Inflammatory Cytokines IL-32 and IL-17 Have Common Signaling Intermediates despite Differential Dependence on TNF-Receptor 1

Department of Internal Medicine, Manitoba Centre for Proteomics and Systems Biology, University of Manitoba, Winnipeg, Manitoba R3E3P4, Canada.
The Journal of Immunology (Impact Factor: 4.92). 06/2011; 186(12):7127-35. DOI: 10.4049/jimmunol.1002306
Source: PubMed


Cytokines IL-32 and IL-17 are emerging as critical players in the pathophysiology of immune-mediated chronic inflammatory diseases. It has been speculated that the molecular mechanisms governing IL-32- and IL-17-mediated cellular responses are differentially dependent on the TNF pathway. In this study, kinome analysis demonstrated that following stimulation with cytokine IL-32, but not IL-17, there was increased phosphorylation of a peptide target corresponding to TNF-R1. Consistent with this observation, blocking TNF-R1 resulted in a suppression of IL-32-induced downstream responses, indicating that IL-32-mediated activity may be dependent on TNF-R1. In contrast, blocking TNF-R1 did not affect IL-17-induced downstream responses. Kinome analysis also implicated p300 (transcriptional coactivator) and death-associated protein kinase-1 (DAPK-1) as signaling intermediates for both IL-32 and IL-17. Phosphorylation of p300 and DAPK-1 upon stimulation with either IL-32 or IL-17 was confirmed by immunoblots. The presence of common targets was supported by results demonstrating similar downstream responses induced in the presence of IL-32 and IL-17, such as transcriptional responses and the direct activation of NF-κB. Furthermore, knockdown of p300 and DAPK-1 altered downstream responses induced by IL-32 and IL-17, and impacted certain cellular responses induced by TNF-α and IL-1β. We hypothesize that p300 and DAPK-1 represent nodes where the inflammatory networks of IL-32 and IL-17 overlap, and that these proteins would affect both TNF-R1-dependent and -independent pathways. Therefore, p300 and DAPK-1 are viable potential therapeutic targets for chronic inflammatory diseases.

