Gupta R K, Schuh R A, Fiskum G, Flaws J A
Program in Toxicology, Department of Epidemiology and Preventive Medicine, University of Maryland, 660 W. Redwood Street, Howard Hall 133B, Baltimore, MD, USA.
Toxicol Appl Pharmacol. 2006 Nov 1;216(3):436-45. doi: 10.1016/j.taap.2006.06.013. Epub 2006 Jul 1.
Methoxychlor (MXC) is an organochlorine pesticide that reduces fertility in female rodents by causing ovarian atrophy, persistent estrous cyclicity, and antral follicle atresia (apoptotic cell death). Oxidative damage resulting from reactive oxygen species (ROS) generation has been demonstrated to lead to toxicant-induced cell death. Thus, this work tested the hypothesis that MXC causes oxidative damage to the mouse ovary and affects mitochondrial respiration in a manner that stimulates ROS production. For the in vitro experiments, mitochondria were collected from adult cycling mouse ovaries, treated with vehicle (dimethyl sulfoxide; DMSO) or MXC, and subjected to polarographic measurements of respiration. For the in vivo experiments, adult cycling CD-1 mice were dosed with either vehicle (sesame oil) or MXC for 20 days. After treatment, ovarian mitochondria were isolated and subjected to measurements of respiration and fluorimetric measurements of H2O2 production. Some ovaries were also fixed and processed for immunohistochemistry using antibodies for ROS production markers: nitrotyrosine and 8-hydroxy-2'-deoxyguanosine (8-OHG). Ovaries from in vivo experiments were also used to measure the mRNA expression and activity of antioxidants such as Cu/Zn superoxide dismutase (SOD1), glutathione peroxidase (GPX), and catalase (CAT). The results indicate that MXC significantly impairs mitochondrial respiration, increases production of H2O2, causes more staining for nitrotyrosine and 8-OHG in antral follicles, and decreases the expression and activity of SOD1, GPX, and CAT as compared to controls. Collectively, these data indicate that MXC inhibits mitochondrial respiration, causes ROS production, and decreases antioxidant expression and activity in the ovary, specifically in the antral follicles. Therefore, it is possible that MXC causes atresia of ovarian antral follicles by inducing oxidative stress through mitochondrial production of ROS.
甲氧滴滴涕（MXC）是一种有机氯农药，它通过导致卵巢萎缩、持续性发情周期和窦状卵泡闭锁（凋亡性细胞死亡）来降低雌性啮齿动物的生育能力。已证明活性氧（ROS）生成所导致的氧化损伤会引发毒物诱导的细胞死亡。因此，本研究检验了以下假设：MXC会对小鼠卵巢造成氧化损伤，并以刺激ROS产生的方式影响线粒体呼吸。在体外实验中，从成年处于发情周期的小鼠卵巢中收集线粒体，用溶剂（二甲基亚砜；DMSO）或MXC处理，然后进行呼吸的极谱测量。在体内实验中，给成年处于发情周期的CD-1小鼠分别注射溶剂（芝麻油）或MXC，持续20天。处理后，分离卵巢线粒体并进行呼吸测量以及H2O2产生的荧光测量。一些卵巢也进行固定，并使用针对ROS产生标志物：硝基酪氨酸和8-羟基-2'-脱氧鸟苷（8-OHG）的抗体进行免疫组织化学处理。体内实验的卵巢还用于测量抗氧化剂如铜/锌超氧化物歧化酶（SOD1）、谷胱甘肽过氧化物酶（GPX）和过氧化氢酶（CAT）的mRNA表达及活性。结果表明，与对照组相比，MXC显著损害线粒体呼吸，增加H2O2的产生，导致窦状卵泡中硝基酪氨酸和8-OHG的染色增多，并降低SOD1、GPX和CAT的表达及活性。总体而言，这些数据表明MXC抑制线粒体呼吸，导致ROS产生，并降低卵巢中，特别是窦状卵泡中的抗氧化剂表达及活性。因此，MXC有可能通过线粒体产生ROS诱导氧化应激，从而导致卵巢窦状卵泡闭锁。
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