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首页> 外文期刊>Aquatic Toxicology >Isolation and characterization of two cytochrome P450 aromatase forms in killifish (Fundulus heteroclitus): differential expression in fish from polluted and unpolluted environments
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Isolation and characterization of two cytochrome P450 aromatase forms in killifish (Fundulus heteroclitus): differential expression in fish from polluted and unpolluted environments

机译：鱼类中两种不同形态的细胞色素P450芳香酶的分离和鉴定：污染和未污染环境中鱼类中的差异表达

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Populations of killifish (Fundulus heteroclitus) persist in many different highly polluted environment indicative of adaptation or tolerance. In this study, we sought to determine whether long term, multigenerational exposures to environmental contaminants has affected reproductively relevant genes and biological processes. A homology cloning strategy was used to isolate the killifish cytochrome P450 aromatase (P450arom, estrogen synthetase) cDNAs. Consistent with previous fish studies, killifish were found to have two P450arom forms, which segregated into A- and B-gene clades and were differentially expressed in brain (B A) and ovary (A B). Comparison of killifish from highly polluted (New Bedford Harbor, NBH) and unpolluted (Scorton Creek, SC) environments revealed no site-related differences in P450arom coding sequences or in overall tissue distribution patterns. As measured by real-time quantitative PCR (QPCR) analysis, however, P450arormB (a known marker of estrogen effect) was approximately two-fold higher in the brain of NBH than of SC fish, a difference seen in reproductively active and inactive males and females. Providing further evidence of exposure to estrogen-like pollutants or metabolites in NBH, vitellogenin (vtg) mRNA and protein were elevated in seasonally active and inactive males, and in reproductively inactive females, when compared to SC fish. By contrast, during the period of reproductive activity, NBH females had a lower gonadosomatic index, lower plasma estrogen, a decreased hepatosomatic index, and reduced vtg expression as compared to SC females, indicating that the female hypothalamic-pituitary-gonadal (HPG)-liver axis is impaired in the polluted environment. As measured by a decrease in plasma androgen (but not GSI), the male HPG axis was impaired in reproductively active NBH versus SC fish. In agreement with reports that NBH killifish are resistant to dioxin-like chemicals (DLC) that activate arylhydrocarbon receptor (AhR) signaling, ovarian P450aromA (a marker of dioxin-like effect in zebrafish embryos) did not differ in SC and NBH fish. In conclusion, the killifish population at the NBH Superfund site maintains a level of reproductive competence in the face of evidence of exposure to estrogen-like pollutants and endocrine disruption.
机译：在许多不同的高度污染的环境中，通常会出现许多奇异的鱼类（Fundulus heteroclitus），这表明它们具有适应性或耐受性。在这项研究中，我们试图确定长期，多代接触环境污染物是否影响了生殖相关基因和生物学过程。同源克隆策略用于分离比目鱼细胞色素P450芳香酶（P450arom，雌激素合成酶）cDNA。与先前的鱼类研究一致，发现kill鱼具有两种P450arom形式，它们分别分为A基因和B基因进化枝，并在脑（B A）和卵巢（A B）中差异表达。比较高污染环境（新贝德福德港，新罕布什尔州）和未污染环境（斯科特顿溪，南卡罗来纳州）环境中的比目鱼，发现在​​P450arom编码序列或总体组织分布方式上没有与地点相关的差异。但是，通过实时定量PCR（QPCR）分析测得，NBH的大脑中P450arormB（雌激素作用的已知标志物）大约比SC鱼高两倍，这在生殖活跃和不活跃的雄性和女性。与SC鱼相比，季节性活动和不活动的雄性以及生殖不活动的雌性中卵黄蛋白原（vtg）mRNA和蛋白质的含量增加，提供了进一步暴露于NBH中类似雌激素污染物或代谢产物的证据。相比之下，与SC雌性相比，NBH雌性在生殖活动期间具有较低的性腺体指数，较低的血浆雌激素，降低的肝体指数和降低的vtg表达，表明雌性下丘脑-垂体-性腺（HPG）-在污染的环境中肝轴受损。通过血浆雄激素（而非GSI）的减少来衡量，与SC鱼相比，雄性HPG轴在生殖活动性NBH中受损。与NBH致死鱼对激活芳烃受体（AhR）信号的二恶英类化学物质（DLC）有抵抗力的报道一致，在SC和NBH鱼中，卵巢P450aromA（斑马鱼胚胎中二恶英样作用的标志）没有差异。总之，面对暴露于类似雌激素的污染物和内分泌干扰的证据，NBH超级基金所在地的kill鱼种群保持了一定水平的繁殖能力。

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