2. Academic Validation
  3. Exploring the Role of DNMT1 in Dental Papilla Cell Fate Specification during Mouse Tooth Germ Development through Integrated Single-Cell Transcriptomics and Bulk RNA Sequencing

Exploring the Role of DNMT1 in Dental Papilla Cell Fate Specification during Mouse Tooth Germ Development through Integrated Single-Cell Transcriptomics and Bulk RNA Sequencing

  • J Oral Biosci. 2024 Jun 26:S1349-0079(24)00147-6. doi: 10.1016/j.job.2024.06.010.
Dahlia Eldeeb 1,  Hiroyuki Okada 2,  Yutaka Suzuki 3,  Masahide Seki 3,  Junichi Tanaka 4,  Kenji Mishima 4,  Ung-Il Chung 5,  Shinsuke Ohba 6,  Hironori Hojo 7
Affiliations

Affiliations

  • 1 Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo; Department of Oral Biology, Faculty of Dentistry, Cairo University.
  • 2 Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo; Department of Orthopaedic Surgery, Graduate School of Medicine, The University of Tokyo.
  • 3 Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo.
  • 4 Division of Pathology, Department of Oral Diagnostic Sciences, School of Dentistry, Showa University.
  • 5 Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo; Department of Bioengineering, Graduate School of Engineering, The University of Tokyo.
  • 6 Department of Tissue and Developmental Biology, Graduate School of Dentistry, Osaka University. Electronic address: ohba.shinsuke.dent@osaka-u.ac.jp.
  • 7 Center for Disease Biology and Integrative Medicine, Graduate School of Medicine, The University of Tokyo; Department of Bioengineering, Graduate School of Engineering, The University of Tokyo. Electronic address: hojo@g.ecc.u-tokyo.ac.jp.
PMID: 38942194 DOI: 10.1016/j.job.2024.06.010
Abstract

Objectives: This study aimed to investigate the regulatory mechanisms governing dental mesenchymal cell commitment during tooth development, focusing on odontoblast differentiation and the role of epigenetic regulation in this process.

Methods: We performed single-cell RNA Sequencing (scRNA-seq) of dental cells from embryonic day 14.5 (E14.5) mice to understand the heterogeneity of developing tooth germ cells. Computational analyses including gene regulatory network (GRN) assessment were conducted. We validated our findings using immunohistochemistry (IHC) and in vitro loss-of-function analyses using the DNA Methyltransferase 1 (DNMT1) inhibitor Gsk-3484862 in primary dental mesenchymal cells (DMCs) isolated from E14.5 mouse tooth germs. Bulk RNA-seq of Gsk-3484862-treated DMCs was performed to identify potential downstream targets of DNMT1.

Results: scRNA-seq analysis revealed diverse cell populations within the tooth germs, including epithelial, mesenchymal, immune, and muscle cells. Using single-cell regulatory network inference and clustering (SCENIC), we identified DNMT1 as a key regulator of early odontoblast development. IHC analysis showed the ubiquitous expression of DNMT1 in the dental papilla and epithelium. Bulk RNA-seq of cultured DMCs showed that Gsk-3484862 treatment upregulated odontoblast-related genes, whereas genes associated with cell division and the cell cycle were downregulated. Integrated analysis of bulk RNA-seq data with scRNA-seq SCENIC profiles was used to identify the potential DNMT1 target genes.

Conclusions: DNMT1 may negatively affect odontoblast commitment and differentiation during tooth development. These findings contribute to a better understanding of the molecular mechanisms underlying tooth development and future development of hard-tissue regenerative therapies.

Keywords

Dental mesenchymal cells (1); Dental papilla; Dnmt1; Tooth development; scRNA-seq.

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