Hepatitis C virus host interactions in micropatterened hepatocyte co-cultures

描述（由申请人提供）：丙型肝炎病毒（HCV）是全球主要的公共卫生问题，估计影响1.2 - 1.7亿人。不幸的是，目前还没有有效的丙肝病毒疫苗，目前的治疗方法是有毒的，效果不一。虽然对HCV的发病机制已经了解了很多，但影响自发清除或持续感染的病毒和宿主因素仍然未知。缺乏可靠的体外和体内模型，忠实地模拟人类自然丙型肝炎病毒感染，继续阻碍丙型肝炎病毒发病机制的研究。尽管肝细胞癌衍生细胞系和永生化肝细胞使我们能够研究HCV的复制，但与原代肝细胞相比，它们在表型和功能上都是异常的。虽然原代肝细胞培养被认为是研究HCV感染和病毒/宿主相互作用的理想模型，但它们的肝脏特异性功能在常规培养系统中迅速下降。最近，肝细胞和成纤维细胞的生物工程微模式共培养（mpcc）的创建允许肝细胞长期培养，保留正常细胞功能（白蛋白生成，尿素合成等）。我们研究的长期目标是更好地了解HCV的发病机制，以便设计特异性和有效的治疗方法。我们假设早期病毒和宿主相互作用影响疾病的结果，为自发清除或慢性感染/肝纤维化奠定了基础。为了深入了解宿主对肝脏急性HCV感染的早期反应，我们建议表征HCV病毒/宿主在原发性人肝细胞mpcc和细胞复杂性增加的mpcc中的相互作用。在特定目标1下，我们将对mpcc中感染和未感染的细胞进行不同表型分析，同时进行基因组和蛋白质组学分析，以确定HCV感染中调节的宿主基因和途径。在特定目标2中，我们将增加mpcc的细胞复杂性，以包括非实质细胞，以评估肝脏中的支持细胞类型是否有助于HCV发病和/或纤维化状态的发展。在特定目标3中，我们将增加mpcc的细胞复杂性，以包括HCV感染期间驻留或募集到肝脏的淋巴细胞，以评估免疫细胞对急性HCV感染的影响。通过使用新型mpcc，我们可以更好地了解HCV的发病机制，这可能指导合理设计抗病毒治疗的发展。开发有效的丙型肝炎病毒抗病毒药物或疫苗将对全球公共卫生产生深远影响。

DESCRIPTION (provided by applicant): The hepatitis C virus (HCV) is a major worldwide public health problem affecting an estimated 120-170 million people. Unfortunately, an effective vaccine for HCV does not yet exist and current therapies are toxic and variably effective. While much has been learned about HCV pathogenesis, the virus and host factors that influence spontaneous clearance or persistent infection remain unknown. The lack of robust in vitro and in vivo models that faithfully mimic natural HCV infection of humans continues to hamper studies of HCV pathogenesis. Though hepatocellular carcinoma derived cell lines and immortalized hepatocytes allow us to study the replication of HCV, they are phenotypically and functionally abnormal as compared to primary hepatocytes. While primary hepatocyte cultures are thought to be an ideal model to study HCV infection and virus/host interactions, their liver specific functions rapidly decline in conventional culture systems. Recently, the creation of bioengineered micropatterned co-cultures (MPCCs) of hepatocytes and fibroblasts have allowed for long term culture of hepatocytes with the retention of normal cell function (albumin production, urea synthesis etc). The long-term goal of our research is to gain a better understanding of HCV pathogenesis in order to design specific and effective therapeutics. We hypothesize that early virus and host interactions influence the outcome of disease setting the stage for spontaneous clearance or chronic infection/liver fibrosis. To gain insight into early host response to acute HCV infection in the liver, we propose to characterize HCV virus/host interactions within primary human hepatocyte MPCCs and in MPCCs of increasing cellular complexity. Under specific aim 1, we will differentially phenotype infected and non-infected cells within MPCCs performing both genomic and proteomic profiling to identify host genes and pathways modulated in HCV infection. In specific aim 2, we will increase the cellular complexity of MPCCs to include non-parenchymal cells in order to assess if supporting cell types in the liver contribute to HCV pathogenesis and/or the development of a profibrotic state. In specific aim 3, we will increase the cellular complexity of MPCCs to include lymphocytes resident or recruited to the liver during HCV infection in order to assess the effects of immune cells on acute HCV infection. Through the use of the novel MPCCs, we may gain a better understanding of HCV pathogenesis, which may guide the development of rationally designed antiviral therapies. The development of effective HCV antivirals or vaccines would have a profound impact on global public health.