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Available from: Ka-Yee Grace Choi, Oct 06, 2015
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    • "In addition, the absence of IL-32γ effects on MCL-1 transcription in neutrophils eliminates certain activation pathways involved in survival such as NF-κB. Although IL-32 has been shown to activate the NF-κB pathway, the presence of NF-κB binding sites described in the promoter region of MCL-1 has not been demonstrated in neutrophils [30], [74], [75], [76]. Therefore, the anti-apoptotic effect of IL-32γ on neutrophils implicates a post-transcriptional modulation in which the p38 MAPK cascade appears to be the main regulator of MCL-1 protein levels. "
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    ABSTRACT: IL-32γ is a multifunctional cytokine involved in various inflammatory and auto-immune diseases in which neutrophils can affect the evolution of these diseases. To persist at inflammatory sites, neutrophils require inhibition of their rapid and constitutive apoptosis, an inhibitory effect that phlogogenic cytokines support. To date, the effects of IL-32γ on neutrophils remain unknown. We demonstrate that IL-32γ delays, in a dose-dependent manner, the spontaneous apoptosis of human blood neutrophils by activating mainly p38 MAPK through rapid p38 phosphorylation. PI3-K and ERK1/2 MAPK are also involved, but to a lesser extent. Most of cytokines that induce retardation of neutrophil apoptosis activate the expression of MCL-1 at both mRNA and protein levels. IL-32γ added to human blood neutrophils in vitro is associated with sustained levels of MCL-1 protein. This effect in neutrophils corresponds to a decrease of MCL-1 protein degradation without any effect on MCL-1 mRNA levels. The sustained levels of MCL-1 induced by IL-32γ are only abrogated by the p38β MAPK inhibitor SB202190. Additionally, IL-32γ induces a reduction in caspase 3 activity in neutrophils. In conclusion, IL-32γ affects human blood neutrophils in vitro by increasing their survival, suggesting that this cytokine could have profound effects on the deleterious functions of neutrophils in several diseases.
    PLoS ONE 10/2014; 9(10):e109256. DOI:10.1371/journal.pone.0109256 · 3.23 Impact Factor
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    • "As a result, kinases represent a top priority of the pharmaceutical industry [9] and are currently the most frequently targeted gene class for cancer therapies, second only to G protein-coupled receptors across all therapeutic areas [10]. Increased appreciation for the intimate link between kinases and health has prompted the development of technologies to characterize the phosphoproteome or kinome [11], including efforts by our group to utilize peptide arrays for low-cost, high-throughput kinomic characterizations [12-16]. "
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    ABSTRACT: Recently, questions have been raised regarding the ability of animal models to recapitulate human disease at the molecular level. It has also been demonstrated that cellular kinases, individually or as a collective unit (the kinome), play critical roles in regulating complex biology. Despite the intimate relationship between kinases and health, little is known about the variability, consistency and stability of kinome profiles across species and individuals. As a preliminary investigation of the existence of species- and individual-specific kinotypes (kinome signatures), peptide arrays were employed for the analysis of peripheral blood mononuclear cells collected weekly from human and porcine subjects (n = 6) over a one month period. The data revealed strong evidence for species-specific signalling profiles. Both humans and pigs also exhibited evidence for individual-specific kinome profiles that were independent of natural changes in blood cell populations. Species-specific kinotypes could have applications in disease research by facilitating the selection of appropriate animal models or by revealing a baseline kinomic signature to which treatment-induced profiles could be compared. Similarly, individual-specific kinotypes could have implications in personalized medicine, where the identification of molecular patterns or signatures within the kinome may depend on both the levels of kinome diversity and temporal stability across individuals.
    BMC Genomics 12/2013; 14(1):854. DOI:10.1186/1471-2164-14-854 · 3.99 Impact Factor
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    • "IL-17 is critical for RA pathogenesis and can induce increased expression of TNFα [25]. To-date, a direct interaction between IL-17 and IL-32 has not been reported, although one study implied their interaction by determining that these cytokines have common signal intermediates, p300 and DAPK-1 [22]. Another study demonstrated that IL-32γ-stimulated DCs could induce a Th17 response when co-cultured with CD4+ T cells [29], implying that IL-32 affects IL-17 production independent of TNFα. "
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    ABSTRACT: Introduction Interleukin (IL)-32 and IL-17 play critical roles in pro-inflammatory responses and are highly expressed in the synovium of patients with rheumatoid arthritis (RA). We investigated the relations between these two cytokines (IL-17 and IL-32) for their ability to induce each other and to stimulate osteoclasts in RA fibroblast-like synoviocytes (FLSs) and T cells. Methods FLSs were isolated through surgical synovectomy obtained from patients with RA or osteoarthritis (OA). Real-time PCR were performed to evaluate the expression of IL-32, IL-17 and osteoclast-related genes. Immunohistochemical staining and tartrate-resistant acid phosphatase (TRAP) staining were performed to determine the distribution of inflammatory cytokines and the presence of osteoclastogenesis. Results IL-17 induced the expression of IL-32 in the FLSs from RA patients, as assessed by microarray. IL-32 production was increased by IL-17. IL-32 in the FLSs from RA patients induced the production of IL-17 in CD4+ T cells. IL-32 and IL-17 were colocalized near TRAP-positive areas in joint specimens. IL-17 and IL-32 synergistically induced the differentiation of osteoclasts, as demonstrated by the expression of osteoclast-related genes. IL-32 and IL-17 also could induce resorption by osteoclasts in a RANKL-dependent manner. Conclusions IL-17 affected the expression of IL-32 in FLSs of RA patients and IL-32 induced the production of IL-17 in CD4+ T cells. Both IL-17 and IL-32 cytokines can reciprocally influence each other's production and amplify the function of osteoclastogenesis in the in RA synovium. Separately, IL-17 and IL-32 each stimulated osteoclastogenesis without RANKL. Together, the two cytokines synergistically amplified the differentiation of osteoclasts, independent of RANKL stimulation.
    Arthritis research & therapy 11/2012; 14(6):R246. DOI:10.1186/ar4089 · 3.75 Impact Factor
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