Liu Yu, Li Shujin, Zhang Senlin, Cao Xiaoma, Zhang Yanmei
Department of Stomatology, Second People's Hospital of Hefei City, Hefei 230011, China. *Corresponding author, E-mail:
Department of Clinical Laboratory, Second People's Hospital of Hefei City, Hefei 230011, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2017 Mar;33(3):315-319.
Objective To investigate the impact of lipopolysaccharide of Porphyromonas gingivalis (Pg-LPS) on the autophagy of human gingival fibroblasts (HGFs). Methods Firstly, HGFs was stimulated with 10 μg/mL Pg-LPS for 12 hours or 24 hours. Rapamycin was used as a positive control. The expression of LC3B was detected by Western blotting and the distribution of autophagosomes was observed by indirect immunofluorescence staining. At the same time, mitochondrial ROS (mtROS) was labeled by MitoSOX Red. The levels of mtROS and mitochondrial autophagy were measured in HGFs after treated with Pg-LPS. Then, T-butyl-4 (BHA), N-acetylcysteine (NAC) and coenzyme Q10 (CoQ10) were used separately to block the ROS and the expression of LC3B in Pg-LPS-treated HGFs was tested by Western blotting. Results After treatment with Pg-LPS, the ratio of LC3BII/LC3BI and the number of autophagic cells significantly increased, and the increase in the 24-hour treatment group was the more obvious than that in the 12-hour treatment group. The mtROS production and mitochondrial autophagy were significantly promoted after Pg-LPS treatment. In addition, CoQ10 effectively reduced Pg-LPS-induced autophagy of HGFs. Conclusion Pg-LPS can induce the autophagy of HGFs by raising mtROS production, and autophagy is involved in the degradation of damaged mitochondria to maintain cellular homeostasis.
目的 探讨牙龈卟啉单胞菌脂多糖(Pg-LPS)对人牙龈成纤维细胞(HGFs)自噬的影响。方法 首先,用10 μg/mL Pg-LPS刺激HGFs 12小时或24小时。雷帕霉素用作阳性对照。通过蛋白质免疫印迹法检测LC3B的表达,并通过间接免疫荧光染色观察自噬体的分布。同时,用MitoSOX Red标记线粒体活性氧(mtROS)。检测Pg-LPS处理后HGFs中mtROS水平和线粒体自噬情况。然后,分别使用叔丁基对苯二酚(BHA)、N-乙酰半胱氨酸(NAC)和辅酶Q10(CoQ10)阻断活性氧,通过蛋白质免疫印迹法检测其对Pg-LPS处理的HGFs中LC3B表达的影响。结果 经Pg-LPS处理后,LC3BII/LC3BI比值和自噬细胞数量显著增加,且24小时处理组的增加比12小时处理组更明显。Pg-LPS处理后mtROS生成和线粒体自噬显著增强。此外,CoQ10可有效降低Pg-LPS诱导的HGFs自噬。结论 Pg-LPS可通过提高mtROS生成诱导HGFs自噬,且自噬参与受损线粒体的降解以维持细胞稳态。