Abstract
Monocytes play a central role in regulating inflammation in response to infection or injury, and during auto-inflammatory diseases. Human blood contains classical, intermediate and non-classical monocyte subsets that each express characteristic patterns of cell surface CD16 and CD14; each subset also has specific functional properties, but the mechanisms underlying many of their distinctive features are undefined. Of particular interest is how monocyte subsets regulate secretion of the apical pro-inflammatory cytokine IL-1β, which is central to the initiation of immune responses but is also implicated in the pathology of various auto-immune/auto-inflammatory conditions. Here we show that primary human non-classical monocytes, exposed to LPS or LPS + BzATP (3’-O-(4-benzoyl)benzyl-ATP, a P2X7R agonist), produce approx. 80% less IL-1β than intermediate or classical monocytes. Despite their low CD14 expression, LPS-sensing, caspase-1 activation and P2X7R activity were comparable in non-classical monocytes to other subsets: their diminished ability to produce IL-1β instead arose from 50% increased IL-1β mRNA decay rates, mediated by Hsp27. These findings identify the Hsp27 pathway as a novel therapeutic target for the management of conditions featuring dysregulated IL-1β production, and represent an advancement in understanding of both physiological inflammatory responses and the pathogenesis of inflammatory diseases involving monocyte-derived IL-1β.
Highlights
Monocytes are a heterogeneous population of immune cells accounting for approximately 10% of the blood mononuclear cell population
We first isolated primary human monocytes from the peripheral blood of healthy donors by fluorescence-activated cell sorting (FACS), and separated them into classical, intermediate, and non-classical subsets based on their relative expression of CD14 and CD16 (Supplementary Fig. 1)
We measured the amounts of IL-1βproduced following a two-step stimulation protocol: initial priming by exposure to the bacterial cell wall component lipopolysaccharide (LPS) to induce the transcription and production of pro-IL-1βs; followed by treatment with BzATP, a synthetic substitute for extracellular ATP that acts via the P2X7 receptor (P2X7R) to stimulate the inflammasome complex/caspase-1 and alter membrane dynamics to promote the maturation and release of IL-1β25–28
Summary
Monocytes are a heterogeneous population of immune cells accounting for approximately 10% of the blood mononuclear cell population. A key role of monocytes is to detect early signs of tissue damage or infection and respond by secreting pro-inflammatory cytokines to alert the immune system of the threat. A study done in human monocytic cell line THP-1 demonstrated the involvement of a complex formed by AUF1 with heat shock protein 27 (Hsp27) in the regulation of IL-1β17 though whether this pathway is important during IL-1βproduction by monocytes is currently unknown. Assuming that the IL-1βmRNA evades degradation, a second level of regulation of IL-1βproduction from immune cells exists during the conversion of immature pro-IL-1βinto mature IL-1β. The studies by Piccini et al.[24] and Ward et al.[20], for example, used monocyte isolation methods leading to enrichment of specific subsets, and drew disparate conclusions on the involvement of endogenous ATP during IL-1βsecretion: we hypothesised that monocyte subsets might produce and release IL-1βvia specialised, distinct, and largely-undefined mechanisms
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