Mini-Livers Derived from Human IPS Cells for Modeling Steatosis and Therapy

源自人 IPS 细胞的微型肝脏用于脂肪变性建模和治疗

• 批准号: 10297425
• 负责人: Deepak Nagrath
• 金额: $ 53.63万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10297425
• 关键词: Adult; Alcoholic Liver Cirrhosis; Animals; Cell Communication; Cell Culture Techniques; Cell model; Cells; Cessation of life; Chronic Disease; Cirrhosis; Clinical; Complex; Cues; Data; Deposition; Detection; Development; Diabetes Mellitus; Disease; Engineering; Environment; Epidemic; Epigenetic Process; Evolution; Excision; Fatty Liver; Fatty acid glycerol esters; Fibroblasts; Fibrosis; Funding; Future; Genetic; Genetic Diseases; Genetic Polymorphism; Goals; Hepatic Stellate Cell; Hepatocyte; Heterogeneity; High Prevalence; Histologic; Human; Human Genetics; In Vitro; Inflammatory; Isotopes; Link; Liver; Liver Dysfunction; Liver Fibrosis; Liver diseases; Mediating; Mesenchymal; Metabolic; Metabolic Diseases; Metabolic Pathway; Metabolism; Modeling; Molecular; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Organ; Organ Culture Techniques; Organ Model; Outcome; Pathogenesis; Pathologic; Patients; Pharmacotherapy; Phenotype; Population; Preventive; Primary carcinoma of the liver cells; Process; Public Health; Rattus; Resources; Role; SIRT1 gene; Single Nucleotide Polymorphism; Source; System; Techniques; Technology; Testing; Therapeutic; Therapeutic Intervention; Time; Tissue Model; Tissues; Variant; Work; alternative treatment; base; cell type; chronic liver disease; comorbidity; engineered stem cells; fatty acid biosynthesis; genetic variant; high risk; histone modification; human disease; improved; induced pluripotent stem cell; knock-down; lifestyle factors; lipidomics; liver metabolism; macrophage; metabolic profile; metabolomics; non-alcoholic fatty liver disease; nonalcoholic steatohepatitis; novel strategies; prevent; response; small hairpin RNA; stem; stem cell model; stressor; success

ABSTRACT/SUMMARY Our long-term goal remains the same and targets to develop human liver tissue model from stem cells, with multi-cellular cues, metabolic functionality, that incorporates dynamic genetic and epigenetic factors and encompasses the spectrum and evolution of human NAFLD to uncover disease mechanism and therapeutics. The objectives of the proposed study are to continue developing human fatty livers engineered with iPS cells, that incorporates the dynamic expression of the epigenetic-related histone modification SIRT1 in different genetic polymorphisms and elucidate their role as metabolic regulators in the development of human fatty liver disease. The central hypothesis to be tested here is that genetic polymorphisms especially the NAFLD-high- risk PNPLA3 rs738409, combined with epigenetic-related changes (SIRT1) can control hepatic metabolism and contribute to the development of human fatty liver. The rationale is that the ability to generate human diseased liver tissue using genetically modifiable iPS cells from different human populations with single- nucleotide-polymorphisms is a powerful resource, that now make it possible, for the first time, to functionally interrogate their role in disease. Importantly, based on our strong preliminary data of targeted metabolomic analysis in isolated human hepatocytes obtained from either liver resections “Normal Human Hepatocytes” or hepatocytes isolated from explanted NASH livers “NASH Human Hepatocytes” carrying either the NAFLD-high risk PNPLA3 I148M genetic variant or wild type we hypothesize that PNPLA3 I148M hepatocytes have high levels of ferroptosis thereby leading to low metabolic capacity to adapt to stressors and continuous loss of hepatocytes. In the newly proposed aim3, we aim to uncover ferroptosis based therapeutic strategies for patients with NAFLD-high risk PNPLA3 I148M genetic variants. The work described here is expected to i) generate human iPS cells carrying shRNA mediated conditional knockdown of SIRT1 in different genetic backgrounds (especially the NAFLD-high-risk PNPLA3 rs738409 variant), ii) develop a novel approach for modeling an organ-like environment to determine the role of the epigenetic related factor SIRT1 and the genetic variants in the development of fibrotic human mini-livers with fatty liver disease and iii) determine the role of genetic polymorphisms in regulating functional metabolism in mini-human iPS-derived liver tissue with fatty liver disease with especial focus on ferroptosis. Understanding this process alone or in relation to epigenetic changes in a human liver tissue model can result in preventive therapeutic strategies for patients with NAFLD-high risk PNPLA3 I148M genetic variants. The results of this work will also have a positive impact by establishing the basis and platform for future sophisticated organ engineering techniques that incorporates several different cell types from the same iPS cell source, these techniques could be applied to study other liver diseases (e.g. metabolic diseases) and is expected to be a major contribution to the fields of stem cells engineering and liver steatosis.

摘要/总结 我们的长期目标保持不变，目标是从干细胞中开发人类肝脏组织模型， 多细胞线索，代谢功能，它结合了动态遗传和表观遗传因素， 涵盖了人类NAFLD的谱和演变，以揭示疾病机制和治疗方法。 这项研究的目的是继续开发用iPS细胞改造的人类脂肪肝， 它结合了表观遗传相关的组蛋白修饰SIRT 1在不同细胞中的动态表达， 基因多态性，并阐明其作为代谢调节剂在人类脂肪肝发生中的作用 疾病这里要检验的中心假设是，遗传多态性，特别是NAFLD高- 风险PNPLA 3 rs738409，结合表观遗传相关变化（SIRT 1）可以控制肝脏代谢 有助于人类脂肪肝的发展。其基本原理是， 使用来自不同人群的可遗传修饰的iPS细胞， 核苷酸多态性是一个强大的资源，现在有可能，第一次，在功能上， 探究它们在疾病中的作用。重要的是，基于我们强有力的靶向代谢组学的初步数据， 在从肝切除“正常人肝细胞”或 从携带NAFLD高表达的NASH肝脏分离的肝细胞“NASH人肝细胞” 风险PNPLA 3 I148 M遗传变异体或野生型，我们假设PNPLA 3 I148 M肝细胞具有高风险， 铁凋亡水平，从而导致低代谢能力，以适应压力和持续损失的 肝细胞在新提出的aim 3中，我们的目标是揭示基于铁凋亡的治疗策略， 患有NAFLD-高风险PNPLA 3 I148 M遗传变异的患者。 本文所述的工作预期i）产生携带shRNA介导的条件性条件诱导分化的人iPS细胞， 在不同遗传背景中SIRT 1的敲低（特别是NAFLD-高风险PNPLA 3 rs738409 变体），ii）开发一种用于模拟器官样环境的新方法以确定 表观遗传相关因子SIRT 1和遗传变异在人小肝纤维化发展中的作用 脂肪肝疾病和iii）确定遗传多态性在调节功能代谢中的作用， 患有脂肪肝疾病的迷你人iPS衍生肝组织，特别关注铁凋亡。理解 该过程单独或与人肝组织模型中的表观遗传变化相关可导致预防性的肝损伤 针对具有NAFLD-高风险PNPLA 3 I148 M遗传变异的患者的治疗策略。的结果 这项工作还将产生积极影响，为未来复杂的机构建立基础和平台， 工程技术，包括几种不同的细胞类型，从同一个iPS细胞来源，这些 技术可以应用于研究其他肝脏疾病（例如代谢性疾病），并有望成为一种 在干细胞工程和肝脂肪变性领域的主要贡献。