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.