The design and analysis of synthetic substrates for embryonic stem cell culture

胚胎干细胞（ES）有能力成为人体数千种不同细胞类型中的任何一种。这种独特的能力意味着它们具有治疗几种严重疾病的巨大潜力，而目前的治疗方法往往是不完整的、短暂的，甚至根本不存在。这些疾病包括糖尿病、阿尔茨海默病和帕金森病，所有这些疾病都是由于年龄、感染或损伤导致的细胞退化或功能障碍造成的。然而，发展这种基于胚胎干细胞的治疗有两个主要障碍，首先，我们对如何获得足够数量的未分化胚胎干细胞并在组织培养中维持它们，其次，如何控制它们的分化以产生特定的细胞类型，我们充其量只是零散的了解。胚胎干细胞要么来自单个细胞，要么来自植入前胚胎中存在的极少数细胞，这意味着它们的数量必须在组织培养中扩增，才能产生临床使用所需的数量。这就产生了第一个障碍，因为为了做到这一点，胚胎干细胞目前必须在其他细胞类型或动物产品存在的情况下培养。这些添加到培养环境中的物质使细胞暴露于潜在的有害病毒或其他感染因子中，这些病毒或因子不仅可以转移给患者，还可以转移给与患者接触的其他人。第二个障碍是不完全了解如何控制胚胎干细胞的分化，因为我们对饲养细胞提供的因素知之甚少。因此，该项目的目的是消除暴露于其他细胞类型及其未定义产物的需要，以便开发一种方法，既可以产生大量适合临床使用的未受污染的胚胎干细胞，也可以使我们确定控制胚胎干细胞分化的机制。我们和其他人的工作表明，影响胚胎干细胞在组织培养环境中的行为的关键因素之一是它们彼此之间以及与细胞生长的基质的接触。我们能够以可控的方式修改材料的表面特性，以生产具有特定化学成分和纳米形貌（小于百万分之一米的表面结构）的基底，并且已经表明，不仅化学成分，而且基底的表面结构确实影响细胞行为。我们假设胚胎干细胞可以在合成底物上繁殖，这些底物设计用于控制与细胞的相互作用，从而消除了对培养基中动物产品的需求。在该项目中，我们将详细研究这些材料的表面特性，并评估在其上培养的胚胎干细胞的行为，以选择一组显示出控制胚胎干细胞行为潜力的底物。然后，我们将彻底研究底物的表面化学和地形以及暴露在底物下的胚胎干细胞的行为，以便我们可以设计特定的表面来优化胚胎干细胞的反应。这也将使我们对ES细胞生长的调节机制和底物表面特性控制其生长的机制有了更深入的了解。这些信息将有助于未来生产适合临床使用的胚胎干细胞。

Embryonic stem (ES) cells have the ability to become any of the thousands of different cell types of the human body. This unique ability means that they have great potential for treating several serious diseases, the current treatments for which are often incomplete, transitory or even non-existent. These include diabetes, Alzheimer's and Parkinson's disease all of which result from cell degeneration or malfunction due to age, infection or injury. However two major obstacles in the development of such ES cell-based treatments are that we still have at best a fragmentary understanding of firstly how to obtain adequate numbers of undifferentiated ES cells and maintain them in tissue culture, and secondly how to control their differentiation in order to produce specific cell types. ES cells are derived from either a single cell or the very small numbers of cells present in the preimplantation embryo, meaning that their numbers have to be amplified in tissue culture in order to produce the amounts that will be necessary for clinical use. This gives rise to the first obstacle because in order to do this, the ES cells currently have to be cultured in the presence of other cell types or animal products. These additions to the culture environment expose the cells to potentially harmful viruses or other infectious agents which could be transferred not only to the patients but also others with whom they come into contact. The second obstacle of not fully understanding how to control the differentiation of ES cells also arises because we know little about the factors supplied by the feeder cells. The aim of this project is therefore to eliminate the need for exposure to other cell types and their undefined products in order to develop a method that will both produce large numbers of uncontaminated ES cells suitable for clinical use and will also allow us to determine the mechanisms that control the differentiation of the ES cells. Work by ourselves and others has shown that one of the crucial factors influencing the behaviour of ES cells in the tissue culture environment is their contact with each other and with the substrate on which the cells grow. We are able to modify the surface properties of materials in controlled ways to produce substrates with specific chemistries and nanotopographies (surface architecture on a scale less than one millionth of a meter), and have shown that not only the chemical composition but also the surface architecture of the substrates does indeed affect cell behaviour. We hypothesise that ES cells can be propagated on synthetic substrates that are designed to control interactions with the cells and thus remove the need for animal products in the culture medium. In the project we will study the surface properties of these materials in detail and evaluate the behaviour of ES cells cultured on them in order to select a set of substrates that show potential to control the behaviour of ES cells. We will then thoroughly investigate the surface chemistry and topography of the substrates and the behaviour of the ES cells exposed to them so that we can design specific surfaces to optimize the ES cell response. This will also allow us to build up an understanding of the mechanism regulating ES cell growth and the mechanisms by which the substrate surface properties control their growth. Such information will facilitate the future production of ES cells suitable for clinical use.