SIGNALING PATHWAYS CONTROLLING NF-KB IN RHEUMATIOID ARTHRITIS

类风湿性关节炎（RA）是一种典型的慢性炎症性疾病。我们在RA动物模型中的初步研究表明，NF-κ B在关节炎滑膜的炎症、细胞凋亡和增殖的调节中起着至关重要的作用。因此，NF-κ B成为RA和其他慢性炎症性疾病治疗干预的非常有吸引力的靶点。抑制NF-κ B活化的最合理的方法是调节控制NF-κ B转录活性的信号级联。两个信号级联，NIK/IKK和p38 MAP激酶通路，在RA中NF-κ B的调节中特别重要。我们对这些通路的生理功能的了解非常有限，主要是由于实验方法不足。本提案的主要目的是确定NIK/IKK和p38信号通路在RA中NF-κ B活化中的生理作用，并评估这些通路在RA主要表现（即炎症、侵袭性滑膜的肿瘤样扩张以及骨和软骨吸收）中的作用，从而评估这些通路作为治疗干预的靶点。我们将采用基因转移技术来剖析NIK/IKK和p38 MAPK在治疗干预中的作用。我们将采用基因转移技术来剖析NIK/IKK和p38 MAPK通路在RA病理中NF-κ B活化中的作用。使用关节内（i.a.）显性阴性（DN）抑制剂的基因转移应该允许清楚地解释这些途径在RA中的作用，并将验证这些途径作为药物发现的靶点。在目的1中，我们将研究NIK/IKK和p38通路在体外滑膜成纤维细胞和单核细胞中NF-κ B活化、炎症和促有丝分裂反应以及细胞凋亡中的作用。接下来，我们将评估NIK/IKK和p38通路在大鼠SCW关节炎中的NF-κ B激活、炎症、增生以及骨和软骨吸收中的作用（AIM 2）。这些数据与人类疾病的相关性将通过使用RA的SCID小鼠/人模型来检查（目的3）。这些实验将确定NF-κ B、NIK/IKK和p38通路在调节人RA滑膜炎症、凋亡和软骨破坏中的作用。我们在动物关节炎中的初步研究的一个意想不到的结果是观察到滑膜中NF-κ B的局部抑制不仅在治疗的关节中而且在未治疗的对侧关节中改善了疾病。这表明通过局部治疗减轻疾病的全身表现是可行的。目的4用于探索这种效应的两种假定机制。我们将研究局部抑制NF-κ B对循环T细胞中促炎和抗炎性Th 1和Th 2亚群平衡的影响，以及神经原性机制。

Rheumatoid arthritis (RA) is a prototype of chronic inflammatory disease. Our preliminary studies in animal models of RA demonstrated crucial involvement of NF-kappaB in regulation of inflammation, apoptosis, and proliferation in the arthritic synovium. Thus, NF-kappaB emerges as very attractive target for therapeutic intervention in RA and other chronic inflammatory conditions. The most logical way to inhibit NF-kappaB activation is to modulate the signaling cascades which controls transcriptional activity of NF-kappaB. Two signaling cascades, the NIK/IKK and p38 MAP kinase pathways, are particularly important in regulation of NF-kappaB in RA. Our knowledge of the physiological function of these pathways is very limited, mainly due to inadequate experimental approaches. The major purpose of this proposal is to determine the physiological role of the NIK/IKK and p38 signaling pathways in activation of NF-kappaB in RA, and to assess the contribution of these pathways in major manifestations of RA, i.e. inflammation, tumor-like expansion of invasive synovium, and bone and cartilage resorption, and thus evaluate these pathways as targets for therapeutic intervention. We will employ gene transfer technology for dissecting the role of the NIK/IKK and p38 MAPK for therapeutic intervention. We will employ gene transfer technology for dissecting the role of the NIK/IKK and p38 MAPK pathways in NF-kappaB activation in the RA pathology. Using intraarticular (i.a.) gene transfer of dominant negative (DN) inhibitors should allow for the clear-cut interpretation of the role of these pathways in RA and will validate these pathways as targets for drug discovery. In Aim 1, we will examine the role of the NIK/IKK and p38 pathways in NF-kappaB activation, inflammatory and mitogenic responses, and apoptosis on the cellular level in synovial fibroblasts and monocytic cells in vitro. Next, we will assess the role of the NIK/IKK and p38 pathways in vivo in NF-kappaB activation, inflammation, hyperplasia, and bone and cartilage resorption in SCW arthritis in rats (AIM 2). The relevance of these data to human disease will be examined by using a SCID mice/human models of RA (Aim 3). These experiments will determine the role of NF-kappaB, and the NIK/IKK and p38 pathways in regulation of inflammation, apoptosis, and cartilage destruction in human RA synovium. One unexpected result of our preliminary studies in animal arthritis was observation that local suppression of NF-kappaB in the synovium ameliorated disease not only in treated, but also in untreated, contralateral joints. This indicates the feasibility of alleviating systemic manifestations of the disease through local treatment. Aim 4 serves to explore two putative mechanisms underlying this effect. We will examine the influence of local suppression of NF-kappaB on the balance of pro- and anti-inflammatory TH1 and Th2 subsets in circulating T cells, and in neurogenic mechanisms.