Celastrol Protects RPE Cells from Oxidative Stress-Induced Cell Death via Activation of Nrf2 Signaling Pathway.
Celastrol Protects RPE Cells from Oxidative Stress-Induced Cell Death via Activation of Nrf2 Signaling Pathway.
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DOI：
10.2174/1566524019666190424131704
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发表时间：
2019
影响因子：
2.5
通讯作者：
Yeqi Zhou;Linbin Zhou;Kewen Zhou;Jingyue Zhang;F. Shang;Xinyu Zhang
中科院分区：
文献类型：
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作者：
Yeqi Zhou;Linbin Zhou;Kewen Zhou;Jingyue Zhang;F. Shang;Xinyu Zhang
目的视网膜色素上皮（RPE）细胞的氧化应激和炎症与年龄相关性黄斑变性（AMD）的发病机制密切相关。 Celastrol 是从雷公藤 (Tripterygium wilfordii) 根中分离出来的天然化合物。雷公藤红素已被证明在多种疾病模型中具有有效的抗炎和抗肿瘤作用。本研究的目的是测试雷公藤红素对 RPE 细胞的抗氧化作用并研究其潜在机制。方法 ARPE-19 细胞单独用过氧化氢 (H2O2) 和甲萘醌或与雷公藤红素联合处理。分别通过CCK-8和TUNEL法检测细胞活力和凋亡。通过Western blotting检测Nrf2及其靶基因GCLM和HO-1的表达。 Nrf2 的敲低是通过用编码 Nrf2 的 shRNA 的慢病毒转染 ARPE-19 细胞来完成的。通过实时定量PCR和Western blotting测定敲低效率。结果 用雷公藤红素处理 ARPE-19 细胞可显着减弱 H2O2 和甲萘醌的毒性作用。雷公藤红素处理增强了转录因子 Nrf2 及其靶标 GCLM 和 HO-1 的表达。 shRNA 敲低 Nrf2 表达部分消除了雷公藤红素的保护作用。 L-丁硫氨酸-S,R-亚砜亚胺 (BSO) 对谷胱甘肽合成的化学抑制完全消除了雷公藤红素对 H2O2 和甲萘醌引起的损伤的保护作用。然而，ZnPPIX 对 HO-1 活性的化学抑制并没有降低雷公藤红素的保护作用。结论 本研究提供的证据表明，雷公藤红素治疗 RPE 细胞通过激活 Nrf2 信号通路和上调 GCLM 表达，对氧化损伤具有有效的保护作用。这一发现表明雷公藤红素可能被用作氧化应激相关眼部疾病（例如 AMD）的潜在治疗剂。
PURPOSE Oxidative stress to retinal pigment epithelial (RPE) cells and inflammation are closely related to the pathogenesis of age-related macular degeneration (AMD). Celastrol is a natural compound isolated from the root of Tripterygium wilfordii. Celastrol has been shown to have potent anti-inflammatory and anti-tumor effects in multiple disease models. The objective of this study was to test the anti-oxidative effects of celastrol in RPE cells and to investigate the underlying mechanisms. METHODS ARPE-19 cells were treated with hydrogen peroxide (H2O2) and menadione alone or in combination with celastrol. Cell viability and apoptosis were examined by CCK-8 and TUNEL assay, respectively. The expression of Nrf2 and its target genes, such as GCLM and HO-1 were determined by Western blotting. The knockdown of Nrf2 was done by transfecting ARPE-19 cells with lentivirus encoding shRNA against Nrf2. The knockdown efficiency was determined by real-time quantitative PCR and Western blotting. RESULTS Treatment of ARPE-19 cells with celastrol significantly attenuated the toxic effects of both H2O2 and menadione. Treatment with celastrol enhanced the expression of transcription factor Nrf2 and its targets, GCLM and HO-1. Knockdown of Nrf2 expression by shRNA partially abolished the protective effects of celastrol. Chemical inhibition of glutathione synthesis by L-buthionine-S,R-sulfoximine (BSO) completely abolished the protective effects of celastrol against H2O2 and menadione-induced damage. However, chemical inhibition of HO-1 activity by ZnPPIX did not reduce the protective effects of celastrol. CONCLUSION This study provides evidence that treatment of RPE cells with celastrol shows potent protective effects against oxidative insults via activation of Nrf2 signaling pathway and upregulation of GCLM expression. This finding suggests that celastrol might be used as a potential therapeutic agent for oxidative stress-related eyes diseases, such as AMD.