MECHANISMS OF 1,3-DINITROBENZENE TESTICULAR TOXICITY

随着人们对化学品产生不利影响的能力的认识不断增强 男性生殖，迫切需要增加我们的知识 关于化学物质造成生殖损害的机制。这个 需求是显而易见的，因为能够从实验室进行外推 动物对人类的影响和改进毒性测试取决于对 生物系统以及毒物是如何扰乱它的。在建议的 研究表明，感兴趣的睾丸毒物类别是 硝基芳香族化合物广泛用作农药、炸药、 工业合成中的药品和化学中间体。这个 模型睾丸毒物和硝基芳香族化合物为1，3-二硝基苯 (1，3-DNB)。利用实验室动物模型，该战略将首先 研究代谢(包括肝脏和睾丸内)在 1，3-DNB的睾丸毒性。哪种代谢物负责 毒性及其与细胞成分相互作用的性质 将代表这项研究的第二阶段。考虑到 从动物研究中获得的信息，最终目标是建立 使用人类的动物模型与人类情境的关系 体外研究中的组织。为了实现这些目标，方法是 将在体内测试假说，然后研究特定的细胞 试管内活动。第一个具体目标是确定 肝脏参与了1，3-DNB睾丸毒性的调节。 初步研究表明1，3-DNB发生了睾丸外事件 睾丸内注射高水平1，3-DNB的毒性 没有导致明显的睾丸损伤。两种情况可能是 可以解释这一结果的原因。首先，肝脏可能 将1，3-DNB代谢成能够在体内循环的物种 血液和睾丸中的毒性。亚硝基苯将是 被调查为最有可能成为循环中有毒物质的候选者 代谢物。第二，在以下情况下可以提出不那么直接的互动 肝脏代谢生成高铁血红蛋白代谢产物(S)建立 缺氧症。睾丸内的低氧状态 会促进1，3-DNB的还原代谢 通过氧化还原提高氧利用率而造成的缺氧症 循环或增加有毒代谢物的形成，如 亚硝基苯。缺氧的放大可能会导致 破坏Sertoli细胞的稳态，导致无法 支持发育中的生殖细胞群体。对于具体目标二，在 体外机制研究将进一步探索体内假说 专注于1)代谢激活的细胞指示物的检测 代谢产物(S)能够进行氧化还原循环，以及2)能力 亲电代谢物亚硝基苯与细胞结合 并破坏细胞内环境的稳定。也是；亚细胞 1，3-DNB中涉及的还原酶的定位和鉴定 将对新陈代谢进行研究。第三个具体目标是比较 人和动物组织代谢1，3-DNB和 预测物种间的相对毒性。

With growing awareness of the capacity of chemicals to adversely affect male reproduction, there is a pressing need to increase our knowledge about the mechanisms by which chemicals cause reproductive damage. The need is apparent since the ability both to extrapolate from laboratory animals to man and to improve toxicity testing depends on knowledge of the biological system and how toxicants perturb it. In the proposed studies, the class of testicular toxicants of interest are the nitroaromatics which are widely used as pesticides, explosives, pharmaceuticals and chemical intermediates in industrial syntheses. The model testicular toxicant and nitroaromatic is 1,3-dinitrobenzene (1,3-DNB). Using laboratory animal models, the strategy will be first to investigate the role of metabolism (both hepatic and intratesticular) in the testicular toxicity of 1,3-DNB. Which metabolite is responsible for toxicity and the nature of its interactions with cellular constituents will represent the second stage of the research. Taking into account the information derived from the animal studies, a final goal is to establish relationship between the animal model and the human situation using human tissue in in vitro studies. To accomplish these objectives, the approach will be to test hypotheses in vivo then investigate specific cellular events in vitro. The first specific aim is to establish what role the liver plays in the modulation of 1,3-DNB testicular toxicity. Preliminary studies have implicated extratesticular events in 1,3-DNB toxicity since intratesticular administration of high levels of 1,3-DNB did not result in significant testicular damage. Two scenarios could be envisioned which could account for this result. Firstly, the liver may metabolize 1,3-DNB to a species which is capable of circulating in the blood and eliciting toxicity in the testes. Nitronitrosobenzene will be investigated as the most likely candidate for a circulating toxic metabolite. Second, a less direct interaction can be proposed where liver metabolism generates methemoglobin-forming metabolite(s) setting up conditions of oxygen deficiency. Low oxygen conditions within the testes would promote reductive metabolism of 1,3-DNB either magnifying the oxygen deficit through an increase in oxygen utilization via redox cycling or increasing the formation of a toxic metabolite such as nitronitrosobenzene. Magnification of the oxygen deficiency could lead to disruption of Sertoli cell homeostasis with a consequent inability to support the developing germ cell population. For specific aim two, in vitro mechanistic studies will explore further the in vivo hypotheses focussing on 1) detection of cellular indicators of metabolic activation to metabolite(s) capable of redox cycling, and 2) the capacity of the electrophilic metabolite nitronitrosobenzene to bind to cellular nucleophiles and disrupt cellular homeostasis. Also; the subcellular localization and identity of the reductases involved in 1,3-DNB metabolism will be investigated. The third specific aim is to compare the capacity of human and animal tissue to metabolize 1,3-DNB and to predict relative toxicity between species.