Tianjin Key Laboratory of Retinal Functions and Diseases, Tianjin Branch of National Clinical Research Center for Ocular Disease, Eye Institute and School of Optometry, Tianjin Medical University Eye Hospital, Tianjin 300384, China.
Department of Neural and Behavioral Sciences, Penn State College of Medicine, Hershey, PA 0850, USA.
Int J Mol Sci. 2023 Feb 10;24(4):3565. doi: 10.3390/ijms24043565.
Caffeic acid phenylethyl ester (CAPE) is an antioxidative agent originally derived from propolis. Oxidative stress is a significant pathogenic factor in most retinal diseases. Our previous study revealed that CAPE suppresses mitochondrial ROS production in ARPE-19 cells by regulating UCP2. The present study explores the ability of CAPE to provide longer-term protection to RPE cells and the underlying signal pathways involved. ARPE-19 cells were given CAPE pretreatment followed by t-BHP stimulation. We used in situ live cell staining with CellROX and MitoSOX to measure ROS accumulation; Annexin V-FITC/PI assay to evaluate cell apoptosis; ZO-1 immunostaining to observe tight junction integrity in the cells; RNA-seq to analyze changes in gene expression; q-PCR to validate the RNA-seq data; and Western Blot to examine MAPK signal pathway activation. CAPE significantly reduced both cellular and mitochondria ROS overproduction, restored the loss of ZO-1 expression, and inhibited apoptosis induced by t-BHP stimulation. We also demonstrated that CAPE reverses the overexpression of immediate early genes (IEGs) and activation of the p38-MAPK/CREB signal pathway. Either genetic or chemical deletion of UCP2 largely abolished the protective effects of CAPE. CAPE restrained ROS generation and preserved the tight junction structure of ARPE-19 cells against oxidative stress-induced apoptosis. These effects were mediated via UCP2 regulation of p38/MAPK-CREB-IEGs pathway.
咖啡酸苯乙酯 (CAPE) 是一种抗氧化剂，最初来源于蜂胶。氧化应激是大多数视网膜疾病的重要致病因素。我们之前的研究表明，CAPE 通过调节 UCP2 抑制 ARPE-19 细胞中线粒体 ROS 的产生。本研究探讨了 CAPE 对 RPE 细胞提供更长期保护的能力以及涉及的潜在信号通路。用 CAPE 预处理 ARPE-19 细胞，然后用 t-BHP 刺激。我们使用 CellROX 和 MitoSOX 进行原位活细胞染色来测量 ROS 积累；用 Annexin V-FITC/PI 测定法评估细胞凋亡；用 ZO-1 免疫染色观察细胞中紧密连接的完整性；用 RNA-seq 分析基因表达的变化；用 q-PCR 验证 RNA-seq 数据；用 Western Blot 检测 MAPK 信号通路的激活。CAPE 显著减少细胞和线粒体 ROS 的过度产生，恢复因 t-BHP 刺激而丧失的 ZO-1 表达，并抑制凋亡。我们还表明，CAPE 逆转了即刻早期基因 (IEGs) 的过度表达和 p38-MAPK/CREB 信号通路的激活。UCP2 的基因或化学缺失在很大程度上消除了 CAPE 的保护作用。CAPE 抑制 ROS 的产生并保持 ARPE-19 细胞的紧密连接结构，防止氧化应激诱导的细胞凋亡。这些作用是通过 UCP2 调节 p38/MAPK-CREB-IEGs 通路介导的。
