Study On The Mechanism Of Nerve Factor Affecting The Process Of Rat Periodontal Tissue Defect Healing

Background and target:It is emphasised currently in periodontal treatment field that how to make an effective "increment" of diseased periodontal tissues, especially on the premise of "qualified" functional alveolar bone, thereby the loose teeth is refastened and exercised normal masticatory function. The diseased periodontal tissue is reflected mainly on the disabled absorption of alveolar bone, meanwhile the structure and function of neurovascular system of the diseased part is broken, considering from the point view of development biology, and innervation might play the most important role during regeneration of periodontal tissue. In the sequential treatment strategies of periodontal disease, alleviating inflammation and controlling disease progression were basically achieved by taking various non-surgical treatments. However, it is always considered as an important task in periodontal field to research and explore the way to promote the regeneration of periodontal tissue especially qualified alveolar bone, not only from clinical practice, but also from theoretical research.In many existing strategies and related research in the treatment of periodontal tissue regeneration, it is more popular to research from seed cells, bio-scaffolds and growth factors, however, due to the etiology of periodontal disease and the complexity of repairing of corresponding tissue, as well as the systemic regulation of body tissue regeneration, the results obtained through these did not reach the aim of periondontal tissue regeneration which we are pursuing. Alveolar bone is considered as the most active bone in metabolism, it is reasonable to deduce that neurovascular system plays an important role in the process of alveolar bone remolding, however, most of existing researches focus on how to get more regeneration tissue, but the regeneration of neurovascular is ignored, compared with physiologic periodontal tissue, the artificial induced regenerative tissues has an obvious shortage on structure, density and stress. Obviously, precision of organism was ignored in the "simple recover" compared with "natural formation". Explore the effects of nerve tissue on the process of periodontal tissue regeneration process has great significance in the field of periodontal tissue regeneration. Currently studies had confirmed that nervous system could be involved in the regulation of bone metabolism through mediated the activity of osteoblast and osteoclast in vitro, which played a critical role in bone remodeling. In recent years, along with the in-depth studies on the influence of center and peripheral nervous system on fracture healing, and found that bone tissue was the nerve target organ, peripheral nervous system played a critical role in fracture healing and bone metabolism.Recently studies found that the peripheral nervous system worked on bone tissue by neuropeptide which secreted by peptidergic nerve, and Calcitonin gene related peptide (which is called CGRP hereafter) is a kind of neural peptide which is widely distributed in bone tissue and mainly located in the active section of bone metabolism, it was found that CGRP effected the fracture healing through directly or indirectly regulating the differentiation and activity of osteoblast and osteoclast. It was first discovered by Hukkanen in rat fracture model experiment, CGRP immunohistochemical staining fibers could be found in newly formed bone tissue, soon after the fuction and mechanism of CGRP became the hot point in fracture healing remodeling studies.Studies showed that the CGRP positive nerve fibers are distributed in the jaw bone and periosteum, suggesting its involvement in the jaw bone forming during growth stage and the later remodeling. In periodontal tissue, calcitonin gene related peptide positive nerve fibers are distributed in molar periodontal tissue, alveolar bone and surrounding arterioles and venules were surrounded by CGRP, suggesting that CGRP positive nerves plays a critical role in neural regeneration during the regeneration of periodontal tissue. In the studies of dental surgery, there were many studies on the CGRP influence on fracture healing, studies showed that CGRP could promote the healing of mandibular fracture.In orthodontic medical studies, CGRP had a kind of regulating function on alveolar bone remodeling during teeth movement. Based on above-mentioned rationales, we speculated that CGRP was an indispensable factor in periodontal tissue regeneration. We have identified that nerves can affect the quality of periodontal tissue regeneration by histological experiment provously. The expression of CGRP was changed during periodontal regeneration process by immunohistochemical observation. In order to further explore the role of CGRP in the process of periodontal tissue regeneration, we will take further study on the mechanism of CGRP in its gene and protein level. Methods:1. Researches on the expression changes of CGRP and SP in rat periodontal tissues after cutting off inferior alveolar nerve.Established rat model of cutting of inferior alveolar verve, took left severing nerve as experimental group, and kept the right nerve as the control group, divided into six groups as0days,3days,7days,14days,21days and28days. Realtime PCR and Western Blot were applied to detect the expression changes of CGRP and SP in rat periodontal tissue.2. Changes of CGRP、SP and bone remolding related factors expression in rats periodontal tissue healing process after cutting inferior alveolar nerveUsed the extraoral approach to establish the cut inferior alveolar nerve accompanied periodontal defect model in rats, the left cut the inferior alveolar nerve in rats accompanied periodontal defects as the experimental group, the opposite side nerve-sparing only do periodontal defect for the control group. Divided into five groups as1week,2weeks,4weeks,6weeks and8weeks. Realtime-PCR and Western Blot were applied to detect the expression changes of CGRP、SP、OPG and RANKL in ratperiodontal tissues.Results:1. Changes of CGRP and SP in rat periodontal tissue after cutting off inferior alveolar nerve.CGRP and SP level of gingiva and alveolar bone were decreasing tendency in early stage then raised to normal level after denervation according to the examination results of Realtime-PCR. There were statistical significantly different among3days,7days and14days groups (P<0.05).The results of Western Blot found that changes of SP protein levels is basically the same as SP mRNA after denervation in rat periodontal tissue, There were statistical significantly different among3days,7days and14days groups(P<0.05),Western Blot detection of CGRP protein is negative.2. Changes of CGRP、SP and bone remolding related factors expression in rats periodontal tissue healing process after cutting inferior alveolar nerveThe level of CGRP expression in experimental group was a downward trend in early stage then began to rise from4weeks and which was still at elevated levels until eighth week. In the control group, the expression level was significantly higher than that in the early stage(P<0.05),then gradually returned to normal levels. The trend of SP expression was similarly to CGRP’s. Loosing of the inferior alveolar nerve led to the OPG sustained especially weak positive expression but RANKL continued strong positive expression.Conclusion:1. CGRP and SP expression are changed in periodontal tissues following inferior alveolar nerve transection in rats.2. CGRP may affect periodontal tissue healing process by influencing the expression of osteogenic factor and osteoclast factor...