Protectins, Resolvins and Maresins - Specialized
The oxygenated metabolites or oxylipins derived mainly from eicosapentaenoic acid (20:5(n-3) or EPA), docosapentaenoic acid (22:5(n-3) or DPA) and especially docosahexaenoic acid (22:6(n-3) or DHA) and termed the "(neuro)protectins, resolvins and maresins" have potent anti-inflammatory and immunoregulatory actions at concentrations in the nanomolar to picomolar range. Their structures have been conserved in evolution from diatoms to flatworms to fish to humans, and they function in all major organ systems. They are produced by dioxygen-dependent oxidation from these omega-3 essential fatty acids, which are the focus of considerable interest among nutritionists because of the perceived beneficial effects for the health of consumers. Although the mechanisms by which such effects are exerted are not entirely clear and evidently involve a multiplicity of factors, it seems likely that their oxygenated metabolites play a significant part. The therapeutic potential of these oxylipins is appreciable, and it is hoped that their biosynthesis can be modulated by dietary supplementation with their fatty acid precursors in addition to by direct administration of individual compounds.
Simple lipoxygenase and cytochrome P450 metabolites of EPA and DHA, as well as non-enzymic isoprostane analogues (neuroprostanes), which have related biological properties, are discussed elsewhere on this site for reasons of practical convenience.
1. Definitions and Key Features
The most important of the protectins and related oxylipins are derived from DHA and are dihydroxylated E,E,Z-docosatrienes, i.e. acyclic lipoxygenase metabolites containing a conjugated triene unit with an E,E,Z‑configuration and flanked by two secondary allylic alcohols (together with other non-conjugated cis-double bonds). However, the nomenclature can be confusing as it has changed as the science has developed. Three novel mediator super-families derived from EPA and DHA are now recognized. The trivial name 'neuroprotectin' was coined by Professor Charles N. Serhan and colleagues for one class of these molecules as they were first found in neuronal tissue, though the term 'protectin' was later preferred when it was realized that they occur in many more animal tissues. Subsequently, oxylipins with similar structural features but formed by different enzymatic routes were characterized and termed 'maresins'. Other oxygenated derivatives of DHA, DPA and EPA were named 'resolvins' or 'resolution-phase interaction products', because these compounds were first encountered in resolving inflammatory exudates (pus).
As this range of lipid mediators has expanded with continuing research (35 at the last count in 2020), they have been collectively termed ‘Specialized Pro-resolving Mediators’ or ‘SPMs’ because of their intimate involvement in the resolution of inflammation, as discussed below. Oxylipins derived from EPA are designated as SPMs of the E series, while those formed from the precursors DHA and DPA are denoted as SPMs of the D series.
The lipoxins, derived from arachidonic acid of the omega-6 family of polyunsaturated fatty acids, are a fourth class of SPMs and indeed were the first to be discovered, but they are discussed elsewhere on this site in the web page dealing with the leukotrienes to which they are structurally and biosynthetically related.
It was long thought that resolution of inflammation was a passive process, occurring simply by the dilution of inflammatory signals from the site of injury or infection, but a concept has now emerged in which acute inflammation is initiated in animal tissues by prostaglandins and leukotrienes, before there is a switch in metabolism to the production of SPMs that end the inflammatory process. The acute inflammatory response is a coordinated process, and both initiation and resolution stages are required to maintain healthy tissues; SPMs are key factors in the latter as is discussed below. As uncontrolled or dysregulated inflammation is now widely recognized to be a unifying phenomenon in many chronic diseases, SPMs and derived molecules are believed to have great therapeutic potential.
The first step in the biosynthesis of the dihydroxylated oxylipins derived from of EPA or DHA is the release of the free acids from linkage in phospholipids by the action of specific phospholipases. Thereafter, there are a number of common steps in the mechanism of oxygenation, starting with an antarafacial hydrogen abstraction at the C3 position in one of the cis,cis-1,4-diene moieties. Then, there is a stereo-selective insertion of molecular oxygen to form a carbon-oxygen bond in a hydroperoxide intermediate, followed by a second hydrogen abstraction leading to an intramolecular nucleophilic attack by the oxygen atom in this intermediate to produce an epoxide. The final step involves hydrolase-assisted nucleophilic addition of water to a cis-double bond in the epoxide intermediate. Sequential actions of COX-2/aspirin, various lipoxygenases and CYP450 oxygenases are required as summarized in the following figure.
Although specific SPMs and SPM clusters were first found in biologically active amounts in human inflammatory exudates, they are now known to be present in most tissues and body fluids. Thus many different molecular forms are have been identified by lipidomic techniques in human peripheral blood and serum, lymph nodes, spleen, brain and breast milk, and measurement of these can often be indicative of disease status.
It should be recognized that establishing the precise stereochemistries of the double bonds and hydroxyl groups in SPMs, features that are essential for their biological functions, has required heroic efforts involving total synthesis of large numbers of possible isomers for structural comparisons with those formed naturally.
