Dataset Information

Monosodium urate crystal-induced pro-interleukin-1? production is post-transcriptionally regulated via the p38 signaling pathway in human monocytes.

ABSTRACT: IL-1? is a key mediator of sterile inflammation in response to endogenous particulates, a type of damage-associated molecular pattern (DAMPs) molecule derived from damaged cells. Despite the well-known role of sterile particulates such as monosodium urate (MSU) crystals as inflammasome inducers in monocytes/macrophages, little is known regarding how pro-IL-1? synthesis is induced under sterile inflammatory conditions. We provide evidence that MSU crystals post-transcriptionally induce the rapid production of pro-IL-1? in human primary monocytes. Metabolic labeling and pull-down assays for newly-synthesized proteins clearly showed that MSU crystals rapidly, within 30?min, induce the synthesis of pro-IL-1? as well as global proteins. Notably, MSU crystal-induced pro-IL-1? synthesis is selectively dependent on the p38 MAPK pathway, whereas global protein synthesis is mediated via the mTOR, ERK1/2, and p38 pathways. Furthermore, inhibition of Mnk1, a substrate of p38, blocked MSU crystal-induced pro-IL-1? synthesis downstream of eIF4E phosphorylation. In addition, the p38 MAPK pathway leading to phosphorylation of MK2 was also critical for stabilization of pro-IL-1? mRNA following MSU stimulation. Our findings demonstrate that post-transcriptional regulation via p38 MAPK plays a central role in the rapid synthesis of pro-IL-1? in response to MSU crystals, which is an essential step for IL-1? production in human monocytes.

SUBMITTER: Chung YH

PROVIDER: S-EPMC5046103 | biostudies-literature | 2016 Oct

REPOSITORIES: biostudies-literature

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Monosodium urate crystal-induced pro-interleukin-1β production is post-transcriptionally regulated via the p38 signaling pathway in human monocytes.

Chung Yeon-Ho YH Kim Dong-Hyun DH Lee Won-Woo WW

Scientific reports 20161003

IL-1β is a key mediator of sterile inflammation in response to endogenous particulates, a type of damage-associated molecular pattern (DAMPs) molecule derived from damaged cells. Despite the well-known role of sterile particulates such as monosodium urate (MSU) crystals as inflammasome inducers in monocytes/macrophages, little is known regarding how pro-IL-1β synthesis is induced under sterile inflammatory conditions. We provide evidence that MSU crystals post-transcriptionally induce the rapid ...[more]

PMID: 27694988

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Project description:The murine air pouch is a bursa-like space that resembles the human synovial membrane. Injection of monosodium urate (MSU) crystals into the pouch elicits an acute inflammatory response similar to human gout. We conducted the present study to identify mRNAs that were highly regulated by MSU crystals in the pouch membrane.Air pouch membranes were meticulously dissected away from the overlying skin. Gene expression differences between MSU crystal stimulated and control membranes were determined by oligonucleotide microarray analysis 9 hours after injection of MSU crystals or buffer only. Differential regulation of selected targets was validated by relative quantitative PCR in time course experiments with dissected air pouch membranes and murine peritoneal macrophages.Eleven of the 12 most highly upregulated mRNAs were related to innate immunity and inflammation. They included mRNAs encoding histidine decarboxylase (the enzyme that synthesizes histamine), IL-6, the cell surface receptors PUMA-g and TREM-1, and the polypeptides Irg1 and PROK-2. IL-6 mRNA rose 108-fold 1 hour after crystal injection, coinciding with a surge in mRNAs encoding IL-1beta, tumour necrosis factor-alpha and the immediate early transcription factor Egr-1. The other mRNAs rose up to 200-fold within the subsequent 3 to 8 hours. MSU crystals induced these mRNAs in a dose-dependent manner in cultured macrophages, with similar kinetics but lower fold changes. Among the downregulated mRNAs, quantitative PCR confirmed significant decreases in mRNAs encoding TREM-2 (an inhibitor of macrophage activation) and granzyme D (a constituent of natural killer and cytotoxic T cells) within 50 hours after crystal injection.This analysis identified several genes that were previously not implicated in MSU crystal inflammation. The marked rise of the upregulated mRNAs after the early surge in cytokine and Egr-1 mRNAs suggests that they may be part of a 'second wave' of factors that amplify or perpetuate inflammation. Transcript profiling of the isolated air pouch membrane promises to be a powerful tool for identifying genes that act at different stages of inflammation.

Project description:Background and objectiveBone erosion is common in patients with gout. The role of neutrophil-derived exosomes in gouty bone erosion remains elusive. This study aimed to investigate the functions of the neutrophil-derived exosomes in the development of bone erosion in gout.MethodsNeutrophil-derived exosomes were collected and assessed by transmission electron microscopy and nanoparticle tracking analysis. Cell counting kit-8 assay was applied to evaluate cell viability, and cell apoptosis was assessed by flow cytometry. In addition, quantitative Real-time PCR and Western blotting were used to determine the expression levels of alkaline phosphatase (ALP), osteoprotegerin (OPG), and receptor activator of nuclear factor-κB ligand (RANKL). Neutrophil-derived exosomes were tagged with PKH67. The miRNA expression profiles of exosomes and human fetal osteoblasts (hFOB) were compared using high-throughput sequencing. Functional miRNAs transfected into hFOB after co-incubation with exosomes were selected and validated by preliminary qPCR.ResultsNeutrophil-derived exosomes were stimulated by monosodium urate (MSU). The exosomes could inhibit the viability of the hFOB, and the expression levels of ALP and OPG were down-regulated, while the expression level of RANKL was up-regulated. However, there was no significant difference in the viability of osteoclasts and the expression of nuclear factor of activated T cells 1. Exosomes were observed in the cytoplasm under a confocal microscopy, confirming that exosomes could be taken up by hFOB. In total, 2590 miRNAs were found, of which 47 miRNAs were differentially expressed. Among the delivered miRNAs, miR-1246 exhibited the highest level of differential expression. The viability of hFOB was reduced by miR-1246 mimics and increased by miR-1246 inhibitors. There was no significant difference in hFOB apoptosis rate between the miR-1246 mimic and miR-1246 inhibitor group. MiR-1246 overexpression decreased the expression levels of ALP and OPG, whereas increasing the expression level of RANKL. In contrast, miR-1246 inhibitor increased the expression levels of ALP and OPG, while decreasing the expression level of RANKL. Neutrophil-derived exosomes stimulated by MSU could increase the expression of miR-1246.ConclusionNeutrophil-derived exosomes stimulated by MSU could inhibit the viability of osteoblasts.

Dataset Information

Monosodium urate crystal-induced pro-interleukin-1? production is post-transcriptionally regulated via the p38 signaling pathway in human monocytes.

Publications

Monosodium urate crystal-induced pro-interleukin-1β production is post-transcriptionally regulated via the p38 signaling pathway in human monocytes.

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