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Potent synergistic effect of IL-3 and TNF on matrix metalloproteinase 9 generation by human eosinophils.

ABSTRACT: TNF (designated as TNF-? under previous nomenclature) is the preeminent activator of MMP-9 generation from a variety of cells including eosinophils. We have previously established that TNF strongly synergizes with IFN-? and IL-4 for eosinophil synthesis of Th1- and Th2-type chemokines respectively. Thus, we sought to determine if TNF-induced synthesis of MMP-9 would be enhanced by the presence of Th1, Th2, or the eosinophil-associated common beta chain (?c) cytokines. Human blood eosinophils were cultured with TNF alone or in combination with either IFN-?, IL-4, IL-3, IL-5, or GM-CSF. Concentrations and activities of MMP-9 in eosinophil culture supernates were measured by ELISA and gelatin zymography, mRNA transcription and stabilization by quantitative real-time PCR, and signaling events by immunoblotting and intracellular flow cytometric analysis. Individually, TNF, GM-CSF, or IL-3, but not IL-4 or IFN-?, induced relatively small (<0.2 ng/ml) but statistically significant quantities of MMP-9. Remarkable synergistic synthesis of MMP-9 (ng/ml levels) occurred in response to TNF plus IL-3, GM-CSF or IL-5, in the order of IL-3>GM-CSF>IL-5. Zymography revealed that eosinophils release MMP-9 in its pro-form. Eosinophil stimulation with the combination of IL-3 plus TNF led to increased steady-state levels of MMP-9 mRNA, prolonged mRNA stabilization, and enhanced activation of ERK1/2 phosphorylation. Inhibition of NF-?B, MEK kinase, or p38 MAP kinase, but not JNK signaling pathways, diminished IL-3/TNF-induced MMP-9 mRNA and protein production. Thus, the synergistic regulation of eosinophil MMP-9 by IL-3 plus TNF likely involves cooperative interaction of multiple transcription factors downstream from ERK, p38, and NF-?B activation as well as post-transcriptional regulation of MMP-9 mRNA stabilization. Our data indicate that within microenvironments rich in ?c-family cytokines and TNF, eosinophils are an important source of proMMP-9 and highlight a previously unrecognized role for synergistic interaction between TNF and ?c-family cytokines, particularly IL-3, for proMMP-9 synthesis.

SUBMITTER: Kelly EA

PROVIDER: S-EPMC3319644 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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Potent synergistic effect of IL-3 and TNF on matrix metalloproteinase 9 generation by human eosinophils.

Kelly Elizabeth A B EA Liu Lin Ying LY Esnault Stephane S Quinchia Johnson Beatriz Helena BH Jarjour Nizar N NN

Cytokine 20120209 2

TNF (designated as TNF-α under previous nomenclature) is the preeminent activator of MMP-9 generation from a variety of cells including eosinophils. We have previously established that TNF strongly synergizes with IFN-γ and IL-4 for eosinophil synthesis of Th1- and Th2-type chemokines respectively. Thus, we sought to determine if TNF-induced synthesis of MMP-9 would be enhanced by the presence of Th1, Th2, or the eosinophil-associated common beta chain (βc) cytokines. Human blood eosinophils wer ...[more]

PMID: 22321809

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Project description:Asthma is often associated with airway eosinophilia, and therapies targeting eosinophils are now available to treat severe eosinophilic asthma. Eosinophilic asthma is often due to a type-2 immune response and production of IL-5, which leads to eosinophilopiesis and recruitment of mature eosinophils in the airways. A concomitant type-2 and type-17 response has been reported in some individuals. IL-17 may be enhanced by IL-1β production and can lead to neutrophilic inflammation. In fact, both eosinophilic and neutrophilic (mixed granulocytic) inflammation are simultaneously present in a large population of patients with asthma. In monocyte/macrophage cell populations, release of mature IL-1β occurs via toll-like receptor ligand-induced activation of the inflammasome. Within the inflammasome, a cascade of events leads to the activation of caspase-1, which cleaves pro-IL-1β protein into a mature, releasable, and active form. We have demonstrated that eosinophils can release IL-1β in a Toll-like receptor ligand-independent fashion. The objective of this study was to determine the mechanisms underlying the production and maturation of IL-1β in cytokine-activated eosinophils. Using eosinophils from circulating blood and from bronchoalveolar lavage fluid after an airway allergen challenge, the present study demonstrates that cytokine-activated eosinophils express and release a bioactive form of IL-1β with an apparent size less than the typical 17 kDa mature form produced by macrophages. Using a zymography approach and pharmacological inhibitors, we identified matrix metalloproteinase-9 (MMP-9) as a protease that cleaves pro-IL-1β into a ~15 kDa form and allows the release of IL-1β from cytokine-activated eosinophils. Therefore, we conclude that activated eosinophils produce MMP-9, which causes the release of IL-1β in an inflammasome/caspase-1-independent manner. The production of IL-1β by eosinophils may be a link between the eosinophilic/type-2 immune response and the neutrophilic/type-17 immune response that is often associated with a more severe and treatment-refractory type of asthma.

Project description:We recently developed novel matrix metalloproteinase-2 (MMP-2) inhibitor small molecules for cardioprotection against ischemia/reperfusion injury and validated their efficacy in ischemia/reperfusion injury in cardiac myocytes. The aim of the present study was to test our lead compounds for cardioprotection in vivo in a rat model of acute myocardial infarction (AMI) in the presence or absence of hypercholesterolemia, one of the major comorbidities affecting cardioprotection. Normocholesterolemic adult male Wistar rats were subjected to 30 min of coronary occlusion followed by 120 min of reperfusion to induce AMI. MMP inhibitors (MMPI)-1154 and -1260 at 0.3, 1, and 3 µmol/kg, MMPI-1248 at 1, 3, and 10 µmol/kg were administered at the 25th min of ischemia intravenously. In separate groups, hypercholesterolemia was induced by a 12-week diet (2% cholesterol, 0.25% cholic acid), then the rats were subjected to the same AMI protocol and single doses of the MMPIs that showed the most efficacy in normocholesterolemic animals were tested in the hypercholesterolemic animals. Infarct size/area at risk was assessed at the end of reperfusion in all groups by standard Evans blue and 2,3,5-triphenyltetrazolium chloride (TTC) staining, and myocardial microvascular obstruction (MVO) was determined by thioflavine-S staining. MMPI-1154 at 1 µmol/kg, MMPI-1260 at 3 µmol/kg and ischemic preconditioning (IPC) as the positive control reduced infarct size significantly; however, this effect was not seen in hypercholesterolemic animals. MVO in hypercholesterolemic animals decreased by IPC only. This is the first demonstration that MMPI-1154 and MMPI-1260 showed a dose-dependent infarct size reduction in an in vivo rat AMI model; however, single doses that showed the most efficacy in normocholesterolemic animals were abolished by hypercholesterolemia. The further development of these promising cardioprotective MMPIs should be continued with different dose ranges in the study of hypercholesterolemia and other comorbidities.

Dataset Information

Potent synergistic effect of IL-3 and TNF on matrix metalloproteinase 9 generation by human eosinophils.

Publications

Potent synergistic effect of IL-3 and TNF on matrix metalloproteinase 9 generation by human eosinophils.

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