Monocyte-derived and M1 macrophages from ankylosing spondylitis patients released higher TNF-\u03b1 and expressed more IL1B in response to BzATP than macrophages from healthy subjects

Maryam Akhtari,Mahdi Mahmoudi,Seyed Jalal Zargar,Mahdi Vojdanian,Ahmadreza Jamshidi

doi:10.1038/s41598-021-96262-2

Maryam Akhtari, Mahdi Mahmoudi + Show 3 more

Open Access

https://doi.org/10.1038/s41598-021-96262-2

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Abstract

Macrophages participate in the pathogenesis of ankylosing spondylitis (AS) by producing inflammatory cytokines. Extracellular adenosine triphosphate (eATP), released during cell stress, acts through purinergic receptors (P2XR and P2YR) and induces inflammatory responses. We investigated the effect of 2ʹ(3ʹ)-O-(4-benzoyl benzoyl) ATP (BzATP) (a prototypic agonist of P2X7R) on the production of inflammatory cytokines in both monocyte-generated (M2-like) and M1 macrophages from patients and controls. Macrophages were differentiated from isolated periphery-monocytes (n = 14 in each group) by macrophage colony-stimulating factor (M-CSF). Using LPS and IFN-γ, macrophages were skewed toward M1 type and were treated with BzATP. Gene expression and protein release of IL-1β, IL-23, and TNF-α were evaluated by real-time PCR and ELISA methods respectively before and after treatment. BzATP significantly increased the protein release of TNF-α and the expression of TNFA and IL1B in monocyte-generated macrophages. Besides, BzATP treatment significantly upregulated IL1B expression, reduced TNFA and IL23A expression, and TNF-α release in M1 macrophages from both groups. Monocyte-generated and M1 macrophages from AS patients released higher TNF-α and expressed more IL1B in response to the same concentration of BzATP treatment respectively. Based on our results, AS macrophages were more sensitive to BzATP treatment and responded more intensively. Besides, the diverse effects of BzATP on monocyte-derived and M1 macrophages in our study may represent the differed inflammatory properties of these two groups of macrophages in response to eATP in the body.

Highlights

Under the normal physiological state, adenosine triphosphate (ATP) exists at a low level in the extracellular space, under cellular stress or damage, it is released in large amounts to the outside m ilieu[1,2]
We measured the level of mRNA expression and protein secretion of IL-23, IL-1β, and tumor necrosis factor (TNF)-α pro-inflammatory factors in monocyte-generated macrophages and M1 macrophages from Ankylosing spondylitis (AS) and healthy individuals
Diverse observed effects of BzATP on monocyte-derived and M1 classically activated macrophages in our study, especially regarding TNF-α and IL-23, may represent the differed inflammatory properties of these two groups of macrophages in response to environmental ATP in the body. It seems that the cytotoxicity activity of BzATP on classically activated macrophages may affect their production of inflammatory cytokines

Summary

Introduction

Under the normal physiological state, adenosine triphosphate (ATP) exists at a low level in the extracellular space, under cellular stress or damage, it is released in large amounts to the outside m ilieu[1,2]. Extracellular nucleotides and their derived nucleosides (such as adenosine) participate in purinergic signaling which is an evolutionarily conserved signaling pathway and is involved in various pathophysiological events including neuromodulation, cell differentiation, migration, immune responses, and inflammation[3]. The level of extracellular ATP is increased and ATP-dependent P2X7R activation induces the inflammatory responses in macrophages. Concerning the importance of macrophages in the inflammatory responses of AS disease, here we investigated the effect of a prototypic agonist of P2X7R (BzATP) on the production of inflammatory cytokine in both monocyte-generated (M2-like) and M1 macrophages from AS patients in contrast to macrophages from healthy subjects

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