Divergent Mechanisms Utilized by SOCS3 to Mediate Interleukin-10 Inhibition of Tumor Necrosis Factor α and Nitric Oxide Production by Macrophages

Abstract

The cytokine interleukin-10 (IL-10) potently inhibits macrophage function through activation of the transcription factor STAT3. The expression of SOCS3 (suppressor of cytokine signaling-3) has been shown to be induced by IL-10 in a STAT3-dependent manner. However, the relevance of SOCS3 expression to the anti-inflammatory effect of IL-10 on macrophages has been controversial. Through kinetic analysis of the requirement for SOCS3 in IL-10 inhibition of lipopolysaccharide (LPS)-stimulated tumor necrosis factor-␣ (TNF␣) transcription and translation, SOCS3 was found to be necessary for TNF␣ expression during the early phase, but not the late phase of IL-10 action. SOCS3 was essential for IL-10 inhibition of LPS-stimulated production of iNOS (inducible nitricoxide synthase) protein and nitric oxide (NO 204 and Tyr 221 were the only structural features of SOCS3 that were necessary in mediating IL-10 inhibition of iNOS protein expression and NO production. These data define SOCS3 as an important mediator of IL-10 inhibition of macrophage activation and that SOCS3 interferes with distinct LPS-stimulated signal transduction events through differing mechanisms.Macrophages form the first line of defense against foreign pathogens by producing pro-inflammatory mediators that either directly attack the invading organism, or recruit and activate other cells of the innate and acquired immune systems (1). Macrophage recognition of pathogens is mediated by pattern-recognition receptors, including the members of the Toll-like receptor family (TLR), 3 which recognize microbial structures forming pathogen-associated molecular patterns (2). A well characterized pathogen-associated molecular pattern is the cell wall component of Gram-negative bacteria called lipopolysaccharide (LPS).Binding of LPS to the TLR4 complex on macrophages leads to rapid macrophage activation. Whereas this inflammatory response is usually helpful and protective to the host, if left unchecked, the excessive amount of inflammatory cytokines produced leads to tissue destruction and pathologies such as septic shock (3).Interleukin-10 (IL-10) is a key physiological negative regulator of macrophage activation (4). Activated macrophages produce various inflammatory mediators including tumor necrosis factor-␣ (TNF␣) and nitric oxide (NO), all of which are suppressed by IL-10 (5, 6). Various animal model studies have substantiated the in vivo importance of an anti-inflammatory role for IL-10. Mice in which the IL-10 gene is deleted develop inflammatory bowel disease (7) and exaggerated immune reactions when challenged with antigen or LPS (8, 9). Administration of IL-10, however, ameliorates disease in models of endotoxemia (10), transplantation (11), and autoimmunity (12). In humans, the presence of elevated endogenous IL-10 is a positive prognostic variable in autoimmune disease (13) and allogeneic transplant patients (14).IL-10 initiates its actions on target cells by binding to the heterodimeric IL-10 receptor that belongs to the class II cytokine receptor famil...

Protective effects of plasmin(ogen) in a mouse model of Staphylococcus aureus–induced arthritis

“…The lysates were adjusted to obtain equal protein concentrations. Western blot analysis was performed as described previously (24), using a goat antibody to mouse IL-10 (AF-417-NA; R&D Systems, Abingdon, UK) and a mouse monoclonal antibody against ␤-actin (Sigma-Aldrich Sweden, Stockholm, Sweden). Horseradish peroxidaseconjugated secondary antibodies directed against goat IgG and mouse IgG were obtained from Bio-Rad (Hercules, CA).…”

Enhanced suppressor of cytokine signaling 3 in arthritic cartilage dysregulates human chondrocyte function

“…Among the various cytokines, IL-10 is a likely candidate responsible for high SOCS-3 levels. IL-10 is known to induce SOCS-3, but not SOCS-1 and SOCS-2, in neutrophils and macrophages (28,29) and is expressed in both healthy and OA chondrocytes (30). Furthermore, SOCS-3 is a strong biomechanical stress response gene, which might explain the enhanced SOCS-3 levels observed in trauma patients (31).…”

Interleukin-10 inhibits tumor necrosis factor-α production in lipopolysaccharide-stimulated RAW 264.7 cells through reduced MyD88 expression

“…The experimental result was consistent with the previous report that IL-10 exclusively inhibits an early stage of LPSinduced TNF-α production. 9 …”

“…In fact, IL-10 is known to inhibit LPSinduced TNF-α production by inhibiting activation of NF-κB 6,7 or p38 MAPK, 8 and by inducing suppressor of cytokine signaling (SOCS)-3. 9 However, the precise mechanism by which IL-10 suppresses LPS-induced TNF-α production remains controversial. In the present study, we studied an inhibitory action of IL-10 on TNF-α production in LPS-stimulated RAW 264.7 macrophage cells.…”