Recellularization of Liver Bioscaffolds

描述（由申请人提供）：美国约有3000万人因不同原因患有肝脏疾病，美国每年约有27,000人因肝脏疾病死亡。目前，唯一确定的治疗肝功能衰竭的方法是原位移植。然而，器官严重短缺，每年约有3000个肝脏短缺。类似的数字影响着大多数器官和组织，目前器官等待名单上的总请求数为10万，并且每年以5%的速度增长。考虑到只有处于原始状态的器官才能移植，原位移植将永远是一个有限的资源。一个更优雅、更长远的解决方案是使用干细胞来开发组织工程替代品。然而，虽然许多体外成功已经被证明，临床成功一直非常有限，由于低细胞活力和长期在体内的功能。主要的差距是缺乏一种理想的可移植支架，这种支架具有细胞附着、分化、功能和血管形成所需的所有必要的微观结构和细胞外线索，迄今为止已被证明难以在体外制造。我们的长期目标是设计用于治疗或治疗相关肝脏疾病的可移植肝移植。这项研究的目的是开发功能性和可移植性的肝移植。这里要验证的中心假设是，从废弃肝脏中提取的天然肝支架可以广泛地重新填充，可以为干细胞来源的肝细胞提供足够的成熟环境，并且这些移植物在体内发挥基本的肝脏功能。这项研究的基本原理是，虽然大多数研究都集中在使用合成生物材料从头开始生产理想的支架，但天然ECM可能包含必要的结构和环境线索，因此提出了一种有希望的，很少探索的替代方法来生产器官移植物，可以垂直推进组织工程领域。干细胞来源的肝细胞和肝脏天然基质的功能性肝移植工程是一项创新的努力，因为它有可能成为肝组织工程的新平台。这里描述的工作预计将i)建立脱细胞肝基质作为肝组织工程的可行支架，ii)生成基于肝脏ecm的成熟方案来生成肝细胞样细胞，以及iii)导致一种新的移植物工程方法来提供辅助肝脏支持。虽然这项工作利用肝脏作为模型器官，但这项工作的结果也将产生积极的影响，为未来复杂的器官工程技术奠定基础，这些技术包括几种不同的细胞类型，并可应用于其他器官（胰腺，肾脏等），并可能最终导致体外整个器官的发展。在此开发的ESC成熟协议有望对干细胞工程领域做出重大贡献。在脱细胞基质中培养肝细胞也可能成为药物研究的新平台。公共卫生相关性：美国约有3000万人因不同原因患有肝脏疾病，美国每年约有27,000人因肝脏疾病死亡。目前，唯一确定的治疗肝功能衰竭的方法是原位移植。然而，器官严重短缺，每年约有3000个肝脏短缺。类似的数字影响着大多数器官和组织，目前器官等待名单上的总请求数为10万，并且每年以5%的速度增长。考虑到只有处于原始状态的器官才能移植，原位移植将永远是一个有限的资源。这项研究的结果有望通过开发一种新的方法，利用来自废弃肝脏的干细胞衍生肝细胞和天然基质进行工程辅助肝移植，从而直接改善公众健康，以治疗肝功能衰竭患者。

DESCRIPTION (provided by applicant): About thirty million people in the US undergo a liver disorder for different causes and about 27,000 deaths are registered annually in the US due to liver disease. At this time, the only definitive treatment of hepatic failure is orthotopic transplantation. However, there is a critical shortage of organs, with a deficit of ~3,000 livers per year. Similar numbers affect most organs & tissues, with the total organ waiting list currently at 100,000 requests and the number increasing by 5% every year. Given that only organs in pristine condition are transplantable, orthotopic transplantation will always remain a limited pool. A more elegant, long-term solution is using stem cells to develop tissue-engineered replacements. However, while many in vitro successes have been demonstrated, clinical success has been very limited due to low cell viability and functionality in the long term in vivo. The major gap is the lack of an ideal transplantable scaffold that has all the necessary microstructure and extracellular cues for cell attachment, differentiation, function and vascularization, which has so far proven difficult to manufacture in vitro. Our long-term goal is to engineer transplantable liver grafts for curing or treating relevant liver diseases. The objective of the proposed study is to develop functional and implantable liver grafts. The central hypothesis to be tested here is that the natural liver scaffold derived from discarded livers can be extensively repopulated, can provide an adequate maturation environment for stem cell derived liver cells, and that these grafts perform the essential hepatic functions in vivo. The rationale of the study is that while most research focuses on producing the ideal scaffold from the ground up using synthetic biomaterials, the native ECM is likely to contain the necessary architecture and environmental cues, hence presents a promising, little explored alternative approach for producing organ grafts which can vertically advance the field of tissue engineering. Engineering of functional liver grafts from stem-cell derived hepatocytes and liver's natural matrix is an innovative endeavor, as it has the potential to become a novel platform for hepatic tissue engineering. The work described here is expected to i) establish decellularized liver matrices as a viable scaffold for hepatic tissue engineering, ii) generate a liver ECM-based maturation protocol to generate hepatocyte-like cells, and iii) lead to a novel graft engineering approach to provide auxiliary hepatic support. While this work utilizes liver as the model organ, the results of this work will also have a positive impact by establishing the basis of future sophisticated organ engineering techniques that incorporate several different cell types and can be applied to other organs (pancreas, kidney, etc.), and may ultimately lead to development of entire organs in vitro. The ESC maturation protocol developed here is expected to be a significant contribution to the field of stem cell engineering. Hepatocyte culture in the decellularized matrix may also prove to be a new platform for pharmaceutical studies. PUBLIC HEALTH RELEVANCE: About thirty million people in the US undergo a liver disorder for different causes and about 27,000 deaths are registered annually in the US due to liver disease. At this time, the only definitive treatment of hepatic failure is orthotopic transplantation. However, there is a critical shortage of organs, with a deficit of ~3,000 livers per year. Similar numbers affect most organs & tissues, with the total organ waiting list currently at 100,000 requests and the number increasing by 5% every year. Given that only organs in pristine condition are transplantable, orthotopic transplantation will always remain a limited pool. The results of this study are expected to directly improve public health by the developing a novel approach for engineering auxiliary liver grafts using stem cell derived hepatocytes and native matrices from discarded livers, in order to treat patients with liver failure.