Suppression of inflammatory and immune responses by the A_2A adenosine receptor: an introduction

Abstract

The purine nucleoside adenosine has been described as a ‘retaliatory metabolite' by virtue of its ability to function in an autocrine manner to modify the activity of a range of cell types following its extracellular accumulation during cell stress or injury. These effects are largely protective and are triggered by the binding of adenosine to any of four G-protein-coupled adenosine receptors. Most of the anti-inflammatory effects of adenosine have been assigned to the adenosine A_2A receptor subtype, which is expressed in many immune and inflammatory cells. In this brief article, we will outline the growing evidence to support the hypothesis that the development of agonists selective for the A_2A receptor is an effective strategy for suppressing the exaggerated inflammatory responses associated with many diseases by virtue of the receptor's ability to inhibit multiple pro-inflammatory signalling cascades.

Introduction

Adenosine is an endogenous purine nucleoside that mediates a wide variety of physiological functions by interaction with four cell-surface receptors (A₁, A_2A, A_2B and A₃; Fredholm et al., 2000, 2001). Numerous studies have highlighted the anti-inflammatory role of the A_2A receptor from studies of receptor distribution on inflammatory cells (Gessi et al., 2000) as well as from observations of inhibitory effects of A_2A receptor-selective agonists in vitro and in vivo and enhanced inflammatory responses in vivo in A_2A receptor-deficient mice (Sitkovsky, 2003; Hasko and Cronstein, 2004).

This article is written as an introduction to the symposium on New Insights into the Anti-inflammatory Effects of A_2AAR Agonists presented at the Life Sciences Meeting in Glasgow on July 11, 2007. Its purpose is to introduce the reader to some key concepts and serve as a basis for other chapters in this journal from the symposium discussing the following: (1) A_2A adenosine receptors in tissue protection from reperfusion injury, by Professor Joel Linden (University of Virginia, USA); (2) Adenosine receptors and asthma, by Professor Clive Page (King's College, London); (3) Moving out and turning tail: restricted collision of the A_2A-adenosine receptor revisited, by Professor Michael Freissmuth (University of Vienna, Austria); and (4) Hypoxia-adenosinergic regulation of immune response and tissue damage by Professor Misha Sitkovsky (Northeastern University, Massachusetts, USA).

Generation of extracellular adenosine

Before considering what is currently known about A_2A receptor activation and its intracellular effects on inflammatory and immune signalling, it is important to consider how the endogenous agonist is generated in vivo. Adenosine is released from all cells upon the degradation of ATP. Consequently, conditions of stress, hypoxia and increased tissue energy expenditure can lead to a rapid >200-fold elevation in tissue adenosine over basal levels (typically <50 nM) (Rivkees et al., 2001). Specifically, ATP is converted to adenosine at sites of inflammation and injury by the action of CD39, an ecto-apyrase, and the subsequent conversion of AMP to adenosine by the ecto-5′-nucleotidase CD73. These enzymes are abundantly expressed on endothelial cells as well as on leucocytes and regulatory T cells (Lennon et al., 1998; Eltzschig et al., 2004; Deaglio et al., 2007). The protective role of this pathway has been elegantly demonstrated by gene-targeting studies in mice, which have shown that the presence of functional CD39 and CD73 is necessary to maintain endothelial barrier function and thus prevent excessive vascular leakage following hypoxia (Eltzschig et al., 2003; Thompson et al., 2004). However, the accumulation of extracellular adenosine is transient due to its conversion to inosine by ecto-adenosine deaminase (ADA) or uptake into endothelial cells via specific transporters and conversion either to inosine by intracellular ADA or to AMP by adenosine kinase. Because adenosine exerts profound auto-/paracrine effects on critical aspects of the immune and inflammatory responses, it is perhaps not surprising that the expression of enzymes involved in its accumulation is subject to tight regulation. For example, both CD39 and CD73 are each strongly induced in vascular endothelial cells upon the onset of hypoxia, and CD73 expression can be potentiated also by interferon (IFN)-α and adenosine itself via receptor-mediated elevation of cyclic AMP (cAMP) (Narravula et al., 2000; Niemela et al., 2004). Thus, control of adenosine accumulation and initiation of protective signalling clearly represents an important adaptive mechanism by which hypoxia-mediated damage to the endothelium can be minimized.

