State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, People's Republic of China.
J Ocul Pharmacol Ther. 2019 Mar;35(2):116-123. doi: 10.1089/jop.2018.0092. Epub 2018 Dec 31.
Celastrol is a triterpenoid quinine methide that exerts important biological effects on a variety of disease models. In this study, we aim to assess the ability of celastrol to inhibit lipopolysaccharide (LPS)-induced inflammation in retinal pigment epithelial (RPE) cells.
Primary cultures of human RPE (HRPE) cells and ARPE-19 cell lines were treated with celastrol alone or in combination with LPS. The cytotoxic effect of celastrol on RPE cells was determined by the CCK-8 assay. Protein and mRNA levels of inflammatory cytokines, including IL-6, IL-8, and MCP-1, were detected by flow cytometry or by real-time fluorescent quantitative PCR, respectively. The levels of phosphorylated intermediates in the NF-κB signaling pathway (such as IκBα/β and p65) and MAPK signaling pathway (p38MAPK, SAPK/JNK, and p42/p44MAPK) were detected by western blotting.
Celastrol significantly inhibited LPS-induced expression of protein and mRNA expression levels encoding the proinflammatory cytokines, IL-6, IL-8, and MCP-1, in both HRPE and ARPE-19 cells. Cell viability and apoptosis assays revealed that celastrol had no apparent cytotoxic effect and it inhibited apoptosis of RPE cells at concentrations of less than 1 μM. Mechanistically, RPE cells that were pretreated with celastrol exhibited a substantial decrease in phosphorylation of the NF-κB pathway regulators, IKKα/β and IκBα, and subsequently inactivated P65, suggesting that celastrol ameliorates LPS-induced inflammation by suppressing the NF-κB signaling pathway.
Our results provide evidence that celastrol is a potent anti-inflammatory agent in RPE cells and it may have potential applications in prevention and treatment of age-related macular degeneration.
雷公藤红素是一种三萜类奎宁甲基化物，对多种疾病模型具有重要的生物学作用。本研究旨在评估雷公藤红素抑制脂多糖（LPS）诱导的视网膜色素上皮（RPE）细胞炎症的能力。
用人 RPE（HRPE）细胞和 ARPE-19 细胞系进行原代培养，用雷公藤红素单独或与 LPS 联合处理。用 CCK-8 法测定雷公藤红素对 RPE 细胞的细胞毒性作用。用流式细胞术或实时荧光定量 PCR 分别检测炎症细胞因子（IL-6、IL-8 和 MCP-1）的蛋白和 mRNA 水平。用 Western blot 检测 NF-κB 信号通路（如 IκBα/β 和 p65）和 MAPK 信号通路（p38MAPK、SAPK/JNK 和 p42/p44MAPK）中磷酸化中间产物的水平。
雷公藤红素显著抑制 LPS 诱导的 HRPE 和 ARPE-19 细胞中促炎细胞因子（IL-6、IL-8 和 MCP-1）的蛋白和 mRNA 表达水平。细胞活力和凋亡试验表明，雷公藤红素在低于 1μM 的浓度下对 RPE 细胞没有明显的细胞毒性作用，并且抑制 RPE 细胞凋亡。机制上，用雷公藤红素预处理的 RPE 细胞中 NF-κB 途径调节剂 IKKα/β 和 IκBα 的磷酸化显著减少，随后 P65 失活，表明雷公藤红素通过抑制 NF-κB 信号通路改善 LPS 诱导的炎症。
我们的结果提供了证据，表明雷公藤红素是 RPE 细胞中的一种有效的抗炎剂，它可能在预防和治疗年龄相关性黄斑变性方面具有应用潜力。
