Abstract
The aim of the present study was to establish the role of platelet‑activating factor acetyl hydrolase (PAF‑AH) in the resolution phase of gout using an established invitro mononuclear cell model. The effects of signalling pathway inhibitors on PAF‑AH secretion, as well as the effects of the common treatments hydrocortisone and colchicine, an antibody against the anti‑inflammatory cytokine transforming growth factorβ1 (TGFβ1), and the transcriptional inhibitor actinomycinD, were also investigated. The effect of recombinant PAF‑AH on cytokine secretion by these cells was also determined. Human peripheral blood‑derived monocytes were isolated and differentiated into macrophages. Monocytes and macrophages were stimulated with monosodium monohydrate urate (MSU) crystals or lipopolysaccharide in the presence or absence of AEG3482 [a c‑Jun N‑terminal kinase (JNK) inhibitor], MG132 (a proteasome inhibitor), hydrocortisone or colchicine. Cultures were then analysed for PAF‑AH secretion using ELISA. A 6‑fold upregulation of PAF‑AH secretion was observed following macrophage exposure to MSU crystals for 24h (29.3±6 vs. 5.4±0.3ng/ml unstimulated; P<0.05). Following 72h, PAF‑AH levels decreased significantly (11.1±1.8; P<0.01). Secretion was further enhanced following pre‑treatment with the JNK protein kinase inhibitor AEG3482 prior to MSU crystal stimulation (P<0.05) and was abrogated when cells were preincubated with actinomycin D or the proteasome inhibitor MG132 (50, 100 and 200µM). The addition of recombinant PAF‑AH (2.5‑10ng/ml) to MSU crystal‑stimulated immature monocyte cultures significantly decreased pro‑inflammatory interleukin (IL)‑1β (unstimulated 687±124vs.stimulated 113±30pg/ml) and IL‑6 secretion (unstimulated 590±50vs.stimulated 182±19pg/ml). Treatment of MSU crystal‑stimulated macrophages with hydrocortisone (2µM) also significantly decreased PAF‑AH release (P<0.05). Neutralising anti‑TGFβ1 addition decreased PAF‑AH dose‑dependently with the highest inhibition observed at 1µg/ml (P<0.05). The results implicated that PAF‑AH may have an anti‑inflammatory role in the resolution phase of gout.
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