A20通过抑制自噬缓解实验性牙周炎牙槽骨吸收

A20 alleviates alveolar bone loss in experimental periodontitis by inhibiting autophagy

目的:研究泛素编辑酶A20在抑制小鼠实验性牙周炎牙槽骨吸收中的作用并探索其潜在机制。方法:将20只小鼠随机分为4组:无牙周炎(Control)组、牙周炎及PBS注射(PBS+P)组、牙周炎及阴性对照腺相关病毒(adeno-associated virus,AAV)注射(AAV+P)组、牙周炎及A20过表达AAV(AAV-A20)注射(A20+P)组。采用丝线结扎联合局部涂菌构建C57BL/6J小鼠实验性牙周炎模型。在小鼠牙龈局部注射AAV-A20以实现A20在牙周组织的过表达。A20免疫荧光染色验证AAV-A20在局部牙周组织的转染效率。微计算机断层扫描(microcomputed tomography,Micro-CT)、苏木素伊红(haematoxylin and eosin,HE)染色及抗酒石酸酸性磷酸酶(tartrate-resistant acid phosphatase,TRAP)染色比较各组小鼠牙槽骨吸收程度及上颌第一第二磨牙之间破骨细胞数目。免疫组织化学染色观察各组小鼠牙周组织核因子-κB受体活化因子配体(receptor activator of nuclear factor-κВligand,RANKL)及自噬相关因子表达。结果:与PBS+P组及AAV+P组相比,A20+P组小鼠牙周组织自噬水平降低(Beclin-1、LC3B表达下降,p62表达上升),RANKL表达下调,上颌第一第二磨牙之间TRAP阳性破骨细胞数目减少,牙槽骨吸收程度减轻。结论:A20通过负向调控自噬缓解小鼠实验性牙周炎牙槽骨吸收,有望成为牙周炎治疗的新靶点。

Objective:This study aims to explore the role of ubiquitin-editing enzyme A20 in inhibiting alveolar bone resorption in mice with experimental periodontitis and the potential mechanism underlying it. Methods:Twenty mice were randomly divided into 4groups:normal group with no periodontitis(control),periodontitis and PBS treatment(PBS+P)group,periodontitis and negative control adeno-associated virus(AAV)treatment(AAV+P)group,periodontitis and AAV targeting A20(AAV-A20)treatment(A20+P)group.Silk ligation combined with local application of Porphyromonas gingivalis(P. gingivalis)suspensions was used to construct the experimental periodontitis model. Gingiva of mice were locally injected with AAV-A20 to achieve A20 overexpression in the periodontal tissue. Immunofluorescence for A20 was used to verify AAV-A20 transfection efficiency in the periodontal tissue.microcomputed tomography(Micro-CT),haematoxylin and eosin(HE)staining and tartrate-resistant acid phosphatase(TRAP)staining were employed to compare the degree of alveolar bone resorption and the number of osteoclasts between the maxillary first and second molar of mice. Expressions of RANKL(receptor activator of nuclear factor-κВ ligand)and autophagy-related molecules in the periodontal tissue of mice were detected by immunohistochemical staining. Results:Compared with the PBS+P and AAV+P group,mice in the A20+P group exhibited decreased autophagic level and RANKL expression in their peridontal tissues. Besides,the number of TRAP positive osteoclasts between the maxillary first and second molar and alveolar bone resorption in mice were also repressed in the A20+P group. Conclusion:A20 alleviates alveolar bone resorption in mice with experimental periodontitis through negative regulation of autophagy,and A20 is expected to be a new target for periodontitis treatment.