Identification of 5-hydroxymethylfurfural in cigarette smoke extract as a new substrate metabolically activated by human cytochrome P450 2A13
Identification of 5-hydroxymethylfurfural in cigarette smoke extract as a new substrate metabolically activated by human cytochrome P450 2A13
复制标题
香烟烟雾提取物中 5-羟甲基糠醛的鉴定作为人细胞色素 P450 2A13 代谢激活的新底物
DOI：
10.1016/j.taap.2018.09.031
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发表时间：
2018-11-15
影响因子：
3.8
通讯作者：
Wang, Shou-Lin
中科院分区：
文献类型：
--
作者：
Ji, Minghui;Zhang, Zhan;Wang, Shou-Lin
细胞色素P450 2A 13（CYP 2A 13）是一种主要在人体呼吸系统表达的肝外酶，据报道可介导香烟烟雾中烟草特异性N-亚硝胺（TSNA）的代谢。本研究旨在鉴定香烟烟雾中CYP 2A 13的其他新底物及其相应的呼吸毒性。在稳定表达CYP 2A 13（B-2A 13）的BEAS-2 B细胞中通过HPLC、GC-MS/MS、NMR和细胞毒性试验分离后，在香烟烟雾提取物（CSE）的4-5 min切片中筛选并鉴定了5-羟甲基糠醛（5-HMF）。体外代谢结果表明，CYP 2A 13介导5-HMF的快速清除，并形成代谢产物5-HMF酸（5-HMFA）。CSE-5-HMF（CSE-5-HMF）在B-2A 13和B-2A 5细胞中显示出与标准5-HMF相似的细胞毒性，该毒性可被8-甲氧基卟啉（8-MOP）（一种β-内酰胺酶抑制剂）抑制。小鼠CYP 2A 5是与CYP 2A 13同源的一种酶，在香烟烟雾中与CYP 2A 13共享许多底物。因此，生成CYP 2A 5（-/-）小鼠以探索CYP 2A 5在5-HMF生物活化中的作用。与CYP 2A 5（-/-）小鼠相比，WT小鼠表现出严重的肺和鼻嗅粘膜组织学损伤，以及支气管肺泡灌洗液中炎性细胞增加和TNF-α和IL-6水平升高。WT小鼠鼻微粒体5-HMFA的形成速度快于CYP 2A 5（-/-）小鼠，且能被8-MOP抑制。本研究首次发现5-HMF是香烟烟雾中人CYP 2A 13的一种新的毒性底物，可能在香烟烟雾诱导的呼吸系统损伤中发挥潜在作用。
Cytochrome P450 2A13 (CYP2A13) is an extrahepatic enzyme mainly expressed in the human respiratory system and is reported to mediate tobacco-specific N-nitrosamines (TSNA) metabolism in cigarette smoke. This study aimed to identify other new substrates of CYP2A13 in cigarette smoke and their corresponding respiratory toxicity. Following separation by HPLC, GC-MS/MS, NMR and cytotoxicity assays in BEAS-2B cells stably expressing CYP2A13 (B-2A13), 5-Hydroxymethylfurfural (5-HMF) was screened and identified in the 4-5 min section of cigarette smoke extract (CSE). In vitro metabolism results showed that CYP2A13 mediated the fast clearance of 5-HMF and formed the metabolite 5-HMF acid (5-HMFA). CSE 5-HMF (CSE-5-HMF) showed cytotoxicity similar to that of standard 5-HMF in B-2A13 and B-2A5 cells, which was inhibited by 8-methoxypsoralen (8-MOP), a CYP enzyme inhibitor. Mouse CYP2A5, a homologous CYP enzyme to CYP2A13, shares many substrates with CYP2A13 in cigarette smoke. Thus, CYP2A5(-/-) mice were generated to explore the role of CYP2A5 in 5-HMF bioactivation. Compared with CYP2A5(-/-) mice, WT mice showed serious histological lung and nasal olfactory mucosa damage, as well as increased inflammatory cells and elevated TNF-alpha and IL-6 levels in bronchoalveolar lavage fluid. Besides, nasal microsomes undertook fast 5-HMFA formation in WT mice than that in CYP2A5(-/-) mice, which could be inhibited by 8-MOP. This study is the first to identify 5-HMF as a new toxic substrate of human CYP2A13 in cigarette smoke, it may play a potential role in cigarette smoke-induced respiratory injuries.