蒋宇霞2,
杨雷2,
应光国1,
史文俊1
1. 华南师范大学环境研究院, 广东省化学污染与环境安全重点实验室, 华南师范大学环境理论化学教育部重点实验室, 广州 510006;
2. 中国科学院广州地球化学研究所有机地球化学国家重点实验室, 广州 510640
作者简介: 马栋栋(1991-),男,硕士研究生,研究方向为环境毒理学,E-mail:madong9110@163.com.
基金项目: 国家自然科学基金资助项目(41807480)；中国博士后科学基金面上项目(2018M643116)；华南师范大学研究生创新计划资助项目(2018LKXM018)中图分类号: X171.5
Effect of Androstadienedione and Androstenedione on Transcription of Genes in Circadian Rhythm and Hypothalamic-pituitary-gonadal Axis in Zebrafish Embryos
Ma Dongdong1,Jiang Yuxia2,
Yang Lei2,
Ying Guangguo1,
Shi Wenjun1
1. Environmental Research Institute, Guangdong Provincial Key Laboratory of Chemical Pollution and Environmental Safety & MOE Key Laboratory of Environmental Theoretical Chemistry, South China Normal University, Guangzhou 510006, China;
2. State Key Laboratory of Organic Geochemistry, Guangzhou Institute of Geochemistry, Chinese Academy of Sciences, Guangzhou 510640, China
CLC number: X171.5
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摘要:雄激素1,4-雄烯二酮(androstadienedione, ADD)和雄烯二酮(androstenedione, AED)主要用于人类和牲畜疾病的预防和治疗。近年来，ADD和AED的大量使用导致其在河流中广泛检出，甚至在多种鱼类体内亦有检出，且浓度较高。ADD和AED已被证实对鱼类具有生殖毒性和发育毒性，但ADD和AED在转录水平上对鱼类的影响鲜有报道。为探究ADD和AED分子水平毒性，本研究考察了斑马鱼胚胎暴露于ADD(4.48、30.0和231 ng L?1)和AED(3.64、21.7和230 ng L?1)144 h后，对其昼夜节律和下丘脑-垂体-性腺轴(hypothalamic-pituitary-gonadal axis, HPG axis)通路中基因转录表达的影响。结果表明，所有浓度的ADD都显著上调了昼夜节律通路中生物钟基因(per1b)、核受体亚族1的D群基因(nr1d2b)、隐花色素基因(cry5)和si:ch211-132b12.7的转录水平，30.0和231 ng L?1的ADD下调了时钟节律调节因子基因(clocka)和芳香烃受体核转录蛋白样基因(arntl2)的转录水平。3.64 ng L?1 AED显著增强了per1b和nr1d2b的转录表达。此外在HPG轴中，30.0 ng L?1 ADD显著降低了促黄体生成素V亚基基因(lhb)的转录表达水平，而3.64 ng L?1 AED显著上调了lhb的转录表达水平。值得注意的是，4.48 ng L?1 ADD和3.64 ng L?1 AED均显著降低了细胞色素P450的11亚族基因(cyp11b)的转录表达水平。上述研究表明，ADD和AED对昼夜节律和HPG轴中相关基因的转录表达有显著性影响，对斑马鱼具有潜在的内分泌干扰风险。
关键词: 1,4-雄烯二酮/
雄烯二酮/
斑马鱼胚胎/
内分泌干扰/
昼夜节律/
下丘脑-垂体-性腺轴
Abstract:Androgen 1,4-androstenedione (ADD) and androstenedione (AED) are mainly used for the prevention and treatment of human and livestock diseases. ADD and AED have been frequently detected in the surface waters at concentrations in the range of ng L?1, and rarely up to μg L?1. In addition, high concentrations of ADD and AED were also detected in a variety of fish. So far, several studies have shown that AED can cause masculinization in female fathead minnow and in mosquitofish and even lead to sex reversal in zebrafish. ADD can reduce the fin length of fish, decrease the egg production and adversely affect the gonadal development. These studies indicate that ADD and AED have reproductive toxicity at physiological level. However, little study focused on the effect of ADD and AED on transcription of genes in the related systems of fish, such as circadian rhythm and hypothalamic-pituitary-gonadal (HPG) axis. Therefore, the aim of the present study was to investigate the effect of ADD and AED on transcription of genes involved in circadian rhythm and HPG axis in zebrafish embryos by qPCR analysis. The zebrafish embryos were exposed to three concentrations of ADD (4.48, 30.0 and 231 ng L?1) and AED (3.64, 21.7 and 230 ng L?1) for 144 h post fertilization (hpf), respectively. The data of qPCR analysis showed that exposure to all three concentrations of ADD significantly increased the transcription of period circadian clock 1b (per1b), nuclear receptor subfamily 1, group D, member 2b (nr1d2b), cryptochrome circadian regulator 5 (cry5) and si:ch211-132b12.7 in the circadian rhythm network. The fold change of per1b, nr1d2b and si:ch211-132b12.7 even reached to 3.19, 2.72 and 3.63 at 4.48 ng L?1, respectively. Exposure to 30 ng L?1 and 230 ng L?1 of ADD suppressed the transcription of clocka (fold change, ~1.29) and aryl hydrocarbon receptor nuclear translocator-like 2 (arntl2) (fold change, ~1.29). In addition, exposure to 3.64 ng L?1 of AED enhanced the transcription of per1b and nr1d2b. For the effect of ADD and AED on HPG axis, exposure to 30 ng L?1 of ADD significantly decreased the transcription of luteinizing hormone, beta polypeptide (lhb), while exposure to 3.64 ng L?1 of AED increased the transcription of lhb. It is noted that exposure to both 4.48 ng L?1 of ADD and 3.64 ng L?1 of AED significantly down-regulated the transcription of cytochrome P450, family 11, subfamily C, polypeptide 1 (cyp11b), which participates in androgen synthesis. Taken together, the results indicated that ADD and AED could affect the transcription of genes belonging to the HPG axis and circadian rhythm after 144 hpf exposure, and suggested that ADD and AED might have potential endocrine disruption effect in fish.
Key words:1,4-androstenedione/
androstenedione/
zebrafish embryo/
endocrine disruption/
circadian rhythm/
hypothalamic-pituitary-gonadal (HPG) axis.
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