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

丙型肝炎病毒(HCV)诱导的肝硬变的发病机制被认为是由宿主免疫系统本身驱动的。KCS代表肝脏中驻留的巨噬细胞群，已有研究表明，在慢性丙型肝炎患者的肝脏中，它们被激活并频率增加。此外，库普弗细胞(KCs)可在体外释放IL-1b和IL-18作为对丙型肝炎病毒的应答。然而，暴露于丙型肝炎病毒的KCs对丙型肝炎病毒复制的直接影响尚不清楚，KCs与肝脏浸润性自然杀伤细胞(NK细胞)之间的相互作用也尚未详细研究。这是令人惊讶的，因为NK细胞代表了参与对病毒感染的先天免疫反应的主要效应细胞群，并且已经描述了依赖Toll样受体(TLR)的KCs的串扰。在慢性丙型肝炎患者中，肝内NK细胞的频率高于外周血，并且比外周血中的NK细胞更活跃。它们表现出功能性的二分法，其特征是IFNG的产生减少和细胞毒性增强，可能是导致肝脏损伤的原因，因为NK细胞的细胞毒性与ALT水平相关。此外，我们最近能够将炎症小体介导的外周血单核细胞分泌IL-18与NK细胞产生的IFNG联系起来。许多以前研究的局限性是只使用血液中的单个核细胞群体。事实上，研究表明，单个核细胞的组成和功能在血液和肝脏中是不同的。因此，只有通过对肝组织来源的细胞进行检测，才能确定肝脏单个核细胞之间的真正相互作用。利用我们科室正在进行的肝胆移植手术，这个项目建议在丙型肝炎病毒复制的背景下研究先天性免疫细胞，特别是在肝脏，这是一个生物学上更相关的部分。在第一步中，我们将利用丙型肝炎病毒复制子系统来分析肝内KCs和肝内NK细胞在应对丙型肝炎病毒时的相互作用。我们将讨论KCs是否发挥直接的抗病毒作用，以及它们是否也通过NK细胞产生的IFNG产生间接的抗病毒作用。在第二部分中，我们将使用慢性丙型肝炎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丙型肝炎患者的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，以及使用来自健康对照组的KCs，肝内NK细胞的功能损害是否可逆。最后，在使用不含干扰素的直接作用抗病毒药物(DAAs)治疗丙型肝炎病毒的一个疗程中，通过前瞻性跟踪观察丙型肝炎病毒阳性患者，我们将研究完全清除病毒对外周天然免疫细胞的影响，比较原位肝移植前后的患者。

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.