Activation of signalling pathways by the A_2A receptor: in vitro studies

The A_2A receptor was originally identified by virtue of its ability to elevate intracellular levels of cAMP via receptor interaction with the heterotrimeric G-protein G_s and subsequent activation of adenylyl cyclase (Linden, 2001). Changes in cAMP levels are translated into pleiotropic intracellular effects by a panel of cAMP-binding effector proteins, which include cyclic nucleotide-gated ion channels, cAMP-dependent protein kinase (PKA) and exchange proteins directly activated by cAMP (Epacs) (Beavo and Brunton, 2002). Signal termination is achieved by hydrolysis of cAMP to 5′-AMP catalysed by the large superfamily of cyclic nucleotide phosphodiesterases (PDEs). A key aspect of cAMP's effects is the generation of intracellular cAMP gradients arising from the opposing effects of adenylyl cyclases and PDEs (Lynch et al., 2006). The ability of distinct regions within the cell to sample these gradients is dictated in part by specific A-kinase anchoring protein scaffolds that localize PDEs, RI and RII regulatory cAMP-binding subunits of PKA and Epacs to defined intracellular compartments. In the case of PKA, binding of cAMP to R subunits releases catalytic C subunits from the PKA holoenzyme and allows phosphorylation of nearby substrates (Baillie et al., 2005).

However, similar to the other three receptors, the A_2A receptor can also activate the extracellular signal-regulated kinase (ERK) pathway; this has been demonstrated in Chinese hamster ovary cells as well as in human embryonic kidney (HEK)-293, PC12 and vascular endothelial cells (reviewed by Schulte and Fredholm, 2003). Intriguingly, multiple mechanisms are responsible that appear to differ between cell types. For example, whereas A_2A receptor activation of ERK in CHO cells occurs via an Src-mediated process that can be blocked by H-89 and mimicked by treatment with the cell-permeable cAMP analogue 8-bromo-cAMP, stimulation of ERK by either endogenous A_2A receptors in human umbilical vein endothelial cells or recombinant A_2A receptors expressed in HEK293 cells appears to occur via a Ras- and Src kinase-dependent mechanism independent of cAMP elevation (Seidel et al., 1999). Identification of the Arf6 guanine nucleotide exchange factor as an A_2A receptor-binding protein revealed that its interaction with the receptor's unique cytoplasmic C-terminal domain was critical for the sustained cAMP-independent ERK activation observed upon receptor expression in HEK293 cells, although not affecting the initial spike of ERK phosphorylation detectable at 5 min (Gsandtner et al., 2005).

Suppression of immune and inflammatory events by the A_2A receptor: in vitro studies

Functional and immunological approaches have shown that A_2A receptors are expressed in specific haematopoietic cell populations. These include CD4⁺ and CD8⁺ T cells and natural killer cells as well as monocytes, macrophages and neutrophils. In contrast, the receptor is largely absent from B cells (Koshiba et al., 1999; Sullivan et al., 2001; Pinhal-Enfield et al., 2003; Zhang et al., 2005; Raskovalova et al., 2006). The majority of the A_2A receptor's inhibitory effects on immune and inflammatory processes in these cell types have been proposed to occur via cAMP-/PKA-dependent pathways, which are known to have wide-ranging effects on immune cell function (Tasken and Stokka, 2006). For example, selective activation of A_2A receptors expressed on lymphokine-activated killer cells using CGS21680 has been shown to suppress perforin- and FasL-mediated cytotoxicity via an RI PKA-dependent pathway (Raskovalova et al., 2006). Also, inhibition of both the oxidative burst response (Sullivan et al., 2001) and VLA-4 induction (Sullivan et al., 2004) in human neutrophils by A_2A receptor-selective agonists can be reversed by the PKA-selective inhibitor H-89 and potentiated by inclusion of rolipram, an inhibitor of PDE-4. Similarly, the inhibitory effect of A_2A receptor agonists on anti-CD3ɛ-mediated tyrosine phosphorylation of ZAP70, a critical step in T-cell activation, could also be reversed by H-89, indicative of a cAMP-/PKA-dependent mechanism (Sevigny et al., 2007). However, an alternative cAMP-activated pathway has recently emerged by which the A_2A receptor can suppress JAK (Janus kinase)–STAT (signal transducer and activator of transcription) pathway activation by cytokines that utilize the transmembrane protein gp130 to trigger downstream signalling. In vascular endothelial cells and embryonic fibroblasts, cAMP-mediated activation of Epac1 results in the induction of the gene encoding ‘suppressor of cytokine signalling-3′ (SOCS-3) (Sands et al., 2006). SOCS-3 is one of the eight SOCS family members (cytokine-inducible SH2 domain protein, SOCS-1 to -7) defined by the presence of a distinct N-terminal region linked to a central SH2 and C-terminal SOCS box domains (Yoshimura et al., 2007). Upon induction, the SH2 domain binds to specific phosphor-Tyr residues present on target cytokine receptors, such as gp130 and the leptin receptor Ob-R (Heinrich et al., 2003) (Figure 1). Once bound, a so-called ‘kinase inhibitory region' within the N-terminal domain of SOCS-3 binds and inhibits the kinase activity of receptor-associated JAKs, thereby inhibiting STAT phosphorylation and activation (Yoshimura et al., 2007) (Figure 1). The exact mechanism by which Epac1 activation triggers SOCS-3 gene transcription remains to be elucidated, but it may provide a common mechanism by which Epac-mediated, PKA-independent effects of cAMP on the induction of genes such as those for AQP-2 and pro-glucagon can be rationalized (Lotfi et al., 2006; Umenishi et al., 2006).