2. Protectins (Neuroprotectins)
In studies of metabolite formation from DHA in brain tissue in response to aspirin treatment, it was shown that new docosanoids were produced that were initially termed ‘neuroprotectins’. Like the leukotrienes, there are three double bonds in conjugation (so they are sometimes termed E,E,Z‑docosatrienes) with six double bonds in total. As it is now recognized that the formation and actions of these docosanoids are not restricted to neuronal tissue, the simpler term ‘protectins’ is preferable. For example, protectin D1 is present in murine inflammatory exudates and lung, in peripheral human blood and exhaled breath condensates, and in a wide range of cell types. The biosynthetic pathway to neuroprotectin NPD1 or protectin D1, as established in murine brain tissue, and human leukocytes and lymphocytes, is illustrated.
Uniquely among SPMs, the initial product is 17S-hydroperoxy-DHA by reaction with 15-lipoxygenase (ALOX15), and this is converted first to a 16(17)-epoxide and then to the 10,17‑dihydroxy-triene unit of 10R,17S-dihydroxy-docosa-4Z,7Z,11E,13E,15Z,19Z-hexaenoic acid, denoted as 10R,17S-DT or PD1 (or NPD1). Each step in the biosynthetic sequence is under precise stereochemical control by enzymes, and the structures of these DHA-derived mediators are highly conserved from fish to humans. Synthesis of PD1 is induced as a response to oxidative stress and/or activation of neurotrophins, and this highly stereospecific structure is essential for biological activity.
Further oxygenation can occur, and 17S-hydroperoxy-DHA can react with 5-lipoxygenase to form 10S,17S-dihydroxy-docosatriene in a double di‑oxygenation reaction, as illustrated, together with some of the 7,17-isomer. In the action of 5-LOX, the role of the accessory protein FLAP, which is so important in leukotriene biosynthesis, appears to require some clarification, but there is a suggestion that cytosolic 5-LOX, uncoupled to FLAP, favours synthesis of pro-resolving mediators over that of leukotriene B4. The intermediate 16,17-epoxy-docosatriene can also undergo non-enzymatic hydrolysis to yield 10R/S,17S-dihydroxy- and 16R/S,17S-dihydroxy-docosatrienes. 22-Hydroxy-PD1 has also been detected in human tissues and has significant biological activity.
Both 22:5(n-3) and 22:5(n-6) fatty acids are good substrates for 15-lipoxygenase. Thus, 22:5(n-3) is first subjected to 17-lipoxygenation and then via an epoxy intermediate to either 10,17-dihydroxy-7Z,11,13,15,19Z-docosapentaenoic acid (designated PD1n-3 DPA) or 16,17-dihydroxy-7Z,10,13,14,19Z-docosapentaenoic acid (PD2n-3 DPA). 22:5(n-6) gives 17S-hydroxy-22:5(n-6) and 10,17S-dihydroxy-22:5(n-6) as the main products. An omega-hydroxylated analogue of PD1 (22‑OH‑PD1n‑3 DPA), produced by the action of a CYP1 monooxygenase, has also been detected in human neutrophils and monocytes. All of these are potent anti-inflammatory agents.
Poxytrins: Unfortunately, an isomer generated from DHA by the action of plant 15-lipoxygenase in a double di-oxygenation reaction was misidentified as PD1 and was made available commercially under that name, leading to some erroneous reports in the literature. This isomer (now designated ‘PDX’) differs in the geometry of double bonds in the conjugated triene, which is E,Z,E for PDX and E,E,Z for PD1, and also in the configuration of carbon 10, which is S in PDX and R in PD1.
On the other hand, PDX is now known to be a minor endogenous metabolite in murine peritonitis, and small amounts have been detected in rat brain, both in free form and esterified to phospholipids. Similar oxylipins with the E,Z,E-conjugated triene motif between two secondary hydroxyl groups have been characterized that are related to leukotrienes (LTB4 and LTBX), and collectively they have been named 'poxytrins' (PUFA oxygenated trienes). They are believed to be produced in animal tissues by sequential oxidation by 15-LOX and perhaps a second lipoxygenase, followed by reduction by cytosolic glutathione peroxidase.
Elovanoids: Very-long chain (C32 and C34) oxylipins produced by the action of elongases, especially ELOV4 - hence the name, on DHA in brain and retina are converted to protectin analogues under conditions of oxidative stress, i.e. to (14Z,17Z,20R,21E,23E,25Z,27S,29Z)-20,27-dihydroxydotriaconta-14,17,21,23,25,29-hexaenoic acid (ELV-N32) and (16Z,19Z,22R,23E,25E,27Z,29S,31Z)-22,29-dihydroxytetratriaconta-16,19,23,25,27,31-hexaenoic acid (ELV-N34). They are low-abundance, high-potency, protective mediators. In contrast to DHA per se, which is located in position sn-2 of phospholipids in these tissues, the precursors of the elovanoids are located in position sn-1. This implies that the stimulus for the release of the precursors and formation of the novel mediators must be somewhat different from that of the protectins.
