Department of Pharmacology, College of Medicine, Chang Gung University, 259 Wen-Hwa 1st Road Kwei-San, Tao-Yuan, Taiwan.
Cell Commun Signal. 2013 Jan 23;11(1):8. doi: 10.1186/1478-811X-11-8.
Endothelin-1 (ET-1) is a proinflammatory mediator and elevated in the regions of several brain injury and inflammatory diseases. The deleterious effects of ET-1 on endothelial cells may aggravate brain inflammation mediated through the regulation of cyclooxygenase-2 (COX-2)/prostaglandin E2 (PGE2) system in various cell types. However, the signaling mechanisms underlying ET-1-induced COX-2 expression in brain microvascular endothelial cells remain unclear. Herein we investigated the effects of ET-1 in COX-2 regulation in mouse brain microvascular endothelial (bEnd.3) cells.
The data obtained with Western blotting, RT-PCR, and immunofluorescent staining analyses showed that ET-1-induced COX-2 expression was mediated through an ETB-dependent transcriptional activation. Engagement of Gi- and Gq-protein-coupled ETB receptors by ET-1 led to phosphorylation of ERK1/2, p38 MAPK, and JNK1/2 and then activated transcription factor NF-κB. Moreover, the data of chromatin immunoprecipitation (ChIP) and promoter reporter assay demonstrated that the activated NF-κB was translocated into nucleus and bound to its corresponding binding sites in COX-2 promoter, thereby turning on COX-2 gene transcription. Finally, up-regulation of COX-2 by ET-1 promoted PGE2 release in these cells.
These results suggested that in mouse bEnd.3 cells, activation of NF-κB by ETB-dependent MAPK cascades is essential for ET-1-induced up-regulation of COX-2/PGE2 system. Understanding the mechanisms of COX-2 expression and PGE2 release regulated by ET-1/ETB system on brain microvascular endothelial cells may provide rationally therapeutic interventions for brain injury or inflammatory diseases.
内皮素-1（ET-1）是一种促炎介质，在几种脑损伤和炎症性疾病的区域中升高。ET-1 对内皮细胞的有害影响可能通过调节环氧化酶-2（COX-2）/前列腺素 E2（PGE2）系统在各种细胞类型中加剧脑炎症。然而，ET-1 诱导脑微血管内皮细胞中 COX-2 表达的信号机制尚不清楚。在此，我们研究了 ET-1 在小鼠脑微血管内皮（bEnd.3）细胞中对 COX-2 调节的影响。
通过 Western blot、RT-PCR 和免疫荧光染色分析获得的数据表明，ET-1 诱导的 COX-2 表达是通过 ETB 依赖性转录激活介导的。ET-1 与 Gi 和 Gq 蛋白偶联的 ETB 受体结合导致 ERK1/2、p38 MAPK 和 JNK1/2 的磷酸化，然后激活转录因子 NF-κB。此外，染色质免疫沉淀（ChIP）和启动子报告基因分析的数据表明，激活的 NF-κB 易位到细胞核并与 COX-2 启动子中的相应结合位点结合，从而开启 COX-2 基因转录。最后，ET-1 上调 COX-2 促进这些细胞中 PGE2 的释放。
这些结果表明，在小鼠 bEnd.3 细胞中，ETB 依赖性 MAPK 级联激活 NF-κB 对于 ET-1 诱导的 COX-2/PGE2 系统上调是必要的。了解 ET-1/ETB 系统调节 COX-2 表达和 PGE2 释放的机制可能为脑损伤或炎症性疾病提供合理的治疗干预。
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