(De-) Oiling Inflammasomes

Abstract

Activation of inflammasome signaling can produce harmful inflammation. In this issue of Immunity, Yan et al (Yan, 2013) suggest that omega-3 fatty acids commonly found in marine oils can suppress activation of NLRP3 and NLRP1b inflammasomes.

Marine-derived omega-3 (ω-3) polyunsaturated fatty acids (FA) such as docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) exhibit anti-inflammatory properties (Oh et al., 2010), but exact mechanisms for these properties are not fully understood. ω-3 FAs inhibit inflammatory cytokines such as IL-1β (Oh et al., 2010), prompting interest in the idea that ω-3 FAs can inhibit inflammasomes. In this issue of Immunity, Yan et al (Yan, 2013) provide insight into details of how such an inhibition may occur.

Inflammasomes are intracellular protein complexes that serve as sensors of microbial and endogenous cellular insults (Davis et al., 2011; Vladimer et al., 2013). Release of inflammasome-derived IL-1 β and IL-18 typically occurs in a two-step process: triggering of a Toll like receptor (TLR) that primes the inflammatory response (“signal 1”) and intracellular danger signals that initiate formation of the inflammasome complex (“signal 2”).

The NLRP3 inflammasome is activated by a large and diverse array of pathogens as well as endogenous and exogenous sterile agonists and danger signals. It is not surprising that NLRP3 has been implicated to play a role in several inflammatory disorders such as gout and atherosclerotic disease by potentially sensing uric acid crystals and cholesterol crystals, respectively (Davis et al., 2011; Duewell et al., 2010). Recent evidence pointed to a role for inflammasomes in initiating obesity-induced inflammation and insulin resistance related to type 2 diabetes (Vandanmagsar et al., 2011) (Henao-Mejia et al., 2012) and other metabolic disorders (Davis et al., 2011). The increase in obesity and chronic inflammatory states underscores the need for clinical therapeutics to prevent and treat inflammation.

Because ω-3 FAs can inhibit pro-inflammatory cytokines such as TNF and IL-1β (Oh et al., 2010) and NLRP3 activation promotes IL-1 β release, Yan et al (Yan, 2013) set out to examine the possibility that thwarting IL-1β release by ω-3 FAs was mediated by blocking inflammasome activation. They confirmed that pretreating cells DHA prior to stimulation blocked caspase-1 activation and IL-1β and IL-18 production. ω-3 FAs EPA and to a lesser extent the plant-derived α-Linolenic acid (ALA) also blocked IL-1β, but ω-6 and ω-9 FAs had no effect, indicating this inhibition is specific for certain ω-3 FAs.

DHA inhibited activation for all NLRP3 agonists tested, but failed to inhibit activation induced by activators of other inflammasome components, such as Salmonella (NLRC4 inflammasomes) or DNA (AIM2 inflammasomes). However, DHA also blocked activation of the NLRP1b inflammasome which is activated by the microbial anthrax lethal toxin. This finding suggested ω-3 FAs as specific inhibitors of NLRP3 and NLRP1b inflammasomes.

The surface-exposed G protein-coupled receptor 120 (GPR120) can mediate anti-inflammatory effects of DHA and EPA (Oh et al., 2010). By inhibiting or decreasing expression of both GPR120 and GPR40, DHA treatment no longer repressed caspase-1 activation and IL-1β release compared to partial restoration of cytokine release when GPR120 or GPR40 were individually inhibited. Therefore, ω-3 FAs signal through GPR120 and GPR40 to inhibit NLRP3 activation.

Additional mechanistic investigations revealed that gene targeting of the GPR120 downstream scaffold protein β-arrestin-2 (ARRB2) led to abrogation of the inhibitory effects of DHA and EPA on NLRP3 and NLRP1b. Cells from animals deficient in ARRB2 showed only partial abrogation of DHA inhibitory activity, indicating that another pathway in addition to GPR120-GPR40-ARRB2 may also be involved in NLR inflammasome inhibition induced by DHA.

