Deciphering the oxidation status of biomolecules in synovial fluids and tissues acting as damage associated molecular patterns (DAMPs) in rheumatoid arthritis

关节炎可能是由退化或自身免疫引起的，在工业化国家是导致残疾和提前退休的主要原因。例如，200多万德国公民被诊断出受到软骨退化和/或一个或多个关节功能丧失的严重影响。因此，卫生保健的社会经济后果和成本是巨大的。更糟糕的是，该病的发病率不断增加。过度产生活性氧基(ROS)，如低合子酸(HOCl)和羟基自由基，是炎症性疾病的重要标志，尤其是类风湿性关节炎(RA)。这些高活性物种有可能修饰几乎所有的生物分子：对蛋白质、糖胺多聚糖(软骨细胞外基质的主要碳水化合物)和磷脂中(多)不饱和脂肪酰基残基的影响可以得到明确的证实。然而，对于氧化修饰的脂类(带有反应醛)与丰富的蛋白质或糖胺多聚糖的氨基反应形成的产物，人们缺乏相当大的了解。此外，人们越来越认识到，被ROS(损伤相关分子模式；DAMPS)修饰的内源性分子激活了伴随着炎症增加的细胞受体。我们假设，这些已知但尚未研究的物质与炎症性疾病的发病机制特别相关，即氧化化合物与炎症性风湿病的发病机制特别相关，因此具有诊断和治疗重要性。我们在不同的复杂程度上处理这些方面。首先，我们将合成氧化和氯化脂肪酸和磷脂。通过高分辨率层析、质谱学和核磁共振谱对这些产物进行详细表征是一项非常具有挑战性的任务，因为方法必须首先适应这些分子。更具挑战性的将是分析氧化脂肪酸/脂类与模拟风湿病相关蛋白质(润滑素、聚集素)活性部位的糖胺多聚糖或多肽之间形成的反应产物。第二，第一个目标中确定的氧化分子类别将被调查风湿性疾病患者的滑液和滑膜提取物中与疾病严重程度的关系。第三，我们将判断被氧化的PL、GAG和蛋白质作为细胞免疫反应的氧化抑制物的相关性。在这项任务中，他们将通过测量典型的炎症标记物，如干扰素-γ和细胞释放的选定细胞因子，在体外研究它们对类风湿滑膜中T细胞、抗原提呈细胞和单核细胞的影响。

Arthritis, which can be due to degeneration or autoimmunity, is a major cause of disability and early retirement in the industrialized countries. For instance, more than 2 million German citizens were diagnosed to be markedly affected by cartilage degradation and/or loss of function of one or more joints. Thus, the socio-economic consequences and costs for health care are immense. Even worse, the incidence of the disease is continuously increasing.The excessive production of reactive oxygen species (ROS), such as hypochorus acid (HOCl) and hydroxyl radicals, is an important hallmark of inflammatory diseases, especially for rheumatoid arthritis (RA). These highly reactive species have the potential to modify nearly all biomolecules: effects towards proteins, glycosaminoglycans (the prime carbohydrates of the extracellular matrix of cartilage) and the (poly)unsaturated fatty acyl residues within phospholipids could be unequivocally substantiated. However, there is a considerable lack of knowledge about the products that are formed, for example, by oxidatively modified lipids (with reactive aldehyde groups) reacting with the amino groups of abundant proteins or glycosaminoglycans. Additionally, it is increasingly recognized that endogenous molecules modified by ROS (damage-associated molecular patterns; DAMPs) activate cellular receptors that are accompanied by increased inflammation. We hypothesize that these known but "non-studied" substances are particularly relevant for the pathogenesis of inflammatory diseases, i.e. that oxidized compounds are particularly relevant for the pathogenesis of inflammatory rheumatic diseases and are thus of diagnostic and therapeutic importance. We deal with these aspects on different levels of complexity. First, we will synthesize oxidized and chlorinated fatty acids and phospholipids. The detailed characterization of these products by high resolution chromatography, mass spectrometry and NMR spectroscopy is a very challenging task, as methods have to be first adapted to these molecules. Even more challenging will be the analysis of reaction products formed between oxidized fatty acids/lipids and glycosaminoglycans or peptides mimicking the active sites of proteins (lubricin, aggrecan) involved in rheumatic diseases. Second, oxidized molecule classes identified in the first aim will be investigated both in synovial fluid and extracts from the synovial membranes from patients suffering from rheumatic diseases in relation to the severity of the disease. Third, we will judge the relevance of identified oxidized PL, GAG, and proteins as oxidative DAMPs on the cellular immune response. For this task, their effects on T cells, antigen presenting cells and monocytes from the rheumatoid synovium will be studied in vitro by measuring typical inflammation markers such as interferon-gamma and selected cytokines that are released by the cells.