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SOCS1 Regulates the IFN but Not NFκB Pathway in TLR-Stimulated Human Monocytes and Macrophages

Prêle, C.M., Woodward, E.A., Bisley, J., Keith-Magee, A., Nicholson, S.E. and Hart, P.H. (2008) SOCS1 Regulates the IFN but Not NFκB Pathway in TLR-Stimulated Human Monocytes and Macrophages. The Journal of Immunology, 181 (11). pp. 8018-8026.

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SOCS1 can regulate TLR-mediated signal transduction, yet mechanistic studies in murine macrophages have been confusing and contradictory. This study has used an adenoviral transfection system to determine the role of SOCS1 in the regulation of TNF-α production by activated human monocytes. Monocytes were infected with AdV-SOCS1 or with an empty vector control, AdV-GFP, for 24 h before activation with the TLR4 ligand, LPS. SOCS1 did not regulate TNF-α mRNA or protein production within the first two hours of TLR4 activation. However, SOCS1 suppressed the sustained production of TNF-α by primary human monocytes and synovial fluid macrophages ex vivo. In addition, SOCS1 regulated the production of IL-6, but not IL-10, by monocytes. Analysis of the early signaling pathway downstream of TLR4 demonstrated that SOCS1 had no regulatory effect on the activation or on the DNA binding capacity of NFκB. The late effects of LPS are mediated in part through the MyD88-independent pathway activating IRF3 and initiating the production of IFN-β. In response to adenoviral infection and before LPS exposure, monocytes expressed enhanced levels of IFN-β and Myxovirus A mRNA, an anti-viral molecule characterizing IFN-β activity. These two genes were reduced in AdV-SOCS1-infected cells. Further, SOCS1 regulated IFN-dependent pathways in LPS-activated cells as evidenced by reduced IFN-β production and STAT1 phosphorylation. Using AdV-infection to dissect SOCS1 control of IFN-dependent pathways, this study suggests that SOCS1-regulation of the IFN-dependent component of the LPS-induced TLR4 signaling pathway may contribute to the down-regulation of inflammatory cytokine production by AdV-SOCS1-infected human monocytes.

Chronic inflammatory diseases such as rheumatoid arthritis, Crohn’s disease, and inflammatory bowel disease are characterized by prolonged and deregulated proinflammatory cytokine production. TNF-α is arguably the most important proinflammatory cytokine driving the inflammatory response, initiating a cascade of events that controls the sequential production of other inflammatory cytokines such as IL-1 and IL-6. Although anti-TNF-α therapies have been successful at reducing inflammation, the side effects associated with such treatments, such as the risk of sepsis, have prompted the development of therapeutic strategies to specifically target the intracellular signaling intermediates responsible for regulating TNF-α production. Development of this approach requires a better understanding of the signaling pathways initiating and regulating TNF-α production in human cells.

TLRs are a family of membrane bound pattern recognition receptors that respond to invading pathogens to initiate an immune response (reviewed in Ref. 1). TLR4 is expressed on macrophages, and expression levels and activation through this receptor is increased in synovial fluid macrophages from patients with rheumatoid arthritis (2). TLRs recognize many endogenous ligands (such as fibronectin/hyaluronan fragments, self-mRNA, and high mobility group box 1) (3). It has been hypothesized that once joint damage occurs, a self-perpetuating process mediated by endogenous ligands through TLR-activation is established, resulting in chronic progressive disease.

The signaling pathways initiated downstream of the TLR, as a consequence of ligand binding, are responsible for driving the production of TNF-α. Specificity of signaling is determined by ligand-specific receptor usage and the differential recruitment of the adaptor proteins MyD88, MyD88-adaptor-like (Mal),3 toll/interleukin-1 receptor domain-containing adaptor protein inducing IFN-β (TRIF), and TRIF-related adaptor molecule (TRAM) which are required for signal amplification (4, 5). For example, the TLR4 ligand LPS initiates two signal transduction pathways, the MyD88-dependent and a MyD88-independent, TRIF-dependent pathway. The early production of TNF-α, i.e., within the first few hours of receptor activation, is due to the rapid activation of the NFκB pathway. NFκB is sequestered in an inactive form in the cytoplasm via its association with the inhibitory κB (IκB) complex. Following activation, NFκB is released into the cytoplasm and translocates to the nucleus where it initiates gene transcription. TLR4 signaling via the MyD88-independent pathway requires the recruitment of TRIF and TRAM and contributes to the sustained production of TNF-α. Activation of this pathway culminates in the activation of IRF3, a member of the IRF family of transcription factors which regulate the expression of type I IFNs such as IFN-α/β (6). The induction of IFN-β production by IRF3 is central to the initiation of the anti-viral immune response (7, 8, 9, 10).

A number of recent reports have suggested that TLR-mediated signaling can be regulated by members of the suppressor of cytokine signaling (SOCS) family of proteins. The SOCS proteins are a family of eight proteins that include cytokine-inducible SH2-domain containing protein and SOCS1–7. SOCS1 was first characterized as a regulator of IFN-γ-induced inflammatory responses (11, 12), but more recent studies have suggested that the regulatory capacities of SOCS1 are not restricted to the regulation of IFN. Recently there has been much interest in the ability of SOCS1 to regulate TLR-mediated signal transduction and the subsequent production of proinflammatory cytokines (13, 14, 15, 16, 17). SOCS1 expression is increased in macrophages following LPS exposure (13). Furthermore, studies in bone marrow-derived macrophages suggest negative regulation of TLR-mediated signaling by SOCS1. The mechanisms underlying SOCS1 regulation of proinflammatory cytokine production remain unclear. Studies in murine macrophages have suggested control of NFκB signaling by SOCS1 (14, 15, 16, 17), whereas others suggest that SOCS1 regulates IRF3-driven processes (13, 18, 19, 20).

The capacity of SOCS1 to regulate TLR-induced proinflammatory cytokine production by activated human monocytes and macrophages has not been investigated. In this study, infection of human monocytes with a SOCS1-expressing adenovirus did not regulate TNF-α mRNA or protein production within the first two hours of TLR activation. However, by 24 h, TLR-induced TNF-α levels were significantly reduced. Our results suggest that this effect may be due to regulation of the MyD88-independent TRIF-dependent pathway. Furthermore, AdV-SOCS1 control of TNF-α production was confirmed ex vivo in primary human macrophages obtained from the synovial fluid of patients with inflammatory arthritis.

Item Type: Journal Article
Publisher: American Association of Immunologists
Copyright: © 2008 by The American Association of Immunologists
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