The antiinflammatory drug sulfasalazine inhibits tumor necrosis factor alpha expression in macrophages by inducing apoptosis

Objective. Sulfasalazine (SSZ) is a commonly used drug in the treatment of inflammatory diseases such as rheumatoid arthritis and Crohn's disease. In both diseases, the proinflammatory cytokine tumor necrosis factor alpha (TN F alpha) plays a prominent role. In these studies, we investigated the mech anism by which SSZ inhibits TNF alpha expression in macrophages and macroph age-like cell lines. Methods. Monocyte-derived macrophages and several macrophage-like cell line s were exposed to SSZ in vitro, and the effect on TNF alpha expression was monitored by reverse transcriptase-polymerase chain reaction and Western bl ot analysis. In addition, the effects of SSZ in vivo were examined by intra peritoneally injecting mice with SSZ, after which peritoneal cells were har vested and examined using various staining methods. Results. Preincubation of macrophages with SSZ, but not with methotrexate, inhibited lipopolysaccharide (LPS)-induced TNF alpha expression. Inhibition of TNF alpha expression by SSZ coincided with the induction of apoptosis, as judged by the appearance of morphologic changes typical of apoptosis, su ch as nuclear condensation and fragmentation. Induction of apoptosis by SSZ was confirmed by TUNEL analysis and by the detection of cleaved U1-70K, a substrate of caspase 3. Intraperitoneal injections of SSZ in mice resulted in the induction of apoptosis of peritoneal cells within a few hours. SSZ-i nduced cleavage of the U1-70K protein was inhibited by Zn2+ and by specific inhibitors of caspases 3 and 8, but not caspases 1 and 9. Interestingly, t he reduced expression of LPS-induced TNF alpha in the presence of SSZ was r estored by inhibition of caspase 8. Conclusion. Inhibition of TNF alpha expression in macrophages by SSZ is due to the induction of apoptosis and involves the activation of caspase 8.