Pathobiology of Liver Injury

肝损伤的病理学

基本信息

批准号：
9208922
负责人：
Harmeet Malhi
金额：
$ 35.78万
依托单位：
MAYO CLINIC ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-23 至 2021-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9208922
关键词：
Address Americas Anabolism Applications Grants Attenuated Award Binding Proteins Biogenesis Biological Assay Boxing Carrier Proteins Ceramides Characteristics Chemotaxis Data Development Disease Disease Progression Enzymes Factor X Fatty Liver Generations Genetic Genetic Transcription Goals Grant Hepatic Hepatocyte Immune In Vitro Inflammation Inflammatory Inositol Knockout Mice Lead Leukocytes Link Lipids Liver Liver diseases Mass Spectrum Analysis Mediating Modeling Molecular Genetics Multivesicular Body Mus Myeloid Cells Nonesterified Fatty Acids Obesity Palmitates Pathogenesis Pathway interactions Process Production Proteins Public Health Recruitment Activity Research Research Personnel Role Signal Transduction Sphingosine-1-Phosphate Receptor Staging Testing United States Up-Regulation Vesicle activating transcription factor base career chronic liver disease endoplasmic reticulum stress exosome extracellular vesicles genetic approach immune activation in vitro Model in vivo in vivo Model inhibitor/antagonist insight intercellular communication lipid mediator liver inflammation liver injury macrophage mouse model nonalcoholic steatohepatitis novel therapeutic intervention receptor research study sensor serine palmitoyltransferase sphingosine 1-phosphate sphingosine kinase steroidogenic acute regulatory protein vesicular release

项目摘要

PROJECT ABSTRACT My long term career objective is to define the mechanisms of liver inflammation in nonalcoholic steatohepatitis (NASH), the most-prevalent chronic liver disease in the United States of America. NASH is characterized by endoplasmic reticulum (ER) stress, which results in activation of the ER stress sensor Inositol Requiring Enzyme-1 alpha (IRE1α), due to the accumulation of toxic lipids within hepatocytes. Macrophage- mediated liver inflammation associated with recruitment of circulating myeloid cells into the liver is also pivotal in NASH. The current proposal links hepatocyte-derived lipid mediators to macrophage-mediated inflammation by proposing that extracellular vesicles (EVs) from lipotoxic hepatocytes recruit macrophages to the liver, resulting in liver injury and inflammation. In preliminary experiments we have observed that lipotoxic hepatocytes (treated with the free fatty acid palmitate) release ceramide-enriched proinflammatory EVs in an IRE1α-dependent manner. Sphingosine 1-phosphate (S1P), derived from ceramide, on these EVs activates its receptor S1P1 on macrophages, which may promote macrophage chemotaxis into the liver. This has led to the central hypothesis that hepatocyte IRE1α regulates PA-induced EV biogenesis, release and lipid cargo (ceramide and ceramide-derived S1P) accumulation, which in turn attracts macrophages into the liver promoting NASH pathogenesis. Therefore, the goals of this proposal are to understand: i) how IRE1α mediates release of ceramide-enriched EVs; ii) how the ceramide-derived lipid mediator, S1P, on PA- stimulated EVs recruits macrophages to the liver; and iii) can lipotoxic EV production and signaling be targeted in vivo to decrease liver inflammation? The proposed experiments will employ complementary in vitro and in vivo models of lipotoxicity and NASH, respectively; and pharmacological, molecular and genetic approaches to address three integrated hypotheses. First we will directly test the hypothesis that palmitate-induced ER stress drives ceramide biosynthesis leading to EV release by a) the IRE1α-activated transcription factor, X-box binding protein-1 (XBP1) upregulation of the ceramide biosynthesis regulating enzyme serine palmitoyltransferase 1 (SPT1), and b) the transfer of ceramide to multivesicular bodies via the ceramide transport protein STARD11. Second we will directly test the hypothesis that S1P on lipotoxic EVs activates macrophage chemotaxis by a) compartmental generation of S1P by sphingosine kinase 2 forming S1P on PA- induced EVs, and b) S1P on EVs activates macrophage chemotaxis via S1P1 receptor. Third we will directly test the hypothesis that interrupting EV release or signaling is salutary in vivo in a NASH mouse model by a) reduction of EV release by IRE1α hepatocyte-specific knockout mice, and b) genetic and pharmacologic inhibition of S1P signaling on macrophages. This R01 grant application by a current K08 awarded early stage investigator will yield mechanistic insights into the processes of macrophage recruitment in NASH, thus identifying potentially druggable targets, e.g., inhibitors of IRE1α, SPT or S1P1 receptor.

项目摘要我的长期职业目标是确定非酒精性肝脏炎症的机制脂肪性肝炎（NASH）是美国最常见的慢性肝病。以内质网 (ER) 应激为特征，导致 ER 应激传感器肌醇激活由于肝细胞内有毒脂质的积累，需要酶 1 α (IRE1α)。与循环骨髓细胞募集到肝脏相关的介导的肝脏炎症也至关重要目前的提议将肝细胞来源的脂质介质与巨噬细胞介导的炎症联系起来。提出脂毒性肝细胞的细胞外囊泡（EV）将巨噬细胞招募到肝脏，导致肝损伤和炎症在初步实验中我们观察到脂毒性。肝细胞（用游离脂肪酸棕榈酸酯处理）释放富含神经酰胺的促炎性 EV IRE1α 依赖性方式，源自神经酰胺的鞘氨醇 1-磷酸 (S1P) 在这些 EV 上激活其功能。巨噬细胞上的受体S1P1，可能会促进巨噬细胞趋化进入肝脏。中心假设是肝细胞 IRE1α 调节 PA 诱导的 EV 生物发生、释放和脂质运输（神经酰胺和神经酰胺衍生的 S1P）积累，进而吸引巨噬细胞进入肝脏因此，本提案的目标是了解： i) IRE1α 是如何发挥作用的。介导富含神经酰胺的 EV 的释放；ii) 神经酰胺衍生的脂质介质 S1P 如何作用于 PA- 受刺激的 EV 将巨噬细胞招募到肝脏；以及 iii) 脂毒性 EV 的产生和信号传导是否可以成为目标体内减少肝脏炎症？拟议的实验将采用体外和体内互补的方法。脂毒性和 NASH 的体内模型；以及药理学、分子和遗传学方法首先，我们将直接检验棕榈酸酯诱导 ER 的假设。压力驱动神经酰胺生物合成，通过 a) IRE1α 激活转录因子 X-box 释放 EV 结合蛋白-1 (XBP1) 上调神经酰胺生物合成调节酶丝氨酸棕榈酰转移酶 1 (SPT1)，以及 b) 通过神经酰胺将神经酰胺转移至多泡体其次，我们将直接检验脂毒性 EV 上的 S1P 激活的假设。 a) 鞘氨醇激酶 2 在 PA 上形成 S1P，产生 S1P 的区室生成，从而导致巨噬细胞趋化诱导 EV，b) EV 上的 S1P 通过 S1P1 受体激活巨噬细胞趋化性。通过以下方式检验以下假设：在 NASH 小鼠模型中，中断 EV 释放或信号传导在体内是有益的： IRE1α 肝细胞特异性敲除小鼠减少 EV 释放，b) 遗传和药理学巨噬细胞上 S1P 信号传导的抑制该 R01 拨款申请由当前 K08 早期授予。阶段研究者将对 NASH 中巨噬细胞招募过程产生机制见解，从而识别潜在的药物靶标，例如 IRE1α、SPT 或 S1P1 受体的抑制剂。