Phosphodiesterase 4 (PDE4) regulation of proinflammatory cytokine and chemokine release from rheumatoid synovial membrane.

BACKGROUND: The cAMP-metabolising enzyme, phosphodiesterase 4 (PDE4), has been implicated in a number of immune responses, including tumour necrosis factor α (TNFα) production. To date, few data have directly addressed whether synovial cytokine and chemokine production is modified by PDE4. OBJECTIVE: Using specific PDE4 inhibitors, roflumilast plus two novel inhibitors, INH 0061 and INH 0062, the authors studied the effect of PDE4 inhibition on proinflammatory cytokine and chemokine release from primary rheumatoid arthritis (RA) synovial digest suspensions and in a macrophage T cell co-culture assay system. RESULTS: All PDE4 inhibitors dose-dependently reduced the release of TNFα from primary synovial membrane cultures (n=5), half maximal inhibitory concentration (IC(50)) 300-30 nM, p<0.05. Similarly, a significant suppression in the release the proinflammatory chemokines, monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein (MIP)-1α, MIP-1ß (IC(50) 300-30 nM) and regulated upon activation normal T-cell expressed and secreted (RANTES) (IC(50) 3 nM) was also observed, p<0.05. While interleukin 1ß was also reduced, it did not achieve an IC(50). These observations were further confirmed in a macrophage T cell co-culture system, demonstrating the importance of PDE4 pathways in regulating cytokine/chemokine release in a cellular interaction implicated in inflammatory synovitis. Subsequent studies using the human monocytic cell line U937 also demonstrated cytokine regulation with PDE4 knockdown utilising a small interfering RNA approach. CONCLUSION: These data provide direct evidence of PDE4-dependent pathways in human RA synovial inflammatory cytokine and chemokine release and may provide a novel approach in treating chronic autoimmune conditions such as RA.

Gene expression profiles in the rat streptococcal cell wall-induced arthritis model identified using microarray analysis.

Experimental arthritis models are considered valuable tools for delineating mechanisms of inflammation and autoimmune phenomena. Use of microarray-based methods represents a new and challenging approach that allows molecular dissection of complex autoimmune diseases such as arthritis. In order to characterize the temporal gene expression profile in joints from the reactivation model of streptococcal cell wall (SCW)-induced arthritis in Lewis (LEW/N) rats, total RNA was extracted from ankle joints from naive, SCW injected, or phosphate buffered saline injected animals (time course study) and gene expression was analyzed using Affymetrix oligonucleotide microarray technology (RAE230A). After normalization and statistical analysis of data, 631 differentially expressed genes were sorted into clusters based on their levels and kinetics of expression using Spotfire profile search and K-mean cluster analysis. Microarray-based data for a subset of genes were validated using real-time PCR TaqMan analysis. Analysis of the microarray data identified 631 genes (441 upregulated and 190 downregulated) that were differentially expressed (Delta > 1.8, P < 0.01), showing specific levels and patterns of gene expression. The genes exhibiting the highest fold increase in expression on days -13.8, -13, or 3 were involved in chemotaxis, inflammatory response, cell adhesion and extracellular matrix remodelling. Transcriptome analysis identified 10 upregulated genes (Delta > 5), which have not previously been associated with arthritis pathology and are located in genomic regions associated with autoimmune disease. The majority of the downregulated genes were associated with metabolism, transport and regulation of muscle development. In conclusion, the present study describes the temporal expression of multiple disease-associated genes with potential pathophysiological roles in the reactivation model of SCW-induced arthritis in Lewis (LEW/N) rat. These findings improve our understanding of the molecular events that underlie the pathology in this animal model, which is potentially a valuable comparator to human rheumatoid arthritis (RA).