Figure 1.

A_2A receptor-mediated inhibition of cytokine receptor activation of the JAK-STAT pathway. Binding of adenosine or synthetic agonists to the A_2A receptor triggers the activation of the stimulatory heterotrimeric G-protein G_s, which in turn activates adenylyl cyclase. The resulting elevation of cAMP levels is sensed by multiple intracellular cAMP-binding effectors, including Epac1. Epac1 functions as a cAMP-activated guanine nucleotide exchange factor for the Rap family of Ras-related small G-proteins, promoting the formation of GTP-bound Rap1a. This initiates transcription of the SOCS-3 gene via a transcription factor (TF) that has yet to be identified. In vascular endothelial cells, SOCS-3 inhibits JAK-mediated tyrosine phosphorylation of STATs after binding to tyrosine-phosphorylated Ob-Rb and gp130 following their activation by leptin and soluble IL-6 receptor α/IL-6 (sIL-6Rα/IL-6) trans-signalling complexes, respectively.

A key molecular mechanism that has emerged as being critical for the inhibitory effects of the A_2A receptor on inflammatory and immune responses is suppression of the nuclear factor-κB (NF-κB) pathway activated by cytokines such as tumour necrosis factor (TNF)-α, IL-1β as well as pathogen-derived Toll-like receptor (TLR) agonists such as lipopolysaccharride (LPS). Interestingly, the molecular basis for this inhibition appears to be cell type-specific. Thus, A_2A receptor expression in rat glioma cells prevents the accumulation of transcriptionally active NF-κB dimers in the nucleus in response to either LPS or TNFα by specifically blocking the phosphorylation and degradation of inhibitor of NF-κB (IκB) proteins, resulting in an abolition of inducible nitric oxide synthase (iNOS) induction in response to LPS or TNFα in combination with IFNγ (Sands et al., 2004). This would suggest that the receptor inhibits a common step in TNFα receptor/TLR signalling at or above the level of IκB kinases (IKKs). Similarly, deletion of the A_2A receptor in macrophages increases the rate and extent to which TNFα triggers the degradation of IκBα, resulting in an enhanced accumulation of multiple κB-regulated transcripts (Lukashev et al., 2004). In contrast, overexpression of the A_2A receptor in vascular endothelial cells blocks the TNFα-stimulated nuclear accumulation of p50/RelA heterodimers, thereby reducing κB-regulated adhesion molecule expression, without altering degradation of IκB proteins (Sands et al., 2004). Thus, presumably A_2A receptor expression is either blocking the nuclear import of NF-κB dimers or accelerating chromosomal region maintenance 1 (CRM1)-dependent export back to the cytoplasm, although experiments utilizing CRM1 inhibitor leptomycin B suggest the former mechanism (EW Strong and TM Palmer, unpublished observations). Interestingly, recent work describing how the adenosine generated during hypoxic preconditioning suppresses cytokine activation of NF-κB has suggested that activation of adenosine receptors inactivates the Skp-cullin-F box-1 E3 ligase complex required for poly-ubiquitylation and degradation of IκBα by promoting the removal of the ubiquitin-like protein Nedd8 from the Cul1 component of the complex, although the identity of the adenosine receptor subtype involved and the relative contributions of cAMP or other signalling pathways have not been determined (Khoury et al., 2007). Nevertheless, such a mechanism could potentially account for the enhanced IκBα degradation observed in macrophages from A_2A receptor-deficient mice, as one would predict that the absence of the A_2A receptor should increase the proportion of active Cul1, thereby potentiating IκBα degradation (Lukashev et al., 2004). Finally, it is important to note that the expression of the A_2A receptor is positively regulated by the pro-inflammatory stimuli whose responses it inhibits. For example, IL-1β, LPS and TNFα each increase A_2A receptor mRNA and protein levels in microvascular endothelial cells, macrophages and THP-1 monocytic cells (Nguyen et al., 2003; Khoa et al., 2004; Murphree et al., 2005). The effect is inhibited, although not abolished, by pre-incubation with an IKK inhibitor (Murphree et al., 2005), indicative of a role for NF-κB in up-regulating A_2A receptor gene transcription. This potentiates the ability of receptor activation to inhibit LPS-stimulated TNFα production in macrophages and also reduces TNFα-mediated up-regulation of vascular endothelial growth factor mRNA in endothelial cells (Nguyen et al., 2003; Murphree et al., 2005). Potentiation of A_2A receptor expression therefore constitutes an additional negative feedback mechanism by which inflammatory responses may be limited in vivo.

