Proteomic analysis of oxidative stress response in human umbilical vein endothelial cells (HUVECs): role of heme oxygenase 1 (HMOX1) in hypoxanthine-induced oxidative stress in HUVECs
Proteomic analysis of oxidative stress response in human umbilical vein endothelial cells (HUVECs): role of heme oxygenase 1 (HMOX1) in hypoxanthine-induced oxidative stress in HUVECs
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DOI：
10.21037/tau.2020.03.11
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发表时间：
2020-03
影响因子：
2
通讯作者：
Pei Zhu;T. Qi;Zhan-sen Huang;Hao Li;Bo Wang;Jianchao Feng;Shuaipeng Ma;Hengjun Xiao;Yu-Xin Tang-Y
中科院分区：
文献类型：
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作者：
Pei Zhu;T. Qi;Zhan-sen Huang;Hao Li;Bo Wang;Jianchao Feng;Shuaipeng Ma;Hengjun Xiao;Yu-Xin Tang-Y
勃起功能障碍（艾德）是糖尿病的常见并发症，影响高达75%的糖尿病男性。虽然糖尿病艾德的病因是多因素的，但内皮功能障碍被认为是其病理生理学的支柱。内皮功能障碍是由高葡萄糖水平和增加的氧化应激对构成血管内皮的内皮细胞的有害影响引起的。目的：研究高糖诱导的氧化应激对细胞蛋白质组的影响，并探讨血红素加氧酶1（heme oxygenase 1，HMOX 1）在其中的作用。方法：采用iTRAQ技术结合液相色谱-串联质谱（LC-MS）分析次黄嘌呤诱导的人脐静脉内皮细胞（human umbilical vein endothelial cells，HUVECs）氧化应激后细胞蛋白质组的变化（LC-MS/MS）。通过Network和京都基因和基因组百科全书（KEGG）途径分析来分析差异表达蛋白（DEP）。进行了进一步的验证试验，以验证HMOX 1的作用。结果HUVECs氧化应激后，有66个DEPs表达上调，76个DEPs表达下调。在这些蛋白质中，我们通过定量逆转录PCR验证了8个失调基因，包括核仁素（NCL），X射线修复交叉互补蛋白6（XRCC 6），泛喹啉-细胞色素C还原酶结合蛋白（UQCRB），非POU结构域包含八聚体结合（NONO），血红素加氧酶1（HMOX 1），核结合蛋白1（NUCB 1），DEK和染色质靶点prmt 1（CHTOP）。此外，使用过表达和基因敲低方法，我们发现HMOX 1对HUVECs的氧化应激反应至关重要。结论HMOX 1与次黄嘌呤诱导的氧化应激反应密切相关。据我们所知，这项研究是第一次概述了HUVECs对氧化应激的反应。本研究结果将有助于详细分析艾德患者内皮功能障碍的发病机制及相关分子机制。
Background Erectile dysfunction (ED) is a well-known complication of diabetes, affecting up to 75% of diabetic men. Although the etiology of diabetic ED is multifactorial, endothelial dysfunction is considered to be a pillar of its pathophysiology. Endothelial dysfunction is caused by the harmful effects of high glucose levels and increased oxidative stress on the endothelial cells that comprise the vascular endothelium. The aim of this study was to identify the proteomic changes caused by high glucose-induced oxidative stress and explore the role of heme oxygenase 1 (HMOX1) in it. Methods The cellular proteomic response to hypoxanthine-induced oxidative stress in human umbilical vein endothelial cells (HUVECs) was analyzed by isobaric tags for relative and absolute quantitation (iTRAQ) combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Differentially expressed proteins (DEPs) were analyzed through Network and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Further validation assays was performed to validate the role of HMOX1. Results The results showed that 66 and 76 DEPs were markedly upregulated and downregulated, respectively, for HUVECs oxidative stress. Among these proteins, we verified eight dysregulated genes by quantitative reverse transcription PCR, including nucleolin (NCL), X-ray repair cross-complementing protein 6 (XRCC6), ubiquinol-cytochrome C reductase binding protein (UQCRB), non-POU domain containing octamer binding (NONO), heme oxygenase 1 (HMOX1), nucleobindin 1 (NUCB1), DEK, and chromatin target of prmt1 (CHTOP). Further, using overexpression and genetic knockdown approaches, we found that HMOX1 was critical for the oxidative stress response in HUVECs. Conclusions We found that HMOX1 was closely related to the oxidative stress response induced by hypoxanthine. To the best of our knowledge, this study is the first overview of the responses of HUVECs to oxidative stress. The findings will contribute to analyses of the detailed molecular mechanisms involved in the pathogenesis of endothelial dysfunction and related molecular mechanisms in ED patients.