Strategies for the development and maturation of functional hepatocytes from hES cells

每年，开发新药的公司都要花费数百万英镑进行测试，以确保他们的新药不仅有效，而且对我们来说也是安全的。尽管如此，新药最大的问题是，有时它们会对肝脏、心脏或大脑造成损害，导致它们被下架。如果相关公司有办法(在早期阶段)使用与正常人体肝脏、心脏和大脑中发现的细胞更接近的细胞来测试新药的安全性，这种情况可能会得到极大改善。肝移植是治疗危及生命的肝病和疾病的有效方法，但捐献器官的供应严重有限，这意味着每年仍有许多人在等待名单上。高效制造功能性肝细胞的新方法将导致这两个领域的改善。许多研究人员，包括我们自己的团队，已经证明有可能将称为干细胞的非专门化细胞转化为肝细胞，更具体地说，是称为肝细胞的细胞。正是我们肝脏中的肝细胞负责在我们一生中清除血液中的有害物质。目前，我们可以从干细胞产生的肝细胞样细胞不如正常的肝细胞，缺乏执行许多肝脏功能的能力。此外，目前使用的将干细胞转化为肝细胞的方法非常昂贵，需要大约4周的时间。在这项提案中，我们希望创造条件，使我们能够更高效、更廉价地将干细胞转化为肝细胞。我们还想测试如何使肝细胞样细胞更接近于在人类中发现的正常成人肝细胞。为了帮助我们做到这一点，我们将制造干细胞，当它们从干细胞发展到肝细胞时，在特定的阶段打开荧光信号(我们称之为记者)。其中一名记者将在大约一周后的部分过程中切换，并将帮助我们改进在肝脏发育早期形成的细胞的生成，我们还希望发现如何增加这些前体细胞的数量。我们还将制造第二个报告干细胞系，只有当细胞发育成成熟的肝细胞时才会打开。我们将使用这一干细胞系来帮助我们解决将干细胞转化为肝细胞样细胞的条件，这些细胞尽可能接近正常的成人肝细胞。我们将通过检查其特性来检查我们产生的肝细胞是否像正常的成人肝细胞。我们将确定它们是否含有仅在肝细胞中发现的蛋白质，我们将研究它们是否含有对药物代谢至关重要的酶，因为这些酶对正常的肝细胞功能非常重要。我们希望我们的结果将定义条件：(I)提高我们将干细胞转化为肝细胞的能力，(Ii)帮助我们扩大我们可以制造的肝脏前体细胞的数量，(Iii)使我们能够产生更像正常成人肝细胞的肝细胞样细胞，以及(Iv)改进将干细胞转化为肝细胞的效率和方法，从而使它们的制造成本更低。从可再生干细胞来源获得成熟的肝细胞样细胞对制造新药的公司将非常有价值，因为这将帮助它们开发对患者应该更安全的药物，并且开发这些药物所需的时间将减少，从而节省资金。干细胞来源的肝细胞不仅对测试新药的安全性有用，而且在未来它们作为移植细胞来源也可能很有价值。因此，我们发现的在生成功能性肝细胞样细胞方面的改进也将有助于科学家和医生开发细胞移植策略和人造肝脏，作为需要肝移植的患者的替代治疗方法。

Each year companies that develop new medicines spend millions of pounds on tests that are meant to make sure their new drugs are not only effective, but also safe for us to take. In spite of this, the single biggest problem with new medicines is that sometimes they cause damage to the liver, heart or brain, which results in them being taken off the market. This situation could be greatly improved if the companies involved had ways of testing (at an early stage) the safety of new medicines using cells that more closely match those found in normal human liver, heart and brain. Liver transplantation is an effective treatment for patients suffering from life-threatening liver diseases and disorders but the supply of donor organs is severely limited meaning that many people remain on the waiting list each year. New ways to efficiently make functional liver cells would lead to improvements in both of these areas. Many researchers, including our own groups, have shown that it is possible to turn unspecialised cells called stem cells into liver cells, more specifically cells called hepatocytes. It is the hepatocytes in our livers that are responsible for removing harmful products from our blood throughout our lives. At the moment, the hepatocyte-like cells we can generate from stem cells are not as good as proper liver cells and lack the ability to carry out many liver functions. Also, the methods currently used to turn stem cells into liver cells are very expensive and take approximately 4 weeks. In this proposal we want to develop conditions that allow us to turn stem cells into liver cells more efficiently and cheaply. We also want to test for ways in which we can make hepatocyte-like cells that more closely resemble the normal adult liver cells found in humans. To help us do this we will make stem cells that turn on fluorescent signals (which we call reporters) at particular stages as they progress from stem to liver cells. One of these 'reporters' will switch on part way through the process, after about a week, and will help us to improve the generation of cells formed during the early stages of liver development and we also hope to discover how we can increase the numbers of these precursor cells. We will also make a second reporter stem cell line will switch on only when the cells have developed into mature liver cells. We will use this stem cell line to help us work out those conditions that turn stem cells into hepatocyte-like cells that are as close as possible to proper adult liver cells. We will examine whether the liver cells we generate are like normal adult liver cells by examining their properties. We will determine if they contain proteins only found in liver cells and we will study if they contain the enzymes important for metabolising medicines, since these are very important for proper liver cell function. We hope that our results will define conditions that: (i) improve our ability to turn stem cells into liver cells, (ii) help us to expand the number of liver precursor cells we can make, (iii) enable us to generate hepatocyte-like cells that are more like proper adult liver cells and (iv) improve the efficiency and methods of turning stem cells into liver cells so that they are cheaper to make. The availability of mature hepatocyte-like cells from a renewable source of stem cells will be extremely valuable to companies making new medicines, since it will help them develop drugs that should be safer for patients and the time taken to develop them will be reduced, saving money. Not only would stem cell-derived liver cells useful for testing the safety of new medicines, but in the future they may also be valuable as a source of cells for transplantation. Therefore, the improvements in generating functional hepatocyte-like cells that we discover will also help scientists and doctors who are developing cell transplant strategies and artificial livers as alternative treatments for patients requiring liver transplants.