MECHANISM OF ALCOHOL TERATOGENESIS--OXIDATIVE STRESS

酒精致畸机制--氧化应激

• 批准号: 2046598
• 负责人: GEORGE I HENDERSON
• 金额: $ 17.27万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-08-01 至 1998-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2046598
• 关键词: alcoholic beverage consumption; antioxidants; cellular respiration; disease /disorder model; embryo /fetus toxicology; ethanol; fetal alcohol syndrome; free radical oxygen; laboratory rat; lipid peroxides; membrane lipids; mitochondria; neurotoxins; oxidative stress; postnatal growth disorder; tissue /cell culture

Maternal ethanol (E) consumption often has a devastating and lasting effect on fetal growth and development, both in humans and in animal models. Yet, the mechanism(s) for this remaining uncertain and no treatment, either preventative or postinsult for the fetal damage has evolved. Previous studies in our laboratory have documented a variety of fetotoxic responses to E using rat models for in vivo exposure and primary cultures of fetal cells in vitro. Recently we reported that in cultured fetal rat hepatocytes (FRH), E blocked replication, induced total and mitochondrial membrane damage, increased production of reactive oxygen species [O2 and H2O2 (ROS)] and membrane lipid peroxidation products in whole cell and mitochondria (M), and reduced GSH pools and ATP synthesis. Importantly, normalizing cell antioxidant status could reverse these measures and restore cell replicative capacity. Preliminary studies included here illustrate similar responses in vivo after a maternal binge exposure as well as in cultured neonatal astrocytes (A) and cardiac myocytes (CM), and documented an E-related inhibition of M respiratory complexes I and IV. Also, administration of allopurinol (AL) to dams concomitant with E, normalized H2O2 production and GSH in fetal brain (but not liver), indicating a xanthine oxidase interaction. Thus, we hypothesize, first, that the underlying mechanism for the toxic effects of E on fetal cells is E-induced generation of ROS, second, that many of these fetotoxic effects can be prevented or mitigated by treatments that optimize oxidant defense systems in fetal tissues, and third, that E damage of M plays an important role in these processes. The models will be Sprague Dawley rats exposed to E from gestation days (GD) 3-20 (chronic) or on GD days 11-13, 14-17, 18-20 (binge). We will focus on fetal brain, heart and liver as known targets fore, and on key cells of these tissues, cultured RFH, A, and CM. Specific Aim 1 is to define the oxidative stress with respect to ROS production, oxidative products in membrane lipids (MDA, dienes, 4-hydroxynonenal), proteins (thiol oxidation, aldehyde adducts), DNA (8-hydroxydeoxyguanosine), antioxidant status (GSH, vitamin E, ascorbate), and activities of SOD, catalase, and glutathione peroxidase. We will determine the rate of onset; duration, degree, and gestational timing of E exposure needed to elicit effects; and the reversibility of these parameters. Specific Aim 2 will develop treatment regimens to prevent or reduce this oxidative stress and its consequences. To this end, we will optimize oxidative defenses in E-exposed fetal tissues by maintaining GSH stores (with N- acetylcysteine, S-adenosylmethionine) and other antioxidants (vitamin E, ascorbate), and by administration of a xanthine oxidase inhibitor (AL). These treatments will be tested individually and in combination to be tailored to specific E exposure patterns and tissue requirements. Specific Aim 3 will test the hypothesis that there are close connections between E-related oxidative stress and M damage and that this is an important factor in the fetotoxicity of E. We will define effects of E on M integrity and function (especially respiratory chain) with respect to onset, exposure pattern, and reversal by antioxidant treatment.

母亲的乙醇 (E) 消耗通常会造成毁灭性且持久的后果 对人类和动物胎儿生长发育的影响 模型。 然而，这种机制仍然不确定，也没有 对胎儿损伤进行预防性或侮辱后治疗 进化了。 我们实验室之前的研究已经记录了多种胎儿毒性 使用大鼠模型进行体内暴露和原代培养对 E 的反应 体外胎儿细胞。 最近我们报道了在培养的胎鼠中 肝细胞 (FRH)，E 阻断复制，诱导总线粒体和线粒体 膜损伤，活性氧 [O2 和 全细胞和细胞中的H2O2 (ROS)]和膜脂过氧化产物 线粒体 (M)，并减少 GSH 池和 ATP 合成。 重要的是， 使细胞抗氧化状态正常化可以逆转这些措施 恢复细胞复制能力。 此处包含初步研究 说明母亲暴饮暴食后体内的类似反应 以及培养的新生儿星形胶质细胞 (A) 和心肌细胞 (CM)， 并记录了 M 呼吸复合物 I 和 E 相关的抑制 四． 此外，还可以同时给母坝施用别嘌呤醇 (AL) E，胎儿大脑（但不是肝脏）中 H2O2 产生和 GSH 标准化， 表明黄嘌呤氧化酶相互作用。 因此，我们首先假设有毒物质的潜在机制 E对胎儿细胞的影响是E诱导ROS的产生，其次， 许多这些胎儿毒性作用可以通过以下方法预防或减轻： 优化胎儿组织中的氧化防御系统的治疗，以及 第三，M的E损伤在这些过程中起着重要作用。 这些模型是斯普拉格道利大鼠，从妊娠期起就暴露于E (GD) 3-20（慢性）或 GD 第 11-13、14-17、18-20 天（暴食）。 我们将 重点关注胎儿大脑、心脏和肝脏作为已知目标，并重点关注关键 这些组织的细胞，培养 RFH、A 和 CM。 具体目标 1 是 定义与 ROS 产生、氧化相关的氧化应激 膜脂产品（MDA、二烯、4-羟基壬烯醛）、蛋白质 （硫醇氧化、醛加合物）、DNA（8-羟基脱氧鸟苷）、 抗氧化状态（GSH、维生素 E、抗坏血酸）和 SOD 活性， 过氧化氢酶和谷胱甘肽过氧化物酶。 我们将确定费率 发病；所需的电子暴露的持续时间、程度和妊娠时间 引发效应；以及这些参数的可逆性。 具体目标 2 将制定治疗方案来预防或减少这种氧化 压力及其后果。 为此，我们将优化氧化 通过维持 GSH 储存（用 N- 乙酰半胱氨酸、S-腺苷甲硫氨酸）和其他抗氧化剂（维生素 E、 抗坏血酸），以及通过施用黄嘌呤氧化酶抑制剂（AL）。 这些治疗方法将单独进行测试，并结合起来进行测试 根据特定的电子暴露模式和组织要求量身定制。 具体目标 3 将检验存在密切联系的假设 E 相关的氧化应激和 M 损伤之间的关系，这是一个 E 胎儿毒性的重要因素。我们将定义 E 的影响 尊重 M 的完整性和功能（尤其是呼吸链） 发病、暴露模式以及抗氧化治疗的逆转。