Synthetic Biomimetic Environment for Improving IVF Embryo Culture

项目摘要 不孕不育是一种普遍的疾病，全世界超过10%的夫妇受到了不孕不育的影响。为了这些 对于夫妇来说，试管受精(IVF)是最好的受孕机会。然而，对于试管受精，总的 一次成功的分娩不到25%。此外，尽管大多数试管婴儿看起来很健康，但报告显示 植入前体外培养过程中的次佳暴露会影响出生后的生长、葡萄糖代谢、 脂肪沉积和血管功能。 人们普遍认为，与精子-卵母细胞体外融合(只需不到10小时)相比 人类)，植入前几天的胚胎体外培养更多地是造成这些表观遗传学的原因 精神错乱。这并不令人惊讶，因为体外和体内环境之间的巨大差异 植入前胚胎。而在过去的几十年里，胚胎培养液，孵化/观察 系统和氧气水平的控制都有了很大的改善，微环境的精细控制 胚胎一直被忽视。例如，植入前胚胎的培养仍然使用传统的2D 组织培养聚苯乙烯表面。因此，在这个项目中，我们将重点放在改进植入前胚胎上 通过提供仿生微环境进行培养。我们之前的研究已经证明了这一重要性 决定人类胚胎干细胞命运的生物物理线索。根据初步研究，我们 假设最佳的基质硬度、液体流动和胚胎滚动将改善胚胎质量 体外培养。为了实现我们的目标，我们将首先询问矩阵刚度的独立作用， 胚胎发育中的液体流动和胚胎滚动(目标1)。此外，为了整合这些线索，我们将开发 一种新型的模拟纤毛输卵管上皮细胞的可驱动纤维基质(AFS) 替代目前的试管受精培养皿。评估AFS对胚胎发育的影响，我们将检查 细胞谱系标记、印记基因的表达模式及其等位基因特异性DNA甲基化 与性别和胚胎细胞类型相关的差异效应。此外，注入频率和 将使用假孕小鼠测试体内胎儿/胎盘的发育(目标2)。 这个项目的长期目标是推进试管受精程序，以提高总体成功率。 并减少与试管受精程序相关的出生后疾病。值得注意的是，这里提出的AFS是生物兼容的， 多功能、易用、全自动化。它与在培养液中生长的胚胎相容。 被油层覆盖的液滴，这是临床上经常使用的。实时监测胚胎生长发育 我们的平台还允许使用延时孵化器。因此，临床医生可以使用AFS的相同方式 适应障碍最小的标准试管受精培养皿。

Project Summary Infertility is a widespread condition, which has affected more than 10% of all couples worldwide. For these couples, in vitro fertilization (IVF) represents the best chance to conceive. However, for IVF, the overall rate of a successful birth is less than 25%. Moreover, while most IVF children appear healthy, reports showed suboptimal exposures during pre-implantation in vitro culture can affect postnatal growth, glucose metabolism, fat deposition, and vascular function. It is generally believed that, compared with sperm-oocyte fusion in vitro (which needs less than 10 hours in human), several days of pre-implantation embryo in vitro culture is more responsible for these epigenetic disorders. This is not a surprise due to the drastic differences between in vitro and in vivo environment for the pre-implantation embryo. While over the past few decades, embryo culture media, incubation/observation systems, and oxygen level controls have been drastically improved, fine-control of the microenvironment of embryos has been overlooked. For example, culture of pre-implantation embryos still utilizes traditional 2D tissue culture polystyrene surfaces. Thus, in this project, we focus on improving the pre-implantation embryo culture by providing a biomimetic microenvironment. Our previous research has demonstrated the importance of biophysical cues in the fate decision of human embryonic stem cells. Based on preliminary studies, we hypothesize that optimal matrix stiffness, fluid flow, and embryo rolling will improve the quality of embryo culture in vitro. To accomplish our objectives, we will first interrogate the independent role of matrix stiffness, fluid flow and embryo rolling in embryo development (Aim 1). Further, to integrate these cues, we will develop a novel Actuatable Fibrillar Substrates (AFS) to mimic ciliated oviductal epithelial cells, as potential replacements for current IVF dishes. The evaluate the effects of AFS on embryo development, we will examine cell lineage markers, expression pattern of imprinted genes with their allele-specific DNA methylation, as well as differential effects associated with gender and cell types of embryo. Further, implantation frequency and fetus/placenta development in vivo will be tested using pseudopregnant mice (Aim 2). The long-term objective of this project is to advance the IVF procedures to improve the overall success rate and reduce postnatal diseases related to IVF procedures. Notably, the AFS proposed here are biocompatible, multi-functional, easy to use, and fully automated. It is compatible with embryos grown in culture medium droplets covered by an oil layer, which are routinely used in clinics. Real-time monitoring embryo growth using time-lapse incubators is also allowed with our platform. Thus, clinicians can use AFS the same way as standard IVF dishes with minimal hurdles of adaptation.