Session Title: Cytokines, Mediators, and Gene Regulation I
Session Type: Abstract Submissions (ACR)
Background/Purpose: Citrullination is a post-translational modification that is the conversion of arginine to citrulline in proteins mediated by peptidylarginine deiminase (PAD). Antibodies directed towards the citrullinated proteins are highly specific for rheumatoid arthritis (RA). We and others have shown that some chemokines including ENA-78/CXCL5 play important roles in the development of RA. We undertook this study to examine whether citrullinated ENA-78/CXCL5 (citENA-78/CXCL5) is detected in RA biological fluids, and if so, what its biological activity is.
Methods: An expression plasmid containing polyhistidine tagged human ENA-78/CXCL5 was transfected into the human embryonic kidney 293T cell line for ENA-78/CXCL5 production. After citrullination of the ENA-78/CXCL5 with rabbit PAD, citENA-78/CXCL5 was used as standard for enzyme linked immunosorbent assay (ELISA). The concentrations of citENA-78/CXCL5 in synovial fluids (SFs) or sera were measured by ELISA using anti-modified citrulline antibody. The citENA-78/CXCL5 levels in RA were compared with osteoarthritis (OA) and other inflammatory rheumatic diseases, and the correlation between the citENA-78/CXCL5 levels and clinical data was analyzed. Monocyte and polymorphonuclear neutrophil (PMN) chemotaxis assays were performed using a 48-well Boyden chamber system to examine the biological activity of citENA-78/CXCL5 compared to ENA-78/CXCL5. C57BL/6 mice were injected intra-articularly with ENA-78/CXCL5 or citENA-78/CXCL5 to induce inflammation and the severity of inflammation was evaluated to compare the biological activity of citENA-78/CXCL5 to ENA-78/CXCL5.
Results: CitENA-78/CXCL5 was significantly higher in RA (n=11, mean±SE; 286.3±68.0 pg/ml) than NL sera (n=15, 1.2±2.6 pg/ml) and higher in RA (n=20, 1126.4±296.6 pg/ml) compared to other inflammatory diseases (n=14, 14.1±8.2 pg/ml) and OA (n=15, 2.3±1.0 pg/ml) SFs. There was no significant correlation between ENA-78/CXCL5 levels and clinical data. On the other hand, there were significant positive correlations between citENA-78/CXCL5 and C-reactive protein (CRP) levels (RA; n=14, r=0.69, p<0.05, RF positive RA; n=10, r=0.86, p<0.05), citENA-78/CXCL5 and erythrocyte sedimentation rate (ESR) (RA; n=15, r=0.77, p<0.05, RF positive RA; n=11, r=0.71, p<0.05). Chemotaxis assays showed that PMN migration in response to citENA-78/CXCL5 was similar to that induced by ENA-78/CXCL5. However, citENA-78/CXCL5 induced monocyte migration, but ENA-78/CXCL5 did not. In vivo, citENA-78/CXCL5 induced more intraarticular inflammation compared to ENA-78/CXCL5.
Conclusion: CitENA-78/CXCL5 was detected in RA biological fluids and the concentrations were significantly higher in RA than OA or other diseases, and correlated with the inflammatory markers CRP and ESR. CitENA-78/CXCL5 induced monocyte migration while ENA-78/CXCL5 did not. CitENA-78/CXCL5 induced more severe joint inflammation compared to ENA-78/CXCL5. This may be attributed to the fact that citENA-78/CXCL5 acquired a monocyte recruiting function that ENA-78/CXCL5 does not have. These results indicate that citENA-78/CXCL5 may have novel inflammatory properties compared to ENA-78/CXCL5 in RA pathogenesis.
P. P. Tak,
J. H. Ruth,
D. L. Baeten,
D. M. Gerlag,
M. A. Amin,
A. E. Koch,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/citrullination-of-ena-78cxcl5-results-in-conversion-from-a-non-monocyte-recruiting-to-a-monocyte-recruiting-chemokine/