Optimization of the engineered 3D hepatic microenvironment enhances pluripotent stem cell derived hepatocyte

项目摘要/摘要 肝病在美国和世界范围内是一个日益严重的临床问题，影响着3000万美国人 导致每年75万人住院，3.6万人死亡。不幸的是，与其他常见疾病不同 例如心血管疾病、肝病及其相关并发症的发病率 越来越多。鉴于肝病患者的稳步上升，对肝移植的需求 在器官供应保持不变的同时继续增加，产生了迫切需要解决的问题 这种器官的匮乏。提出的器官移植的替代方案是细胞移植，而不是人类 肝细胞也同样很难找到来源。因此，目前还没有其他方法可以获得健壮性 功能性肝细胞正在显著阻碍科学研究和更有效的发展 临床疗法；我们的工作解决了这一重大研究和临床差距。多能干细胞来源 肝细胞样细胞经常被认为是器官移植的潜在替代细胞来源，并被认为是 药学和科研平台。然而，目前无法衍生出稳健和完全成熟的 肝细胞限制了目前生产的细胞的临床和科学用途。多能干细胞的命运和 肝细胞的成熟由复杂的内部信号和外部信号阵列决定 微环境。不幸的是，我们目前的分化平台不能模仿细胞、细胞外 基质(ECM)，体内肝脏微环境的3D复杂性以及ECM细胞的作用 而分化过程中的细胞间相互作用在很大程度上仍不清楚。我们的中心假设是 3D微环境和随后的细胞相互作用(例如细胞-ECM和细胞-细胞相互作用)决定了 细胞命运决定，正是通过这些因素，胎儿细胞类型成熟到终末分化 单元类型。在最近的工作中，我们小组开发了强大的工具来系统和高效地操作 三维微环境。这项提议的中心目标是识别和检查重要的诱导细胞 定向多能干细胞在体外分化为表型的相互作用 和功能成熟的肝细胞。我们将系统地检查和鉴定细胞-细胞和细胞- 细胞外基质的相互作用在调节胎儿到成人肝细胞的转变中发挥作用，并将结合这些发现 到我们目前的分化和组织工程构建中。这些优化的3D多细胞球体 平台将使科学研究和开发更有效的临床疗法成为可能。

Project Summary/Abstract Liver disease is a growing clinical problem in the United States and worldwide affecting 30 million Americans resulting in 750,000 hospitalizations and 36,000 deaths yearly. Unfortunately, unlike other common diseases such as cardiovascular diseases, the incidence of liver disease and its associated complications are increasing. Given the steady rise of patients with liver disease, the demand for liver transplantation has continued to increase while the supply of organs has remained unchanged creating a pressing need to address this organ scarcity. Proposed alternatives to organ transplantation are the transplantation of cells but human hepatocytes are similarly difficult to source. As a result, there are currently no alternatives for obtaining robust functional hepatocytes which is significantly hampering scientific study and the development of more effective clinical therapies; our work addresses this significant research and clinical gap. Pluripotent stem cell derived hepatocyte-like cells are often cited as a potential alternative cell source for organ transplants and as a platform for pharmaceutical and scientific study. However, the current inability to derive robust and fully mature hepatocytes limits the clinical and scientific utility of currently produced cells. Pluripotent stem cell fate and hepatocyte maturation is determined by the complex array of internal signals and external cues from the microenvironment. Unfortunately our current differentiation platforms do not mimic the cellular, extracellular matrix (ECM), and 3D complexities of the in vivo hepatic microenvironment and therefore the role of ECM-cell and cell-cell interactions in the differentiation process remains largely unknown. Our central hypothesis is that the 3D microenvironment and subsequent cellular interactions (e.g. cell-ECM and cell-cell interactions) dictates cell fate decisions and it is through these factors by which fetal cell types mature into terminally differentiated cell types. In recent work our group has developed robust tools to systematically and efficiently manipulate the 3D microenvironment. The central goal of this proposal is to identify and examine important inductive cellular interactions for directed pluripotent stem cell differentiation in vitro to enable the generation of phenotypically and functionally mature hepatocytes. We will systematically examine and identify the role cell-cell and cell- ECM interactions play in regulating the fetal to adult hepatocyte transition and will incorporate these findings into our current differentiation and tissue engineered constructs. These optimized 3D multicellular spheroids platforms will enable scientific study and the development of more effective clinical therapies.