A Model for Human Liver Fibrosis

人类肝纤维化模型

• 批准号: 10685178
• 负责人: GARY A PELTZ
• 金额: $ 77.23万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10685178
• 关键词: Address; Adhesives; Adult; Animals; Appearance; Autosomal Recessive Polycystic Kidney; Biochemical; Cause of Death; Cells; Chemicals; Cirrhosis; Collagen; Collagen Fiber; Cues; Cytometry; Data; Development; Disease; Drug Combinations; Engineering; Etiology; Evaluation; Extracellular Matrix; FDA approved; Fibrosis; Genes; Genetic; Genetic Diseases; Genomics; Hepatic; Human; Joubert syndrome; Ligands; Liver; Liver Fibrosis; Liver diseases; Mechanics; Mediating; Methods; Microscopy; Modeling; Monitor; Mutation; Myofibroblast; Nonesterified Fatty Acids; Organoids; Outcome; Pathogenesis; Pathologic; Pathway interactions; Patients; Pharmaceutical Preparations; Production; Property; Research; Scheme; Synthesis Chemistry; System; Testing; Therapeutic; Thick; Time; Tissues; Viral; antifibrotic treatment; base; causal variant; cell type; chronic liver disease; chronic liver injury; drug candidate; high dimensionality; human stem cells; imaging modality; induced pluripotent stem cell; insight; live cell imaging; metabolomics; nonalcoholic steatohepatitis; novel; novel therapeutics; response; single-cell RNA sequencing; transcriptomics; viscoelasticity

Abstract Liver fibrosis is a pathological condition that results from extracellular matrix (ECM) accumulation in response to chronic liver injury and is a major global cause of death in adults (~1M per year) due to inadequate therapeutic options. To address this limitation, we have developed human hepatic organoid models that enable hypothesis- driven, mechanistic evaluation of novel drug candidates for treatment of liver fibrosis. One model is produced by engineering iPSC to express a common causative mutation for Autosomal Recessive Polycystic Kidney Disease (ARPKD). ARPKD organoids develop the key hallmarks of hepatic fibrosis: they accumulate thick collagen fibers; and have a marked increase in collagen-producing myofibroblasts whose transcriptomic profile is like those present in liver tissues obtained from patients with commonly occurring (acquired) forms of liver fibrosis (viral- induced cirrhosis and advanced non-alcoholic steatohepatitis, NASH). We also developed a NASH organoid fibrosis model; along with two live cell imaging methods for monitoring for the appearance of collagen fibers and collagen producing cells. We hypothesize that since the fibrosis that develops in this human, multi-lineage, hepatic organoid resembles that in patients with congenital and acquired forms of liver fibrosis, it can be used to advance liver fibrosis research and to discover and characterize anti-fibrotic therapies. In Aim 1, ARPKD and NASH organoids are used to develop a novel platform for assessing the anti-fibrotic efficacy of 10 agents whose mechanism of action is relevant to liver fibrosis, and to identify drug combinations with increased anti-fibrotic efficacy. Since nine are FDA-approved drugs, but none are currently used to treat liver fibrosis, these studies could have significant translational importance. In Aim 2, these models evaluate the fibrogenic effect of ECM cues using a novel, fully chemically defined, biosynthetic matrix. ECM changes are widely thought to promote fibrotic remodeling. A novel, synthetic chemistry scheme enables tuning of the key mechanical (stiffness, viscoelasticity) and biochemical (cell-adhesive ligand identity) matrix properties. ARPKD and NASH organoids grown in synthetic matrices will enable us to examine the effects that matrix cues have on fibrosis, and this analysis includes single cell RNA sequencing (scRNA-Seq). In Aim 3, to identify common pathogenetic drivers that are shared among congenital and acquired forms of liver fibrosis, we extend our modeling approach to generate and characterize organoid models for Joubert Syndrome Related Disorder (JSRD), which is a multi- system genetic disease that causes liver fibrosis in some cases, and we characterize a unique NASH organoid model. JSRD liver disease cannot be modeled in animals. JSRD and NASH organoids and isogenic controls will be analyzed using scRNA-Seq, high-dimensional time of flight mass cytometry (CyTOF) and two semi-targeted metabolomic methods. JSRD organoids will also be used to test the anti-fibrotic effects of 10 drugs (Aim 1) and for characterizing ECM effects on fibrosis (Aim 2). This data will provide important information about shared mechanisms that mediate liver fibrosis of different etiologies.

抽象的 肝纤维化是一种病理状况，是由细胞外基质（ECM）累积响应的病理状况 由于治疗不足 选项。为了解决这一局限性，我们开发了人类肝癌模型，以实现假设 - 新型药物候选物治疗肝纤维化的驱动，机械评估。一种模型由 工程IPSC表达常染色体隐性多囊性肾脏疾病的常见原因突变 （ARPKD）。 ARPKD类器官发展了肝纤维化的关键标志：它们积累了厚厚的胶原蛋白纤维； 并在产生胶原蛋白的成肌纤维细胞中显着增加，其转录组轮廓就像那样 存在于从常见发生（获得）形式的肝纤维化（病毒 - 病毒）患者中获得的肝组织 诱发肝硬化和晚期非酒精性脂肪性肝炎，纳什）。我们还开发了纳什的类器官 纤维化模型；以及两种活细胞成像方法，用于监测胶原纤维的外观和 胶原蛋白产生细胞。我们假设，自从这种人类的多条道中发展的纤维化， 肝癌类似于先天性和获得形式的肝纤维化的患者，可以用来用于 预先肝纤维化研究并发现和表征抗纤维化疗法。在AIM 1，ARPKD和 NASH类器官用于开发一个新型平台，用于评估10种药物的抗纤维化功效 作用机制与肝纤维化有关，并鉴定抗纤维化增加的药物组合 功效。由于九种是FDA批准的药物，但目前没有用于治疗肝纤维化，因此这些研究 可能具有重大的翻译重要性。在AIM 2中，这些模型评估了ECM的纤维化作用 使用一种新颖的化学定义，生物合成基质的提示。 ECM的变化被广泛认为促进 纤维化重塑。一种新颖的合成化学方案，可以调整关键机械（刚度， 粘弹性）和生化（细胞粘合剂配体身份）基质特性。 ARPKD和NASH类器官 在合成矩阵中生长将使我们能够检查基质提示对纤维化的影响，这 分析包括单细胞RNA测序（SCRNA-SEQ）。在AIM 3中，以识别常见的致病驱动器 在先天性和获得形式的肝纤维化中共享，我们将建模方法扩展到 生成和表征乔伯特综合征相关疾病（JSRD）的器官模型，这是一种多 在某些情况下会引起肝纤维化的系统遗传疾病，我们表征了独特的NASH类器官 模型。 JSRD肝病不能在动物中建模。 JSRD和NASH类器官以及等源控制将 使用SCRNA-SEQ，飞行质量细胞仪（Cytof）的高维时间和两个半目标进行分析 代谢组方法。 JSRD类器官也将用于测试10种药物的抗纤维化作用（AIM 1）和 为了表征ECM对纤维化的影响（AIM 2）。这些数据将提供有关共享的重要信息 介导不同病因的肝纤维化的机制。