Rider Cynthia V, LeBlanc Gerald A
Department of Environmental and Molecular Toxicology, North Carolina State University, Raleigh, North Carolina 27695, USA.
Toxicol Sci. 2006 Oct;93(2):443-9. doi: 10.1093/toxsci/kfl067. Epub 2006 Jul 19.
Hemoglobin accumulation in daphnids is an important adaptive response that is regulated by at least two distinct molecular pathways: an endocrine pathway stimulated by terpenoid hormones and an oxygen-sensing pathway involving the hypoxia-inducible factor. We found that the herbicide atrazine elevated hemoglobin levels in Daphnia magna and hypothesized that atrazine induced hemoglobin in daphnids through the hormonal regulatory pathway. This hypothesis was tested by modeling the combined effects of atrazine and the terpenoid hormone mimic pyriproxyfen on hemoglobin mRNA levels assuming the same mechanism of action (concentration addition model) and alternatively, assuming different mechanisms of action (response addition model). Model predictions were then compared to experimental assessments of the combined action of these two chemicals on hemoglobin mRNA levels. Changes in hemoglobin expression were evaluated using real-time RT PCR with primers specific to each of three D magna hemoglobin genes (dhb1, dhb2, and dhb3). Both atrazine and pyriproxyfen significantly elevated levels of the hb2 gene product, while having little effect on hb1 and hb3 gene products. Induction of dhb2 by combinations of atrazine and pyriproxyfen did not conform to the concentration addition predictions. Rather, dhb2 induction by these binary combinations was highly consistent with response addition model predictions. These results indicate that atrazine does not induce hemoglobin through the terpenoid hormone-signaling pathway. Results from this study demonstrate that mixtures modeling can be used to assess a chemical's mechanism of action and that atrazine likely stimulates hemoglobin accumulation through the oxygen-sensing pathway.
水蚤体内血红蛋白的积累是一种重要的适应性反应，至少受两条不同的分子途径调控：一条是由萜类激素刺激的内分泌途径，另一条是涉及缺氧诱导因子的氧感应途径。我们发现除草剂阿特拉津可提高大型溞体内的血红蛋白水平，并推测阿特拉津通过激素调节途径诱导水蚤产生血红蛋白。通过模拟阿特拉津和萜类激素类似物吡丙醚对血红蛋白mRNA水平的联合作用来验证这一假设，一种假设是作用机制相同（浓度相加模型），另一种假设是作用机制不同（反应相加模型）。然后将模型预测结果与这两种化学物质对血红蛋白mRNA水平联合作用的实验评估结果进行比较。使用针对大型溞三个血红蛋白基因（dhb1、dhb2和dhb3）各自的特异性引物，通过实时RT-PCR评估血红蛋白表达的变化。阿特拉津和吡丙醚均显著提高了hb2基因产物的水平，而对hb1和hb3基因产物影响很小。阿特拉津和吡丙醚组合对dhb2的诱导不符合浓度相加预测。相反，这些二元组合对dhb2的诱导与反应相加模型预测高度一致。这些结果表明，阿特拉津并非通过萜类激素信号通路诱导血红蛋白。本研究结果表明，混合物建模可用于评估化学物质的作用机制，且阿特拉津可能通过氧感应途径刺激血红蛋白的积累。
