Department of Biology/Toxicology, Ashland University, Ashland, OH 44805, USA.
Environ Toxicol Pharmacol. 2004 Mar;16(1-2):13-23. doi: 10.1016/j.etap.2003.08.002.
Single-chemical and mixture concentration-response curves generated using a frog embryo model were examined for value in assessing whether chemicals exert toxic effects at the same or at different molecular sites of action. Toxicity tests were conducted on a series of osteolathyrogens, i.e. chemicals that inhibit cross-linking of developing connective tissue fibers. Induction of osteolathyrism, which manifests as lesions in the notochord of exposed tadpoles, has several possible molecular sites of action, including agent-cofactor reactivity during the enzyme-mediated cross-linking process. UV-VIS spectrophotometry of osteolathyrogen-cofactor reactivity (i.e. in vitro analysis) was coupled with the 96-h frog embryo mixture toxicity assay (i.e. in vivo toxicity) to compare molecular sites of action for several osteolathyrogens with the combined osteolathyritic effects of the agents. Single-chemical concentration-response curves were used to calculate theoretical curves for the dose-addition model of combined effect. Slope and EC(50) values for both theoretical and experimental mixture curves were then generated to statistically examine the hypothesis that agents with shared sites of action have dose-response curve (DRC) slopes that are similar when given alone and in combination, and slope and EC(50) values that, when administered together, are consistent with those calculated for dose-addition. For combinations of cofactor-binding agents (semicarbazide, thiosemicarbazide, aminoacetonitrile), slope values were generally similar with additivity quotients near 1.0 (1.0=dose-additive) and combined osteolathyritic effects that were consistent with dose-addition. None of these were true for combinations that included agents that did not show rapid cofactor binding (β-aminopropionitrile, methyleneaminoacetonitrile). The results suggest that DRC analysis could be a useful tool for delineating common or different molecular sites of toxic action and that the approaches used warrant further study for evaluating the mechanistic basis for combined effects of toxicants.
使用蛙胚模型生成的单一化学物质和混合物浓度-反应曲线，用于评估化学物质是否在相同或不同的分子作用部位发挥毒性作用。进行了一系列骨生成抑制剂的毒性测试，即抑制发育中结缔组织纤维交联的化学物质。骨生成抑制剂诱导的骨生成抑制症，表现为暴露的蝌蚪脊索中的病变，有几个可能的分子作用部位，包括在酶介导的交联过程中，试剂-辅因子的反应性。骨生成抑制剂-辅因子反应性的 UV-VIS 分光光度法（即体外分析）与 96 小时蛙胚混合物毒性测定（即体内毒性）相结合，比较了几种骨生成抑制剂的分子作用部位与试剂的联合骨生成抑制作用。单一化学物质浓度-反应曲线用于计算联合效应的剂量相加模型的理论曲线。然后生成理论和实验混合物曲线的斜率和 EC（50）值，以统计检验具有共同作用部位的试剂在单独和组合时的剂量-反应曲线（DRC）斜率相似，并且当一起给药时，斜率和 EC（50）值与计算的剂量相加一致的假设。对于辅因子结合试剂（氨基脲、硫代氨基脲、氨基乙腈）的组合，斜率值通常相似，加性商接近 1.0（1.0=剂量相加），并且联合骨生成抑制作用与剂量相加一致。对于包括没有显示快速辅因子结合的试剂（β-氨基丙腈、亚甲基氨基乙腈）的组合，这些都不成立。结果表明，DRC 分析可能是一种有用的工具，用于描绘共同或不同的毒性作用分子部位，并且所使用的方法值得进一步研究，以评估毒物联合效应的机制基础。
