Role of LAP in controlling liver homeostasis

LAP 在控制肝脏稳态中的作用

• 批准号: BB/W002450/1
• 负责人: Naiara Beraza
• 金额: $ 72.24万
• 依托单位: Quadram Institute
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FW002450%2F1
• 关键词: Role; LAP; controlling; liver; homeostasis

Dead cell clearance is a fundamental physiological process essential for tissue homeostasis throughout life, and for efficient responses to multicellular tissue damage; generally termed phagocytosis, it is achieved by professional (immune, myeloid) and non-professional (epithelial, endothelial) phagocytic cells that recognise, internalise and degrade dying cells and extracellular debris.Recently, the novel LC3-dependent phagocytosis (LAP) pathway was described which mediates engulfment of extracellular components and their intracellular processing and degradation. Our research team (co-I: Wileman) has defined the molecular components regulating LAP and how this preserves brain homeostasis and prevents Alzheimer's Disease during old age. Our team also dissected cell-specific LAP functions in other tissues, showing how LAP activation affects sensitivity to influenza virus infection in the lung.To advance our understanding of cell clearance in the liver, we have generated a transgenic mouse in which LAP can be depleted specifically in hepatocytes. This unique experimental tool enables us to dissect the, as yet unknown, role of LAP in hepatocytes and its influence on liver function.This is particularly important since the liver is constantly challenged with antigens and bacterial products circulating from the intestine, as well as an influx of dying lymphocytes from blood circulation. These promote low-grade apoptosis of hepatocytes which are replaced by new hepatocytes after proliferation, a regenerative process tightly regulated by crosstalk with resident macrophages. In response to excessive hepatocyte death after liver injury, this self-regulated interaction can become overwhelmed leading to chronic inflammation and disease progression.Remarkably, despite the well-known capacity of hepatocytes to engulf dead cells, the underpinning mechanisms remain undefined, supporting the urgent need for our research aimed at characterising LAP in hepatocytes and defining its influence on liver function and response to injury.Our pilot results using hepatocyte-specific LAP-deficient mice show that absence of LAP protects the liver from hepatocellular death, inflammation and fibrosis. This indicates that LAP has a specific function in hepatocytes, different from that recently described in myeloid cells infiltrating the liver, and supporting the key role of LAP in hepatocyte modulation of liver response to challenge.Building on our exciting pilot work, here we propose a programme of research that integrates in vitro and in vivo work in murine liver cells and tissue samples, to define the specific role of LAP in hepatocytes for regulation of liver function and immunity.First, we will: define what activates LAP in hepatocytes; characterise the molecular machinery of the LAP pathway; and determine the fate of intracellular cargo in hepatocytes using in vitro techniques that are well-established in our labs.Next, we will determine how hepatocyte-LAP controls liver responses to injury and inflammation in vivo using our unique genetically-modified mice and experimental models, also well-established in our groups. Exploiting the expertise of our team, we will precisely dissect how hepatocyte-LAP activation can influence on other individual liver cell type function during responses to injury using high-throughput 10x single cell sequencing technology.Ultimately, we will apply our expertise in the mechanisms mediating liver regeneration to define the role of LAP in restoration of liver function after challenge and preservation of liver homeostasis.Through this programme of research, we will deliver new fundamental mechanistic insights into the role of hepatocyte-LAP in controlling liver responses to injury and in preventing disease progression. This will enable us to develop future strategies that will reduce hepatocyte injury and thus prevent and treat disease and preserve life-long health.

死细胞清除是一个基本的生理过程，对整个生命过程中的组织稳态和对多细胞组织损伤的有效反应至关重要;一般称为吞噬作用，它是由专业人员完成的。（免疫，骨髓）和非专业（上皮，内皮）吞噬细胞，识别，内化和降解死亡细胞和细胞外碎片。最近，描述了新的LC 3依赖性吞噬作用（pG）途径，其介导细胞外组分的吞噬及其细胞内加工和降解。我们的研究团队（co-I：Wileman）已经定义了调节脑内平衡的分子成分，以及这如何在老年时保持大脑内稳态并预防阿尔茨海默病。我们的研究小组还分析了其他组织中细胞特异性的β-内酰胺酶功能，显示β-内酰胺酶激活如何影响肺对流感病毒感染的敏感性。为了进一步了解肝脏中的细胞清除，我们已经产生了一种转基因小鼠，其中β-内酰胺酶可以特异性地在肝细胞中耗尽。这一独特的实验工具使我们能够剖析目前尚不为人所知的肝细胞中的HSP 70的作用及其对肝功能的影响。这一点尤其重要，因为肝脏不断受到来自肠道的抗原和细菌产物的挑战，以及来自血液循环的垂死淋巴细胞的涌入。这些促进肝细胞的低度凋亡，其在增殖后被新的肝细胞取代，这是一种通过与驻留巨噬细胞的相互作用紧密调节的再生过程。在肝损伤后，由于肝细胞过度死亡，这种自我调节的相互作用可能变得不堪重负，导致慢性炎症和疾病进展。值得注意的是，尽管肝细胞吞噬死细胞的能力众所周知，但其基础机制仍不明确，支持我们的研究的迫切需要，旨在表征肝细胞中的肝细胞凋亡，并确定其对肝功能和对损伤的反应的影响。使用肝细胞特异性LAP缺陷小鼠的结果显示，没有LPS保护肝脏免于肝细胞死亡、炎症和纤维化。这表明，在肝细胞中的特异性功能，不同于最近描述的骨髓细胞浸润肝脏，并支持肝细胞的肝细胞调节肝脏反应的关键作用，以我们令人兴奋的试点工作的基础上，在这里，我们提出了一个研究计划，整合在体外和体内的工作，在小鼠肝细胞和组织样本，为了明确肝细胞中β-淀粉样蛋白在调节肝功能和免疫方面的特殊作用，我们首先要：明确什么激活了肝细胞中β-淀粉样蛋白，阐明β-淀粉样蛋白途径的分子机制，并使用我们实验室中成熟的体外技术确定肝细胞内货物的命运。接下来，我们将使用我们独特的基因修饰小鼠和实验模型来确定肝细胞凋亡如何控制肝脏对体内损伤和炎症的反应，这些模型也在我们的小组中得到了很好的建立。利用我们团队的专业知识，我们将使用高通量10倍单细胞测序技术精确剖析肝细胞活化如何在对损伤的反应期间影响其他个体肝细胞类型的功能。最终，我们将运用我们在肝再生机制方面的专业知识，确定在挑战后恢复肝功能和维持肝脏稳态方面的作用。通过这个项目，通过这项研究，我们将为肝细胞凋亡在控制肝脏对损伤的反应和预防疾病进展中的作用提供新的基本机制见解。这将使我们能够制定未来的策略，减少肝细胞损伤，从而预防和治疗疾病并保持终身健康。

项目成果

Control of infection by LC3-associated phagocytosis, CASM, and detection of raised vacuolar pH by the V-ATPase-ATG16L1 axis.

• DOI: 10.1126/sciadv.abn3298
• 发表时间: 2022-10-28
• 期刊: Science advances
• 影响因子: 13.6