Hardt Miriam, Mayr Alexandra, Kutschera Eric, Marciniak Jana, Küchler Erika Calvano, Kirschneck Christian, Deschner James, Jäger Andreas, Beisel-Memmert Svenja
Department of Orthodontics, University Hospital Bonn, Medical Faculty, Bonn, Germany.
School of Dentistry, Universidade Tuiuti do Paraná, Curitiba, Brazil.
Braz Dent J. 2024 Dec 6;35:e245944. doi: 10.1590/0103-6440202405944. eCollection 2024.
Autophagy is a self-digestion mechanism of cells, which is related to cell stress. It enables cell survival by maintaining cellular homeostasis or initiates cell death. This study aimed to investigate the intracellular signaling of pressure-induced autophagy regulation in human periodontal ligament (PDL) cells and to analyze the involvement of Rho kinases (ROCK) and reactive oxygen species (ROS) in particular. Human PDL cells were treated with the ROCK inhibitor Y-27632 and the ROS scavenger N-acetylcysteine (NAC) in combination with pressure magnitudes of 2, 6, and 8 g/cm2 over 16 hours. Cells treated with rapamycin served as a positive control and untreated cells as a control group. The Cyto-ID® Autophagy Detection Kit was used for flow cytometric analysis. Statistical analysis was performed using ANOVA and post-hoc tests. The results show that the pressure-induced autophagy was affected differently by the two inhibitors (p<0.05). The application of Y-27632 led to a significant reduction in autophagy in all pressure groups. The application of NAC led to reduced autophagy at pressures of 2 g/cm2 and 6 g/cm2. At 8 g/cm2, this effect was no longer present. In the control group, autophagy was significantly reduced by Y-27632 and significantly increased by NAC. Our data suggest that both Rho-kinase and reactive oxygen species could influence pressure-induced autophagy regulation in PDL cells.
自噬是细胞的一种自我消化机制，与细胞应激相关。它通过维持细胞内稳态使细胞存活，或引发细胞死亡。本研究旨在探究人牙周膜（PDL）细胞中压力诱导的自噬调节的细胞内信号传导，尤其分析Rho激酶（ROCK）和活性氧（ROS）的参与情况。将人PDL细胞与ROCK抑制剂Y-27632和ROS清除剂N-乙酰半胱氨酸（NAC）联合处理，压力大小分别为2、6和8 g/cm²，处理时间为16小时。用雷帕霉素处理的细胞作为阳性对照，未处理的细胞作为对照组。使用Cyto-ID®自噬检测试剂盒进行流式细胞术分析。采用方差分析和事后检验进行统计分析。结果表明，两种抑制剂对压力诱导的自噬影响不同（p<0.05）。Y-27632的应用导致所有压力组的自噬显著降低。NAC的应用导致在2 g/cm²和6 g/cm²压力下自噬减少。在8 g/cm²时，这种效果不再存在。在对照组中，Y-27632使自噬显著降低，而NAC使自噬显著增加。我们的数据表明，Rho激酶和活性氧都可能影响PDL细胞中压力诱导的自噬调节。
