Immune (dys-)regulation by the PD-1/PD-L checkpoint pathway in chronic liver disease

慢性肝病中 PD-1/PD-L 检查点通路的免疫（失调）调节

• 批准号: MR/X009904/1
• 负责人: Evangelos Triantafyllou
• 金额: $ 98.38万
• 依托单位: Imperial College London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FX009904%2F1
• 关键词: Immune; dys; regulation; PD; checkpoint

Chronic liver disease (CLD) is a global healthcare burden, affecting over 840 million people worldwide and has a rising incidence. Irrespective of the cause of CLD, persistent damage to the liver (e.g., alcohol) leads to the progressive formation of scar tissue (called fibrosis), loss of liver function, and may ultimately result in cirrhosis (scarring). The acute development of major clinical complications and/or infections in cirrhotic patients is called acute decompensation of cirrhosis. Non-alcoholic fatty liver disease (NAFLD), another common aetiology of CLD, is present in 25% of adult population globally. NAFLD is associated with obesity and type 2 diabetes and arises when too much fat is deposited in the liver. This is linked to fibrosis, progression to cirrhosis and increased risk of liver cancer. Treatment options are limited so there is a huge clinical need to develop new effective therapies.The immune system plays a crucial role in CLD progression. Macrophages are immune cells present throughout our body. Liver macrophages express different proteins on their surface via which they interact ("cross-talk") with other cells or sense signals from tissue damage and the deposition of fat or scar tissue. Hence, macrophages play a central part in regulating liver injury. As disease advances to cirrhosis, these signals may reduce the function of immune cells, thus increasing patients' susceptibility to microbial infections and mortality risk.Our recent work studied a crucial cell signalling pathway via which immune cells use the PD-1 and PD-L proteins on their surface to communicate. We showed that this pathway controls the function of liver macrophages, and its therapeutic targeting can improve macrophage anti-microbial responses after acute liver injury. We now intend to study this pathway in more detail, to understand how it controls the function of immune cells after chronic liver injury. To achieve this, we will perform series of experiments using modern technological approaches in human and mouse samples. Firstly, we will use scientific techniques called flow cytometry and multispectral imaging and apply them on blood and liver tissue samples of CLD patients. This will allow us to detect which immune cells express the PD-1 and PD-L surface proteins. We will next use cell culture and flow cytometry to study how these proteins' interactions alter immune cell function. We will use patient plasma and stool samples to perform cell culture experiments to identify the signals that increase PD-1/PD-L protein levels. We will then use therapeutic drugs that inhibit these signals, aiming to restore the function of immune cells from CLD patients (e.g., to enhance antimicrobial defence). Secondly, we aim to identify which liver cells express PD-1 and PD-L proteins over time during chronic liver injury. To do this, we will use mouse models of liver fibrosis and fatty liver disease and perform flow cytometry on cells isolated from the blood and livers of mice. We will next assess if liver injury improves or gets worse when the PD-L protein isn't produced. To achieve this, we will use genetically-modified mice whose liver macrophages do not express the PD-L protein. We will perform a new technique called single-cell RNA sequencing on liver cell samples, and this will tell us how immune cells' function changes after injury. Finally, having identified which signals increase PD-1/PD-L protein levels, we will use drugs to block these signals and study if their therapeutic targeting can reduce liver fibrosis and/or improve the antimicrobial defence mediated by liver macrophages.Overall, by completing this project we will increase our understanding of how the PD-1/PD-L pathway controls immune cell function and therefore regulates liver injury. This will provide new opportunities for therapeutic intervention, discovery of prognostic biomarkers for CLD progression, and ultimately enhance patient outcomes.

慢性肝病（CLD）是全球医疗保健负担，全球影响超过8.4亿人，发病率上升。不论CLD的原因如何，对肝脏（例如，酒精）的持续损害会导致疤痕组织的进行性形成（称为纤维化），肝功能的丧失，并最终可能导致肝硬化（SCARRING）。肝硬化患者重大临床并发症和/或感染的急性发展称为肝硬化的急性代表。非酒精性脂肪肝疾病（NAFLD）是CLD的另一种常见病因，全球25％的成人人群中存在。 NAFLD与肥胖症和2型糖尿病有关，当肝脏沉积过多时会出现。这与纤维化，肝硬化的进展和肝癌风险增加有关。治疗方案有限，因此有巨大的临床需要开发新的有效疗法。免疫系统在CLD进展中起着至关重要的作用。巨噬细胞是我们整个体内存在的免疫细胞。肝巨噬细胞在其表面上通过其相互作用（“交叉对话”）与其他细胞或从组织损伤和脂肪或疤痕组织沉积的感觉信号。因此，巨噬细胞在调节肝损伤方面起着核心作用。随着疾病的发展，这些信号可能会降低免疫细胞的功能，从而提高患者对微生物感染和死亡率风险的敏感性。我们最近的工作研究了一种至关重要的细胞信号通路，免疫细胞使用PD-1和PD-L蛋白在其表面上使用PD-1和PD-L蛋白进行传播。我们表明，该途径控制着肝巨噬细胞的功能，其治疗性靶向可以改善急性肝损伤后巨噬细胞抗微生物反应。现在，我们打算更详细地研究该途径，以了解它如何控制慢性肝损伤后免疫细胞的功能。为了实现这一目标，我们将使用人和小鼠样品中的现代技术方法进行一系列实验。首先，我们将使用称为流式细胞术和多光谱成像的科学技术，并将其应用于CLD患者的血液和肝组织样本。这将使我们能够检测哪些免疫细胞表达PD-1和PD-L表面蛋白。接下来，我们将使用细胞培养和流式细胞术来研究这些蛋白质的相互作用如何改变免疫细胞功能。我们将使用患者血浆和粪便样品进行细胞培养实验，以确定增加PD-1/PD-L蛋白水平的信号。然后，我们将使用抑制这些信号的治疗药物，旨在恢复CLD患者免疫细胞的功能（例如，增强抗菌防御）。其次，我们旨在确定在慢性肝损伤期间，随着时间的流逝，哪些肝细胞表达PD-1和PD-L蛋白。为此，我们将使用肝纤维化和脂肪肝病的小鼠模型，并对从小鼠的血液和肝脏分离的细胞进行流式细胞仪。接下来，我们将评估未产生PD-L蛋白时肝损伤是否会改善或恶化。为了实现这一目标，我们将使用遗传改性的小鼠，其肝巨噬细胞不表达PD-L蛋白。我们将在肝细胞样品上执行一种称为单细胞RNA测序的新技术，这将告诉我们免疫细胞损伤后免疫细胞的功能如何变化。最后，在确定了哪些信号增加了PD-1/PD-L蛋白水平之后，我们将使用药物阻止这些信号并研究其治疗性靶向是否可以减少肝纤维化和/或改善由肝巨噬细胞介导的抗菌防御，从而通过完成该项目来完成对PD-1/PD-L Pathway pathway Conlation Sunder Sunder Sunder Sunder river的理解，从而受到liver river的影响。这将为治疗干预，发现CLD进展的预后生物标志物提供新的机会，并最终增强患者的预后。