley-Liss, Inc. KEY WORDS: ODONTOBLAST; NERVE GROWTH FACTOR; TM14; MINERALIZATION E xperiments exploring the pathways of tooth formation are often used as a model system for organogenesis [Thesleff et al., 1996]. Tooth germ development is based on the reciprocal interactions between epithelial and mesenchymal components and on subsequent cell differentiation, morphogenesis, and extracellular matrix formation. Our understanding of this complex biological process and of the regulating signaling pathways has been rapidly expanding recently. Novel aspects of tooth development, with respect to cell–cell, cell–matrix, and matrix–matrix interactions, shed light on the cascade mechanism of organogenesis, which has been thoroughly investigated during the last several decades. Extending our knowledge of the biochemical and cellular com- ponents of dental development represents a promising approach for increased biological applications including tissue engineering. Although tooth regeneration has been addressed using different strategies, all such attempts share a common feature, that is, that odontogenic capacity is inevitably necessary to generate the tooth structure. This potential has been recognized in isolated epithelial cells [Komine et al., 2007], mesenchymal cells from developing teeth [Yu et al., 2006], oral primordial cells [Bhattacherjee et al., 2007], and even in non-dental cells [Ohazama et al., 2004] when successful recombination with tooth germ components could be achieved. Maintaining the odontogenic potential of various stem cells derived from dental progenitor cells or other tissues (e.g., bone marrow cells, embryonic cells, and neural cells) [Modino and Sharpe, 2005] seems to be a key factor during tooth regeneration experiments. It is generally accepted that odontogenic capacity switching from the dental epithelium to the dental mesenchyme during the early stage of tooth formation is a result of sequential and reciprocal biochemical communication. Regulatory molecules, growth factors, transcription factors, and cell adhesion molecules have been intensively studied in terms of the tissue-specific cross-talk, but the signaling network is still not yet fully understood. Recently, some neurotrophic factors have been shown to be involved in the differentiation of dental epithelial cells [Yoshizaki et al., 2008] as well as pulp cells [Mizuno et al., 2007]. Members of the neurotrophin family have been found to be expressed in the developing tooth germ [Nosrat et al., 2002] and play an important Journal of Cellular Biochemistry ARTICLE Journal of Cellular Biochemistry 106:539–545 (2009) 539 Grant sponsor: Japan Society for the Promotion of Science, Ministry of Education, Culture, Sports, Science and Technology of Japan; Grant number: 18300159. *Correspondence to: Szilvia Arany, Department of Biochemistry, Akita University School of Medicine, Hondo 1-1-1, Akita 010-8543, Japan. E-mail: aszilvia@med.akita-u.ac.jp Received 11 September 2008; Accepted 5 November 2008  DOI 10.1002/jcb.22006  2009 Wiley-Liss, Inc. Published online 23 January 2009 in Wiley InterScience (www.interscience.wiley.com).