title: Differentiation of Odontoblast-Like Cells From Mouse Induced Pluripotent Stem Cells by Pax9 and Bmp4 Transfection.
abstract: The field of tooth regeneration has progressed in recent years, and human tooth regeneration could become viable in the future. Because induced pluripotent stem (iPS) cells can differentiate into odontogenic cells given appropriate conditions, iPS cells are a potential cell source for tooth regeneration. However, a definitive method to induce iPS cell-derived odontogenic cells has not been established. We describe a novel method of odontoblast differentiation from iPS cells using gene transfection. We generated mouse iPS cell-derived neural crest-like cells (iNCLCs), which exhibited neural crest markers. Next, we differentiated iNCLCs into odontoblast-like cells by transfection of Pax9 and Bmp4 expression plasmids. Exogenous Pax9 upregulated expression of Msx1 and dentin matrix protein 1 (Dmp1) in iNCLCs but not bone morphogenetic protein 4 (Bmp4) or dentin sialophosphoprotein (Dspp). Exogenous Bmp4 upregulated expression of Msx1, Dmp1, and Dspp in iNCLCs, but not Pax9. Moreover, cotransfection of Pax9 and Bmp4 plasmids in iNCLCs revealed a higher expression of Pax9 than when Pax9 plasmid was used alone. In contrast, exogenous Pax9 downregulated Bmp4 overexpression. Cotransfection of Pax9 and Bmp4 synergistically upregulated Dmp1 expression; however, Pax9 overexpression downregulated exogenous Bmp4-induced Dspp expression. Together, these findings suggest that an interaction between exogenous Pax9- and Bmp4-induced signaling modulated Dmp1 and Dspp expression. In conclusion, transfection of Pax9 and Bmp4 expression plasmids in iNCLCs induced gene expression associated with odontoblast differentiation, suggesting that iNCLCs differentiated into odontoblast-like cells. The iPS cell-derived odontoblast-like cells could be a useful cell source for tooth regeneration.
  
  
  CONCLUSIONS
  It has been reported that induced pluripotent stem (iPS) cells differentiate into odontogenic cells by administration of recombinant growth factors and coculture with odontogenic cells. Therefore, they can be potential cell sources for tooth regeneration. However, these previous methods still have problems, such as usage of other cell types, heterogeneity of differentiated cells, and tumorigenicity. In the present study, a novel method to differentiate iPS cells into odontoblast-like cells without tumorigenicity using gene transfection was established. It is an important advance in the establishment of efficient methods to generate homogeneous functional odontogenic cells derived from iPS cells.
authors.affiliation: Division of Orthodontics and Dentofacial Orthopedics, Tohoku University Graduate School of Dentistry, Miyagi, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Seki
authors.affiliation: Division of Orthodontics and Dentofacial Orthopedics, Tohoku University Graduate School of Dentistry, Miyagi, Japan.
authors.forename: Nobuo
authors.initials: N
authors.lastname: Takeshita
authors.affiliation: Division of Orthodontics and Dentofacial Orthopedics, Tohoku University Graduate School of Dentistry, Miyagi, Japan.
authors.forename: Toshihito
authors.initials: T
authors.lastname: Oyanagi
authors.affiliation: Division of Orthodontics and Dentofacial Orthopedics, Tohoku University Graduate School of Dentistry, Miyagi, Japan.
authors.forename: Shutaro
authors.initials: S
authors.lastname: Sasaki
authors.affiliation: Division of Orthodontics and Dentofacial Orthopedics, Tohoku University Graduate School of Dentistry, Miyagi, Japan.
authors.forename: Ikuko
authors.initials: I
authors.lastname: Takano
authors.affiliation: Division of Orthodontics and Dentofacial Orthopedics, Tohoku University Graduate School of Dentistry, Miyagi, Japan.
authors.forename: Masakazu
authors.initials: M
authors.lastname: Hasegawa
authors.affiliation: Division of Orthodontics and Dentofacial Orthopedics, Tohoku University Graduate School of Dentistry, Miyagi, Japan t-yamamo@m.tohoku.ac.jp.
authors.forename: Teruko
authors.initials: T
authors.lastname: Takano-Yamamoto
mesh_terms: D000818:Animals; D055415:Bone Morphogenetic Protein 4 / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation; D060846:Cell Engineering; D064987:Cell- and Tissue-Based Therapy; D016326:Extracellular Matrix Proteins / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D057026:Induced Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D051957:MSX1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D051379:Mice; D058953:Neural Stem Cells / Q000166:cytology / Q000378:metabolism*; D009804:Odontoblasts / Q000166:cytology / Q000378:metabolism*; D009805:Odontogenesis / Q000235:genetics*; D051764:PAX9 Transcription Factor; D051761:Paired Box Transcription Factors / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection
chemical_list: C525810:Bmp4 protein, mouse; D055415:Bone Morphogenetic Protein 4; C513308:Dmp1 protein, mouse; D016326:Extracellular Matrix Proteins; D051957:MSX1 Transcription Factor; C484084:Msx1 protein, mouse; D051764:PAX9 Transcription Factor; D051761:Paired Box Transcription Factors; C488759:Pax9 protein, mouse
keywords: Cytokines; Differentiation; Induced pluripotent stem cells; Transcriptional factors