The critical role of inflammatory processes in health and disease has long been recognized,1 yet the detailed molecular mechanisms and biological events that regulate the progression and resolution of inflammation remain of interest. A number of recent investigations have provided strong evidence that the resolution of inflammation is not a passive process, as believed earlier.2–4 Instead, resolution is a biosynthetically active process, regulated by biochemical mediators and receptor-signaling pathways, and driven by specialized pro-resolving mediators (SPM). In particular, following a number of findings by Serhan and his group, the authors and their collaborators introduced and systematically investigated a number of SPM derived from polyunsaturated fatty acids (PUFA), including lipoxins, E-series resolvins, D-series resolvins, protectins/neuroprotectins, and, most recently, maresins. This review summarizes efforts on the resolvins and protectins with an emphasis on the corresponding biochemical pathways. Additional reviews covering different aspects of these anti-inflammatory and pro-resolving lipid mediators,5 including immunology,6,7 pathology,8 biochemistry,9 pharmacology10 and chemistry11 are also available.6–9,12
1.1 The Inflammatory Response and the Resolution of Inflammation
Localized acute inflammation is part of the host’s normal protective response to tissue injury and infection by invading microbial pathogens.1 Although this inflammatory response to a range of harmful stimuli is protective to the host, if kept uncontrolled it can result in a wide range of acute, chronic and systemic inflammatory disorders. Indeed, some of the most common and difficult to treat diseases are linked to excessive, uncontrollable, or chronic inflammation, including: cardiovascular disease, rheumatoid arthritis, periodontal disease, asthma, diabetes, inflammatory bowel disease (IBD), as well as neurological disorders such as Alzheimer’s disease and age-related macular degeneration (AMD).1,13,14 Although the involvement of inflammatory pathways in the initiation of all of these diseases is well established, the specific role by which inflammation contributes to their pathogenesis is not fully understood. The recent findings that the resolution of inflammation is an active process2–4,15 have provided new insights and created new paradigms for understanding and treating these conditions.
The key role of a number of lipid mediators in the initiation of the inflammatory response and the subsequent progression towards resolution is illustrated in Fig. 1. Among the first signaling events following microbial infection or tissue injury, is the release of pro-inflammatory lipid mediators, such as leukotrienes (LTs) and prostaglandins (PGs) that launch a series of signaling cascades with the ultimate goal of destroying the invading pathogens and repairing the damaged tissue.16–18 Thus, the biosynthesis and release of the potent chemotactic agent leukotriene B4 (LTB4) promotes the recruitment of neutrophils (PMNs) to the inflamed tissue, while the formation of prostaglandins E2 and D2 (PGE2 and PGD2)15 further accelerates the inflammatory process, ultimately resulting in a condition of acute inflammation. Despite its critical host-protective function, acute inflammation is not sustainable over a prolonged period of time, giving rise to disruptive conditions of chronic inflammation that may be responsible for the pathogenesis of a wide range of diseases, that can be attributed to a failure of resolution.19 Typically, the therapeutic treatment of such conditions involves the inhibition of pro-inflammatory mediators, but in many cases such approaches are often not very effective.
The recognition of the proactive nature of the resolution of inflammation has revealed alternative therapeutic paradigms based on resolving acute inflammation and preventing the onset of chronic inflammation.19 Indeed, a number of endogenous lipid mediators identified are able to act in this manner, suggesting a lipid mediator class switching3 from the initial actions of pro-inflammatory lipid mediators, i.e. leukotriene and prostaglandins, to the anti-inflammatory and pro-resolving actions of lipoxins, resolvins, protectins and maresins, which are discussed in this review. Each family of these pro-resolving mediators exert specialized actions, including blocking neutrophil recruitment, promoting the recruitment and activation of monocytes, as well as mediating the nonphlogistic phagocytosis and lymphatic clearance of apoptotic neutrophils by activated macrophages. Eventually, through the combined actions of these mediators, the resolution of inflammation is completed and homeostasis is reached. In this regard, it was important to introduce precise definitions for resolution mechanisms and indices19–21 that also permitted the first information that certain agents can be resolution-toxic.22–24
1.2 Lipid Mediator Pathways in Inflammation and Resolution: The Roles of Polyunsaturated Fatty Acids
The key role of several new classes of lipid mediators for regulating the resolution of inflammation has recently emerged by several recent discoveries,5–9,11,12 including extensive collaborative efforts.25–64 In particular, it was discovered that a number of novel lipid mediators derived from polyunsaturated fatty acids (PUFA) that are endogenously generated during inflammation have potent anti-inflammatory actions and serve as specialized pro-resolving lipid mediators (SPM)8,10 and are able to promote the resolution of inflammation.7,12,15 Several families of new lipid-derived SPM have been recently identified and characterized. The biosynthetic and signaling pathways of these pro-resolving molecules, as well as their relationships with selected pro-inflammatory mediators are outlined in Fig. 2.
