Oxidative stress and inflammation play important roles in the development of diabetes mellitus. p-Synephrine, the primary pharmacologically active protoalkaloid in Citrus species, has been popularly consumed as a dietary supplement...
The odontogenic differentiation of dental pulp stem cells (DPSCs), which is vital for tooth regeneration, was regulated by various functional molecules. In recent years, a growing body of research has shown that miRNAs play a crucial role in the odontogenic differentiation of human dental pulp stem cells (hDPSCs). However, the mechanisms by which miRNAs regulated odontogenic differentiation of hDPSCs remained unclear, and the application of miRNAs in reparative dentin formation in vivo was also rare. In this study, we first discovered that miR-3074-3p had an inhibitory effect on odontogenic differentiation of hDPSCs and antagomiR-3074-3p-conjugated PEI-AuNPs effectively promoted odontogenic differentiation of hDPSCs in vitro. AntagomiR-3074-3p-conjugated PEI-AuNPs was further applied to the rat pulp-capping model and showed the increased formation of restorative dentin. In addition, the results of lentivirus transfection in vitro suggested that FKBP9 acted as the key target of miR-3074-3p in regulating the odontogenic differentiation of hDPSCs. These findings might provide a new strategy and candidate target for dentin restoration and tooth regeneration.
Background Dental pulp stem cells (DPSCs) play a crucial role in dentin-pulp complex regeneration. Further understanding of the mechanism by which DPSCs remain in a quiescent state could contribute to improvements in the dentin-pulp complex and dentinogenesis. Methods TSC1 conditional knockout (DMP1-Cre+; TSC1f/f, hereafter CKO) mice were generated to increase the activity of mechanistic target of rapamycin complex 1 (mTORC1). H&E staining, immunofluorescence and micro-CT analysis were performed with these CKO mice and littermate controls. In vitro, exosomes were collected from the supernatants of MDPC23 cells with different levels of mTORC1 activity and then characterized by transmission electron microscopy and nanoparticle tracking analysis. DPSCs were cocultured with MDPC23 cells and MDPC23 cell-derived exosomes. Alizarin Red S staining, ALP staining, qRT‒PCR, western blotting analysis and micro-RNA sequencing were performed. Results Our study showed that mTORC1 activation in odontoblasts resulted in thicker dentin and higher dentin volume/tooth volume of molars, and it increased the expression levels of the exosome markers CD63 and Alix. In vitro, when DPSCs were cocultured with MDPC23 cells, odontoblastic differentiation was inhibited. However, the inhibition of odontoblastic differentiation was reversed when DPSCs were cocultured with MDPC23 cells with mTORC1 overactivation. To further study the effects of mTORC1 on exosome release from odontoblasts, MDPC23 cells were treated with rapamycin or shRNA-TSC1 to inactivate or activate mTORC1, respectively. The results revealed that exosome release from odontoblasts was negatively correlated with mTORC1 activity. Moreover, exosomes derived from MDPC23 cells with active or inactive mTORC1 inhibited the odontoblastic differentiation of DPSCs at the same concentration. miRNA sequencing analysis of exosomes that were derived from shTSC1-transfected MDPC23 cells, rapamycin-treated MDPC23 cells or nontreated MDPC23 cells revealed that the majority of the miRNAs were similar among these groups. In addition, exosomes derived from odontoblasts inhibited the odontoblastic differentiation of DPSCs, and the inhibitory effect was positively correlated with exosome concentration. Conclusion mTORC1 regulates exosome release from odontoblasts to inhibit the odontoblastic differentiation of DPSCs, but it does not alter exosomal contents. These findings might provide a new understanding of dental pulp complex regeneration.
Dental pulp stem cells (DPSCs) are the crucial part of dentin-pulp complex regeneration. A further understanding of the mechanism of how DPSCs stay in a quiescent state can contribute to improvements in dentin-pulp complex and dentinogenesis. Here, our study discovered that mTORC1 activation in odontoblasts resulted in thicker dentin and higher dentin volume/tissue volume of molars and led to increased exosomes marker expression levels of CD63 and Alix. In vitro, we confirmed that mTORC1 inactivation in odontoblasts reverses the inhibition of DPSC odontoblastic differentiation when cocultured with odontoblasts. Besides, our study indicated that exosomes derived from mTORC1-activated or mTORC1-inactivated MDPC23 both inhibited the odontoblastic differentiation of DPSCs and at same concentration. The miRNA sequence of exosomes derived from shTSC1-tansfected MDPC23 or rapamycin-treated MDPC23 or non-treated MDPC23 indicated that the majority of the miRNA was similar. Furthermore, we discovered that the exosomes derived from odontoblasts inhibited the odontoblastic differentiation of DPSCs, and the inhibitory effect was positively correlated with exosomes concentration. Taken together, mTORC1 regulates the exosomes release by odontoblasts to inhibit the odontoblastic differentiation of DPSCs but not the exosomes contents. These findings might provide a new understanding for dental-pulp complex regeneration.
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