Organelle Dysfunction in Liver Inflammation

肝脏炎症中的细胞器功能障碍

基本信息

批准号：
9204405
负责人：
Harmeet Malhi
金额：
$ 7.95万
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：
2016-01-15 至 2017-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9204405
关键词：
Address Anabolism Award Biogenesis Biological Assay Cartoons Cell Communication Cells Ceramides Chemicals Chemotaxis Clinical Investigator Award Data Development Disease Progression Endosomes Enzymes Evolution Factor X Fatty Liver Foundations Functional disorder Funding Future Genetic Transcription Goals Grant Hepatic Hepatocyte Immune In Vitro Inflammation Inflammatory Injury Inositol Lead Link Lipids Liver Liver diseases Macrophage Activation Mass Spectrum Analysis Measurement Mediating Mediator of activation protein Mentors Modeling Molecular Genetics Motion Mus Nonesterified Fatty Acids Obesity Organelles Palmitates Pathogenesis Pathway interactions Pharmacology Process Production Proteins Public Health Recruitment Activity Regulation Research Research Proposals Rodent Model Role Signal Transduction Specificity Sphingolipids Stress Systems Biology Testing Transcriptional Regulation Up-Regulation Vesicle Work base bindin career endoplasmic reticulum stress experimental study extracellular vesicles feeding genetic approach immune activation in vitro Model in vivo in vivo Model inhibitor/antagonist injured insight liver inflammation macrophage metabolomics nonalcoholic steatohepatitis novel therapeutic intervention public health relevance response sensor targeted treatment transcription factor vesicular release

项目摘要

DESCRIPTION (provided by applicant): My long term career objective is to define the mechanisms of liver inflammation in nonalcoholic steatohepatitis (NASH). The current proposal views this via a systems biology approach, focusing on defining the cross-talk between lipotoxic hepatocytes and proinflammatory macrophages at a cellular level; and the endoplasmic reticulum stress sensor Inositol Requiring Enzyme-1 alpha (IRE1α) and extracellular vesicle (EV) biogenesis at a sub-cellular level. Signals from injured hepatocytes serve to recruit macrophages to the liver, activate proinflammatory pathways in resident and recruited macrophages and thus set in motion an inflammation-injury feed-forward loop. In preliminary experiments we have observed that lipotoxic hepatocytes (treated with the toxic free fatty acid palmitate) release proinflammatory extracellular vesicles; these vesicles are released in an IRE1α-dependent manner; and are enriched in ceramides. Furthermore, the ceramide enrichment also occurs in an IRE1α-dependent manner. Our preliminary observations have led to the central hypothesis that IRE1α regulates palmitate-induced extracellular vesicle biogenesis and cargo, thereby leading to the release of proinflammatory extracellular vesicles, which in turn activate macrophages thus promoting NASH pathogenesis. Therefore, the goals of this proposal are to understand: i) how IRE1 mediates release of lipotoxic EVs; ii) what imparts specificity t lipotoxic EV cargo, and iii) how are lipotoxic EVs proinflammatory? The proposed experiments will employ complementary in vitro and in vivo models of lipotoxicity and NASH, respectively; and chemical, pharmacological, molecular and genetic approaches to address the specific aims to test the hypotheses that: i) IRE1α and the IRE1α-activated transcription factor, X- box bindin protein-1 (XBP-1) drive the release of lipotoxic hepatocyte EVs via increased biogenesis of ceramides, and ii) IRE1α-mediated ceramide transport is necessary for EV release, and iii) IRE1α-dependent proinflammatory EV release in vivo leads to macrophage recruitment and liver inflammation. To address these hypotheses the applicant has become adept at EV isolation and characterization, macrophage isolation, assays of macrophage activation and chemotaxis, transcriptional regulation, metabolomics by mass spectrometry and in vivo rodent models of conditional deletion of IRE1. With funding through this R03 small grant program for K08 awardees the applicant hopes to gain additional preliminary data to propel this research proposal to an R01 proposal in the near future. This work represents a coherent and logical extension of the applicant's earlier work on endoplasmic reticulum stress in the pathogenesis of NASH and represents a significant step forward in independence from previous research mentors. The applicant has established a network consisting of Dr. Gregory J. Gores as her primary mentor, and Dr. Vijay Shah and Dr. Nicholas LaRusso as collaborators. Our results will yield mechanistic insights into the processes of macrophage activation and recruitment in NASH, thus identifying potentially druggable targets.

描述（由适用提供）：我的长期职业目标是定义非酒精性脂肪性肝炎（NASH）中肝炎的机制。当前的提案通过系统生物学方法观察了这一点，重点是定义脂肪毒性肝细胞和促炎性巨噬细胞之间的串扰。以及需要酶-1α（IRE1α）和细胞外蔬菜（EV）生物发生的内质网应激传感器肌醇。受伤的肝细胞的信号可用于将巨噬细胞募集到肝脏，激活居民的促炎途径并招募巨噬细胞，从而使感染受伤的进料向前环。在初步实验中，我们观察到脂蛋白毒性肝细胞（用有毒的游离脂肪酸棕榈酸酯处理）释放促炎细胞外蔬菜。这些蔬菜以IRE1α依赖性方式释放。并富含神经酰胺。此外，神经酰胺富集也以IRE1α依赖性方式发生。我们的初步观察结果导致了一个中心假设，即IRE1α调节棕榈酸酯引起的细胞外蔬菜和货物，从而导致促炎细胞外蔬菜的释放，从而激活巨噬细胞，从而促进nash发病机理。因此，该提案的目标是理解：i）IRE1如何介导脂肪毒性电动汽车的释放； ii）什么赋予脂肪毒性EV的特异性，以及iii）脂肪毒性电动汽车如何促进？所提出的实验将分别在体外和体内脂肪毒性和NASH的体内模型。 and chemical, pharmaceutical, molecular and genetic approaches to address the specific aims to test the hypotheses that: i) IRE1α and the IRE1α-activated transcription factor, X-box bindin protein-1 (XBP-1) drive the release of lipotoxic hepatocyte EVs via increased biogenesis of ceramides, and ii) IRE1α-mediated ceramide transport is necessary for EV release, and iii) IRE1α依赖性促炎性EV在体内导致巨噬细胞募集和肝注射。为了解决这些假设，适用的已适用于EV分离和表征，巨噬细胞分离，暗杀巨噬细胞激活和趋化性，转录调控，质谱法代谢组和IRE1条件缺失的体内啮齿动物模型。申请人希望通过此R03小型赠款计划为K08获奖者提供资金，该申请人希望在不久的将来获取该研究建议的其他初步数据。这项工作代表了申请人在NASH发病机理中早期关于内质网应激的工作的连贯和逻辑扩展，这代表了与以前的研究导师独立的重要一步。该应用程序已经建立了一个由Gregory J. Gores博士作为她的主要导师的网络，Vijay Shah博士和Nicholas Larusso博士担任合作者。我们的结果将产生对NASH中巨噬细胞激活和募集过程的机械见解，从而确定了潜在的可药物靶标。