Plasma kininogen and kininogen-cleaving proteases in arthritis

关节炎中的血浆激肽原和激肽原裂解蛋白酶

• 批准号: 8514307
• 负责人: Yi Wu
• 金额: $ 7.65万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8514307
• 关键词: Ablation; Address; Adult; Animals; Arthritis; Attenuated; Autoantibodies; Bradykinin; Chronic; Cleaved cell; Clinical Research; Coupling; Development; Disease; Dissection; Etiology; Event; Factor XI; Factor XII; Future; Generations; Genes; Genetic; Goals; High-Molecular-Weight Kininogen; Human; Immune response; Immune system; In Vitro; Indium; Inflammatory; Investigation; Kallikrein-Kinin System; Kininogens; Kinins; Knockout Mice; Knowledge; Mediator of activation protein; Messenger RNA; Modeling; Molecular; Mouse Strains; Mus; Pathogenesis; Patients; Peptide Hydrolases; Plasma; Plasma Kallikrein; Play; Prekallikrein; Production; Prostate-Specific Antigen; Rattus; Reaction; Regulation; Research; Resistance; Rheumatoid Arthritis; Role; Series; Serine Protease; Severities; Surface; System; Testing; Therapeutic; United States; Wild Type Mouse; base; chemokine; chronic pain; cytokine; enzyme substrate; human disease; improved; in vivo; inhibitor/antagonist; innovation; insight; joint injury; monocyte; mouse model; mutant; novel; novel therapeutics; public health relevance; receptor; research study

DESCRIPTION (provided by applicant): Experimental and clinical studies have suggested that the activation of plasma kallikrein-kinin system (KKS) is involved in the pathogenesis f arthritis. The KKS consists of 3 serine proteases, prekallikrein (PK), factor XII (FXII), facto XI (FXI), and a non-enzymatic co-factor, high-molecular-weight kininogen (HK). HK plays an important role in the assembly of this system on activation surfaces. In vitro, the active form of PK, FXII and FXI cleave HK to generate bradykinin and kinin-free HK (called as HKa). Both bradykinin and HKa possess multiple proinflammatory functions. Elevated activities of the KKS components and increased cleavage of HK have been detected in plasma from patients with RA and animals bearing arthritis. In Lewis rat model of arthritis, either HK deficiency or the blockad of HK cleavage inhibits arthritis. Our long term goal is to determine the pathogenic role of the KKS in arthritis and the underlying mechanisms. Understanding the regulation of arthritis by the KKS should provide novel insights into the pathogenesis of arthritis and the systemic complications. In our preliminary studies, we have generated a new mouse strain of HK deficiency (Kng1-/-) by deleting the Kng1 gene. We found that Kng1-/- mice display reduced arthritis. In this proposal, we hypothesize that HK and HK-cleaving proteases play an important role in the pathogenesis of arthritis. Because the KKS of the mouse, compared to the rat, more closely resembles that of the human, our Kng1-/- mouse model becomes a critical approach for evaluating the role of the KKS in arthritis. In this application, our hypothesis will be tested through the following specific aims: In Aim 1, we plan to compare the severity of joint inflammation, the levels of cytokines/chemokines and the activities of the KKS components in plasma, and the expression of cytokine mRNA in monocytes between wild type and Kng1-/- mice. We will examine whether genetic ablation of HK confers protection of arthritis, down regulates cytokines/chemokines production, and decreases the in vivo activation of the KKS. This study will reveal the essential role of HK in the pathogenesis of arthritis. In Aim 2, we will determine which proteases are responsible for HK cleavage in arthritis. Although the active form of plasma kallikrein, FXII and FXI can cleave H in purified systems, no studies have ever been performed to demonstrate which proteases cleave HK in vivo, especially in the setting of arthritis. We have obtained the knockout mice lacking each protease of the KKS, which are in the same genetic background with the Kng1-/- mice. We will determine the requirement of each protease for HK cleavage in vivo. This proposal which utilizes a series of genetically deficient mouse models is highly innovative, because it is the first genetic dissection of the role of HK and HK-cleaving proteases in arthritis. The proposed studies will improve in depth understanding of the contribution of the KKS to the pathogenesis of arthritis, reveal new targets to battle RA, and initiate further studie of the underlying cellular and molecular mechanisms.

描述（由申请人提供）：实验和临床研究表明，血浆激肽释放酶-激肽系统（KKS）的激活参与关节炎的发病机制。KKS由3种丝氨酸蛋白酶组成：前激肽释放酶（PK）、因子XII（FXII）、因子XI（FXI）和非酶促辅因子高分子量激肽原（HK）。 HK在活化表面上组装该系统中起着重要作用。在体外，PK、FXII和FXI的活性形式裂解HK以产生缓激肽和无激肽HK（称为HKa）。缓激肽和HKa都具有多种促炎功能。 在RA患者和患有关节炎的动物的血浆中检测到KKS组分的活性升高和HK的裂解增加。在刘易斯大鼠关节炎模型中，HK缺乏或HK切割阻断抑制关节炎。我们的长期目标是确定KKS在关节炎中的致病作用及其潜在机制。了解KKS对关节炎的调节，将为关节炎的发病机制和全身并发症提供新的见解。 在我们的初步研究中，我们已经产生了一个新的小鼠品系的HK缺陷（Kng 1-/-）通过删除Kng 1基因。我们发现Kng 1-/-小鼠显示关节炎减少。 在这个提议中，我们假设HK和HK裂解蛋白酶在关节炎的发病机制中起重要作用。 由于小鼠的KKS与大鼠相比更接近于人类的KKS，因此我们的Kng 1-/-小鼠模型成为评估KKS在关节炎中作用的关键方法。在本申请中，我们的假设将通过以下具体目标进行测试：在目标1中，我们计划比较野生型和Kng 1-/-小鼠之间关节炎症的严重程度、血浆中细胞因子/趋化因子的水平和KKS组分的活性以及单核细胞中细胞因子mRNA的表达。我们将研究是否遗传消融HK赋予关节炎的保护，下调细胞因子/趋化因子的生产，并减少在体内激活的KKS。本研究将揭示HK在糖尿病发病机制中的重要作用， 关节炎 在目标2中，我们将确定哪些蛋白酶负责关节炎中的HK裂解。 虽然血浆激肽释放酶的活性形式FXII和FXI可以在纯化的系统中切割H，但从未进行过研究来证明哪些蛋白酶在体内切割HK，特别是在关节炎的情况下。我们获得了KKS基因各蛋白酶缺失的基因敲除小鼠，它们与Kng 1-/-小鼠具有相同的遗传背景。我们将确定每种蛋白酶在体内切割HK的需求。这项利用一系列遗传缺陷小鼠模型的提议是高度创新的，因为它是第一次对HK和HK切割蛋白酶在关节炎中的作用进行遗传解剖。这些研究将有助于深入了解KKS在关节炎发病机制中的作用，揭示对抗RA的新靶点，并启动对潜在细胞和分子机制的进一步研究。