Interleukin‐18 enhances monocyte tumor necrosis factor α and interleukin‐1β production induced by direct contact with T lymphocytes: Implications in rheumatoid arthritis

Abstract

Objective At sites of inflammation, T cells exert pathologic effects through direct contact with monocyte/macrophages, inducing massive up-regulation of interleukin-1 (IL-1) and tumor necrosis factor α (TNFα). We examined the regulatory effects of IL-18 on monocyte activation by direct contact with T lymphocytes in rheumatoid arthritis (RA). Methods Activated T cells were isolated from RA synovial fluid. Resting T cells and monocytes were isolated from peripheral blood mononuclear cells. RA synovial T cells or phytohemagglutinin (PHA)–stimulated T cells were fixed by paraformaldehyde and then cocultured with monocytes at a ratio of 4:1. Levels of TNFα, IL-1β, IL-10, and IL-18 were measured by enzyme-linked immunosorbent assay. Expression of adhesion molecules, IL-18 receptor, and TNF receptors was analyzed by flow cytometry. Expression of NF-κB p65, phosphorylated IκBα, and phosphatidylinositol 3-kinase (PI 3-kinase) p110 was analyzed by Western blotting. Results IL-18 dose-dependently enhanced the production of IL-1β and TNFα, but not IL-10, by monocytes following contact with RA synovial T cells or PHA-prestimulated T cells. NF-κB inhibitors N-acetyl-L-cysteine and Bay 11-7085 and PI 3-kinase inhibitor LY294002 inhibited the enhancing effects of IL-18, but MAPK p38 inhibitor SB203580, ERK inhibitor PD98059, and JNK inhibitor SP600125 did not. Increased levels of NF-κB in the nucleus, phosphorylated IκB, and PI 3-kinase were confirmed in monocytes cocultured with PHA-prestimulated T cells, and the levels were further increased by stimulation with IL-18. Neutralizing antibody to IL-18 inhibited monocyte activation induced by direct contact with PHA-prestimulated T cells. Via cell–cell contact, PHA-prestimulated T cells increased autocrine production of IL-18 by monocytes, which was mediated by activation of the NF-κB and PI 3-kinase pathways, and up-regulated the expression of the IL-18 receptor in monocytes. IL-18 up-regulated the expression of the TNF receptors vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1) on monocytes. Blocking the binding of the TNF receptors VCAM-1 or ICAM-1 on monocytes to their ligands on stimulated T cells suppressed the IL-18–enhanced production of TNFα and IL-1β in monocytes induced by contact with PHA-prestimulated T cells. Conclusion IL-18 augments monocyte activation induced by contact with activated T cells in RA synovitis, which is dependent on activation of the NF-κB and PI 3-kinase pathways. IL-18 up-regulates the expression of the TNF receptors VCAM-1 and ICAM-1 on monocytes, which mediate the enhancing effects of IL-18 on T cell–monocyte contact.