Full Length ArticleOdontoblast death drives cell-rich zone-derived dental tissue regeneration
Introduction
The bone tissue is constantly resorbed by osteoclasts and replaced with the newly formed bone tissue by osteoblasts [1,2]. Although the average lifetime of mature osteoblasts is limited, their counts are continuously maintained by bone marrow mesenchymal stem and progenitor cells (BM-MSPCs), which exhibit the potential for self-renewal and differentiation into mesenchymal lineages in the bone marrow [[3], [4], [5], [6]]. Studies indicate that dental pulp tissue also contains a multipotent stromal population, known as dental pulp stem and progenitor cells (DPSCs) [[7], [8], [9]]. Although controversial, several reports suggest that the DPSC population is identified as neural-glial antigen 2+ pericytes [10], glioma-associated oncogene 1+ cells [11], or proteolipid protein 1+ glial cells [12] in the incisors and alpha-smooth muscle actin (αSMA)+ perivascular cells [13] or Axin2+ cells [14] in the molars. In contrast to osteoblasts, dentin-forming odontoblasts reside within the dental tissue for a long term; therefore, DPSCs do not differentiate into odontoblasts in healthy fully formed teeth [15]. However, when odontoblast death occurs as a result of severe tooth damage, the DPSCs proliferate and differentiate into odontoblast-like cells that contribute to reparative dentin formation [10,11,14,16,17]. However, how odontoblast death is associated with the mechanisms underlying reparative dentin formation remains unclear.
The cell-rich zones (CZs) adjacent to the odontoblast layer in the dental pulp have been reported to contain odontoblast progenitor cells [[18], [19], [20]], which can be specifically detected in Nestin (Nes) reporter mice, wherein the gene encoding green fluorescent protein (GFP) is driven by the enhancer element of the second intron of Nes and minimal promoter of the heat shock protein-68 gene [21,22]. Furthermore, BM-MSPCs can be labeled based on Nes-GFP expression in another Nes-GFP line, which expresses GFP under the control of the 5.8-kb promoter and 1.8-kb fragment of the second intron of Nes [[23], [24], [25]]. Although these reports suggest that the Nes-GFP lines are useful for detecting the precursor population of hard tissue-forming cells, whether the Nes-GFP-positive population contributes to dental tissue regeneration and neoformation of odontoblast-like cells in the dental pulp remains elusive.
The biologically active N-terminal 1–34 peptide of the human parathyroid hormone [PTH (1–34)] exerts a bone anabolic activity and is therefore used clinically for the treatment of patients with osteoporosis. Previous studies have suggested that one of the mechanisms underlying the therapeutic action of PTH (1–34) is the positive regulation of BM-MSPC osteoblastogenesis [[26], [27], [28]]. The type I PTH/PTH-related peptide (PTHrP) receptor (PTH1R) has been reported to be expressed by diverse dental mesenchymal cell types, including odontoblasts, dental pulp cells, and dental follicle cells, during the neonatal period and contributes to root formation [29,30]. These reports suggest that the activation of additional signaling pathways downstream of PTH1R may positively affect the recovery of damaged dental tissue; however, experimental validation is still required.
Therefore, to understand the mechanistic details of dental tissue regenerative processes, herein, we induced Cre/LoxP-mediated odontoblast-specific cell death and found that this process significantly affects the dental pulp environment in triggering tissue regeneration.
Section snippets
Experimental animals
C57BL/6 mice were purchased from Japan SLC (Shizuoka, Japan). B6.FVB-Tg(Col1a1-cre)1Kry mice (Col1(2.3)-Cre) (RBRC0503) [31] were purchased from RIKEN BRC (Ibaraki, Japan). C57BL/6-Gt(ROSA)26Sortm1(HBEGF)Awai/J (iDTR) (JAX007900) mice were purchased from Jackson Laboratory (Bar Harbor, ME, USA). Col1(2.3)-GFP [32] mice were generated in one of the authors' laboratories (K.M.). Nes-GFP [23,25] mice were kindly provided by G. Enikolopov from Stony Brook University. All mice used for the
Odontoblasts are regenerated after depletion
To induce cell death specifically in the odontoblasts using a Cre/LoxP-based strategy, we confirmed the odontoblast-specific expression pattern of GFP in type I collagen α [Col1(2.3)]-GFP mice, in which GFP is expressed under the control of a 2.3-kb fragment of the Col1 promoter [29,32,40]. Consistent with previous reports, GFP expression was only observed in odontoblasts in the maxillary first molars of Col1(2.3)-GFP mice (Fig. 1A, arrows: Col1(2.3)-GFP+ odontoblasts). Based on these
Discussion
The immature odontoblast population is believed to contribute to the formation of reparative dentin that will replenish the depleted odontoblasts as a result of severe dental tissue damage; however, this regenerative process is still unclear. Targeted cell depletion approaches using genetically modified mice provide a method to understand the in vivo events related to tissue environment modifications, specifically those caused by cell death. Our comprehensive analyses revealed that pulp
Conclusion
In conclusion, while our study demonstrates that the activation of the PTH1R signaling cascade positively regulates reparative dentin formation, the mediators of the upregulation of PTH1R in response to Cre/LoxP-mediated odontoblast death or physical pulp-exposed dentin damage remain to be elucidated. Furthermore, it is still unclear whether the upregulation of PTH1R is also induced in damaged dental pulp tissue in humans. In addition, it is necessary to evaluate the age or sex differences in
Data availability
RNA-sequencing data that support the findings of this study have been deposited in the DDBJ Sequenced Read Archive under the accession number DRA010453.