Anti-inflammatory potential of the A_2A receptor: in vivo evidence

The evidence that activation of the A_2A receptor in vitro leads to suppression of inflammatory responses is overwhelming as reviewed above. Three lines of experimental evidence targeting the A_2A receptor directly in animal models of inflammation–tissue damage should support the in vitro evidence if: (1) administration of an A_2A receptor-selective antagonist enhances inflammation following induction of an inflammatory response, (2) inflammation is exaggerated in animals with targeted deletion of the A_2A receptor and (3) application of selective agonists reduces inflammation and tissue damage.

Exacerbation of tissue damage and inflammation with A_2A receptor antagonists has been shown in a variety of inflammatory models. For instance, treatment of mice with the selective A_2A receptor antagonist ZM241385 enhanced liver injury and inflammation in response to concanavalin A, Pseudomonas aeruginosa and carbon tetrachloride (Ohta and Sitkovsky, 2001; Chan et al., 2006). Similarly, ZM241385 prevented both the anti-inflammatory effects and the increased survival rates induced by low-dose ketamine administration, which promotes adenosine accumulation, in mice where sepsis was induced by LPS or Escherichia coli (Mazar et al., 2005). In the mouse lung, treatment with ZM241385 enhanced lung neutrophilia in response to intratracheal administration of LPS (Thiel et al., 2005). However, in other models, ZM241385 did not enhance inflammation/tissue damage in response to an inflammatory stimulus (see, for example, Peirce et al., 2001; Fozard et al., 2002). This may reflect either varying amounts of endogenous adenosine in the different models or, alternatively, administration of a near-maximal dose of inflammatory stimulus, which would make it difficult to observe any further enhancement with an A_2A receptor-selective antagonist.

Studies in animals with targeted deletion of the A_2A receptor largely support the conclusions from experiments using ZM241385. For instance, mice deficient in the A_2A receptor displayed an exaggerated inflammatory response and cytokine release in response to a wide variety of inflammatory insults, such as concanavalin A- and carbon tetrachloride-induced liver damage, and LPS-induced inflammation in an air pouch model (Ohta and Sitkovsky, 2001; Chan et al., 2006). Mice lacking the A_2A receptor were recently reported to have exaggerated lung inflammation in response to sensitization and inhalation with ragweed antigen (Nadeem et al., 2007). In addition, adoptive transfer studies employing inflammatory cells from mice lacking the A_2A receptor have also supported the concept that this receptor functions as a physiological anti-inflammatory mechanism (Yang et al., 2006).

The above-mentioned two groups of studies, in which signalling via the A_2A receptor has been inhibited, support the concept that in an inflammatory environment the A_2A receptor functions as a ‘physiological brake' on inflammatory processes. A number of reviews have been written, highlighting the concept that the A_2A receptor is involved in inflammation (e.g., Sitkovsky, 2003; Sitkovsky et al., 2004; Hasko and Cronstein, 2004). From a therapeutic perspective, these data also suggest that administration of selective A_2A receptor agonists should inhibit the excessive inflammation and tissue damage associated with disease. There are many publications supporting the view that agonists of the A_2A receptor can inhibit inflammation in a wide variety of in vivo models. In addition, CGS21680, the first reported selective agonist at the A_2A receptor (Hutchinson et al., 1989), and the more recently developed ATL-146e (Sullivan et al., 2001) have also shown a broad spectrum of anti-inflammatory and tissue-protective effects in many different animal models of inflammation (Akkari et al., 2006; and see below).

