Regulation of hepatic NKT cells by CDld+ liver cells

描述（由申请人提供）：全球有近2亿人患有丙型肝炎病毒（HCV），其中绝大多数为慢性肝炎，美国有近2%的人感染HCV，这是一个重大的公共卫生挑战，目前美国每年约有9000人死亡。慢性肝炎经常导致肝硬化，这反过来又是肝细胞癌（HCC）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）中的作用正在出现，但对关键的肝脏免疫亚群随进展的变化知之甚少。适应性hcv特异性T细胞主要集中在具有先天样自然杀伤细胞（NK）和NKT的肝脏中，约占正常和病变肝脏白细胞的50%。肝脏NKT参与了对急性病毒感染的保护，但可能被HCV破坏。肝NKT细胞活化的调控机制尚不清楚。本研究旨在确定丙型肝炎患者CD1d+肝细胞调节主要的异质人群肝脏NKT的机制。基于我们的研究结果，我们假设在慢性丙型肝炎患者中，大量异质人群肝脏NKT不是具有保护作用，而是促进炎症和纤维化，这被认为是HCC发展的先决条件。我们提出肝脏CD1d可以为肝脏NKT细胞提供新的脂质配体。我们有证据表明肝NKT的配体，我们从CD1d+细胞中分离出选择性刺激iNKT的部分。这些数据提供了原理证明，我们可以获得具有配体活性的NKT分数。该提案将利用我们在人类肝脏和控制性NKT、生理性CD1d+呈递肝细胞方面的独特专业知识，以及我们和合作者开发的试剂，用于鉴定肝脏NKT激活组分和脂质库中的潜在配体，为CHC和HCC潜在的新型治疗干预措施的临床前开发提供靶点。考虑到我们已经发现的慢性NKT刺激的有害影响，这将使我们有可能通过新的分子(s)来操纵肝脏NKT，从而获得新的治疗方法，以帮助延缓丙型肝炎肝硬化和HCC的进展。建立人类肝脏cd1d反应性NKT细胞的代表性小组，并测试候选小分子对肝脏NKT细胞与血液不变性NKT细胞的特异性激活或抑制作用。1一个。制备富集的肝脏NKT板，并对其进行功能表征，以与iNKT一起使用。1 b。从一个选择性文库中确定哪些iNKT配体也激活肝脏NKT，哪些抑制。目标2。确定人类肝脏NKT对肝脏与其他CD1d+靶细胞的特异性，并确定特异性激活或抑制肝脏NKT的肝脏CD1d+细胞组分。2 a。确定富集的肝脏NKT细胞相对于iNKT细胞是否对肝脏或其他CD1d+靶细胞具有明显的精细特异性。2 b。鉴定洗脱的肝脏CD1d配体中特异性激活肝脏NKT和/或iNKT的部分。

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.