Modeling alcohol toxicity in human hepatocytes

模拟人类肝细胞的酒精毒性

• 批准号: 10442515
• 负责人: Ype Peter De Jong
• 金额: $ 38.14万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-05 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10442515
• 关键词: Alcoholic Hepatitis; Alcoholic Liver Cirrhosis; Alcoholic Liver Diseases; Alcohols; Animals; Biological Models; Cell Transplantation; Cells; Cessation of life; Cirrhosis; Complex; Disease; Engineering; Engraftment; Environmental Risk Factor; Ethanol; Ethanol Metabolism; Ethanol toxicity; Failure; Generations; Genetic; Genetic Predisposition to Disease; Genomics; Goals; Hepatocyte; Human; Human Genetics; Immunodeficient Mouse; Impairment; In Vitro; Individual; Injury; Lentivirus Vector; Liver; Liver diseases; Liver parenchyma; Long-Term Effects; Modeling; Mus; Natural regeneration; Organoids; Outcome Study; Patients; Persons; Phenotype; Pluripotent Stem Cells; Positioning Attribute; Primary carcinoma of the liver cells; Proliferating; Protocols documentation; Research; Rodent; Rodent Model; Role; Source; System; Technology; Testing; Therapeutic Uses; Toxic effect; Transplantation; United States; Variant; advanced system; alcohol effect; alcohol exposure; alcohol misuse; alcohol response; base; design; fatty liver disease; feeding; genetic manipulation; genetic variant; improved; in vivo; induced pluripotent stem cell; insight; liver cell proliferation; liver inflammation; liver injury; mouse model; overexpression; reconstitution; response; species difference; stem cell therapy; tool

Project Summary Alcohol contributes to nearly half the liver related deaths in the United States, mostly from cirrhosis and hepatocellular cancer. Even though large numbers of people misuse alcohol, only a small fraction develops liver disease. A better understanding of the mechanisms that contribute to hepatocyte damage from alcohol is clearly warranted. Mouse models are one tool to further our understanding of the role of alcohol on hepatocytes. However, in part due to species differences and because human genetic predispositions cannot be modeled in mice, ethanol studies in mice have been of limited translational use. An alternative approach to study the effects of alcohol on human hepatocytes are liver chimeric mice. These models are based on the transplantation of primary human hepatocytes into immunodeficient mice with liver injury, after which the human cells proliferate and repopulate the mouse liver parenchyma. There are many limitations with these models, one of them being that only very healthy hepatocytes engraft. Hepatocytes from patients with inflamed livers or cirrhosis generally fail to engraft, which has limited our ability to create disease-specific chimeric models. Possible solutions to this problem would be to genetically manipulate primary hepatocytes or to create chimeric mice with pluripotent stem cells. However, manipulating primary human cells has remained challenging and pluripotent stem cells have largely failed to reconstitute liver chimeric models. We have recently created protocols to efficiently engraft a liver chimeric model with induced pluripotent stem cells or with primary cells that were genetically altered in culture These protocol leads to chimeric animals in which the majority of the liver has been humanized. In addition we can isolate large numbers of human hepatocytes from these mice to create primary human hepatocyte cultures that are stable for months. These advances, combined with a new organoid system, provides us with unique tools to study various hepatocyte responses to alcohol. We here propose to combine these advances to test the effects of alcohol on human hepatocytes. We will use both primary human hepatocytes and pluripotent stem cell-derived hepatocytes to establish systems of alcohol toxicity in hepatocytes, and test a genetic variant that predisposes humans to develop alcoholic hepatitis. In addition the new organoid technology will allow us to test the effects of alcohol on hepatocyte regeneration. At the completion of these studies we will have established systems with which the effects of long- term alcohol toxicity on human hepatocytes can be modeled. These systems will further advance the creation of patient-specific chimeric models for alcoholic liver disease, and may advance the use of stem cell therapies for therapeutic use in patients.

项目摘要 在美国，与肝脏有关的死亡中有近一半是由酒精造成的，主要是死于肝硬变 和肝细胞癌。即使大量的人滥用酒精，只有一小部分人 会患上肝病。更好地了解导致肝细胞损伤的机制 戒酒显然是有根据的。 小鼠模型是进一步了解酒精对肝细胞作用的工具之一。 然而，部分原因是物种的差异，以及人类的遗传倾向无法建模 在小鼠身上，乙醇研究在小鼠身上的翻译用途有限。另一种研究方法 酒精对人肝细胞的影响是肝嵌合小鼠。这些模型基于 原代人肝细胞移植到免疫缺陷小鼠肝损伤后 人类细胞在小鼠肝实质中增殖并重新填充。这些都有很多局限性 模型，其中之一是只有非常健康的肝细胞移植。慢性阻塞性肺疾病患者的肝细胞 发炎的肝脏或肝硬变通常不能移植，这限制了我们创造疾病特异性的能力 嵌合模型。这个问题的可能解决方案是对原代肝细胞进行遗传操作。 或者创造具有多潜能干细胞的嵌合鼠。然而，操纵原代人类细胞已经 仍然具有挑战性和多潜能的干细胞在很大程度上未能重建肝脏嵌合模型。 我们最近已经制定了有效的方案来移植诱导的肝脏嵌合模型 多能干细胞或在培养中基因改变的原代细胞，这些方案导致 大部分肝脏已人性化的嵌合动物。此外，我们还可以分离出大型 这些小鼠的人肝细胞的数量，以建立原代人肝细胞培养，这些细胞是 稳定了几个月。这些进展与一种新的有机系统相结合，为我们提供了独特的工具来 研究各种肝细胞对酒精的反应。 我们建议结合这些进展来测试酒精对人肝细胞的影响。 我们将使用原代人类肝细胞和多能干细胞来源的肝细胞来建立 酒精对肝细胞的毒性系统，并测试一种易于人类患上酒精中毒的基因变异 酒精性肝炎。此外，新的有机化合物技术将允许我们测试酒精对 肝细胞再生。 在这些研究完成后，我们将建立一个系统，通过这些系统，长期- 长期酒精对人肝细胞的毒性可以被模拟。这些系统将进一步推进 酒精性肝病患者特异性嵌合模型的建立，并可能促进干细胞的使用 用于患者治疗的细胞疗法。