Hepatocyte-derived extracellular vesicles in alcoholic liver disease

酒精性肝病（ALD）在美国仍然是一种毁灭性的疾病， 全球死亡率估计每年超过70万的其他国家。慢性乙醇 消耗导致肝细胞中的脂质积累及其细胞器应激， 炎症和纤维化在ALD进展中的作用。肝细胞和非肝细胞之间的串扰 实质细胞-包括肝巨噬细胞（HM）和肝星状细胞（HSC）-是 这一过程至关重要。然而，参与的分子机制和信号通路， 脂质超载肝细胞和非实质细胞之间的串扰仍然存在 不完全理解。特别是，这种串扰促进或 调节从慢性无症状酒精性脂肪性肝炎（ASH）向致命酒精性脂肪性肝炎的转变 肝炎（AH）是设计针对这种破坏性疾病的有效新型治疗方法的中心问题， 疾病在专注于细胞间通讯研究的同时， 受损的肝细胞释放细胞外囊泡（EV），这些EV在肝细胞中循环， 血液在ALD小鼠模型中。EV被有效地内化到靶细胞中并将其转移到靶细胞中。 货物包括miRNA。后者是EV中包裹的miRNA的关键机制 EV-miRNAs作为“功能性胞外RNA”调节靶细胞中的蛋白质翻译， 细胞EV还包含称为损伤相关分子模式的无菌危险信号 （DAMP）。我们发现，来自AH小鼠肝细胞的EV富含 线粒体DNA（mtDNA），其有助于炎症过程的激活。此外，我们的 比较AH与先前慢性（c）ASH小鼠肝脏的RNA-seq数据的途径分析， 将EV鉴定为最显著上调的途径之一。基于这些结果，我们 提出了一个中心假设，即具有定量和定性不同货物的电动汽车是 从AH与cASH前受损肝细胞中释放，以诱导独特的细胞串扰 在AH病理表型的发生中。我们还假设血浆EV分析 用作液体肝活检，为ALD严重程度的诊断和分期提供条形码。到 调查这些假设，我们的提案有以下具体目标。1)确定 EV及其来源于肝细胞的货物作为条形码的作用， 在鼠模型和人ALD中将无症状cASH转化为AH。2)剖析其中的机制 参与cASH向AH转变中EV介导的细胞与细胞通讯。解决 这些核心问题，我们已经组建了一个具有EV生物学专业知识的MPI调查小组， ALD病理学，独特动物模型的开发，人类ALD和RNA治疗。

Alcoholic liver disease (ALD) continues to be a devastating disease in the United States and many other countries with the global mortality estimated to exceed 0.7 million per year. Chronic ethanol consumption results in lipid accumulation in hepatocytes and their organelle stress leading to inflammation and fibrosis in the progression of ALD. The crosstalk between hepatocytes and non- parenchymal cells - including hepatic macrophages (HMs) and hepatic stellate cells (HSCs) - is crucial to this process. However, the molecular mechanisms and signaling pathways involved in the crosstalk between lipid overloaded hepatocytes and non-parenchymal cells, remain incompletely understood. In particular, the mechanisms by which this crosstalk facilitates or regulates a transition from chronic asymptomatic alcoholic steatohepatitis (ASH) to fatal alcoholic hepatitis (AH) is a central issue for designing efficacious novel treatments for this devastating disease. While focused on the investigation of cell-to-cell communication we recently revealed that damaged hepatocytes release extracellular vesicles (EVs) and these EVs circulate in the blood in mouse models of ALD. EVs are efficiently internalized into target cells and transfer their cargo including miRNAs. The later is a key mechanism by which encapsulated miRNAs in EVs (EV-miRNAs) serve as “functional extracellular RNAs” to regulate protein translation in target cells. EVs also contain sterile danger signal known as damage-associated molecular patterns (DAMPs). We found that EVs derived from hepatocytes from AH mice are enriched in mitochondrial DNA (mtDNA) that contribute to activation of an inflammatory process. Further, our pathway analysis of RNA-seq data comparing AH vs. preceding chronic (c)ASH mouse livers, identifies EV as one of the most significantly upregulated pathways. Based on these results, we propose the central hypothesis that EVs with a quantitatively and qualitatively distinct cargo are released from damaged hepatocytes in AH vs. preceding cASH to induce unique cellular crosstalk in the genesis of the AH pathologic phenotype. We also hypothesize that plasma EV analysis serves as a liquid liver biopsy providing a barcode for diagnosis and staging of ALD severity. To investigate these hypotheses our proposal has the following SPECIFIC AIMS. 1) Determine the role of EVs and their cargo derived from hepatocytes as barcodes for a transition from asymptomatic cASH to AH in murine models and human ALD. 2) Dissect the mechanisms involved in EV-mediated cell-to-cell communication in the cASH to AH transition. To address these central issues, we have put together a MPI investigative team with expertise in EV biology, ALD pathology, development of unique animal models, human ALD, and RNA therapeutics.