Investigation of the anti-inflammatory mechanisms in vivo have supported many of the observations made in vitro and have provided evidence for the anti-inflammatory effects of A_2A receptor agonists in a variety of target tissues, as discussed below.

Liver

Concanavalin A induces polyclonal T-cell activation and subsequently liver cell death (Tiegs et al., 1992). The suggested mechanism of injury involves CD4⁺ T cells with secretion of cytokines such as TNFα, IFNγ and IL-6 (Gantner et al., 1995; Ohta and Sitkovsky, 2001). TNFα and IL-6 are both known to be important for induction of liver cell damage, apoptosis and necrosis. CGS21680 and ATL-146e inhibit both liver damage and the associated elevation in serum cytokine levels induced by concanavalin A administration, suggesting that inhibition of cytokine production by T cells contributed to the protective effect observed (Odashima et al., 2006). This conclusion was supported by the observations of Ohta and Sitkovsky (2001), who demonstrated that concanavalin A-induced liver damage was enhanced in mice lacking the A_2A receptor.

Kidney

A tissue protection mechanism via an inhibitory action on CD4⁺ T cells has also been supported by in vivo studies examining the protective effects of ATL-146e on ischaemia–reperfusion (IR) damage in the mouse kidney (Day et al., 2003, 2006). In these studies, the protective effect of ATL-146e was inhibited in mice whose bone marrow had been ablated and reconstituted with bone marrow from A_2A receptor-deficient animals. Using adoptive transfer techniques, the mechanism of protection of ATL-146e was shown to be due to an action of A_2A receptor agonists on CD4⁺ T cells and involved an inhibition of IFNγ release.

Lungs

In rats and mice, intratracheal or intranasal administration of CGS21680 has been shown to suppress lung inflammation in response to pro-inflammatory stimuli such as antigen (ovalbumin) and involved inhibition of neutrophil, eosinophil, macrophage and lymphocyte infiltration (Fozard et al., 2002; Bonneau et al., 2006). In the study by Fozard et al. (2002), the effects of CGS21680 were inhibited by the selective A_2A receptor antagonist ZM241385, although this agent did not appear to exaggerate inflammation induced by antigen itself. In contrast, CGS21680 was much less effective in inhibiting lung inflammation in response to LPS–N-formyl-methionine-leucine-phenylalanine administration in mice, but it did appear to inhibit neutrophil activation, as assessed by elastase release (Bonneau et al., 2006). Thiel et al. (2005) recently reported that the deleterious effect of oxygen therapy on lung inflammation and survival in mice was prevented by treatment with CGS21680, and several publications have also documented the inhibitory effects of A_2A receptor agonists on lung injury and inflammation in response to IR (Hasko et al., 2006).

Heart

Agonists at the A_2A receptor reduce infarct size and inflammation in studies of IR in canine and mouse models of myocardial infarction. Initial studies in canine models demonstrated that inhibition of P-selectin induction, neutrophil infiltration and other indices of inflammation accompanied the reduction in infarct size (Glover et al., 2005, 2007). Subsequent studies in mouse models of cardiac IR have highlighted the potential central role of CD4⁺ T cells in orchestrating several aspects of the inflammatory process, such as neutrophil infiltration, in response to IR. For instance, infarct size in response to IR is smaller in Rag1 knockout mice, which lack mature lymphocytes, and is increased by adoptive transfer of CD4⁺ T cells from control animals and those lacking the A_2A receptor but not in those T cells lacking the ability to secrete IFNγ. Importantly, activation of the A_2A receptor reduced both infarct size and inflammation in Rag1 knockout mice reconstituted with mature T cells but not in those reconstituted with CD4⁺ T cells lacking the A_2A receptor (Yang et al., 2006).

Overwhelmingly, therefore, the anti-inflammatory/tissue-protective effects of the A_2A receptor have been repeatedly demonstrated in several organs in a wide variety of in vivo model systems. Indeed, some anti-inflammatory medications have been proposed to derive some of their clinical benefit by promoting adenosine release (Cronstein et al., 1991; Gadangi et al., 1996; Hwang et al., 2001), and data from animal models suggest that this triggers an essential activation of the A_2A receptor (Cronstein et al., 1993; Montesinos et al., 2007).

