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，是由于细胞外基质（ECM）积聚而引起的 慢性肝损伤是全球成人死亡的一个主要原因（每年约 100 万人），原因是治疗不充分 选项。为了解决这一局限性，我们开发了人类肝脏类器官模型，该模型可以实现以下假设： 对治疗肝纤维化的新候选药物进行驱动的机制评估。一种型号是由 改造 iPSC 表达常染色体隐性多囊肾病的常见致病突变 （ARPKD）。 ARPKD 类器官形成肝纤维化的关键特征：它们积累厚的胶原纤维； 并且产生胶原蛋白的肌成纤维细胞显着增加，其转录组谱如下 存在于从患有常见（获得性）肝纤维化（病毒性）的患者获得的肝组织中 诱发性肝硬化和晚期非酒精性脂肪性肝炎 (NASH)。我们还开发了 NASH 类器官 纤维化模型；以及两种用于监测胶原纤维外观的活细胞成像方法和 产生胶原蛋白的细胞。我们假设，由于人类多谱系中出现纤维化， 肝类器官类似于先天性和后天性肝纤维化患者的肝类器官，可用于 推进肝纤维化研究并发现和表征抗纤维化疗法。在目标 1 中，ARPKD 和 NASH 类器官用于开发一个新平台，用于评估 10 种药物的抗纤维化功效，这些药物的抗纤维化功效 作用机制与肝纤维化相关，并确定具有增强抗纤维化作用的药物组合 功效。由于九种药物是 FDA 批准的药物，但目前没有一种用于治疗肝纤维化，这些研究 可能具有重大的翻译重要性。在目标 2 中，这些模型评估了 ECM 的纤维化作用 使用一种新颖的、完全化学定义的生物合成基质来提供线索。人们普遍认为 ECM 的变化可以促进 纤维化重塑。一种新颖的合成化学方案能够调整关键机械（刚度、 粘弹性）和生化（细胞粘附配体特性）基质特性。 ARPKD 和 NASH 类器官 在合成基质中生长将使我们能够检查基质线索对纤维化的影响，这 分析包括单细胞 RNA 测序 (scRNA-Seq)。目标 3：确定常见的致病驱动因素 先天性和后天性肝纤维化共有这些特征，我们将建模方法扩展到 生成并表征 Joubert 综合征相关疾病 (JSRD) 的类器官模型，这是一种多 在某些情况下会导致肝纤维化的系统遗传疾病，我们描述了一种独特的 NASH 类器官 模型。 JSRD 肝病无法在动物中建模。 JSRD 和 NASH 类器官和同基因对照将 使用 scRNA-Seq、高维飞行时间质谱流式细胞仪 (CyTOF) 和两种半靶向技术进行分析 代谢组学方法。 JSRD 类器官还将用于测试 10 种药物的抗纤维化作用（目标 1）和 用于表征 ECM 对纤维化的影响（目标 2）。该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。