Dentin, which composes most of the tooth structure, is formed by odontoblasts, long-lived post-mitotic cells maintained throughout the entire life of the tooth. In mature odontoblasts, however, cellular activity is significantly weakened. Therefore, it is important to augment the cellular activity of mature odontoblasts to regenerate physiological dentin; however, no molecule regulating the cellular activity of mature odontoblasts has yet been identified. Here, we suggest that copine-7 (CPNE7) can reactivate the lost functions of mature odontoblasts by inducing autophagy. CPNE7 was observed to elevate the expression of microtubule-associated protein light chain 3-II (LC3-II), an autophagy marker, and autophagosome formation in the pre-odontoblast and mature odontoblast stages of human dental pulp cells. CPNE7-induced autophagy upregulated DSP and DMP-1, odontoblast differentiation and mineralization markers, and augmented dentin formation in mature odontoblasts. Furthermore, CPNE7 also upregulated NESTIN and TAU, which are expressed in the physiological odontoblast process, and stimulated the elongation of the odontoblast process by inducing autophagy. Moreover, lipofuscin, which progressively accumulates in long-lived post-mitotic cells and hinders their proper functions, was observed to be removed in recombinant CPNE7-treated mature odontoblasts. Thus, CPNE7-induced autophagy reactivated the function of mature odontoblasts and promoted the formation of physiological dentin in vivo. On the other hand, the well-known autophagy inducer, rapamycin, promoted odontoblast differentiation in pre-odontoblasts but did not properly reactivate the function of mature odontoblasts. These findings provide evidence that CPNE7 functionally reactivates mature odontoblasts and introduce its potential for dentinal loss-targeted clinical applications.
Alveolar bone is both morphologically and functionally different from other bones of the axial or peripheral skeleton. Because of its sensitive nature to external stimuli including mechanical stress, bone loss stimuli, and medication‐related osteonecrosis of the jaw, alveolar bone rendering is seen as an important factor in various dental surgical processes. Although multiple studies have validated the response of long bone to various factors, how alveolar bone responds to functional stimuli still needs further clarification. To examine the characteristics of bone in vitro, we isolated cells from alveolar, femur, and tibia bone tissue. Although primary cultured mouse alveolar bone‐derived cells (mABDCs) and mouse long bone‐derived cells (mLBDCs) exhibited similar osteoblastic characteristics, morphology, and proliferation rates, both showed distinct expression of neural crest (NC) and epithelial–mesenchymal interaction (EMI)‐related genes. Furthermore, they showed significantly different mineralization rates. RNA sequencing data demonstrated distinct transcriptome profiles of alveolar bone and long bone. Osteogenic, NC‐, and EMI‐related genes showed distinct expression between mABDCs and mLBDCs. When the gene expression patterns during osteogenic differentiation were analyzed, excluding several osteogenic genes, NC‐ and EMI‐related genes showed different expression patterns. Among EMI‐related proteins, BMP4 elevated the expression levels of osteogenic genes, Msx2, Dlx5, and Bmp2 the most, more noticeably in mABDCs than in mLBDCs during osteogenic differentiation. In in vivo models, the BMP4‐treated mABDC group showed massive bone formation and maturation as opposed to its counterpart. Bone sialoprotein expression was also validated in calcified tissues. Overall, our data suggest that alveolar bone and long bone have different responsiveness to EMI by distinct gene regulation. In particular, BMP4 has critical bone formation effects on alveolar bone, but not on long bone. © 2020 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.
Aim Once the periodontal ligament (PDL) is damaged, it is difficult to regenerate its characteristic structure. Copine7 (CPNE7) reportedly plays a functional role in supporting periodontal attachment and PDL alignment. Here we demonstrate the regulatory mechanism of CPNE7 coordination with cytoskeleton reorganization and cementum attachment protein (CAP)‐mediated attachment in PDL regeneration. Materials and Methods The expression and localization of CPNE7, α‐TUBULIN, ACTIN, and microtubule associated protein tau (TAU) were investigated in vitro. The effects of recombinant CPNE7 (rCPNE7) and CPNE7‐derived peptides (CPNE7‐DP) on the regulation of CAP were analysed in vitro, and PDL repair capacity was analysed in vivo. Results CPNE7 co‐localized with F‐ACTIN and induced α‐TUBULIN expansion to the edge of human PDL cells (hPDLCs). ACTIN and α‐TUBULIN protein expressions were not elevated in rCPNE7‐treated hPDLCs. rCPNE7 elevated the protein expression of TAU, which co‐localized with F‐ACTIN and α‐TUBULIN. Replantation studies on mice revealed that well‐attached and well‐aligned PDLs were repaired in the rCPNE7 group. CPNE7‐DP directly up‐regulate the expression of CAP in vitro and promote PDL regeneration in three‐wall defect canine models in vivo. Conclusions Our findings suggest that CPNE7 helps in PDL repair by supporting PDL alignment through TAU‐mediated cytoskeleton reorganization and direct regulation of CAP‐mediated PDL attachments of PDLCs.
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