Rho Kinases and Reactive Oxygen Species in Autophagy Regulation by Pressure in Periodontal Ligament Cells

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.