The Mechanism Of Sensory Nervous System Supporting Homeostasis In The Rat Incisor

Tooth formation is a long-term continuous process,not only in embryonic period,and continue until after birth.Peripheral nerve-associated glia cells generate multipotent mesenchymal stem cells that produce pulp cells and odontoblasts and play an important role in self-renewal and repair of a tooth.The interactions between epithelial and mesenchymal cells makes the rat incisor tooth continuously growing,therefore,offers an excellent model to address the self-renewal and repair of a tooth.Self-renewal of the incisor epithelium is supported by a group of quiescent epithelial stem cells in the cervical loop region and the continuous turnover of odontoblasts and pulp cells are supported by dental pulp stem cells(rDPSCs)within the mesenchyme located near the neurovascular bundle(NVB).The incisor is innervated by sensory nerve derived from inferior alveolar nerve(IAN)and sympathetic nerves derived from the superior cervical ganglion(SCG).The mechanism of nervous system supporting homeostasis in self-renewal and repair of a tooth are poorly understood.To address this issue,we established the animal model of denervated incisor tooth and identified the morphological and histological changes of incisor tooth.We further determined the biological performance of rDPSCs gained from the denervated incisor tooth.Eventually,we explored the molecular mechanism of nervous system supporting homeostasis in rat incisor.This research provide experimental basis for the mechanism of sensory nervous system supporting homeostasis in tooth.Part one: The study on the morphological and histological changes of denervated incisor toothObject: To clarify the roles sensory and sympathetic nerves played respectively in supporting homeostasis in the rat incisor.Method: Establish the animal model of sectioning the unilateral sensory nerve(IANx),sectioning the unilateral sympathetic nerve(SCGx),sectioning the unilateral sensory nerve and sympathetic nerve(IANx+SCGx)and the sham surgical group.The rats were sacrificed at the second week after surgical and incisors were studied by micro-CT.HE and Masson staining were further performed to observe the changes of incisors.Result: The incisors of IANx group and IANx+SCGx group turned chalky 2 weeks after denervation.The micro-CT findings revealed that irregular high-density calcifications existed in pulp and the formation of enamel and dentin reduced.HE and Masson staining of IANx group and IANx+SCGx group showed the disorder structure of pulp,degeneration of odontoblasts and reduced formation of enamel and dentin.Conclusion: The sensory nerve supports the homeostasis in the rat incisor and the sympathetic nerve seems to be of no importance of it.Part two: The study on biological properties of rat dental pulp stem cells(rDPSCs)after sectioning the sensory nerveObject: To explore the biological properties of r DPSCs obtained from rats sectioned the sensory nerve.Method: rDPSCs from IANx group and sham group were obtained and cultivated to detect molecular surface markers,clone formation ability,proliferation ability,multi-directions differentiation ability and apoptosis.Result: Both rDPSCs from IANx group and sham group could positively express the mesenchymal surface markers CD29 and CD105,while the hematopoietic marker CD34 was negative.The rDPSC of IANx group having significantly lower clonogenic potential,proliferative potential,osteogenesis and adipogenic differentiation potential but higher level of apoptosis.Conclusion: The biological behaviour of rDPSCs could be regulated by sensory nervous system.Part three: The study on the mechanism of sensory nervous system supporting homeostasis in the rat incisor through Wnt/?-Catenin signaling pathwayObject: To explore the mechanism of sensory nervous system supporting homeostasis in the rat incisor through Wnt/?-Catenin signaling pathway.Method: Q-PCR and Western Blot were used to determine the expression level of Wnt/?-Catenin signaling pathway related genes and protein after losting the innervation of sensory nerve.The Wnt/?-Catenin signaling pathway of rDPSCs from IANx group was activated by the agonist LiCl.Then,the clone formation ability,proliferation ability,multi-directions differentiation ability and apoptosis were tested compared with the rDPSCs from IANx group and sham group.Result: Denervation led to downregulation of the Wnt signaling pathway and the agonist LiCl could activated Wnt/?-Catenin signaling pathway.The rDPSCs from IANx group exhibit stronger proliferative potential,osteogenesis and adipogenic differentiation potential after activated by LiCl but still weaker than the sham group.Conclusion: Wnt/?-Catenin signaling pathway participate in the mechanism of sensory nervous system supporting homeostasis in the rat incisor.