Effects of diclofenac on the expression of Nrf2 and its downstream target genes in mosquito fish (Gambusia affinis)
Effects of diclofenac on the expression of Nrf2 and its downstream target genes in mosquito fish (Gambusia affinis)
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双氯芬酸对食蚊鱼Nrf2及其下游靶基因表达的影响
DOI：
10.1016/j.aquatox.2017.04.008
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发表时间：
2017-07-01
影响因子：
4.5
通讯作者：
Li, Kaibing
中科院分区：
文献类型：
--
作者：
Bao, Shuang;Nie, Xiangping;Li, Kaibing
双氯芬酸（DCF）是广泛使用的非甾体抗炎药之一。近年来，该药物已在水生环境中普遍检出。然而，其对鱼类的潜在不利影响和氧化应激毒性机制仍不清楚。在本研究中，我们首先克隆了一些关键氧化应激相关基因的关键部分序列，其中包括NF-E2相关因子2（Nrf2）、NAD（P）H：醌氧化还原酶（NQO1）、谷氨酸半胱氨酸连接酶催化亚基（GCLC）、铜锌超氧化物歧化酶（SOD2）、过氧化氢酶（CAT）、α-谷胱甘肽S-转移酶(GSTA) 和食蚊鱼 (Gambusia affinis) 中的 UDP-葡萄糖醛酸基转移酶 (UGT)。我们还推导了Nrf2的氨基酸，然后分别构建了Nrf2、NQO1和GCLC的系统发育树。结果表明，G. affinis 与其他硬骨鱼物种（例如剑尾鱼和网纹鱼）之间存在较高的同一性百分比。然后在环境相关浓度DCF暴露(0 μmol L-1、1.572 x 10(-3) mu mol L-1、1.572 x 10(-2) mu mol L-1、0.1572 μmol L-1和1.572 μmol L-1)下研究这些基因的转录表达和部分相关酶活性24 h和168 h。 Nrf2的表达在24小时被抑制，但在168小时被诱导，在DCF暴露下表现出显着的时间和/或剂量效应关系。在其下游靶基因中也发现了类似的观察结果。然而，在相同浓度的DCF暴露下相同时间下，Nrf2介导的抗氧化酶活性表现出不同的表现。 DCF暴露168 h后，基因表现出明显的诱导趋势，但酶活性和MDA含量无明显变化。总体而言，在 DCF 暴露下，蚊鱼的 mRNA 反应比酶变化更敏感。
Diclofenac (DCF) is one of widely used non-steroidal anti-inflammatory drugs. Recently, this drug has been universally detected in aquatic environment. However, its potential adverse effects and oxidative stress toxic mechanisms on fish remain unclear. In the present study, we first cloned the crucial partial sequences of some key oxidative stress related genes, which include NF-E2-related factor 2 (Nrf2), NAD(P)H: quinoneoxidor-eductase (NQO1), glutamate-cysteine ligase catalytic subunit (GCLC), Cu-Zn superoxide dismutase (SOD2), catalase (CAT), alpha-glutathione S-transferase (GSTA), and UDP-glucuronosyltransferases (UGT) in mosquito fish (Gambusia affinis). We also deduced amino acids of Nrf2 and then constructed the phylogenetic trees of Nrf2, NQO1 and GCLC, respectively. Results showed that a high identity percentage was founded between G. affinis and other bony fish species, such as Xiphophorus maculates and Poecilia reticulate. The transcriptional expression of these genes and partly related enzymes activities were then investigated under the included environmental relevant concentration DCF exposure (0 mu mol L-1, 1.572 x 10(-3) mu mol L-1, 1.572 x 10(-2) mu mol L-1, 0.1572 mu mol L-1 and 1.572 mu mol L-1) for 24 h and 168 h. The expression of Nrf2 was inhibited at 24 h but induced at 168 h, exhibiting a significant time and/or dose-effect relationship under DCF exposure. Similar observation was found in its downstream target genes. However, Nrf2-mediated antioxidant enzymes activities displayed differently under the same concentration of DCF exposure for the same time. Under DCF exposure for 168 h, the genes exhibited dramatic induction trend, but there were no significant changes in enzyme activities and MDA content. Overall, mRNA responses were more sensitive than enzyme changes in mosquito fish under DCF exposure.