Nanoparticle Transport to Oocytes and Toxicological Consequences in Fathead Minnows

该奖项是根据2009年美国复苏和再投资法案（公法111-5）资助的。 一旦内化，纳米材料有可能进入血液，在那里它们可能对包括卵巢在内的内脏器官构成风险。 新出现的文献和当前应用中的初步证据表明，纳米材料可能能够进入卵母细胞，可能影响胚胎发育。 有必要了解使纳米材料运输到卵母细胞的基本机制，以及随后的自然生物过程的改变。目前理解的是，纳米材料的生理行为在很大程度上受到吸附的蛋白质层的影响，这些蛋白质层在暴露于生物介质几秒钟后形成。繁殖活跃的雌鱼血清中的卵黄前体蛋白，卵黄原蛋白水平很高（高达20毫克/毫升）。 这种蛋白质是一种大的磷脂糖蛋白，能够通过各种管道与纳米材料结合，包括高负电荷区域和高疏水性区域。 卵黄原蛋白天然地靶向于发育中的卵母细胞，并通过卵黄原蛋白特异性受体输入这些卵母细胞。该项目的总体假设是，卵黄原蛋白结合的纳米材料将被输入到发育中的卵母细胞中的卵黄颗粒中，并最终与溶酶体合并。 此外，在到达该隔室时，某些纳米材料可以溶解并渗入细胞中，对发育中的胚胎造成毒性。 为了解决这些假设，研究人员有3个具体目标：（1）确定表面改性对鱼类卵巢培养物中纳米材料的吸收，细胞分布和保留的影响。(2)表征暴露于雄性和雌性黑头呆鱼血浆的纳米材料的蛋白质表面涂层。(3)测定纳米粒子在体内的分布和生殖毒性。为了实现这些目标，他们将使用模型发光纳米颗粒，即染料掺杂的二氧化硅和精心选择的尺寸分布和表面性质的量子点，这将使这些材料在体外和体内的运动的详细分析。拟议的研究将提供关键的见解纳米颗粒的性质对他们的易位卵母细胞的作用。 预计这一信息在开发鱼类纳米毒性预测模型以及减轻纳米材料暴露于环境的潜在不利后果的方法方面非常重要。 研究生，本科生和高中生在粒子科学与工程和生物学之间的接口的跨学科培训将是研究计划的重要组成部分。 东南联盟研究生教育和教授（SEAGEP），本科生（REU）计划的相关研究经验，并在佛罗里达大学建立少数民族奖学金计划将被利用，以确保研究计划的多样性。 研究结果将在同行评审的期刊上发表，并在国家和国际会议上传播。 研究人员还计划与国家纳米技术资源，如亚利桑那州立大学的纳米技术社会中心和赖斯的ICON合作，将研究成果传播给科学和教育界，以及整个社会。 学生还将参加公共宣传计划（例如，UF的工程博览会）为他们提供了一个渠道，让公众参与纳米科学的兴奋和环境管理的重要性。 UF的格雷厄姆公共政策中心将参与，以确保研究结果适当传达给决策者。 拟议的研究将提供介导纳米颗粒运输到卵母细胞的因素的基础知识，并提供可能导致鱼类胚胎改变的机制的见解。

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).0853707DenslowGrowing evidence suggests that many nanomaterials are taken up into fish species through the gills or digestive track. Once internalized, there is a potential for nanomaterials to cross into the blood stream, where they could pose a risk to internal organs including the ovary. Emerging literature and preliminary evidence in the current application suggest that nanomaterials may be capable of entering oocytes, potentially impacting embryonic development. There is a need to understand fundamental mechanisms that enable nanomaterial transport to oocytes, and the subsequent alterations of natural biological processes. It is currently understood that the physiological behavior of nanomaterials is largely impacted by adsorbed proteinaceous layers, which form upon seconds of exposure to biological media. Female fish that are actively reproducing have high serum levels (up to 20 mg/ml) of the egg yolk precursor protein, vitellogenin. This protein is a large phospholipoglycoprotein that has the capacity to bind to nanomaterials through various conduits, including areas of high negative charge and areas of high hydrophobicity. Vitellogenin is naturally targeted to developing oocytes and is imported into these oocytes via a vitellogenin-specific receptor. The overall hypothesis of this project is that vitellogenin-bound nanomaterials will be imported into yolk granules in developing oocytes and eventually merge with lysosomes. Furthermore, upon reaching this compartment certain nanomaterials could dissolve and leach into the cell causing toxicity for developing embryos. To address these hypotheses, the researchers have 3 specific aims: (1) Determine the effect of surface modification on uptake, cellular distribution and retention of nanomaterials in fish ovarian cultures. (2) Characterize the protein surface coating of nanomaterials exposed to male and female fathead minnow plasma. (3) Determine the distribution and reproductive toxicity of nanoparticles in vivo. To achieve these aims they will use model luminescent nanoparticulates, namely dye doped silica and quantum dots of carefully chosen size distributions and surface properties that will enable detailed analysis of the movement of these materials in vitro and in vivo. The proposed research will provide critical insights into the role of nanoparticulate properties on their translocation to oocytes. This information is anticipated to be highly important in the development of predictive models for nanotoxicity in fish species, and methods to mitigate potentially adverse outcomes of nanomaterial exposure to the environment. Cross-disciplinary training of graduate, undergraduate, and high school students at the interface between particle science and engineering and biology will be an essential part of the research program. The Southeast Alliance for Graduate Education and the Professoriate (SEAGEP), related Research Experience for Undergraduates (REU) programs, and established minority fellowship programs at the University of Florida will be leveraged to ensure diversity in the research program. Research results will be published in peer-reviewed journals and disseminated at national and international meetings. The investigators also plan to collaborate with national nanotechnology resources such as the Center for Nanotechnology in Society at Arizona State University and ICON at Rice to disseminate research results to the scientific and educational communities, and to society at large. The students will also participate in public outreach programs (e.g., UF's Engineering Fair)providing a conduit for them to engage the general public about the excitement of nanoscience and the importance of environmental stewardship. UF's Graham Center for Public Policy will be engaged to ensure that research findings are appropriately communicated to policy makers. The proposed research will provide fundamental knowledge of factors that mediate nanoparticle transport to oocytes, and offer insights on mechanisms that may lead to alterations in fish embryos.