Role of Kupffer Cell and NK Cell Interactions in Hepatitis C Virus Infektion

丙型肝炎病毒（HCV）诱导的肝硬化的发病机制被认为是由宿主免疫系统本身驱动的。KCs代表肝脏中的常驻巨噬细胞群，研究表明，它们在慢性hcv感染患者的肝脏中被激活并频率增加。此外，库普弗细胞（KCs）在体外响应HCV，释放IL-1b和IL-18。然而，暴露于HCV的KCs对HCV复制的直接影响尚不清楚，KCs与肝脏浸润性NK细胞在HCV复制反应中的相互作用尚未得到详细研究。这是令人惊讶的，因为NK细胞代表了一个主要的效应细胞群，参与了对病毒感染的先天免疫反应，并且已经描述了toll样受体（TLR）依赖性与KCs的串扰。在慢性HCV感染患者中，肝内NK细胞的频率更高，与外周血中的NK细胞相比，它们更活跃。它们表现出一种功能上的二分法，其特征是IFNg的产生减少和细胞毒性增强，这可能导致肝损伤，因为NK细胞的细胞毒性与ALT水平相关。此外，我们最近能够将外周血单核细胞炎症小体介导的IL-18分泌与NK细胞来源的IFNg产生联系起来。许多先前研究的一个局限性是只使用来自血液的单核细胞群。事实上，研究表明，血液和肝脏中单核细胞的组成和功能是不同的。因此，肝单核细胞之间的真正相互作用只能通过肝组织细胞的检测来确定。利用我科正在进行的肝胆移植手术的优势，本项目拟研究先天免疫细胞在HCV复制的背景下，特异性地在生物学上更相关的肝区进行研究。在第一步，我们将利用HCV复制子系统分析肝驻留KCs和肝内NK细胞在HCV应答中的相互作用。我们将解决KCs是否发挥直接抗病毒作用以及它们是否也通过NK细胞衍生的IFNg产生间接抗病毒作用的问题。在第二部分，我们将通过使用慢性hcv感染患者的肝组织来证实我们的观察结果。我们将确定来自慢性HCV感染患者的KCs是否降低了tnfa介导的（直接）和NK细胞介导的（间接）抗病毒作用，以及通过使用健康对照的KCs，肝内NK细胞的功能损伤是否可逆。最后，通过对HCV+患者在不含ifn的直接作用抗病毒药物（DAAs）治疗HCV过程中的前瞻性随访，我们将研究完全去除病毒对OLT前后患者外周血固有免疫细胞的影响。

The pathogenesis of Hepatitis C Virus (HCV)-induced liver cirrhosis is thought to be driven by the host immune system itself. KCs represent the resident macrophage population in the liver and it has been shown that they are activated and increased in frequency in the liver of chronic HCV-infected patients. Furthermore, in response to HCV, Kupffer cells (KCs) release IL-1b and IL-18 in vitro. However, a direct effect of HCV-exposed KCs on HCV replication is still unknown and the interaction between KCs and liver-infiltrating Natural Killer (NK) cells in response to HCV-replication is not studied in detail yet. This is surprising, since NK cells represent a major effector cell population involved in innate immune responses to viral infections and a Toll-like receptor (TLR)-dependent crosstalk with KCs has been described. In patients with chronic HCV infection the frequency of intrahepatic NK cells is higher and they are more activated compared to those in the peripheral blood. They display a functional dichotomy characterized by reduced production of IFNg and enhanced cytotoxicity, probably contributing to the liver injury as NK cell cytotoxicity correlates to ALT levels. Furthermore, we were recently able to link inflammasome-mediated IL-18 secretion by peripheral blood monocytes with NK cell derived IFNg production. A limitation of many previous studies is the use of mononuclear cell populations only from the blood. Indeed, it has been shown that composition and function of mononuclear cells differs between blood and liver. Therefore, the true interaction between liver mononuclear cells can only be determined through testing of cells derived from liver tissue. Taking advantage of hepatobiliary transplant surgery being performed in our department, this project is proposing to study innate immune cells in the context of HCV replication specifically in the liver, the biological more relevant compartment. In a first step, we are going to analyze the interplay of liver resident KCs and intrahepatic NK cells in response to HCV by using the HCV replicon system. We will address the question if KCs exert a direct antiviral effect and whether they also have an indirect antiviral effect via NK cell-derived IFNg production. In a second part, we will confirm our observation by using liver tissue from chronic HCV-infected patients. We will establish whether KCs from patients with chronic HCV infection have reduced TNFa-mediated (direct) and NK-cell mediated (indirect) antiviral effects and if the functional impairment of intrahepatic NK cells is reversible by using KCs from healthy controls. Finally, by following HCV+ patients prospectively during a course of HCV treatment with IFN-free direct-acting antivirals (DAAs), we will study the effect of a complete removal of the virus on peripheral innate immune cells comparing pre and post OLT patients.