High Yield Human Hepatocyte Isolation

描述（由申请人提供）：美国约有30，000人死于终末期肝脏疾病。许多患者病情严重，无法耐受肝移植，即使移植 据指出，有活力的捐赠器官严重短缺，只有28%的等待名单上的病人接受移植。由于缺乏可靠的人肝细胞来源，生物人工肝（BAL）装置、细胞移植和组织工程肝替代品等器官移植新替代品的开发和转化研究受到严重阻碍。然而，大量未使用的人类细胞存在于废弃的供体肝脏中，例如心脏死亡后的供体（DCD），由于热缺血损伤而不能移植，这显著降低了移植成功率。最近的研究，包括我们小组的研究，表明缺血和其他边缘肝脏可以通过体外机器灌注（MP）进行修复。这种轻微受损的肝脏可以被挽救和修复用于移植的估计数量为~ 6，000;更多的肝脏可能受损太严重而不能移植，但仍然可以作为细胞来源。人类细胞的这一巨大来源尚未被探索。我们的长期目标是开发机器灌注作为治疗目前未使用的供体肝脏的一种方法，并增加供体肝脏的可用性， 移植、细胞疗法和研究。本STTR验证研究的目的是确定因过度缺血而不适合移植的人肝脏作为高质量原代肝细胞的可靠来源。这里要测试的中心假设是机器灌注预处理增加了从缺血人类肝脏中恢复肝细胞的产量。从不合格的人类供体肝脏中提高肝细胞的活产率是一项创新性的奋进，因为它利用了我们小组开发的新机器灌注方法，并将大鼠模型中的肝细胞产率提高了25倍。鉴定可靠的人肝细胞来源将通过允许在早期和所有水平的药物测试中使用人肝细胞来显著改变药物发现和测试的实践，减少对毒理学动物模型的依赖，从而降低药物开发的成本。以低成本获得高质量的人肝细胞也将消除实现基于细胞的肝衰竭疗法的主要瓶颈。

DESCRIPTION (provided by applicant): End-stage liver disease claims about 30,000 lives in the US. Many patients become too ill to tolerate liver transplantation, and even if transplantation were indicated, there is a severe shortage of viable donor organs and only 28% of the wait-listed patients receive transplants. Development and translational studies with novel alternatives to organ transplantation, such as Bioartificial Liver (BAL) devices, cell transplantation and tissu engineered liver substitutes are severely bottlenecked by the lack of a reliable source of viable human hepatocytes. However, a vast pool of unused human cells lies in the discarded donor livers, such as Donors after Cardiac Death (DCD), which are not transplanted due to warm ischemic injury, which significantly reduces transplant success. Recent studies, including those from our group, demonstrate that ischemic and other marginal livers can be reconditioned through extracorporeal Machine Perfusion (MP). The estimated number of such marginally damaged livers that could be rescued and reconditioned for transplantation is ~ 6,000; many more are likely to be too damaged for transplantation, but still viable as a cell source. This immense source of human cells remains unexplored. Our long-term goal is to develop machine perfusion as a way to treat currently unused donor livers and increase donor liver availability for transplantation, cell-based therapeutics and research. The objective of this STTR validation study is to establish human livers disqualified for transplantation because of excessive ischemia, as a reliable source of high quality primary hepatocytes. The central hypothesis to be tested here is that machine perfusion pre-treatment increases the yield of hepatocyte recovery from ischemic human livers. Enhancing the viable yield of hepatocytes from disqualified human donor livers is an innovative endeavor as it capitalizes on the new machine perfusion method developed by our group and increases hepatocyte yield by 25 fold in the rat model. Identifying a reliable human hepatocyte source would dramatically alter the practice in drug discovery and testing by allowing use of human hepatocytes early and at all levels of pharmaceutical testing, reduce reliance on animal models for toxicology, and therefore lead to reduced cost for drug development. Availability of high quality human hepatocytes for a low cost would also eliminate a major bottleneck in realization of cell-based therapies for liver failure.