Department of Ophthalmology, The First Affiliated Hospital of Kunming Medical University, Kunming, 650032, China; Department of Ophthalmology, The Affiliated Hospital of Yunnan University, Kunming, 650021, China.
Department of Pediatric Ophthalmology, The Affiliated Hospital of Yunnan University, Kunming, 650021, China.
Exp Eye Res. 2023 Jul;232:109496. doi: 10.1016/j.exer.2023.109496. Epub 2023 May 31.
Diabetes retinopathy (DR) is one of the most common microvascular complications of diabetes. Retinal pigment epithelial (RPE) cells exposed to a high glucose environment experience a series of functional damages, which is an important factor in promoting the progression of DR. Acteoside (ACT) has strong antioxidant and anti-apoptotic properties, but the mechanism of ACT in DR is not completely clear. Therefore, the purpose of the present study was to explore whether ACT inhibits the damage to RPE cells in a high glucose environment through antioxidative effects to alleviate the DR process. The DR in vitro cell model was constructed by treating RPE cells with high glucose, and the DR in vivo animal model was constructed by injecting streptozotocin (STZ) into the peritoneal cavity of mice to induce diabetes. The proliferation and apoptosis of RPE cells were detected by CCK-8 and flow cytometry assays, respectively. The expression changes in Nrf2, Keap1, NQO1 and HO-1 were evaluated by qRT‒PCR, Western blot and immunohistochemistry analyses. The MDA, SOD, GSH-Px and T-AOC contents were detected by kits. The changes in ROS and nuclear translocation of Nrf2 were observed by immunofluorescence assays. HE staining was used to measure the thickness of the outer nuclear layer (ONL) of the retina, and TUNEL staining was used to detect the number of apoptotic cells in the retinas of mice. In the present study, ACT effectively ameliorated outer retina damage in diabetic mice. In high glucose (HG)-induced RPE cells, ACT treatment had the following effects: improved proliferation, decreased apoptosis, inhibited Keap1 expression, promoted the nuclear translocation and expression of Nrf2, upregulated NQO1 and HO-1 (the target genes of Nrf2) expression, decreased ROS concentration, and increased the levels of the SOD, GSH-Px and T-AOC antioxidant indicators. However, knockdown of Nrf2 reversed the above phenomena, which indicated that the protective function of ACT in HG-induced RPE cells are closely related to Nrf2. In summary, the present study demonstrated that HG-induced oxidative stress injury is inhibited by ACT in RPE cells and the outer retina through the Keap1/Nrf2/ARE pathway.
糖尿病性视网膜病变 (DR) 是糖尿病最常见的微血管并发症之一。暴露于高糖环境中的视网膜色素上皮 (RPE) 细胞经历一系列功能损伤，这是促进 DR 进展的重要因素。獐牙菜苦苷 (ACT) 具有很强的抗氧化和抗凋亡作用，但 ACT 在 DR 中的作用机制尚不完全清楚。因此，本研究旨在探讨 ACT 是否通过抗氧化作用抑制高糖环境对 RPE 细胞的损伤，从而缓解 DR 进程。通过用高糖处理 RPE 细胞构建 DR 体外细胞模型，通过腹腔注射链脲佐菌素 (STZ) 构建 DR 体内动物模型诱导糖尿病。通过 CCK-8 和流式细胞术分别检测 RPE 细胞的增殖和凋亡。通过 qRT-PCR、Western blot 和免疫组化分析评估 Nrf2、Keap1、NQO1 和 HO-1 的表达变化。通过试剂盒检测 MDA、SOD、GSH-Px 和 T-AOC 含量。通过免疫荧光法观察 ROS 和 Nrf2 核转位的变化。HE 染色测量视网膜外核层 (ONL) 的厚度，TUNEL 染色检测小鼠视网膜中凋亡细胞的数量。在本研究中，ACT 有效改善了糖尿病小鼠的外视网膜损伤。在高糖 (HG) 诱导的 RPE 细胞中，ACT 处理具有以下作用：改善增殖，减少凋亡，抑制 Keap1 表达，促进 Nrf2 的核转位和表达，上调 NQO1 和 HO-1（Nrf2 的靶基因）表达，降低 ROS 浓度，并提高 SOD、GSH-Px 和 T-AOC 抗氧化指标的水平。然而，Nrf2 的敲低逆转了上述现象，这表明 ACT 在 HG 诱导的 RPE 细胞中的保护作用与 Nrf2 密切相关。综上所述，本研究表明，ACT 通过 Keap1/Nrf2/ARE 通路抑制 HG 诱导的 RPE 细胞和外视网膜的氧化应激损伤。