1.2.1 Pro-inflammatory Lipid Mediators from Arachidonic Acid
At the onset of the inflammatory response, phospholipase enzymes (e.g. cPLA2) acting on phospholipids mediate the release of free polyunsaturated fatty acids (PUFA), including arachidonic acid (AA) and the omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). These fatty acids initiate a series of biosynthetic pathways involving lipoxygenases (human 5-LO, 65,66 12-LO and 15-LO) and cyclooxygenases, particularly the inflammation-induced isoform COX-2. Among the resulting PUFA-derived hydroxylated metabolites are several series of potent lipid mediators that exert their specialized inflammation-related actions by acting on specific G-protein coupled receptors (GPCR). Activation of these receptors by these lipids directly affects the expression levels of multiple enzymes, chemokines, cytokines, and growth factors that play dominant roles in inflammation and resolution.
Arachidonic acid has long been known to be involved in the initiation phase of inflammation, by forming pro-inflammatory leukotrienes (LTB4 and LTC4) and prostaglandins (PGE2 and PGD2) that govern the early events in host defense.16 Each of these potent molecules acts via its perspective GPCRs that are present in the membranes of the relevant cell types, and triggers the expression of inflammatory enzymes (5-LO,65,66 COX-2,67–70), chemokines and cytokines (IL-6, IL-8) that initiate and accelerate inflammation. Thus, activation of BLT1/2 by LTB4 leads to increased neutrophil recruitment, while activation of CysLT1/2 by LTC4 leads to airway smooth muscle contraction. Similarly, activation of the EP1–4 receptors by PGE2 and the DP1/2 receptors by PGD2 leads to multiple pro-inflammatory actions. The inhibition of key pro-inflammatory enzymes (e.g. 5-LO,65,66, COX-267–70) and the development of antagonists for key pro-inflammatory receptors (e.g. BLT1/2, CysLT1/2)71 has been extensively investigated as a means of developing therapeutics for inflammatory disorders, some of which are now in clinical use.
1.2.2 Anti-inflammatory and Pro-resolving Lipid Mediators from Arachidonic Acid
Notably, despite its central role in the initiation and progression of inflammation, AA is also involved in the biosynthesis of anti-inflammatory and pro-resolving lipid mediators.72,73 Most prominent are the lipoxins5,11 (LXA4 and LXB4), which are formed in humans to a large extent via transcellular biosynthesis through the sequential actions of 5-LO and 12-LOX with LTA4 as an intermediate or 15-LOX-initiated interactions with 5-LOX-bearing cells. Of interest, blocking or reducing LTA4 hydrolase given an increase in endogenous LXA4 biosynthesis and an anti-inflammatory response.74 The epimeric aspirin-triggered lipoxins,29,33,34 (AT-LXA4 and AT-LXB4) have similar actions as the lipoxins, but are formed from COX-2 in the presence of aspirin or via the P450 pathway. The biosynthetic pathways and biological functions of the lipoxins (LX) and aspirin-triggered lipoxins (ATL) have been described in several reviews.5,11,75 For reviews on the lipoxins and their analogs, design and synthesis, see Petasis et al.11 and Guilford et al.76 The discovery that LX undergo inactivation catalyzed by the enzyme 15-prostaglandin dehydrogenase (15-PGDH), led us to the first rational design and synthesis of a series of stable lipoxin analogs11,58 that retain all of the biological actions and sub-nanomolar potency of the lipoxins. These LX analogs served as key molecular probes for extensive investigations on the biological actions of the lipoxins, and provided the basis for the first LX-based lead compounds for clinical development.11,76 Both LXA4 and AT-LXA4 act on ALXR/FPR2 (for a recent review, see Ref. 77) and GPR32, and exhibit potent anti-inflammatory and pro-resolving actions.63 A prostaglandin-type pro-resolving lipid mediator is 15-deoxy-prostaglandin J2 (15dPGJ2),24,78 which is formed from PGD2. PGE2 and PGD2 can both switch the phenotype of human neutrophils from leukotrienes to lipoxins and resolution.3,20
1.2.3 New Pro-resolving Lipid Mediators from Omega-3 Fatty Acids
While enzymatic oxygenation of AA, an omega-6 fatty acid, generates both pro-inflammatory and pro-resolving lipid mediators, analogous pathways with the corresponding omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) generally lead to the formation of anti-inflammatory, pro-resolving and cytoprotective lipid mediators. Using an unbiased lipidomics and systems approach, the enzymatic oxygenation of omega-3 fatty acids, Serhan and collaborators identified, characterized and elucidated families of novel pro-resolving lipid mediators from EPA and DHA.6–9,12 These include the E-series resolvins (RvE1 and RvE2) 2,45 derived EPA, as well as the D-series resolvins (RvD1, RvD2, RvD3, RvD4, RvD5, RvD6),4,55,60 the neuroprotectins/protectins (NPD1/PD1),51,79–81 and the maresins (MaR1),82 which are derived from DHA. Additional series of lipid mediators formed in the presence of aspirin were also discovered, including the D-series resolvins4 and protectins and their aspirin-triggered forms,79 all of which are biosynthesized from PUFA and aspirin-acetylated COX-2 that acts as a modified dioxygenase inserting a molecule of oxygen with opposite stereochemistry.
