Eye Institute of Xiamen University, Xiamen, Fujian, China.
Medical College of Xiamen University, Xiamen, Fujian, China.
Invest Ophthalmol Vis Sci. 2018 Jul 2;59(8):3286-3293. doi: 10.1167/iovs.18-24057.
Oxidative stress is a major pathogenesis of certain ocular surface diseases. This study investigated the association of oxidative stress and cellular autophagy in corneal epithelium.
We applied hydrogen peroxide (H2O2) to induce oxidative damage to cultured human corneal epithelial (HCE) cells and rat corneas. Cell viability, Western blotting of caspase 8, and TUNEL staining were conducted to measure the cellular injury. The production of reactive oxygen species (ROS) was measured and the levels of the following marker and key factors of ROS were also measured to detect oxidative stress: 3-nitrotyrosine, nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4), superoxide dismutase, catalase, and glutathione S-transferase P. The following key factors of autophagy were measured: LC3, beclin 1, Atg 12, and P62. We also applied an agonist of autophagy, rapamycin, in the experiment.
Cellular injury and oxidant damage were induced after exposure to H2O2 in HCE cells and rat corneas, such as increases of cell death and production of ROS; upregulation of a ROS generation enzyme, NOX4; and downregulation of degradation factors of ROS, superoxide dismutase, catalase, and glutathione S-transferase P. However, the process of cellular autophagy was suppressed by the measurements of LC3, beclin 1, Atg 12, and P62. Furthermore, application of rapamycin antagonized the cellular and oxidant injury induced by H2O2 but increased the level of autophagy in HCE cells.
The oxidative stress of corneal epithelium is associated with the inhibition of cellular autophagy.
氧化应激是某些眼表疾病的主要发病机制。本研究探讨了角膜上皮细胞的氧化应激与细胞自噬之间的关系。
我们应用过氧化氢（H2O2）诱导培养的人角膜上皮（HCE）细胞和大鼠角膜发生氧化损伤。通过细胞活力测定、半胱天冬酶 8 的 Western blot 分析和 TUNEL 染色来评估细胞损伤。检测活性氧（ROS）的产生，并检测以下 ROS 标志物和关键因子的水平来检测氧化应激：3-硝基酪氨酸、烟酰胺腺嘌呤二核苷酸磷酸氧化酶 4（NOX4）、超氧化物歧化酶、过氧化氢酶和谷胱甘肽 S-转移酶 P。还测量了自噬的以下关键因素：LC3、beclin 1、Atg 12 和 P62。我们还在实验中应用了自噬激动剂雷帕霉素。
H2O2 处理 HCE 细胞和大鼠角膜后，细胞损伤和氧化剂损伤被诱导，例如细胞死亡增加和 ROS 产生增加；ROS 生成酶 NOX4 上调；ROS 降解因子超氧化物歧化酶、过氧化氢酶和谷胱甘肽 S-转移酶 P 下调。然而，LC3、beclin 1、Atg 12 和 P62 的测量表明细胞自噬过程受到抑制。此外，雷帕霉素的应用拮抗了 H2O2 诱导的细胞和氧化剂损伤，但增加了 HCE 细胞的自噬水平。
角膜上皮的氧化应激与细胞自噬的抑制有关。