The biochemical mechanism(s) of protection in vivo have also supported the in vitro observations. For instance, mice lacking the A_2A receptor display an enhanced release of TNFα and other cytokines, which is associated with increased gene transcription in response to injection of LPS. NF-κB is the major transcription factor responsible for production of pro-inflammatory cytokines in immune cells and, as described previously, studies in A_2A receptor-deficient mice have demonstrated an enhanced NF-κB activation associated with elevated IKK-mediated phosphorylation and proteasomal degradation of IκBα (Lukashev et al., 2004). In wild-type mice, administration of CGS21680 and ZM241385 were able to inhibit and enhance, respectively, CpG-mediated increases in inflammatory gene transcripts, presumably by modulating activation of the NF-κB pathway (Lukashev et al., 2004). Influences on the mitogen-activated protein kinase activation pathways and, in particular, activation of ERK1/2 have also been investigated, as these have been suggested to play a role in cell protection. For example, studies in lung have demonstrated that administration of A_2A receptor agonists induce a robust activation of the ERK pathway that is associated with reduced levels of pro-apoptotic markers (Rivo et al., 2007).

Clinical potential of A_2A receptor agonists

Despite all the overwhelming pre-clinical evidence that A_2A receptor-selective agonists can reduce inflammation and tissue damage, the most advanced agents (Regadenson from CV Therapeutics and Apadenoson from Adenosine Therapeutics) are actually in late-phase development as imaging agents/cardiac stress agents for coronary artery disease. This is based on the principle that activation of the A_2A receptor in the coronary circulation produces a profound vasodilation. Thus, infusion with a compound with a longer half-life (in the order of a few minutes) and better selectivity for the A_2A receptor than adenosine, which is currently utilized for such applications, would be expected to have a better and safer clinical profile in these conditions (Cerqueira, 2004).

Why then have the anti-inflammatory and tissue-protective properties of A_2A receptor agonists not yet been adequately tested in clinical trials? The A_2A receptor has a widespread tissue/cell-type distribution, and therefore activating this receptor might lead to unwanted side effects. For instance, the A_2A receptor mediates inhibition of platelet aggregation, hypotension and a variety of effects within the CNS, some of which involve interactions with dopamine D₂ receptors (see Ledent et al., 1997; Schwarzschild et al., 2006). Thus, chronic treatment of inflammatory conditions with an A_2A receptor-selective agonist might have an unacceptable side-effect liability. In support of this concept, Fozard et al. (2002) demonstrated that, following intratracheal administration, inhibition of lung inflammation by CGS21680 was accompanied by falls in blood pressure, and these authors speculated that strategies would have to be developed to target the action of adenosine A_2A receptor agonists within the lung to avoid the observed systemic side effects. Not withstanding the above consideration, at least two companies (Adenosine Therapeutics and Pfizer) are reported to be testing the anti-inflammatory potential of A_2A receptor-selective agonists in a variety of clinical situations.

Concluding remarks

Many studies have now established the ability of selective A_2A receptor activation to repress the exaggerated immune and inflammatory responses associated with many diseases. Although several high-affinity agonists with strong selectivity for the A_2A receptor have now been developed and tested in several animal models, a major aim in future will be to assess whether these will have a sufficiently low side-effect liability to be suitable for systemic administration in patients. The identification of adenosine as a critical factor that accumulates in tumours in response to hypoxia, and which prevents their destruction by inhibiting anti-tumour T-cell function via A_2A receptor activation (Lukashev et al., 2007), would also suggest that selective antagonists would be beneficial in many types of cancers. Finally, although the vast majority of the A_2A receptor's effects appear to be mediated via elevation of cAMP, unravelling the detailed molecular mechanisms by which it suppresses specific pro-inflammatory signalling pathways will be essential if we are to (1) understand why A_2A receptor activation is such a powerful strategy for turning off immune and inflammatory responses and (2) devise ways to minimize unwanted side effects of receptor activation arising from initiation of cAMP-independent signalling pathways.

Glossary

ADA

adenosine deaminase

cAMP

cyclic AMP

CRM1

chromosomal region maintenance 1

Epac

exchange protein directly activated by cAMP

ERK

extracellular signal-regulated kinase

HEK

human embryonic kidney

IκB

inhibitor of NF-κB

IFN

interferon

IKK

IκB kinase

IL

interleukin

IR

ischaemia–reperfusion

JAK

Janus kinase

LPS

lipopolysaccharride

NF-κB

nuclear factor-κB

NK

natural killer

PDE

phosphodiesterase

PKA

cAMP-dependent protein kinase

SOCS

suppressor of cytokine signalling

STAT

signal transducer and activator of transcription

TLR

Toll-like receptor

TNFα

tumour necrosis factor-α

TNFR

TNFα receptor