CRediT authorship contribution statement
Lijuan Zhao: Formal analysis, Investigation, Methodology, Visualization, Writing – review & editing. Shinichirou Ito: Investigation, Visualization. Atsushi Arai: Investigation. Nobuyuki Udagawa: Investigation. Kanji Horibe: Investigation, Methodology. Miroku Hara: Investigation, Methodology, Visualization. Daisuke Nishida: Investigation. Akihiro Hosoya: Methodology. Rinya Masuko: Investigation, Methodology, Visualization. Koji Okabe: Investigation. Masashi Shin: Investigation, Visualization.
Declaration of competing interest
R.M. is an employee of JEOL Ltd., Tokyo, Japan. All other authors declare that they have no conflicts of interest with the contents of this article.
Acknowledgments
We would like to thank Asahi Kasei Pharma Corporation (Tokyo, Japan) for providing human PTH (1–34) and G. Enikolopov (Stony Brook University, Stony Brook, NY) for the Nes-GFP mice. We would also like to thank N. Takahashi (Matsumoto Dental University, Nagano, Japan), A. Yamaguchi (Tokyo Dental College, Tokyo, Japan), and R. Takao-Kawabata (Asahi Kasei Pharma Corporation) for useful advice and comments regarding our work. We appreciate the technical assistance provided by Y. Jing, Y. Mengyu,
Funding
This work was supported by Grants-in-Aid from the Japan Society for the Promotion of Science (JSPS) KAKENHI (20H03853 and 17H04374 to T.M.), Private University Branding Project of Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan: Tokyo Dental College Branding Project for Multidisciplinary Research Center for Jaw Disease (MRCJD) (T.M.), The Science Research Promotion Fund from PMAC (T.M.), Takeda Science Foundation (T.M.), and The JSBMR Frontier Scientist Grant (T.M.).
References (56)
- et al.
Endogenous bone marrow MSCs are dynamic, fate-restricted participants in bone maintenance and regeneration
Cell Stem Cell
(2012) - et al.
Osterix marks distinct waves of primitive and definitive stromal progenitors during bone marrow development
Dev. Cell
(2014) - et al.
Leptin-receptor-expressing mesenchymal stromal cells represent the main source of bone formed by adult bone marrow
Cell Stem Cell
(2014) - et al.
Secretion of shh by a neurovascular bundle niche supports mesenchymal stem cell homeostasis in the adult mouse incisor
Cell Stem Cell
(2014) - et al.
Dynamics of bromodeoxyuridine label-retaining dental pulp cells during pulpal healing after cavity preparation in mice
J. Endod.
(2013) - et al.
Parathyroid hormone directs bone marrow mesenchymal cell fate
Cell Metab.
(2017) - et al.
Cyclic AMP stimulates somatostatin gene transcription by phosphorylation of CREB at serine 133
Cell
(1989) - et al.
Modulation of osteoclast differentiation and function by the new members of the tumor necrosis factor receptor and ligand families
Endocr. Rev.
(1999) - et al.
Osteoclast differentiation and activation
Nature
(2003) - et al.
A subset of chondrogenic cells provides early mesenchymal progenitors in growing bones
Nat. Cell Biol.
(2014)
The application of bone morphogenetic proteins to dental tissue engineering
Nat. Biotechnol.
Making a tooth: growth factors, transcription factors, and stem cells
Cell Res.
Dental mesenchymal stem cells
Development
Dual origin of mesenchymal stem cells contributing to organ growth and repair
Proc. Natl. Acad. Sci. U. S. A.
Glial origin of mesenchymal stem cells in a tooth model system
Nature
αSMA-expressing perivascular cells represent dental pulp progenitors in vivo
J. Dent. Res.
Axin2-expressing cells differentiate into reparative odontoblasts via autocrine Wnt/β-catenin signaling in response to tooth damage
Sci. Rep.
Ten Cate’s Oral Histology: Development, Structure, and Function
A quiescent cell population replenishes mesenchymal stem cells to drive accelerated growth in mouse incisors
Nat. Commun.
Odontoblast differentiation
Int. J. Dev. Biol.
Cell dynamics in the pulpal healing process following cavity preparation in rat molars
Histochem. Cell Biol.
Thy-1-positive cells in the subodontoblastic layer possess high potential to differentiate into hard tissue-forming cells
Histochem. Cell Biol.
In vitro neurogenesis by progenitor cells isolated from the adult human hippocampus
Nat. Med.
Nestin expression is differently regulated between odontoblasts and the subodontoblastic layer in mice
Histochem. Cell Biol.
Neural stem and progenitor cells in nestin-GFP transgenic mice
J. Comp. Neurol.
Mesenchymal and haematopoietic stem cells form a unique bone marrow niche
Nature
Nestin-based reporter transgenic mouse lines
Methods Mol. Biol.
Parathyroid hormone regulates fates of murine osteoblast precursors in vivo
J. Clin. Invest.
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