Synovial mast cells in inflammatory arthritis

描述(申请人提供)：类风湿性关节炎(RA)患者关节中活化的肥大细胞(MC)数量增加，在20世纪80年代增加了MC参与关节炎关节炎症、蛋白分解损伤和/或重塑的可能性。利用MC缺陷小鼠，我们和其他人随后证明，这些免疫细胞确实对实验性炎症性关节炎起到了重要作用。然而，Fc3RIII和C5a激活的滑膜MC产生的对关节产生不利影响的相关介质尚未确定。小鼠MC蛋白水解酶(MMCP)6和mMCP-7是染色体17A3.3类胰蛋白酶家族的成员，它们以同型和异型四聚体的形式优先储存在MCs的分泌颗粒中，与含肝素的丝氨酸蛋白多糖结合。人类mMCP-6的同源基因是hTryptase-21。我们产生了具有酶活性的mMCP-6、mMCP-7和hTryptase-21。我们最近还创造了近交系转基因C57BL/6小鼠，它们在mMCP-6、mMCP-7和肝素的表达上存在差异。利用后一组小鼠，我们发现缺乏类胰蛋白酶7-肝素复合体的C57BL/6小鼠在两种不同的炎症性关节炎小鼠模型中减少了炎症，并减少了软骨中蛋白多糖的损失。拟议研究的总体目标是使用互补的方法在分子水平上确定小鼠和人四聚体形成类胰蛋白酶在RA和这种自身免疫性疾病的小鼠模型中的作用。我们的转基因小鼠品系、重组类胰蛋白酶和其他关键的蛋白酶特异性试剂将用于拟议的研究，以确定MC限制性四聚体形成人和小鼠类胰蛋白酶在不同的体外和体内研究中如何在多个水平上影响成纤维细胞样滑膜细胞(FLS)和软骨细胞，以及这些间充质细胞如何影响MC。将进行更多的研究，以确定分泌的mMCP-6和hTryptase-21如何在关节炎关节，甚至在MC本身中受到负调控。控制MC内具有酶活性的类胰蛋白酶表达的因素和机制尚未确定。为此，我们最近发现，FLS来源的IL-33诱导MC在mRNA和蛋白水平上增加其类胰蛋白酶的表达。因此，建议进行更多的机制研究，以了解IL-33如何控制MC中mMCP-6和hTryptase-21基因的转录和/或其转录的稳定性。结合起来，这些研究有望为MC和滑膜生理学提供新的见解，并阐明人类炎症性关节炎的新疾病机制。 公共卫生相关性：通过研究小鼠的疾病模型，我们发现一种称为肥大细胞的细胞谱系可以促进类风湿性关节炎(RA)的炎症。现在，我们发现肥大细胞制造的一种蛋白质，称为类胰蛋白酶，是肥大细胞影响疾病的方式之一。我们正在研究类胰蛋白酶影响的机制，以更好地了解如何针对这一新途径治疗类风湿性关节炎。

DESCRIPTION (provided by applicant): The presence of elevated numbers of activated mast cells (MCs) in the joints of patients with rheumatoid arthritis (RA) raised the possibility in the 1980s that MCs participate in the inflammation, proteolytic damage, and/or remodeling of the arthritic joint. Using MC-deficient mice, we and others subsequently demonstrated that these immune cells do indeed contribute substantially to experimental inflammatory arthritis. Nevertheless, the relevant mediators produced by the Fc3RIII- and C5a-activated MCs in the synovium that adversely affect the joint had not been identified. Mouse MC protease (mMCP) 6 and mMCP-7 are members of the chromosome 17A3.3 family of tryptic proteases that are preferentially stored in the secretory granules of MCs as homotypic and heterotypic tetramers ionically bound to heparin-containing serglycin proteoglycans. The human ortholog of mMCP-6 is hTryptase-21. We generated enzymatically active mMCP-6, mMCP-7, and hTryptase-21. We also recently created inbred transgenic C57BL/6 mouse strains that differ in their expression of mMCP-6, mMCP-7, and heparin. Using the latter mice, we showed that C57BL/6 mice that lack tryptase7heparin complexes have reduced inflammation and diminished loss of aggrecan proteoglycans from their cartilage in two different mouse models of inflammatory arthritis. The overall objectives of the proposed studies are to use complementary approaches to determine at the molecular level the roles of mouse and human tetramer-forming tryptases in RA and mouse models of this autoimmune disorder. Our genetically modified mouse strains, recombinant tryptases, and other key protease-specific reagents will be used in the proposed studies to determine how MC-restricted tetramer-forming human and mouse tryptases affect fibroblast-like synoviocytes (FLS) and chondrocytes at multiple levels in varied in vitro and in vivo studies, as well as how these mesenchymal cells affect MCs. Additional studies will be carried out to determine how exocytosed mMCP-6 and hTryptase-21 are negatively regulated in the arthritic joint, and even in MCs themselves. The factors and mechanisms that control the expression of enzymatically active tryptases inside MCs have not been identified. To this end, we recently discovered that FLS-derived IL-33 induces MCs to increase their expression of tryptases at the mRNA and protein levels. Thus, additional mechanistic studies are proposed to understand how IL-33 controls the transcription of the mMCP-6 and hTryptase-21 genes in MCs and/or the stability of their transcripts. In combination, these studies promise to provide novel insights into both MC and synovial physiology, as well as elucidate novel disease mechanisms in human inflammatory arthritis. PUBLIC HEALTH RELEVANCE: We have discovered that a cell lineage called the mast cell can contribute to inflammation in rheumatoid arthritis (RA) by studying mouse models of disease. Now, we have found that a protein made by mast cells, called tryptase, is among the means by which mast cells impact disease. We are working out the mechanisms that tryptase impacts to understand better how to target this novel pathway in treating people with RA.