Regulation of hepatic NKT cells by CDld+ liver cells

描述(申请人提供)：全球有近2亿人感染丙型肝炎病毒，其中绝大多数是慢性肝炎，在美国，近2%的人感染了丙型肝炎病毒，这是一个重大的公共卫生挑战，美国每年约有9000人死亡。目前。慢性肝炎常导致肝硬变，而肝硬变又是肝细胞癌的主要危险因素。虽然特异性免疫在急性和慢性丙型肝炎(CHC)中的作用正在显现，但对关键的肝脏免疫亚群随着疾病进展而变化的情况知之甚少。适应性丙型肝炎病毒特异性T细胞集中在肝脏中，具有先天类自然杀伤细胞(NK)和NKT，约占正常和病变肝白细胞的50%。肝脏NKT参与了对急性病毒感染的保护作用，但可能被丙型肝炎病毒所取代。对肝脏NKT细胞激活的调控知之甚少。这项建议是为了确定我们首次从功能上定义的人类肝脏NKT的主要和异质性群体受丙型肝炎CD1d+肝细胞调节的机制。根据我们的发现，我们假设在慢性丙型肝炎中，这种巨大的异质性群体的NKT非但没有起到保护作用，反而促进了炎症和纤维化，被认为是肝细胞癌发生的先决条件。我们认为，肝脏CD1d可以为肝脏NKT细胞提供新的脂质配体。我们有证据表明S是肝脏NKT的配体，我们已经从CD1d+细胞中分离出选择性刺激iNKT的部分成分。这些数据为我们获得具有配体活性的NKT组分提供了原理证明。这项建议将利用我们在人类肝脏和控制NKT、生理CD1d+提呈肝细胞以及我们和合作者开发的试剂方面的独特专业知识，从脂库中识别肝脏NKT激活部分和潜在配体，为临床前开发潜在的慢性丙型肝炎和肝癌的潜在治疗措施提供靶点(S)。鉴于我们已经发现的慢性NKT刺激的有害影响，这将使我们能够通过新分子(S)转向新的治疗方法来潜在地操纵肝脏NKT，以帮助延缓丙型肝炎进展为肝硬变和肝癌。目的1.建立具有代表性的人肝CD1d反应性NKT细胞小组，并测试候选小分子对肝脏NKT细胞和血液不变NKT细胞的特异性激活或抑制。1A.制备与我们的iNKT一起使用的浓缩肝脏NKT面板并对其功能进行表征。1B.确定选择性文库中的哪些iNKT配体也激活肝脏NKT，以及哪些抑制。目的2.确定人肝NKT相对于其他CD1d+靶细胞的特异性，确定特异性激活或抑制肝NKT的肝CD1d+细胞组分。2A。确定与iNKT细胞相比，富集的肝脏NKT细胞对肝脏CD1d+靶细胞是否具有明显的良好特异性。2B。鉴定特异性激活肝脏NKT和/或iNKT的洗脱的肝脏CD1d配体的组份。 与公共卫生相关：全世界有近2亿人患有丙型肝炎，其中许多人将发展为感染的并发症，包括肝硬化和肝细胞癌。这项建议是为了研究和操纵人类肝脏中的一个主要免疫群体--NKT细胞，这些细胞虽然在急性感染中具有保护作用，但似乎有助于炎症和纤维化损害，从而导致肝脏疾病的进展。

DESCRIPTION (provided by applicant): Worldwide, nearly 200 million people have hepatitis C virus (HCV), the great majority of which have chronic hepatitis, nearly 2% in the U.S. are infected with HCV, a major public health challenge with ~9,000 U.S. deaths / yr. currently. Chronic hepatitis frequently leads to cirrhosis, which in turn is the major risk factor for hepatocellular carcinoma (HCC). Although roles of specific immunity in acute and chronic hepatitis C (CHC) are emerging, little is known about key liver immune subset changes with progression. Adaptive HCV-specific T cells are concentrated in liver with innate-like natural killer (NK) and NKT, which represent to ~50% of normal and diseased liver leukocytes. Hepatic NKT are implicated in protection against acute viral infection, but maybe subverted by HCV. Regulation of activation of hepatic NKT cells is poorly understood. This proposal is to determine mechanisms by which the major and heterogeneous population of human hepatic NKT, that we first functionally defined, are regulated by CD1d+ liver cells in hepatitis C. Based on our findings, we hypothesize that in chronic hepatitis C, this large heterogeneous population of liver NKT, instead of being protective, promote inflammation and fibrosis, believed to be prerequisites for development of HCC. We propose that hepatic CD1d can present novel lipid ligands to liver NKT cells. We have evidence for ligand(s) for hepatic NKT and we have isolated fractions from CD1d+ cells that stimulate iNKT selectively. These data provide proof-of-principle that we can obtain fractions with ligand-like activity for NKT. This proposal will exploit our unique expertise with human hepatic and control NKT, physiological CD1d+ presenting liver cells, and reagents we and collaborators have developed to identify hepatic NKT- activating fractions and potential ligands from lipid libraries, to provide target(s) for preclinical development of potential novel therapeutic interventions in CHC and HCC. Given deleterious effects of chronic NKT stimulation we have uncovered, this will enable us to potentially manipulate hepatic NKT through novel molecule(s) toward novel therapeutics to help delay progression to cirrhosis and HCC in hepatitis C. Aim 1. Establish a representative panel of human hepatic CD1d-reactive NKT cells and test candidate small molecules for specific activation or inhibition of hepatic NKT cells vs. blood invariant NKT cells. 1a. Prepare and functionally characterize enriched hepatic NKT panels for use alongside our iNKT. 1b. Determine which iNKT ligands from a selective library also activate hepatic NKT and which inhibit. Aim 2. Determine specificity of human hepatic NKT for hepatic vs. other CD1d+ target cells, and define hepatic CD1d+ cell fractions that specifically activate or inhibit hepatic NKT. 2a. Determine whether enriched hepatic NKT cells have distinct fine specificity for hepatic vs. other CD1d+ target cells relative to iNKT cells. 2b. Identify fractions of eluted liver CD1d ligands that specifically activate hepatic NKT and/or iNKT. PUBLIC HEALTH RELEVANCE: Worldwide nearly 200 million people have hepatitis C and many of these will progress to develop complications of the infection, including cirrhosis and hepatocellular carcinoma. This proposal is to study and manipulate a major immune population in the human liver, 'NKT' cells, which while protective in acute infections appear to contribute to inflammatory and fibrotic damage leading to liver disease progression.