Developing a renewable and dissectible human liver for the study of HBV/HCV infection

开发可再生、可解剖的人类肝脏用于研究 HBV/HCV 感染

• 批准号: 10686216
• 负责人: Xianfang Wu
• 金额: $ 48.3万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-18 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10686216
• 关键词: 3-Dimensional; Acceleration; Acute Liver Failure; Address; Affect; Age; Antiviral Agents; Area; CRISPR/Cas technology; Cell Communication; Cell Culture Techniques; Cell Differentiation process; Cell Line; Cells; Cellularity; Chronic Hepatitis B; Cirrhosis; Clinical; Coculture Techniques; Complex; DNA sequencing; Development; Diagnostic; Disease; Disease Progression; Endothelial Cells; Epidemiology; Fibrosis; Foundations; Future; Genes; Genetic; Genetic Variation; Goals; Hepatic; Hepatic Stellate Cell; Hepatitis A; Hepatitis B; Hepatitis B Infection; Hepatitis B Therapy; Hepatitis B Vaccines; Hepatitis B Virus; Hepatitis C; Hepatitis C Therapy; Hepatitis C co-infection; Hepatitis C virus; Hepatitis D; Hepatitis E virus; Hepatitis Viruses; Hepatocyte; Heterogeneity; Human; IL18 gene; In Vitro; Individual; Infection; Kupffer Cells; Link; Liver; Liver diseases; Macrophage; Mediating; Methods; Modeling; Molecular; Natural Immunity; Outcome; Patients; Persons; Population; Primary carcinoma of the liver cells; Reporting; Research; Risk; Sampling; Severities; Single Nucleotide Polymorphism; System; Technology; Time; Tissue Engineering; Translating; Treatment outcome; United States; Viral; Viremia; Virus; Work; aging population; career; cell type; chronic infection; chronic liver disease; co-infection; disease phenotype; epigenomics; genetic variant; genome wide association study; hemodynamics; high risk; human model; human pluripotent stem cell; improved; in vivo; individual variation; infection risk; innovation; insight; liver transplantation; mortality; novel; post-doctoral training; response; stem; stem cells; three dimensional cell culture; tool; transcriptomics; transmission process

Project Summary HBV and HCV infections are among the leading causes of chronic liver disease worldwide. In the United States, it is estimated that over 850 thousand people are currently infected with HBV and more than 2.4 million for HCV. Despite the availability of highly effective HBV vaccine and HCV treatment, the mortality and the burden associated with HBV and HCV are nevertheless increasing as individuals with existing infections advance into more advanced stages including fibrosis, cirrhosis, and HCC. Moreover, HCV-infected patients, even though cured of viremia, remain at a significantly elevated risk for advanced liver diseases. Due to their shared modes of transmission and epidemiological features, HBV and HCV frequently coexist in patients in endemic areas or among subjects at high risk of infection. Coinfection is usually more complex than monoinfection, leading to an accelerated disease progression and complicated viral interaction for their treatment. Despite the severity of HBV and HCV infections, the mechanisms by which they lead to liver disease, of how coinfection causes an increased severity and risk for complications, and of how HBV and HCV interact in the liver of coinfected patients that may lead to reactivation of HBV upon the cure of HCV remains largely unclear. This stems, in part, from the lack of relevant human models that recapitulate key disease phenotypes and permit detailed mechanistic studies that could answer these questions. The primary research goal of this proposal is to establish a novel, high fidelity multicellular in vitro liver model to address the aforementioned gaps. In this system, we plan to coculture human pluripotent stem cells (hPSCs)-derived hepatocytes, hepatic stellate cells, macrophages, and endothelial cells. Unlike existing traditional cell culture models, we propose to create an innovative three dimensional platform that mimics the liver’s multicellularity and microenvironment, by culturing these four cell types in a configuration that resembles the liver’s in vivo spatial organization, its hemodynamics, and its cell-cell interaction. To distinguish cell type- specific responses, we will develop methods to purify individual cell types for downstream analysis. With this multicellular culture platform, we will pursue our long-range objectives, across which we capitalize on the unique features of this novel platform to address questions that cannot be adequately answered with any existing in vitro human-relevant system. These include the mechanisms of continued disease progression following the cure of HCV infection, an accelerated disease progression with HBV/HCV coinfection, and the interaction between HBV/HCV that leads to HBV reactivation following the cure of HCV infection (Aim 1). In Aim 2, we will combine hPSCs with gene editing to understand how genetic variants affect the course of disease and treatment following HBV and HCV infection. These studies will not only provide new insights that could inform development of effective diagnostics and treatments for HBV/HCV infection, but also lead to technology advance that open up new avenues for future studies focusing on other liver diseases.

项目摘要 乙肝病毒和丙型肝炎病毒感染是全球慢性肝病的主要原因之一。在美国， 据估计，目前有超过85万人感染了乙肝病毒，超过240万人感染了丙型肝炎病毒。 尽管有高效的乙肝疫苗和丙型肝炎治疗，但死亡率和负担 与乙肝病毒和丙型肝炎病毒相关的人仍在增加，因为现有感染的人进入 更晚期，包括纤维化、肝硬变和肝细胞癌。此外，感染丙型肝炎病毒的患者，即使 病毒血症治愈后，罹患晚期肝病的风险仍显著增加。由于它们的共享模式 在传播和流行病学特征方面，在流行区或高发区的患者中经常同时存在乙肝病毒和丙型肝炎 在感染高危人群中。混合感染通常比单一感染更复杂，导致 加速疾病进展和复杂的病毒相互作用为他们的治疗。 尽管乙肝病毒和丙型肝炎病毒感染的严重性，但它们导致肝脏疾病的机制 合并感染如何导致严重程度和并发症风险增加，以及乙肝病毒和丙型肝炎病毒如何在 在治愈丙型肝炎病毒后，混合感染患者的肝脏可能会导致乙肝病毒的重新激活，目前仍不清楚。 这在一定程度上源于缺乏相关的人类模型来概括关键的疾病表型并允许 可以回答这些问题的详细的机械研究。 本方案的主要研究目标是建立一种新型的、高保真的体外多细胞肝脏。 模型，以解决上述差距。在这个系统中，我们计划共同培养人类多能干细胞 (HPSCs)来源的肝细胞、肝星状细胞、巨噬细胞和内皮细胞。与现有的不同 传统的细胞培养模型，我们建议创建一个创新的三维平台，模仿 肝脏的多细胞和微环境，通过将这四种细胞培养成类似于 肝脏的活体空间组织、血流动力学和细胞与细胞的相互作用。为了区分细胞类型- 对于特定的反应，我们将开发方法来纯化单个细胞类型，以便进行下游分析。 有了这个多细胞培养平台，我们将追求我们的长远目标，我们将通过这些目标来实现 关于这个新颖平台的独特功能，以解决任何无法充分回答的问题 存在与人体有关的体外系统。其中包括疾病持续发展的机制。 随着丙型肝炎病毒感染的治愈，乙肝病毒/丙型肝炎病毒混合感染的疾病进展加快， 在治愈丙型肝炎病毒感染后，导致乙肝病毒重新激活的乙肝病毒/丙型肝炎病毒之间的相互作用(目标1)。在AIM 2，我们将把hPSCs与基因编辑结合起来，以了解基因变异如何影响疾病的进程和 乙肝病毒和丙型肝炎病毒感染后的治疗。这些研究不仅将提供新的见解，可能会 开发有效的乙肝病毒/丙型肝炎病毒感染的诊断和治疗方法，但也导致技术进步 这为未来专注于其他肝病的研究开辟了新的途径。