Baier-Anderson C, Anderson RS
Program in Toxicology and the Center for Environmental Science, Chesapeake Biological Laboratory, University of Maryland, P.O. Box 38, 20688, Solomons, MD, USA
Aquat Toxicol. 2000 Aug 1;50(1-2):85-96. doi: 10.1016/s0166-445x(99)00092-2.
Chlorothalonil (TCIN) is the most commonly applied fungicide in the USA, with substantial use in the Chesapeake Bay area. Little is known about the sublethal toxicity of TCIN to fish, but since it is structurally similar to the immunotoxicant pentachlorophenol, the potential for immunomodulation exists. Previous studies have indicated that in vitro exposure of macrophages to TCIN modulates immunostimulated reactive oxygen species (H(2)O(2)/hypochlorous acid) and NADPH production in striped bass (Morone saxatilus). The goals of this study were to determine if TCIN inhibits superoxide (O(2)(-)) production by macrophage NADPH oxidase, to examine the role of cellular sulfhydryl groups in TCIN-induced macrophage dysfunction, and to identify the extent to which lipid peroxidation contributes to the observed toxic effects. The results of lucigenin-augmented chemiluminescence assays indicated that TCIN suppressed both baseline and stimulated O(2)(-) production in a dose-dependent manner. Similar results were obtained using both the particulate stimulant zymosan and the lipid-soluble stimulant phorbol 12-myristate-13-acetate. Inhibition of glutathione synthesis by pre-treatment with buthionine sulfoximine (BSO) enhanced the suppression of O(2)(-) production. The protection of sulfhydryl groups by culturing macrophages with dithiothreitol (DTT) reduced TCIN-induced macrophage dysfunction. TCIN did not initiate lipid peroxidation in macrophages, as measured by the thiobarbituric acid reactive substances (TBARS) assay, nor did pre-treatment with BSO potentiate lipid peroxidation. Because the observed TCIN-induced suppression of O(2)(-) was modulated by altering cellular sulfhydryl status with BSO and DTT, it is possible that toxicity results from the inhibition of NADPH oxidase activity by TCIN binding to its functional sulfhydryl groups.
百菌清（TCIN）是美国应用最广泛的杀菌剂，在切萨皮克湾地区大量使用。关于TCIN对鱼类的亚致死毒性知之甚少，但由于其结构与免疫毒性物质五氯苯酚相似，存在免疫调节的可能性。先前的研究表明，巨噬细胞在体外暴露于TCIN会调节条纹鲈（Morone saxatilus）免疫刺激产生的活性氧（H₂O₂/次氯酸）和NADPH的产生。本研究的目的是确定TCIN是否抑制巨噬细胞NADPH氧化酶产生超氧化物（O₂⁻），研究细胞巯基在TCIN诱导的巨噬细胞功能障碍中的作用，并确定脂质过氧化在观察到的毒性作用中的贡献程度。光泽精增强化学发光分析结果表明，TCIN以剂量依赖的方式抑制基线和刺激的O₂⁻产生。使用颗粒刺激剂酵母聚糖和脂溶性刺激剂佛波酯12 - 肉豆蔻酸酯13 - 乙酸酯均获得了类似结果。用丁硫氨酸亚砜胺（BSO）预处理抑制谷胱甘肽合成可增强对O₂⁻产生的抑制作用。用二硫苏糖醇（DTT）培养巨噬细胞对巯基的保护可减轻TCIN诱导的巨噬细胞功能障碍。通过硫代巴比妥酸反应物质（TBARS）分析测定，TCIN不会引发巨噬细胞中的脂质过氧化，用BSO预处理也不会增强脂质过氧化。由于观察到的TCIN诱导的O₂⁻抑制可通过用BSO和DTT改变细胞巯基状态来调节，因此毒性可能是由于TCIN与其功能性巯基结合抑制了NADPH氧化酶活性所致。
