Behnisch Peter Alexander, Hosoe Kazunori, Sakai Shin-ichi
Life Science Research Laboratories, Kaneka Corporation, 1-8 Miyamae-Machi, Hyogo Takasago 676-8688, Japan.
Environ Int. 2003 Sep;29(6):861-77. doi: 10.1016/s0160-4120(03)00105-3.
Recently, several countries agreed to adopt the Stockholm convention on persistent organic pollutants (POPs). One future obligation will be to add other POPs as new evidence becomes available. In vitro cell-based bioassays offer a rapid, sensitive, and relatively inexpensive solution to screen possible POP candidates. In the present study, we investigated the aryl hydrocarbon (Ah)-receptor activity of several dioxin-like POPs by using the Micro-EROD (Ethoxy-Resorufin-O-Deethylase) and DR-CALUX (Dioxin-Responsive-Chemical Activated Luciferase gene eXpression) bioassays, which are two state-of-the-art methods. The Micro-EROD system used in our study utilizes a wild-type rat liver cell line (rat liver H4IIEC3/T cells), while the DR-CALUX bioassay consists of a genetically modified rat hepatoma H4IIE cell line that incorporates the firefly luciferase gene coupled to dioxin-responsive elements (DREs) as a reporter gene. In the case of the DR-CALUX bioassay, we used an exposure time of 24 h, whereas we used a 72-h exposure time in the Micro-EROD bioassay. The aim of this study was to compare conventional dioxin-like POPs (such as polychlorinated dibenzodioxins and -furans, PCDD/Fs and coplanar polychlorinated biphenyls, PCBs) with several other classes of possible candidates to be added to the current toxicity equivalent factor (TEF) model in the future. Therefore, this study compares in vitro CYP1A1 (Micro-EROD bioassay) and firefly luciferase induction (DR-CALUX bioassay) in several mixed polyhalogenated dibenzodioxins and -furans (PXDD/Fs; X=Br, Cl, or F), alkyl-substituted polyhalogenated dibenzodioxins and -furans (PMCDD/Fs; M=methyl), polyhalogenated biphenyls (PXBs, X=Br, Cl ), polybrominated diphenyl ethers (PBDEs), pentabromophenols (PBPs), and tetrabromobisphenol-A (TBBP-A). We also evaluate congener-specific relative potencies (REPs) and efficacies (% of TCDD(max)) and discuss the dose-response curves of these compounds, as well as the dioxin-like potency of several other Ah-receptor agonists, such as those of the polyaromatic hydrocarbons (PAHs) and polychlorinated naphthalenes (PCNs). The highest REP values were found for several PXDD/F congeners, followed by some coplanar PXBs, trichlorinated PCDD/Fs, PAHs, PBDE-126, 1-6-HxCN, and some brominated flame retardants (TBBP-A). These in vitro investigations indicate that further research is necessary to evaluate more Ah-receptor agonists for dioxin-like potency.
最近，几个国家同意采用《关于持久性有机污染物的斯德哥尔摩公约》。未来的一项义务将是随着新证据的出现增加其他持久性有机污染物。基于细胞的体外生物测定法为筛选可能的持久性有机污染物候选物提供了一种快速、灵敏且相对廉价的解决方案。在本研究中，我们使用微EROD（乙氧基-间苯二酚-O-脱乙基酶）和DR-CALUX（二噁英反应性化学激活荧光素酶基因表达）生物测定法，这两种都是最先进的方法，研究了几种二噁英类持久性有机污染物的芳烃（Ah）受体活性。我们研究中使用的微EROD系统利用野生型大鼠肝细胞系（大鼠肝脏H4IIEC3/T细胞），而DR-CALUX生物测定法由一个基因改造的大鼠肝癌H4IIE细胞系组成，该细胞系整合了与二噁英反应元件（DREs）偶联的萤火虫荧光素酶基因作为报告基因。对于DR-CALUX生物测定法，我们使用24小时的暴露时间，而在微EROD生物测定法中我们使用72小时的暴露时间。本研究的目的是将传统的二噁英类持久性有机污染物（如多氯二苯并二噁英和多氯二苯并呋喃、PCDD/Fs以及共平面多氯联苯、PCBs）与未来可能添加到当前毒性当量因子（TEF）模型中的其他几类候选物进行比较。因此，本研究比较了几种混合多卤代二苯并二噁英和多卤代二苯并呋喃（PXDD/Fs；X = Br、Cl或F）、烷基取代的多卤代二苯并二噁英和多卤代二苯并呋喃（PMCDD/Fs；M = 甲基）、多卤代联苯（PXBs，X = Br、Cl）、多溴二苯醚（PBDEs）、五溴苯酚（PBPs）和四溴双酚-A（TBBP-A）的体外CYP1A1（微EROD生物测定法）和萤火虫荧光素酶诱导（DR-CALUX生物测定法）。我们还评估了同系物特异性相对效力（REPs）和效力（TCDD(max)的百分比），并讨论了这些化合物的剂量反应曲线，以及几种其他Ah受体激动剂的二噁英样效力，如多环芳烃（PAHs）和多氯萘（PCNs）的效力。在几种PXDD/F同系物中发现了最高的REP值，其次是一些共平面PXBs、三氯PCDD/Fs、PAHs、PBDE - 126、1 - 6 - HxCN和一些溴化阻燃剂（TBBP-A）。这些体外研究表明，有必要进一步研究以评估更多Ah受体激动剂的二噁英样效力。
