In Vitro Liver Models to Investigate the Progression of Liver Fibrosis

摘要 肝纤维化，其由慢性肝损伤连同细胞外基质的积聚引起。 基质（ECM）蛋白是几种慢性肝病的特征。肝脏的动态变化 微环境（LME）被广泛认为是肝纤维化进展的关键参与者， 治疗反应。LME成分，包括实质（肝细胞）和非实质（肝细胞）之间的相互作用 实质细胞（肝窦内皮细胞[LSEC]、肝星状细胞、枯否细胞），信号传导 分子（配体-胶原蛋白、层粘连蛋白）和来自ECM的机械信号，已经被牵连在 肝纤维化的进展。星状细胞活化是肝纤维化的标志;然而， 肝细胞和LSEC的功能尚未被广泛理解。此外， LME组分调节肝功能，并且对各种信号级联反应知之甚少，因此限制了对LME的研究。 肝病的最佳诊断和治疗方案的发展（例如，酒精性肝病， 非酒精性脂肪性肝病、非酒精性脂肪性肝炎以及B和C型肝炎）。因此有 迫切需要开发体外模型，模拟动态LME组件，并有效地研究其 在肝纤维化中的作用为了研究LME对细胞信号传导的直接影响，必须使用体外肝脏 模型来模拟肝纤维化的基本复杂性和动态性， 翻译效度该应用程序的目标是使用多学科的方法来开发三个 独立的体外肝模型来研究不同的LME组分（肝细胞-LSEC相互作用， ECM硬度，配体类型和密度）调节肝细胞和LSEC功能，这些LME的作用是什么 在肝纤维化的进展中起作用。拟议研究的具体目的是：1） 研究肝细胞-LSEC相互作用对肝功能的影响，2）研究肝细胞-LSEC相互作用的变化如何影响肝功能， 硬度改变肝细胞功能，以及3）确定配体类型/密度在调节肝细胞中的作用 功能这项工作将在利用体外肝脏模型的能力方面提供重大进展， 准确描述正常与疾病状态下的肝功能和代谢，尤其是 LME调节肝脏功能的发展和维持。重要的是，这种模式是创新的，因为它 将按时间顺序模拟纤维化阶段，类似于临床条件，并拥有一个环境， 比动物模型更可控更系统预计该项目将产生逐步的影响 肝纤维化和相关领域的研究，因为保留LME的肝脏模型的可用性将 促进了解大海洋生态系统活动的分子机制， 维持肝脏生物学。

ABSTRACT Liver fibrosis, which results from chronic liver damage in conjunction with the accumulation of extracellular matrix (ECM) proteins, is characteristic of several chronic liver diseases. Dynamic changes to the liver microenvironment (LME) are widely recognized as a critical participant in liver fibrosis progression and therapeutic responses. LME components, including interactions between parenchymal (hepatocytes) and non- parenchymal cells (liver sinusoidal endothelial cells [LSECs], hepatic stellate cells, kupffer cells), signaling molecules (ligands-collagen, laminin), and mechanical cues from ECM, have been implicated in the progression of liver fibrosis. Stellate cells activation is the hallmark of liver fibrosis; however, the effect on hepatocytes and LSECs function has not been extensively understood. Also, the mechanisms by which the LME components regulate liver function and various signaling cascades are poorly understood, thus limiting the development of optimal diagnosis and treatment regimes for liver diseases (e.g., alcoholic liver disease, nonalcoholic fatty liver disease, non-alcoholic steatohepatitis, and hepatitis B and C). Therefore, there is a critical need to develop in vitro models that simulate the dynamic LME components and effectively study their role in liver fibrosis. To study the direct effects of LME on cell signaling, it is imperative to use in vitro liver models to simulate the fundamental complexity and dynamism of liver fibrosis and to achieve greater translational validity. The goal of this application is to use a multidisciplinary approach to develop three independent in vitro liver models to study how different LME components (hepatocytes-LSEC interactions, ECM stiffness, ligand type and density) regulate hepatocyte and LSEC function and what role these LME components play in the progression of liver fibrosis. The specific aims of the proposed study are to: 1) investigate the effect of hepatocytes-LSEC interaction(s) on hepatic function, 2) investigate how variation in stiffness alters hepatic cell function, and 3) determine the role of ligand type/density in regulating liver cell function. This work will provide a significant advancement in the ability to utilize in vitro liver models to accurately describe the liver function and metabolism in normal versus diseased states, and especially how the LME regulate the development and maintenance of liver function. Importantly, this model is innovative as it will chronologically emulate the fibrosis stage, is similar to clinical conditions, and boasts an environment that is more controlled and systematic than animal models. This project is expected to have a progressive impact on the study of liver fibrosis and related fields because the availability of a liver model that retains LME will facilitate understanding of the molecular mechanisms that underlie LME activities in mechanisms critical for the maintenance of liver biology.