AUTHOR=Miao Xinchao , Niibe Kunimichi , Fu Yunyu , Zhang Maolin , Nattasit Praphawi , Ohori-Morita Yumi , Nakamura Takashi , Jiang Xinquan , Egusa Hiroshi 

TITLE=Epiprofin Transcriptional Activation Promotes Ameloblast Induction From Mouse Induced Pluripotent Stem Cells via the BMP-Smad Signaling Axis

JOURNAL=Frontiers in Bioengineering and Biotechnology

VOLUME=Volume 10 - 2022

YEAR=2022

URL=https://www.frontiersin.org/journals/bioengineering-and-biotechnology/articles/10.3389/fbioe.2022.890882

DOI=10.3389/fbioe.2022.890882

ISSN=2296-4185

ABSTRACT=The genetic modification of induced pluripotent stem cells (iPSCs) holds promise for their directed differentiation into ameloblasts, which are usually lost after tooth eruption. Ameloblast differentiation is regulated by multiple signaling molecules, such as bone morphogenetic proteins (BMPs). Epiprofin (Epfn), a transcription factor, is expressed in the dental epithelium during tooth development. However, it remains unknown whether and how Epfn activation promotes ameloblast induction in mouse iPSCs. Here, we generated doxycycline-inducible Epfn-expressing mouse iPSCs (Epfn-iPSCs). Ameloblasts, which are characterized by positive staining for keratin 14 and amelogenin and alizarin red S staining, were successfully derived from Epfn-iPSCs based on a stage-specific induction protocol, which involved the induction of the surface ectoderm, dental epithelial cells, and ameloblasts at stages 1, 2, and 3, respectively. Epfn activation by doxycycline at stages 2 and/or 3 decreased cell proliferation and promoted ameloblast differentiation, along with the upregulation of p-Smad1/5/9, a key regulator of the BMP-Smad signaling pathway. Gene analysis of the BMP-Smad signaling pathway-associated molecules revealed that Epfn activation decreased follistatin (a BMP inhibitor) expression at stage 2, whereas it increased BMP2/4/7 expression at stage 3. Perturbations in the ameloblast differentiation process were observed when the BMP-Smad signaling pathway was blocked by a BMP receptor inhibitor (LDN-193189). Simultaneous LDN-193189 treatment and Epfn activation largely reversed the perturbations in ameloblast induction, with partial recovery of p-Smad1/5/8 expression, suggesting that Epfn activation promotes ameloblast induction from mouse iPSCs partially by upregulating BMP-Smad activity. These results reveal the potential regulatory networks between Epfn and the BMP-Smad pathway and suggest that Epfn is a promising target for inducing the development of ameloblasts, which can be used in enamel and tooth regeneration.