Interestingly, ARRB2 interacted with NLRP3 and NLRP1b, and not NLRC4 or AIM2, providing a likely reason why DHA does not inhibit NLRC4 or AIM. DHA and EPA treatment promoted interaction between NLRP3 and ARRB2 via GRP120 and GRP40. All together, these results demonstrate that ARRB2 acts downstream of GPR120 and GPR40 to inhibit inflammasome activation by binding to NLRP3 or NLRP1b.

Interestingly, ω-3FAs play a more global role in inhibiting inflammatory cytokines and subsequently inflammatory disease in an NLRP3-dependent manner in vivo. Mice fed a high fat diet (HFD) develop characteristics of type 2 diabetes (T2D). HFD-fed mice supplemented with DHA showed remarkable decreases in T2D symptoms compared to normal diet-treated mice that were not further reduced in NLRP3-deficient animals, indicating that DHA prevents HFD-induced metabolic disorder in a NLRP3-dependeent manner.

In summary, Yan et al (2013) identified a mechanism for how ω-3FAs exert their anti-inflammatory properties via inhibition of inflammasome activation. It is noteworthy that ω-3FAs may inhibit this activation at several steps, by hampering both “signal 1” and “signal 2”, as ω-3FA inhibition of NF-kB signaling has also been reported (Glass and Olefsky, 2012). Thus, several inflammatory signaling pathways may be affected. This study elegantly ties together the interplay between dietary components and known inflammatory pathways (Figure 1) and could have implications for the prevention and treatment of inflammatory disorders.

Figure 1. Activation and inhibition of NLRP3 inflammasome signaling.

Activation of the NLRP3 inflammasome is a two-signal process. “Signal 1” occurs by stimulation of toll-like receptors (TLRs) by microbial or endogenous ligands leading to NF-κB-dependent upregulation of the pro-IL-1β. “Signal 2” is provided by a wide array of NLRP3 inflammasome activators such as nigericin, anthrax lethal toxin, β-amyloid, islet amyloid polypeptide and cholesterol and monosodium irate crystals (not shown). Activated NLRP3 binds to pro-caspase-1 via the adaptor protein ASC, leading to caspase-1 cleavage of pro- IL-1β and IL-18. Yan et al. (2013) show that the mechanism for ω-3FA inhibition of NLRP3-mediated inflammation involves ω-3FAs interacting with the GPR120 and GPR40 receptors. Subsequently, the downstream scaffold protein ARRB-2 binds to GPR120 and GPR40 and the complex is internalized. There are reports that this pathway inhibits “signal 1” at the level of TAB1 and TAK1 kinases to inhibit NF-κB (Glass and Olefsky, 2012). Yan et al. (2013) also demonstrate here how “signal 2” is inhibited when ARRB-2 directly associates with NLRP3 leading to inhibition of pro-inflammatory cytokine release, and inhibition of inflammation.

One important issue raised by this study is whether ω-3FAs would have the same effect in human obesity and inflammasome-driven inflammation as in mouse studies, and if the intestinal microbiota has a role in this setting (Henao-Mejia et al., 2012). Also, treating human patients with anti-inflammatory agents can increase susceptibility to a number of infections, and it remains to be seen if the same effect is observed with ω-3FAs. It should be noted that mouse studies suggest both increased and decreased host resistance to infections following fish oil feeding of mice (Anderson and Fritsche, 2002), although this issue is incompletely understood. Supporters of ω-3FAs promote their therapeutic potential for several disease conditions, but some larger meta-studies have failed to show beneficial effects of ω-3FAs in conditions like heart disease (Rizos et al., 2012), thought to be influenced by ω-3FAs. One reason for variability between different human population studies may be that they may vary in how the FAs are provided and how studies are controlled. Are the same effects achieved upon simple dietary changes, ω-3FAs provided as an oil supplement, or as purified ω-3FAs? More research is obviously needed to further establish the role of ω-3FAs in inhibiting inflammation in human disease. In an important step forward, Yan et al provide us with a framework of mechanistic insight into how inflammation could potentially be reduced in a variety of inflammatory states.

Cod liver oil, anyone?