NF-kB activation provides the potential link between inflammation and hyperplasia in the arthritic joint (arthritisyapoptosisytumor necrosis factor ayFasygene therapy)

The transcription factor NF-kB is a pivotal regulator of inf lammatory responses. While the activation of NF-kB in the arthritic joint has been associated with rheumatoid arthritis (RA), its significance is poorly understood. Here, we examine the role of NF-kB in animal models of RA. We demonstrate that in vitro, NF-kB controlled expression of numerous inf lammatory molecules in synoviocytes and protected cells against tumor necrosis factor a (TNFa) and Fas ligand (FasL) cytotoxicity. Similar to that observed in human RA, NF-kB was found to be activated in the synovium of rats with streptococcal cell wall (SCW)-induced arthritis. In vivo suppression of NF-kB by either proteasomal inhibitors or intraarticular adenoviral gene transfer of super-repressor IkBa profoundly enhanced apoptosis in the synovium of rats with SCWand pristane-induced arthritis. This indicated that the activation of NF-kB protected the cells in the synovium against apoptosis and thus provided the potential link between inf lammation and hyperplasia. Intraarticular administration of NF-kB decoys prevented the recurrence of SCW arthritis in treated joints. Unexpectedly, the severity of arthritis also was inhibited significantly in the contralateral, untreated joints, indicating beneficial systemic effects of local suppression of NF-kB. These results establish a mechanism regulating apoptosis in the arthritic joint and indicate the feasibility of therapeutic approaches to RA based on the specific suppression of NF-kB. Inflammation and hyperplasia of the synovium are the hallmarks of rheumatoid arthritis (RA) (1). The normal synovium is a delicate tissue lining the joint capsule; however, in RA, the synovium transforms into an aggressive, tumor-like structure called the pannus (1, 2). It is a common belief that inflammation promotes hyperplasia in the synovium, but the underlying mechanisms for this are largely unknown. Impaired regulation of apoptosis has been associated with RA (3–5). Experimental evidence suggests that tumor necrosis factor a (TNFa)and Fas-mediated apoptosis is important in the regulation of hyperplasia in the synovium (5, 6). The human RA synovium contains a significant proportion of apoptotic cells (7, 8), and elevated expression of TNFa protein and functional FasL in the synovium is a common feature of RA (1, 9). Gene expression is transcriptionally controlled by inducible transcription factors. The transcription factor NF-kB (reviewed in ref. 10) is particularly important in the regulation of inflammation. In unstimulated cells, NF-kB is retained in the cytoplasm through an interaction with inhibitory proteins known as IkB. Cell stimulation causes degradation of IkB, leading to nuclear translocation of NF-kB and activation of transcription. Inducers of NF-kB include interleukin (IL)-1, TNFa, platelet-derived growth factor, lipopolysaccharide, oxidative stress, and viral products. In turn, NF-kB can activate transcription of IL-1, TNFa, IL-6, IL-8, the adhesion molecules ICAM-1 (intercellular adhesion molecule-1), VCAM-1 (vascular cell adhesion molecule 1), E-selectin, the growth factor granulocyteymacrophage colony-stimulating factor, and inducible nitric oxide synthase. Since NF-kB is activated in human RA synovium (11, 12), and the list of inducers and targets of NF-kB almost perfectly matches the profile of mediators of inflammation in RA, this suggests an important role of NF-kB in the control of inflammation in the synovium. Furthermore, the recent discovery of a protective function of NF-kB against the cytotoxicity of TNFa (13–16) suggests the involvement of this transcription factor in the regulation of apoptosis in the synovium. Here, we used animal models of RA to examine the relationship between inflammation, activation of NF-kB, and apoptosis in the synovium. We demonstrate that in primary synovial fibroblasts, NF-kB is required for induction of multiple inflammatory molecules, including IL-1b and TNFa. Specific suppression of NF-kB potentiated TNFaand FasLmediated apoptosis. Similar to that in human RA, NF-kB was found activated in the synovium of rats with streptococcal cell wall (SCW) arthritis. Suppression of NF-kB by intraarticular (i.a.) administration of proteasomal inhibitors and i.a. adenoviral gene transfer of super-repressor IkBa (srIkBa) profoundly enhanced apoptosis in the synovium of rats with SCWand pristane-induced arthritis. This indicated that activation of NF-kB by inflammation inhibited apoptosis in the synovium, thereby potentially contributing to hyperplasia. Administration (i.a.) of NF-kB decoy oligodeoxynucleotides (ODNs) significantly inhibited the severity of recurrent SCW arthritis in treated joints. Unexpectedly, the severity of arthritis also was inhibited in untreated, contralateral joints, indicating systemic therapeutic effects of local treatment. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked ‘‘advertisement’’ in accordance with 18 U.S.C. §1734 solely to indicate this fact. © 1998 by The National Academy of Sciences 0027-8424y98y9513859-6$2.00y0 PNAS is available online at www.pnas.org. Abbreviations: RA, rheumatoid arthritis; SCW, streptococcal cell wall-induced arthritis; Ad, adenoviral; PG-APS, peptidoglycan– polysaccharide complex; TNFa, tumor necrosis factor a; FasL, Fas ligand; IL, interleukin; i.a., intraarticular; ODN, oligodeoxynucleotide; CMV, cytomegalovirus; wt, wild type; mut, mutated; EMSA, electrophoretic mobility-shift assay; TUNEL, terminal deoxynucleotidyltransferase-mediated UTP end labeling; Ab, antibody; GFP, green fluorescent protein. §To whom reprint requests should be addressed at: 222 Lineberger Comprehensive Cancer Center, CB 7295, University of North Carolina, Chapel Hill, NC 27599. ¶To whom reprint requests should be addressed at: 4109 Thurston Arthritis Research Center, CB 7280, University of North Carolina, Chapel Hill, NC 27599. e-mail: [email protected].