金纳米颗粒跨越胎盘屏障转运的机制研究

It should be the basic and key issues on nanosafety and nanomedicine fields that how nanoparticles cross the placental barrier to get fetuses in pregnancy. With the rapid development of nanotechnology in biomedical field, the risk of human exposure to nanomaterials has increased. Studies on evaluation of the transplacental transfer ability of nanomaterials and the potential toxicity of exposure to nanomaterials during pregnancy are causing widespread concern of scientists of all countries. So far only very few studies have investigated whether nanoparticles pass the human placenta, and there is virtually no information on the transplacental uptake and transfer mechanisms involved. In the research supported by NSFC Youth Science Fund (31000452), we explored the transplacental transfer properties of gold nanoparticles on different pregnant ages, and assessed the influence of different physical and chemical properties including the size, surface composition and charge of the nanoparticles in these processes. In the current project, we will focus on transplacental uptake and transfer mechanisms for gold nanoparticles. On different levels of in vivo animal, ex vivo perfused human placenta and in vitro cell culture models, we will detect gold nanoparticles transport rate, transport volume and their interaction with the main biological interfaces within placenta when they pass through placental membrane barrier. We will focuses on identifying the selection of transport pathways for gold nanoparticles between the active transport, passive diffusion and various forms of endocytosis pathway, as well as the the influence of nanoparticle physical and chemical properties on these selections. Through the above research, we would interpret the cellular and molecular mechanisms for both transplaceta transfer and placental shielding of nanoparticles, providing new ideas for developing drug delivery strategy in pregnancy and avoiding fetal injury from nanoparticle exposure. These research would also provide support for reasonable extrapolation of animal experiment data on nanoparticles transplacental transfer to human beings.

纳米颗粒如何跨越胎盘屏障向胎儿转运,是孕期纳米生物安全性和纳米生物医学领域的基础性和关键性问题。在刚刚结题的青年基金项目中，我们系统地研究了金纳米颗粒在小鼠孕期不同时段跨胎盘转运的特性，以及尺度、表面修饰等对于金纳米颗粒胎盘转运的影响。后续课题研究中，我们将针对金纳米颗粒的胎盘转运机制进行系统深入的探索。在孕期动物、人离体胎盘组织和体外培养胎盘细胞等不同的层次，检测金纳米颗粒跨越胎盘膜屏障的转运速率、转运量及其与胎盘内部主要生物界面的相互作用，着重分析和鉴定金纳米颗粒穿越胎盘细胞膜时，在主动转运、被动扩散以及各种形式的内吞途径之间的选择，以及材料不同的理化特性对其转运途径选择的影响。通过上述研究,阐释金纳米颗粒跨越胎盘转运或被胎盘屏蔽的细胞分子机制,为发展针对孕期母体和胎儿健康问题的新型药物输运策略和有效防止纳米颗粒孕期暴露对胎儿的潜在影响提供新的研究思路与途径。

金纳米颗粒（GNPs）是在成像、检测、药物转运等方面具有巨大应用潜力的纳米材料，但是GNPs对于妊娠期母体病生理状态和胎儿发育的影响,以及GNPs跨越胎盘膜转运的细胞机制研究尚属空白，极大阻碍了GNPs在母婴医学领域的研究发展。本项目在胎盘细胞细胞、妊娠动物和人终末期灌流胎盘等不同的生物学层次，建立了GNPs跨越胎盘屏障的母胎转运研究模型，系统研究了GNPs由母体向胎儿转运的特性、影响因素、相关机制以及GNPs跨胎盘转运对于妊娠和胎儿发育的影响。通过ICP-MS为基础的摄取率检测、以转运蛋白和转运通道为目标的转运途径干预、透射电镜解析，以及细胞转运相关蛋白表达分析等系列研究手段相结合，解析、鉴定GNPs穿越胎盘细胞屏障的可能途径和关键作用靶位。研究首次证实了13纳米以下较小尺度GNPs是以团聚状态通过细胞吞噬进行入胞转运，转运不涉及clathrin/caveolae介导的细胞吞饮，并提示了30纳米以上GNPs跨越胎盘细胞膜转运的诸多特性。同时在孕期小鼠动物模型上完成了不同尺度GNPs通透胎盘屏障的差异比较研究，发现小鼠胚胎11.5天（E11.5）是决定4.5纳米以上尺度GNPs能否无障碍跨越胎盘的时间窗口。在小鼠妊娠早期（对应于人的第一个“三月期”），30纳米GNPs可通过不成熟的胎盘直接进入胚胎，干预和抑制早期胚胎细胞向外胚层的分化，导致胚胎停育和流产，GNPs对于胎儿发育的影响遵循“全或无”模式，妊娠晚期暴露于母体循环的30纳米GNPs不影响胎儿发育。在此基础上进行了针对于胎盘滋养细胞的高亲和性抗体的筛选工作，获得了两种胎盘滋养细胞BeWo B24亚株高亲和性抗体。设计构建了人终末期胎盘的灌流系统，用以后续纳米颗粒跨越人胎盘转运特性和安全性评价。.上述系列研究首次阐释了金纳米颗粒跨越胎盘滋养层细胞膜的细胞机制以及金纳米颗粒干预胚胎细胞分化发育的细胞分子机制，具有重要的理论意义。系列研究成果为以颗粒尺度为主要考量维度进行孕期纳米药物设计奠定了基础，并且为孕期可能面临的纳米材料暴露的安全性与评价提供了有价值的参考数据。