Study On Autophagy In HPDLCs During Orthodontic Tooth Movement

Autophagy (comes from the Greek words: auto means "self" and phagein means "toeat", autophagy means "self-eating") is a kind of basic decomposition, cell deathmechanism. During this process, the degradation of unnecessary or dysfunctionalorganelles is accomplished by lysosomes. Autophagy is the essential mechanism for thecell metabolism regulation and cell viability maintenance. In the condition of disease, suchas tumor and infection, autophagy shows two kinds of effects. On one hand, autophagy isan adaptive response, it can provide necessary support for the survival of tumor cells, andpromote the development of the disease. On the other hand, autophagy can induceprogrammed pathological cell death and hinder the progress of the disease. It has beensuggested that some factors, such as endoplasmic reticulum(ER) stress, immune cellactivation, oxidative stress and infection, stimulate autophagy, can induce autophagy.Remodeling changes in paradental tissues are thought to be essential in orthodontictooth movement. During the orthodontic tooth movement, periodontal ligament whichconnects the tooth and alveolar bone plays an important role. Periodontal ligament fibroblasts are the major cells in the periodontal ligament. When orthodontic forcetransferred to periodontal ligament, periodontal ligament fibroblasts would secrete avariety of cytokines which could induce the remodeling of paradental tissues. Manyresearchers suggest that in the process of the orthodontic tooth movement, the periodontalspace would generate a pressure side. On the pressure side, the periodontal ligament isunder compression, which reduces the number of capillaries, cuases occlusion and partialdisintegration of blood vessels and ischaemia. In this condition, periodontal ligamentfibroblasts are in a starvation environment. So, the question is whether the starvationinduced the autophagy of periodontal ligament fibroblasts, and decreased the number ofcells? This mechanism directly related to the paradental tissues remodeling and the rootabsorption in orthodontic treatment. So, this study described the effects of vascularchanges in periodontal ligament during the orthodontic tooth movement, and theautophagy activity of human periodontal ligament fibroblasts in starvation. This study isconsisted of two aspects:1. Morphological and histological changes of periodontal tissue during toothmovement.Objective: After building the model of orthodontic tooth movement on rats, weanalysised histological changes of periodontal tissue during tooth movement. We alsoanalysised the changes of the environmental condition of periodontal ligament cells at thesame time.Methods: Rats were divided into4groups, the normal control group, theorthodontic1day group, the orthodontic3day group and the orthodontic7day group. Thecontrol group was subjected to no disposal. In the orthodontic group, a NiTi-coil springwas applied to the first upper molar of the right upper jaw, and draw first molar to themesial with30g force, the NiTi-coil springs were kept1day,3day,7day, then removed themolar and the corresponding surrounding tissues of the molar. After embedded in paraffin,the samples were further processed into5mm-thick sections for the following hematoxylineosin staining.Results: The results show that（1）The periodontal ligament width change: In the normal control group, the periodontal ligament width of distal and mesial ratio is1.012±0.124. In the orthodontic1day,3day,7day group, with the extension of time, theratio is increased (P <0.05), and in the orthodontics7day group, the radio is more than3times.(2) The vascular of periodontal ligament change: Compare with the contral group,the number of the vascular of periodontal ligament decreased and the vascular lumencompressed in orthodontic1day group and the orthodontic3day group. But, the numberof the vascular of periodontal ligament increased and the vascular lumen expand in theorthodontic7day group.(3) The number of periodontal ligament cells change: Comparedwith the contral group, the number of the periodontal ligament cells significantlydecreased in orthodontic1day group (P <0.05) and the orthodontic3day group (P <0.05). With the extension of time, this decrease trend is becoming more obvious. But,compared with the control group, the number of the periodontal ligament cellssignificantly increased in the orthodontic7day group (P <0.05).Conclusion: The orthodontic force could induce occlusion and partial disintegrationof blood vessels aroud periodontal ligament of the pressure side. And the periodontalligament fibroblasts death was due to it in the hypoxic and starvation environment.2. The effects of starvation on autophagy of human periodontal ligament fibroblastsObjective: To study whether starvation could induce the autophagy of periodontalligament.Methods: periodontal ligament fibroblasts were harvested through primary culturing.Periodontal ligament fibroblasts were divided into normal control group and starvationgroup. In the starvation group, EBSS was used and its treating time was1h,2h,3h,4h and6h respectively. The formation of the autophagosome in periodontal ligament fibroblastswas observed through Transmission Electron Microscope. Furthermore, the expression ofautophagy related protein LC3in the human periodontal ligament fibroblasts was alsochecked through western blot and immunocytochemistry.Results: The autophagosomes and the expression of LC3in the cells treated withEBSS were more than that in the control cells, especially in the2h,3h and4h EBSStreated group. Results of western blot revealed that the ratio of LC3-II/LC3-I in the starvation group was significantly higher than that of normal control group (P <0.05). Inthe starvation group, the ratio of LC3-II/LC3-I elevated after1h and reached its highestpoint in4h(P <0.05), then decreased after6h (P <0.05).Conclusion: The starvation condition could induce the autophagy of the humanperiodontal ligament fibroblasts and the autophagy might play a certain role in thereconstruction procedure caused by the human periodontal ligament fibroblastsThis study showed that the orthodontic force can induce occlusion and partialdisintegration of blood vessels in the pressure side of periodontal ligament. The death ofhuman periodontal ligament fibroblasts was due to the hypoxic and starvationenvironment. The vitro experiment showed that starvation environment could lead to theincrease of the number of autophagosomes and the expression of LC3.In the tissue remodeling process of the pressure side of periodontium during theorthodontic tooth movement, all the changes above are the adaptation change of cells,Furthermore, these adaptation changes might be an important mechanism of cells death inthe pressure side.