1.3 The Beneficial Roles of Omega-3 Fatty Acids in Inflammation and Resolution
The discovery of resolvins, protectins and maresins is of particular significance, since these potent lipid mediators provided the first molecular basis for the many health benefits attributed to the omega-3 fatty acids EPA and DHA, which are abundant in fish and are widely used as dietary supplements. These essential fatty acids have long been associated with beneficial effects in human health and in the prevention of various diseases,83 including inflammation,84 immunomodulation,85 autoimmune diseases,84 rheumatoid arthritis,86 cardiovascular diseases,87,88 Alzheimer’s disease and other neurodegenerative diseases, type-2 diabetes,89 and cancer.90
Although the specific molecular mechanisms for these health benefits have remained unclear, the recent collaborative efforts by our team have provided the first detailed mechanistic insights for the likely pathways involved. The distinct properties of EPA and DHA to form primarily pro-resolving lipid mediators may explain their well-known beneficial health effects. These novel pathways may also explain some of the beneficial effects of aspirin, since they generate epimeric lipid mediators that are more metabolically stable and longer lasting.
1.4 Time-course of the Pro-inflammatory and Pro-resolving Lipid Mediator Pathways
During the inflammatory response, the biosynthesis of PUFA-derived lipid mediators increases with time (Fig. 3). The initial mediators (prostaglandins and leukotrienes) are produced within seconds and minutes and regulate the edema and postcapillary events at the site following the recruitment of neutrophils. With time (hours up to days), increasing leukocyte recruitment with the non-phlogistic appearance of monocytes and macrophages leads to a growing amount of mediators involving these cells (Fig. 3). The net result of this timeline is that initially mostly pro-inflammatory mediators (LTs, PGs) are produced, while at a later time, a lipid mediator class switching results in the production of pro-resolving mediators (LXs, Rvs, PDs) that promote resolution. For recent reviews on the cellular mechanisms, see Refs. 73,91,92 The release of AA and mobilization of EPA and DHA from circulation,57 and the formation of the pro-resolving mediators by these substrates also follows a time course that favors initial inflammatory response and host defense, followed by resolution and return to homeostasis. In the event that the pro-resolving lipid mediators are not produced or are defective, the acute inflammatory process remains unresolved and escalates into a state of chronic inflammation that promotes fibrosis and is associated with the pathogenesis of multiple diseases. Table 1 lists the many in vivo systems where a key enzyme is genetically manipulated in the biosynthesis of proresolving mediators including AA-derived LX and DHA-derived PD1 (see below).
1.5 Specialized Pro-resolving Lipid Mediators: Potential Therapeutics for Inflammation
Given the biological profiles of the new pro-resolving mediators, the pathways that drive the formation and actions of these molecules provide a new paradigm for treating inflammatory diseases. Indeed, our recent efforts on the identification, structural characterization, total synthesis, and biological investigation of selected members and structural analogs of these PUFA-derived lipid mediators have revealed their potent actions (nM and pM range) in a variety of cell types in vitro, as well as in many in vivo models of inflammatory diseases.7 These include: dermal inflammation,29,30 dorsal air pouch,2,4,33 peritonitis,21,44 periodontitis,36,42,50 colitis and intestinal inflammation,31,45 asthma and airway inflammation,93,94 cystic fibrosis,95,96 acute lung injury,59 ischemia-reperfusion injury,27 kidney injury,97 glomerulonephritis,38,98 ischemic-stroke,80 retinal degeneration,62,64 and Alzheimer’s disease.99 Resolvins of the D series and PD1 have been found to be important in protecting the host from obesity-induced insulin resistance and hepatic steatosis in murine models in vivo.100 Also, RvD1 improves insulin sensitivity by resolving the chronic inflammation that is associated with obesity. In this murine system, Hellmann et al found that RvD1 increased adiponectin production and decreased IL-6 as well as reduced the presence of crown-like structures within adipose tissue.101 Together, these results point to an important action of resolvins and protectins in the regulation of inflammation in fat tissues, which is an emerging area of global public health and concern.100,101 RvE1 also reduces inflammation in ocular tissues, reducing the number of macrophages in dry eye murine models and in the symptoms of dry eye.102 As a result of these studies, several synthetic SPM derived from our work are currently in clinical trials, including lipoxin analogs, as well as resolvin E1 and RvE analogs for ocular103–105 and systemic indications including lung, kidney, skin and bowel diseases.45,106–109 RvE1 and RvD1 each regulate and reduce pain110,111, as do the lipoxins112, which opens new means to control pain signals.113 Taken together, these findings demonstrate the profound importance and great therapeutic potential of these novel lipid mediator molecules, listed in Table 2.