EPIGENETIC TRANSGENERATIONAL ENDOCRINE DISRUPTOR ACTIONS

描述（由申请人提供）：环境毒素的跨代效应，如内分泌干扰物，显着放大这些化合物的影响和健康危害。这些化合物作用的跨代性质表明，在种系上存在表观遗传效应。本研究旨在探讨这种跨代表观遗传现象对男性生殖的影响。研究表明，内分泌干扰物会导致精子数量和生育能力异常，从而影响男性生殖。对内分泌干扰物暴露最敏感的时期之一是胚胎发育时期。本研究的目的是探讨内分泌干扰物对男性生殖的作用机制，重点是睾丸发育。一个啮齿动物模型（即大鼠）系统被用来提供深入了解内分泌干扰物作用的机制方面。测试的内分泌干扰物模型是甲氧基氯，其代谢产物是弱雌激素和抗雄激素化合物，vinclozolin是一种抗雄激素化合物。因此，这些模型内分泌干扰物允许考虑雌激素和抗雄激素内分泌干扰物的作用。目的是深入了解内分泌干扰物对男性生殖的分子、细胞和生理（即系统生物学）作用。经检验的假设是，子宫内短暂胚胎暴露于内分泌干扰物会影响胚胎睾丸转录组，并通过表观遗传效应（例如DNA甲基化）导致异常生殖细胞分化，进而影响成年生精能力和男性生育能力，并且这种表型通过种系跨代传播。已有研究表明，甲氧基氯和vinclozolin在睾丸形态发生时影响胚胎睾丸的发育，导致成体生殖细胞凋亡增加。有趣的是，观察结果表明，这种异常的精子发生是跨代的，可能是由于生殖系DNA甲基化的改变，通过内分泌干扰物的表观遗传作用。初步研究还表明，在内分泌干扰物作用下，旁分泌生长因子基因表达的改变直接影响睾丸发育。内分泌干扰物引起的睾丸发育和生殖细胞分化异常，部分原因可能是睾丸转录组控制不当和生殖细胞发育异常。验证上述假设的实验方法包括以下具体目的：1)研究内分泌干扰物对睾丸发育和配子体发生的跨代影响。2)通过分析睾丸和种系的表观遗传效应（如DNA甲基化），确定内分泌干扰物的作用机制。3)确定胚胎短暂暴露于内分泌干扰物对胚胎和出生后睾丸转录组的作用。从拟议的研究中获得的信息将确定环境毒素（即vinclozolin和甲氧基氯）如何通过对胚胎睾丸转录组和种系DNA甲基化产生不利影响而损害男性生育能力。这些研究的完成将有助于深入了解胚胎暴露于内分泌干扰物对成年男性生殖功能的跨代影响的机制方面。拟议的研究将用于推断和深入了解内分泌干扰物对人类发育、生殖和健康的影响。跨代表观遗传内分泌干扰物对男性生殖的作用的新观察严重影响了这些化合物作为环境毒素的潜在危害。本研究将彻底确立这一现象，并阐明其分子机制。

DESCRIPTION (provided by applicant): Transgenerational effects of environmental toxins, such as endocrine disrupters, significantly amplify the impact and health hazards of these compounds. The transgenerational nature of the actions of these compounds suggests an epigenetic effect on the germ-line. The current proposal is designed to investigate this transgenerational epigenetic phenomenon on male reproduction. Endocrine disrupters have been shown to influence male reproduction by causing abnormal sperm numbers and fertility. One of the most sensitive periods to endocrine disrupter exposure is during embryonic development. The objective of the proposed research is to investigate the mechanism of action of model endocrine disrupters on male reproduction with a focus on testis development. A rodent model (i.e. rat) system is used to provide insight into the mechanistic aspects of endocrine disrupter action. The model endocrine disruptors tested are methoxychlor that has metabolites that are both weak estrogenic and anti-androgenic compounds, and vinclozolin, which is an anti-androgenic compound. Therefore, these model endocrine disruptors allow consideration of both estrogenic and anti-androgenic endocrine disrupter actions. The objective is to obtain insight into the molecular, cellular and physiological (i.e. systems biology) actions of endocrine disruptors on male reproduction. The hypothesis tested is that transient embryonic in utero exposure to an endocrine disruptor influences the embryonic testis transcriptome and through epigenetic effects (e.g. DNA methylation) results in abnormal germ cell differentiation that subsequently influences adult spermatogenic capacity and male fertility and that this phenotype is transgenerational through the germ-line. Previous studies have shown that methoxychlor and vinclozolin can effect embryonic testis development at the time of testis morphogenesis and that this causes an increase in germ cell apoptosis in the adult. Interestingly, observations suggest this abnormal spermatogenesis is transgenerational and may be due to altered DNA methylation of the germ-line through an epigenetic action of the endocrine disrupter. Preliminary studies have also demonstrated that altered gene expression of paracrine growth factors directly influence testis development at the time of endocrine disrupter action. Abnormal testis development and germ cell differentiation caused by endocrine disruptors may in part be due to inappropriate control of the testis transcriptome and abnormal germ cell development. The experimental approach to test the above hypothesis consists of the following specific aims: 1) Investigate the transgenerational effects of endocrine disruptors on testis development and gametogenesis. 2) Determine the mechanism of endocrine disrupter actions through analysis of epigenetic effects (e.g. DNA methylation) on the testis and germ-line. 3) Determine the actions of transient embryonic exposure to endocrine disruptors on the embryonic and postnatal testis transcriptome. Information obtained from the proposed research will determine how environmental toxins (i.e. vinclozolin and methoxychlor) may impair male fertility by adversely affecting the embryonic testis transcriptome and germ-line DNA methylation. The completion of these studies will provide insight into the mechanistic aspects of how embryonic exposure to endocrine disruptors cause a transgenerational effect on adult male reproductive function. The proposed research will be used to extrapolate and provide insight into the impact of endocrine disruptors on human development, reproduction and health. The novel observations of transgenerational epigenetic endocrine disrupter actions on male reproduction critically impacts the potential hazards of these compounds as environmental toxins. The proposed research will thoroughly establish this phenomenon and elucidate the molecular mechanisms involved.