Godard Céline A J, Smolowitz Roxanna M, Wilson Joanna Y, Payne Roger S, Stegeman John J
Woods Hole Oceanographic Institution, Biology Department, Woods Hole, Massachusetts 02543, USA.
Toxicol Sci. 2004 Aug;80(2):268-75. doi: 10.1093/toxsci/kfh124. Epub 2004 Mar 31.
Marine mammals can accumulate environmental contaminants in their blubber at concentrations harmful to laboratory animals. Induction of the cytochrome P450 1A1 (CYP1A1) enzyme is widely used as a biomarker of exposure and molecular effects in animal species, yet the validity of this biomarker has not been established in marine mammals. In vivo studies are generally precluded in these protected species, but skin biopsies (epidermis and dermis) can be collected in a minimally invasive way. We developed an in vitro assay using skin biopsy slices to examine CYP1A1 protein induction in marine mammals in response to chemical exposure. Skin biopsies from sperm whale (Physeter macrocephalus) were exposed for 24 h to beta-naphthoflavone (BNF), a prototypical CYP1A1 inducer, and CYP1A1 induction was detected by immunohistochemical staining in endothelial cells, smooth muscle cells, and fibroblasts. Biopsy slices were exposed to a range of BNF concentrations (0.6-600 microM), and a significant concentration-effect relationship was observed in both endothelial and smooth muscle cells (p = 0.05). This is the first study using skin biopsy slices to examine exposure of cetacean tissue to a CYP1A1 inducer. It demonstrates a causal relationship between chemical exposure and CYP1A1 induction and therefore validates the use of CYP1A1 expression in skin biopsies as a biomarker in cetaceans. Our protocol can be adapted to the investigation of chemicals, mixtures, concentrations, incubation times, or biological endpoints of choice. This should prove particularly relevant for these and other protected species that cannot be studied in the laboratory.
海洋哺乳动物的鲸脂中会累积环境污染物，其浓度对实验动物有害。细胞色素P450 1A1（CYP1A1）酶的诱导作用被广泛用作动物物种暴露和分子效应的生物标志物，但这种生物标志物在海洋哺乳动物中的有效性尚未得到证实。对于这些受保护物种，一般无法进行体内研究，但可以以微创方式采集皮肤活检样本（表皮和真皮）。我们开发了一种体外检测方法，使用皮肤活检切片来检测海洋哺乳动物在化学物质暴露后CYP1A1蛋白的诱导情况。将抹香鲸（Physeter macrocephalus）的皮肤活检样本暴露于典型的CYP1A1诱导剂β-萘黄酮（BNF）中24小时，通过免疫组织化学染色在内皮细胞、平滑肌细胞和成纤维细胞中检测到了CYP1A1的诱导。将活检切片暴露于一系列BNF浓度（0.6 - 600 microM）下，在内皮细胞和平滑肌细胞中均观察到了显著的浓度 - 效应关系（p = 0.05）。这是第一项使用皮肤活检切片检测鲸类组织暴露于CYP1A1诱导剂的研究。它证明了化学物质暴露与CYP1A1诱导之间的因果关系，因此验证了将皮肤活检中CYP1A1的表达用作鲸类生物标志物的有效性。我们的方案可适用于对化学物质、混合物、浓度、孵育时间或所选生物终点的研究。这对于这些以及其他无法在实验室中进行研究的受保护物种应该特别有用。
