| Objective: After orthodontic treatment of malocclusion,the balance of the original oral and jaw system is broken,and the moving teeth and jaws tend to return to their original state,that is,the recurrence of malocclusion.At present,the common methods for preventing recurrence in clinical practice all have shortcomings.Therefore,how to keep the orthodontic mobile teeth in a stable and ideal position and shorten the retention time has become an urgent problem for orthodontists.The main reason for the recurrence is that the reconstruction of the periodontal ligament and alveolar bone of the teeth has not been completed.Bone remodeling is mainly completed by the mutual coupling of osteoblasts,osteoclasts and mesenchymal stem cells(MSC).The dynamic balance of bone remodeling is the basic guarantee for the skeletal system to maintain its shape and perform normal functions.Among them,MSC plays an important role in regulating bone remodeling.Odonto-derived stem cells are a type of MSC with multi-differentiation potential from oral and maxillofacial tissues.Compared with bone marrow mesenchymal stem cells,tooth-derived stem cells have the advantages of low immunogenicity,superior cell proliferation and osteogenic differentiation.Therefore,studying the effect of odontogenic stem cells on bone remodeling and exploring the molecular biological mechanism of odontogenic stem cells regulating bone remodeling is of great significance for preventing,delaying or even blocking recurrence after orthodontics.Exosomes are nano-sized vesicles secreted by cells.More and more evidences show that exosomes play an important role in the process of bone remodeling,and the specific proteins,m RNA,miRNA,etc.carried by them may be the key to the role of exosomes in regulating bone remodeling.In addition,exosomes are stable in nature,easy to store,low in immunogenicity,and easier to deliver between cells.Therefore,the use of exosomes to regulate the reconstruction and repair of local tissues has a better prospect.In recent years,the role of micro RNAs in exosomes in signal communication between cells has attracted more and more attention from researchers.Researchers have discovered that miRNAs carried by stem cell exosomes can repair rat skull defects and promote bone tissue regeneration.Whether dental stem cell exosomes can participate in the reconstruction of local alveolar bone is still unclear.mTOR is a mammalian target of rapamycin(mTOR),which is a serine/threonine kinase.mTOR is involved in a variety of pathological and physiological processes.The latest research found that mTOR is also essential for bone remodeling.The previous research of our group found that the inhibition of PI3K/Akt/mTOR signal pathway was accompanied by the slowdown of tooth movement.It is speculated that the inhibition of mTOR may help to inhibit the recurrence of orthodontic tooth movement.And found that local injection of periodontal ligament stem cell exosomes around the teeth in the recurrence stage can inhibit their recurrence.Therefore,we speculate that local injection of periodontal ligament stem cell exosomes may inhibit mTOR,thereby promoting the mechanism of bone tissue remodeling,and achieve the effect of inhibiting recurrence.Therefore,it is of great clinical significance to clarify the mechanism of periodontal ligament stem cell exosomes inhibiting the recurrence of teeth after treatment,and the application of PDLSC-Exo to orthodontic tooth movement and inhibiting recurrence has not been reported.In summary,this project intends to use exosomes as an entry point to explore the regulatory effects of periodontal ligament stem cell exosomes and their biologically active substances on the remodeling of alveolar bone around orthodontic mobile teeth and related molecular mechanisms,in order to solve the clinical problem Recurrence after orthodontic treatment provides a theoretical basis.Methods:(1)Obtain periodontal ligament stem cells through primary cell culture,and identify the cells by flow cytometry and induced differentiation experiments;obtain periodontal ligament stem cell exosomes by ultracentrifugation,and use Western blot,Transmission electron microscopy and NTA are used to characterize exosomes.Observe the endocytosis of exosomes into recipient cells by confocal microscope.(2)Establish a rat orthodontic tooth model.During the recurrence of orthodontic tooth movement,local injection of periodontal ligament stem cell exosomes is used to record the recurrence of orthodontic teeth through impressions,and analyze the recurrence distance and recurrence rate.Hematoxylin and eosin staining was used to observe the effect of periodontal ligament stem cell exosomes on periodontal tissues,and real-time quantitative PCR was used to detect the expression of osteoclast-related genes in periodontal tissues.At the cellular level,explore the effect of periodontal ligament stem cell exosomes on the osteogenic capacity,proliferation capacity and migration capacity of cells.(3)Through bioinformatics research methods,excavate and verify the possible high expression of miRNA in periodontal ligament stem cell exosomes.In vitro experiments to explore the effects of target miRNA and exosomal miRNA on the osteogenic ability of cells.Using bioinformatics research methods again,predict the possible target genes of miRNA,and verify them through in vivo and in vitro experiments.Results:(1)The periodontal ligament stem cells were successfully extracted through primary cell culture,and successfully verified by flow cytometry and osteogenic adipogenic differentiation experiments;the periodontal ligament stem cell exosomes were successfully extracted by ultracentrifugation,and It has been characterized by various means.Under a confocal microscope,it is clear that periodontal ligament stem cell exosomes can enter recipient cells through endocytosis.(2)Successfully established a rat orthodontic movement model.Animal experiments show that periodontal ligament stem cell exosomes can inhibit the movement of orthodontic teeth in rats,reduce the recurrence distance and recurrence rate.Periodontal ligament stem cell exosomes can increase the expression level of osteogenesis-related genes in local periodontal tissues.Cell experiments show that periodontal ligament stem cell exosomes can promote the osteogenic differentiation ability of cells,and can also promote the proliferation and migration ability of osteoblasts.(3)Successfully excavated the database and verified the high expression of miR-100-5p in the exosomes of periodontal ligament stem cells in exosomes.Through in vitro experiments,it is clear that miR-100-5p and exosomal miR-100-5p can promote osteogenic differentiation of cells.Successfully predicted and verified the target gene of miR-100-5p as mTOR through the database.Conclusion: Periodontal ligament stem cell exosomes miR-100-5p can adjust the improvement of the local alveolar bone of orthodontic mobile teeth through mTOR,and inhibit the recurrence after orthodontic treatment. |
