Functional characters of non-coding RNAs in alcoholic liver injury

描述（由申请人提供）： 项目摘要酒精性肝脏疾病（ALD）--从酒精性脂肪肝到酒精性肝纤维化和肝硬化- -占西方国家所有慢性肝脏疾病的50%以上，并且占每年死亡人数的5%。美国，包括我们的退伍军人群体。研究表明，虽然所有重度饮酒者都表现出脂肪肝（脂肪肝）的迹象，但只有10%至35%的酗酒者会发展为肝脏炎症，高达20%会发展为肝硬化。大约15%的美国酗酒者最终会患上酒精性肝病，这是全球肝病和肝脏相关死亡的主要原因之一。酒精性肝病的临床谱包括酒精性脂肪肝、酒精性脂肪性肝炎、酒精性肝硬化和肝细胞癌风险增加。ALD的病理机制涉及酒精及其毒性代谢物对肝脏中各种细胞类型的直接作用之间的复杂相互作用、活性氧（ROS）的诱导、炎症级联的上调以及肝脏中的其他细胞特异性作用。非编码RNA（ncRNA）分子领域代表了生物学的一个范式转变，远离了生物学的中心法则，即RNA分子仅仅是DNA和蛋白质合成之间的信使。然而，超过90%的细胞产生的RNA分子并不是注定要被翻译成蛋白质。相反，这些ncRNA可以作为酶，信号分子和转录因子。微小RNA（microRNAs，miRNAs）是近年来发现的一类调控细胞转录组和蛋白质组的小分子非编码RNA。miRNAs在健康和疾病中塑造组织和器官的分化和功能方面发挥着至关重要的作用。最近的几项研究提供了令人信服的证据，表明乙醇敏感的miRNA确实是调节主开关。更具体地说，miRNAs控制耐受性的发展，这是乙醇成瘾的关键组成部分。其他滥用药物也靶向一些乙醇敏感的miRNAs，这表明成瘾的共同生化机制。因此，了解ALD中ncRNA调控的精确机制对于开发治疗干预变得非常关键，因为如果及时给予适当的支持，衰竭的肝脏可能会恢复。我们的长期目标是阐明导致ALD的ncRNA异常细胞间通讯的机制。在本申请中，我们提出了系统的研究乙醇和内毒素（脂多糖，LPS）依赖的ncRNA作为ALD标记物，通过集中在四个特定的目标：首先，我们将确定功能性乙醇依赖的miRNA参与细胞存活在人类肝星状细胞，肝细胞和胆管细胞。其次，我们将明确LPS调控的miRNAs在人肝星状细胞和肝实质细胞中的作用以及相关的重塑机制。第三，我们将评估特定转录的超保守区（T-UCRs）和miR- 181/let-7家族之间的相互作用，并确定这种相互作用在肝胆上皮和星状细胞中ALD特性中的作用。第四，我们将确定酒精和LPS依赖性ncRNA对酒精性脂肪性肝炎小鼠模型体内ALD进展的影响。miRNA作为体外肝细胞存活、转化和重塑的重要调节因子，以及其上游调节剂和与T-UCR的相互作用的鉴定将提供对改变的ncRNA表达参与ALD进展的深入了解，并在动物模型中测试ALD的新治疗方法。

DESCRIPTION (provided by applicant): Project Summary Alcoholic liver diseases (ALD) -- ranging from alcoholic fatty liver to alcohol-induced liver fibrosis and cirrhosis - - account for more than 50% of all chronic liver diseases in western countries and are responsible for 5% of the deaths occurring annually in the United States including our Veteran population. Studies have shown that while all heavy drinkers display signs of hepatitis steatosis (fatty liver); only 10% to 35% of alcoholics develop hepatic inflammation, with up to 20% progressing to cirrhosis. About 15% of U.S. alcoholics eventually will develop alcoholic liver diseases, one of the leading causes of liver diseases and liver-relate death worldwide. The clinical spectrum of alcoholic liver disease includes alcoholic fatty liver, alcoholic steatohepatitis, alcoholic cirrhosis, and increased risk of hepatocellular carcinoma. The pathological mechanisms of ALD involves complex interactions between the direct effects of alcohol and its toxic metabolites on various cell types in the liver, induction of reactive oxygen species (ROS), upregulation of the inflammatory cascade, and other cell-specific effects in the liver. The field of non-coding RNA (ncRNA) molecules represents a paradigm shift in biology, away from the central dogma of biology which places RNA molecules as mere messengers between DNA and protein synthesis. However, more than 90% of the RNA molecules made by a cell are not destined to be translated into proteins. Instead, these ncRNAs can act as enzymes, signaling molecules and transcriptional factors. MicroRNAs (miRNAs) are small ncRNAs that have been recently identified as master regulators of the cellular transcriptome and proteome. miRNAs play a crucial role in shaping the differentiation and function of tissues and organs in both health and disease. Several recent studies have provided compelling evidence showing that ethanol-sensitive miRNAs are indeed regulatory master-switches. More specifically, miRNAs control the development of tolerance, a crucial component of ethanol addiction. Other drugs of abuse also target some ethanol-sensitive miRNAs suggesting that common biochemical mechanisms underlie addiction. Therefore, the knowledge of the precise mechanisms of ncRNA regulation in ALD becomes very critical to develop therapeutic interventions since the failing liver may revive if given proper support in a timely manner. Our long-term goal is to clarify the mechanisms underlying the abnormal intercellular communication of ncRNAs that lead to ALD. In this application, we propose the systematic investigation of ethanol and endotoxin (lipopolysaccharide, LPS) dependent ncRNAs as ALD markers by focusing on four specific aims: First, we will identify the functional ethanol-dependent miRNAs involved in cell survival in human hepatic stellate cells, hepatocytes and cholangiocytes. Second, we will define the role of LPS regulated miRNAs in human hepatic stellate and parenchymal cells and the related remodeling mechanisms. Third, we will evaluate the cross-talk between specific transcribed ultra conserved regions (T-UCRs) and miR- 181/let-7 families, and define the role of this interaction in ALD properties in hepatobiliary epithelial and stellate cells. Fourth, we will determine the effects of ethanol and LPS dependent ncRNAs on the progression of ALD in a mouse model of alcoholic steatohepatitis in vivo. The identification of miRNAs as an important regulator of hepatic cell survival, transformation and remodeling in vitro, as well as their upstream modulators and interaction with T-UCRs will provide insight into the involvement of altered ncRNA expression in contributing to ALD progression and test novel therapeutic approaches for ALD in animal models.