Innate Immunity in Experimental Arthritis of Kininogen

激肽原实验性关节炎的先天免疫

• 批准号: 6838311
• 负责人: Robert W Colman
• 金额: $ 27.39万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-10 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6838311
• 关键词: clinical research; cytokine; disease /disorder model; gene targeting; genetic susceptibility; genetically modified animals; human subject; immunity; inflammation; kallikreins; kininogens; laboratory mouse; laboratory rat; molecular weight; monoclonal antibody; monocyte; neutrophil; pathologic process; patient oriented research; peptidoglycan; point mutation; polysaccharides; rheumatoid arthritis

DESCRIPTION (provided by applicant): Rheumatoid arthritis (RA) is an immtmologically mediated, familial, chronically relapsing disease eventually resulting in joint destruction. Understanding inflammatory mediators is critical because these molecules provide approaches for pharmacologic blockade by specific inhibitors and receptor antagonists. Recent rodent models of arthritis have shown that bacterial cell wall polymers can induce acute and chronic arthritis in genetically susceptible rat strains. Parenterally injected peptidoglycanpolysaccharide complexes (PG-APS) induce a systemic inflammatory response consisting of erosive polyarthritis, hepatic granulomas, and anemia. We have shown that experimental inflammatory arthritis is associated with in vivo activation of the kallikrein-kinln system (KKS) in Lewis rats and is selectively inactivated in these susceptible Lewis rats, but not in resistant Buffalo rats. A specific kallikrein inhibitor decreases both the KKS activation and acute inflammatory changes (edema, neutrophil infiltration) and prevented arthritis and the systemic complications (splenomegaly, hepatomegaly, leukocytosis and the acute phase reaction) in the PG-APS model. We now focus on the role of innate immune mechanisms in models of rodent inflammatory arthritis, specifically the interaction of the KKS with monocytes and neutrophils resulting in release of cytokines. We explore two major hypothesis-using approaches new to this laboratory. First, we hypothesize that the lack of high molecular weight kininogen (HK), which is required for activation of plasma kallikrein, is the substrate for release of bradykinin and will modulate the acute and chronic arthritis and systemic inflammation produced by PG-APS in genetically susceptible Lewis rats. We have found that these rats have a single point mutation $511N leading to N-glycosylation which increases the susceptibility of HK to proteolysis. We will examine the mechanisms by which accelerated release of bradykinin and cleaved HK (HKa) in Lewis rats will augment inflammatory cytokine release by the innate immune system, particularly macrophages, stimulated through cellular receptors for bradykinin and HKa on leukocytes. This hypothesis will be explored further using a rat line with a single point mutation in kininogen leading to a block in hepatic secretion and severe plasma deficiency, which we have placed onto the susceptible Lewis genotype. We will also use the human HLA-B27 transgenic rat as a model of "spontaneous" chronic arthritis, which we will treat with a monoclonal antibody to HK. Second, we will compare the degree of inflammation in Lewis vs. Fischer rats with variable degrees of kininogen cleavage I during experimental arthritis and systemic inflammation induced by PG-APS. We will test whether modulation of inflammation by] recombinant HK polypeptides, synthetic peptides, or antibodies to that protein, which are known to inhibit binding of HK or HKa to] leukocytes, can prevent many of the inflammatory changes. We will examine the role of HKa and bradykinin using knockouts of their binding proteins, uPAR and B2R, in murine arthritis and systemic inflammation. If these approaches result in a substantial decrease in arthritis and systemic inflammation, it could lead to the control of rheumatic diseases with potentially fewer side effects than current therapeutic approaches.

描述（由申请人提供）： 类风湿性关节炎（RA）是一种免疫介导的、家族性、慢性复发性疾病，最终导致关节破坏。了解炎症介质至关重要，因为这些分子提供了通过特定抑制剂和受体拮抗剂进行药理学阻断的方法。最近的关节炎啮齿动物模型表明，细菌细胞壁聚合物可以在遗传易感的大鼠品系中诱发急性和慢性关节炎。肠胃外注射肽聚糖多糖复合物（PG-APS）可诱导全身炎症反应，包括糜烂性多关节炎、肝肉芽肿和贫血。我们已经证明，实验性炎症性关节炎与 Lewis 大鼠体内激肽释放酶-激肽系统 (KKS) 的激活有关，并且在这些易感 Lewis 大鼠中选择性失活，但在耐药性 Buffalo 大鼠中则不然。在 PG-APS 模型中，特定的激肽释放酶抑制剂可减少 KKS 激活和急性炎症变化（水肿、中性粒细胞浸润），并预防关节炎和全身并发症（脾肿大、肝肿大、白细胞增多和急性期反应）。我们现在关注先天免疫机制在啮齿类炎症性关节炎模型中的作用，特别是 KKS 与单核细胞和中性粒细胞的相互作用，导致细胞因子的释放。我们探索了本实验室的两种主要假设使用方法。首先，我们假设血浆激肽释放酶激活所需的高分子量激肽原 (HK) 的缺乏是缓激肽释放的底物，并将调节遗传易感性 Lewis 大鼠中 PG-APS 产生的急性和慢性关节炎以及全身炎症。我们发现这些大鼠有一个单点突变 $511N，导致 N-糖基化，从而增加了 HK 对蛋白水解的敏感性。我们将研究Lewis大鼠中缓激肽和裂解HK (HKa)的加速释放将增强先天免疫系统（特别是巨噬细胞）通过白细胞上的缓激肽和HKa细胞受体刺激的炎症细胞因子释放的机制。我们将使用激肽原单点突变导致肝分泌受阻和严重血浆缺乏的大鼠系进一步探讨这一假设，我们已将其置于易感的 Lewis 基因型上。我们还将使用人类 HLA-B27 转基因大鼠作为“自发性”慢性关节炎模型，并用 HK 单克隆抗体进行治疗。其次，我们将比较具有不同程度的激肽原裂解 I 的 Lewis 大鼠与 Fischer 大鼠在实验性关节炎和 PG-APS 诱导的全身炎症过程中的炎症程度。我们将测试重组HK多肽、合成肽或该蛋白质的抗体（已知这些抗体可抑制HK或HKa与白细胞的结合）对炎症的调节是否可以预防许多炎症变化。我们将通过敲除 HKa 和缓激肽的结合蛋白 uPAR 和 B2R，研究它们在小鼠关节炎和全身炎症中的作用。如果这些方法能够显着减少关节炎和全身炎症，那么它可能会控制风湿性疾病，并且比目前的治疗方法副作用可能更少。