toll样受体信号通路调节缺氧应激诱导的人微血管内皮细胞成纤维细胞生长因子而非血管内皮生长因子- a

Rukhsar Akhtar, Husain Tahir, E. Stewart, R. Wei, Imran Mohammed, W. Amoaku
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引用次数: 1

摘要

视网膜疾病是全世界不可逆转失明的主要原因。低氧应激下,toll样受体(TLR)信号机制(MyD88和TRIF)在人微血管内皮细胞(HMEC-1)生成促血管生成生长因子中的作用尚不清楚。HMEC-1在常氧(5% CO2, 37℃)和低氧(1% O2, 5% CO2, 94% N2)条件下孵育;在37°C)条件下分别发酵2、6、24和48小时。为了进行TLR通路分析,我们使用药物抑制剂(Pepinh-MyD88和Pepinh-TRIF)对HMEC-1进行预处理,并在常氧和缺氧条件下进行处理。采用定量聚合酶链反应(qPCR)、ELISA和Western blot方法检测血管内皮生长因子-a (VEGF-A)、成纤维细胞生长因子(FGF-2)、缺氧诱导因子1-α (HIF1-α)的基因和蛋白表达。流式细胞术检测TLR3和TLR4的表达水平。在缺氧条件下,VEGF-A和FGF-2水平呈时间依赖性升高。抑制MyD88和TRIF信号通路可降低FGF-2水平,但不能调节HMEC-1分泌VEGF-A。阻断已知的调节因子内皮素受体(ETR)对HMEC-1的VEGF-A分泌也没有影响。总的来说,这项研究为治疗致盲性视网膜疾病提供了靶向TLR信号通路的概念验证。
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Toll-Like Receptor Signalling Pathways Regulate Hypoxic Stress Induced Fibroblast Growth Factor but Not Vascular Endothelial Growth Factor-A in Human Microvascular Endothelial Cells
Retinal diseases are the leading causes of irreversible blindness worldwide. The role of toll-like receptor (TLR) signalling mechanisms (MyD88 and TRIF) in the production of pro-angiogenic growth factors from human microvascular endothelial cells (HMEC-1) under hypoxic stress remains unexplored. HMEC-1 was incubated under normoxic (5% CO2 at 37 °C) and hypoxic (1% O2, 5% CO2, and 94% N2; at 37 °C) conditions for 2, 6, 24, and 48 h, respectively. For TLR pathway analysis, HMEC-1 was pre-treated with pharmacological inhibitors (Pepinh-MyD88 and Pepinh-TRIF) and subjected to normoxia and hypoxia conditions. Gene and protein expressions of vascular endothelial growth factor-A (VEGF-A), fibroblast growth factor (FGF-2), hypoxia inducible factor 1-alpha (HIF1-α) were performed using quantitative polymerase chain reaction (qPCR), ELISA, and Western blot methodologies. Levels of TLR3 and TLR4 were analysed by flow cytometry. Under hypoxia, levels of VEGF-A and FGF-2 were elevated in a time-dependent fashion. Inhibition of MyD88 and TRIF signalling pathways decreased FGF-2 levels but failed to modulate the secretion of VEGF-A from HMEC-1. Blocking a known regulator, endothelin receptor (ETR), also had no effect on VEGF-A secretion from HMEC-1. Overall, this study provides the proof-of-concept to target TLR signalling pathways for the management of blinding retinal diseases.
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来源期刊
Journal of International Translational Medicine
Journal of International Translational Medicine MEDICINE, RESEARCH & EXPERIMENTAL-
自引率
0.00%
发文量
317
审稿时长
8 weeks
期刊介绍: Journal of International Translational Medicine (JITM, ISSN 2227-6394), founded in 2012, is an English academic journal published by Journal of International Translational Medicine Co., Ltd and sponsored by International Fderation of Translational Medicine. JITM is an open access journal freely serving to submit, review, publish, read and download full text and quote. JITM is a quarterly publication with the first issue published in March, 2013, and all articles published in English are compiled and edited by professional graphic designers according to the international compiling and editing standard. All members of the JITM Editorial Board are the famous international specialists in the field of translational medicine who come from twenty different countries and areas such as USA, Britain, France, Germany and so on.
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