Dept. of Orthodontics, Affiliated Stomatology Hospital of Southwest Medical University, Luzhou 646000, China.
Orofacial Reconstruction and Regeneration Laboratory, Southwest Medical University, Luzhou 646000, China.
Hua Xi Kou Qiang Yi Xue Za Zhi. 2023 Jun 1;41(3):260-268. doi: 10.7518/hxkq.2023.2022382.
This work aimed to investigate the molecular mechanism of cyclic tensile stress (CTS) stimulating autophagy in human periodontal ligament cells (hPDLCs).
hPDLCs were isolated and cultured from normal periodontal tissues. hPDLCs were loaded with tensile stress by force four-point bending extender to simulate the autophagy of hPDLCs induced by orthodontic force du-ring orthodontic tooth movement. XMU-MP-1 was used to inhibit the Hippo signaling pathway to explore the role of the Hippo-YAP signaling pathway in activating hPDLC autophagy by tensile stress. The expression levels of autophagy-related genes (Beclin-1, LC3, and p62) in hPDLCs were detected by real-time quantitative polymerase chain reaction. Western blot was used to detect the expression levels of autophagy-related proteins (Beclin-1, LC3-Ⅱ/LC3-Ⅰ, and p62) and Hippo-YAP pathway proteins (active-YAP and p-YAP) in hPDLCs. Immunofluorescence was used to locate autophagy-related proteins (LC3-Ⅱand p62) and Hippo-YAP pathway proteins (active-YAP) of hPDLCs.
CTS-activated autophagy in hPDLCs and expression of autophagy-related proteins initially increased and then decreased; it began to increase at 30 min, peaked at 3 h, and decreased (<0.05). CTS increased the expression of active-YAP protein and decreased the expression of p-YAP protein (<0.05). When XMU-MP-1 inhibited the Hippo-YAP signaling pathway (<0.05), active-YAP protein was promoted to enter the nucleus and autophagy expression was enhanced (<0.05).
The Hippo-YAP signaling pathway is involved in the regulation of autophagy activation in hPDLCs under CTS.
本研究旨在探讨循环张应力（CTS）刺激人牙周膜细胞（hPDLCs）自噬的分子机制。
从正常牙周组织中分离和培养 hPDLCs。使用四点弯曲拉伸扩展器对 hPDLCs 施加张力，模拟正畸力作用下牙周膜细胞自噬。使用 XMU-MP-1 抑制 Hippo 信号通路，探讨张力激活 hPDLC 自噬过程中 Hippo-YAP 信号通路的作用。采用实时定量聚合酶链反应检测 hPDLC 中自噬相关基因（Beclin-1、LC3 和 p62）的表达水平。Western blot 检测 hPDLC 中自噬相关蛋白（Beclin-1、LC3-Ⅱ/LC3-Ⅰ和 p62）和 Hippo-YAP 通路蛋白（活性-YAP 和 p-YAP）的表达水平。免疫荧光定位 hPDLC 中自噬相关蛋白（LC3-Ⅱ和 p62）和 Hippo-YAP 通路蛋白（活性-YAP）。
CTS 激活了 hPDLC 中的自噬，自噬相关蛋白的表达最初增加，然后减少；30min 时开始增加，3h 时达到峰值，然后减少（<0.05）。CTS 增加了活性-YAP 蛋白的表达，减少了 p-YAP 蛋白的表达（<0.05）。当 XMU-MP-1 抑制 Hippo-YAP 信号通路时（<0.05），促进活性-YAP 蛋白进入细胞核，增强自噬表达（<0.05）。
在 CTS 作用下，Hippo-YAP 信号通路参与调节 hPDLC 自噬的激活。
