The xanthine oxidase–NFAT5 pathway regulates macrophage activation and TLR‐induced inflammatory arthritis

Abstract

NFAT5 (nuclear factor of activated T cells), a well-known osmoprotective factor, can be activated by isotonic stimuli such as Toll-like receptor (TLR) triggering. However, it is unclear how NFAT5 discriminates between isotonic and hypertonic stimuli to produce different functional and molecular outcomes. Here, we identified a novel XO-ROS-p38 MAPK-NFAT5 pathway (XO is xanthine oxidase, ROS is reactive oxygen species) that is activated in RAW 264.7 macrophages upon isotonic TLR stimulation. Unlike what is seen under hypertonic conditions, XO-derived ROS were selectively required for the TLR-induced NFAT5 activation and NFAT5 binding to the IL-6 promoter in RAW 264.7 macrophages under isotonic conditions. In mouse peritoneal macrophages and human macrophages, TLR ligation also induced NFAT5 activation, which was dependent on XO and p38 kinase. The involvement of XO in NFAT5 activation by TLR was confirmed in RAW 264.7 macrophages implanted in BALB/c mice. Moreover, allopurinol, an XO inhibitor, suppressed arthritis severity and decreased the expression of NFAT5 and IL-6 in splenic macrophages in C57BL/6 mice. Collectively, these data support a novel function of the XO-NFAT5 axis in macrophage activation and TLR-induced arthritis, and suggest that XO inhibitor(s) could serve as a therapeutic agent for chronic inflammatory arthritis.Keywords: Inflammatory arthritis r Innate immunity r NFAT5 r Reactive oxygen species r Xanthine oxidase Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionThe nuclear factor of activated T cells (NFAT) family is a group of five versatile transcription factors that are crucial to the develCorrespondence: Dr. Wan-Uk Kim e-mail: wan725@catholic.ac.kr opment and function of the immune system. Among the five members of the NFAT family (NFAT1-5), NFAT5 is unique in that it is the only protein that responds to hypertonicity [1]. NFAT5 is very similar to a Drosophila transcription factor involved * These authors contributed equally to this work. 2722 Nam-Hoon Kim et al. Eur. J. Immunol. 2014. 44: 2721-2736 in the defense against hypertonicity [2], indicating that it may represent an ancient prototype among members of the NFAT family. Indeed, the ability to guard cells against osmotic damage is commonly required in invertebrates as well as in vertebrates including humans. Accordingly, the role of NFAT5 is most pronounced in tissues constantly exposed to osmotic stress, such as kidney medulla epithelia, intestine epithelia, skin epidermis, and cornea [3]. Interestingly, NFAT5 is also activated in nonhypertonic conditions. Biologic processes that accompany NFAT5 activation include integrin-mediated carcinoma invasion [4], migration and differentiation of myoblasts [5], and T-cell proliferation [6]. Additionally, we have demonstrated for the first time that NFAT5 activity is increased by tumor necrosis factor alpha (TNF-α) and interleukin (IL) 1β stimulation under isotonic conditions [7]. NFAT5-deficient mice show a marked r...

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“…Upon activation, macrophages can produce high quantities of ROS via XOD, thereby causing local endothelial dysfunction, further impairment of endoneurial blood flow, and myelin degeneration. Overexpression of XOD is actually necessary step for macrophages activation and regulation of proinflammatory mediators and chemokines secretion [31, 32]. Furthermore, ROS generated by local macrophages via XOD may induce nerve axonal loss and demyelination, as demonstrated in a murine model of neuroinflammation [33].…”

Elementary immunology: Na+ as a regulator of immunity

“…Second, increases of 40 mM in effective osmolytes to a final osmolality of 380 mOsm/kg are unlikely to induce a direct effect on pathogen survival given that Escherichia coli can very well tolerate osmolalities of 400 mOsm/ kg [64]. Third, p38/mitogen-activated protein kinase (MAPK) and TonEBP/NFAT5 are both induced by NaCl-mediated osmotic stress and by stimulation with the pro-inflammatory bacterial cell-wall component LPS [65–67]. These observations already imply that osmoprotective and inflammatory responses might be intertwined.…”

Suppression of NFAT5-mediated Inflammation and Chronic Arthritis by Novel κB-binding Inhibitors

“…For example, NFAT5 induces the expression of toll-like receptor (TLR)-mediated inflammatory genes in macrophages in a tonicity-independent manner (Buxadé et al, 2012, Kim et al, 2013). High salt and TLR ligation activate distinct sets of downstream target genes in a NFAT5-dependent manner (Kim et al, 2014). While ROS are essential for this, their source differs depending on the context: mitochondria for high salt and xanthine oxidase for TLR (Kim et al, 2014).…”