Catabolism: Protectin D1 is subjected to rapid β-oxidation from the carboxyl end of the molecule by cytochrome P450 enzymes, and the intermediates 2,3‑dinor‑PD1 and 2,3,4,5‑tetranor-PDI have been characterized; this process may limit the concentration of PD1 in the systemic circulation and prevent its excretion in urine.
3. Aspirin-Triggered Protectins
An important additional route to protectin formation produces the 17R-epimer, i.e. 10R,17R-dihydroxy-docosa-4Z,7Z,11E,13E,15Z,19Z-hexaenoic acid, and requires the intervention of aspirin (acetylsalicylic acid), the mode of action of which was described in our web page on prostaglandins. In brief, aspirin blocks the catalytic site of COX-1 by acetylating it irreversibly, but it can only partially block that of COX-2. The acetylated COX-2 retains lipoxygenase-like activity similar to that of 15-lipoxygenase, but with the oxygen insertion in the R- rather than S-configuration as is the case with lipoxygenases; the initial oxygenation reaction is not catalysed by unacetylated COX-2 or COX-1. With arachidonate as substrate, the 15R-HETE is converted to lipoxins, the first specific lipid mediators known to initiate resolution of inflammation. In effect, low-dose aspirin jump starts the resolution phase. Note that unlike aspirin, most non-steroidal anti-inflammatory drugs inhibit cyclooxygenases reversibly and can delay complete resolution.
Production of 'aspirin-triggered' resolvins (see below) and then of protectins was first identified in resolving exudates and brain in mice and is now well established for human cells. Once more, the anti-inflammatory effects of the 10R,17R-isomer are dependent on the precise stereochemistry, as the 10S,17R‑dihydroxy-docosatriene is essentially inactive in vivo.
(R)-Resolvins (aspirin-triggered): Resolvins are produced from EPA, DPA and DHA, and the trans-cellular mechanism is illustrated below for those derived from EPA. Biosynthesis of the 18R (illustrated) and 18S resolvins of the E series (name derived from EPA) has elements in common with the synthesis of the epi‑lipoxins, leukotrienes and of course the protectins.
In vascular endothelial cells derived from blood vessels during inflammation, the cyclooxygenase enzyme COX-2 that has been acetylated by aspirin introduces an 18R‑hydroperoxy group into the EPA molecule (see previous section); this can also be accomplished by a cytochrome P450 oxygenase. The product is reduced to the corresponding hydroxy compound and transferred to a neighbouring leukocyte before a 5S-hydroperoxy group is introduced into the molecule by the action of 5‑lipoxygenase (ALOX5), as in the biosynthesis of the leukotrienes. A further reduction step produces 15S,18R‑dihydroxy-EPE or resolvin E2 (RvE2). Alternatively, the 5S‑hydroperoxy,18R-hydroxy-EPE intermediate is converted to a 5,6-epoxy fatty acid in polymorphonuclear neutrophils in humans and eventually to 5S,12R,18R-trihydroxy-6Z,8E,10E,14Z,16E-eicosapentaenoic acid or resolvin E1 (RvE1) by an enzyme required for the biosynthesis of leukotrienes in leukocytes, i6 .e. leukotriene A4 hydrolase (LTA4H). There is also a resolvin E3 (RvE3), 17,18-dihydroxy-5Z,8Z,11Z,13E,15E-EPE, synthesised by eosinophils via the 12/15‑lipoxygenase pathway.
DHA is converted to 17R-resolvins by a similar aspirin-triggered COX-2 mechanism to the previous (in the absence of aspirin, COX-2 in human microvascular endothelial cells converts DHA to 13S-hydroxy-DHA). Thence, enzymatic epoxidation generates either 7S,(8)-epoxy or 4S,(5)-epoxy intermediates, which are acted upon by 5-lipoxygenase with the aid of the 5-lipoxygenase-activating protein (FLAP - see again our web page on leukotrienes) to yield the resolvins. The former produces the aspirin-triggered resolvins D1 and D2 illustrated ('D' nomenclature derived from DHA), while the latter produces the aspirin-triggered resolvins D3 and D4 with all containing a 17R‑hydroxyl group. AT-RvD1 is 7S,8R,17R-trihydroxy-docosa-4Z,9E,119E,13Z,159E,19Z-hexaenoic acid.
(S)-Resolvins: The highly specific stereochemistry of resolvin E1 is required for activation of a ligand-specific receptor and thence for its biological activity down to picomolar concentrations. However, epimeric 18S-resolvins are also produced in vivo by related biosynthetic pathways, with the first step catalysed by 15-LOX as for protectins, and these have their own distinctive biological activities. Similarly, the precursor 18(R)- and 18(S)-hydroxy-EPEs have been shown to have anti-inflammatory effects in vitro.
In an alternative reaction in the absence of aspirin in human whole blood, isolated leukocytes and glial cells, 15-lipoxygenase (ALOX15) generates 17S‑hydroxy-DHA as the initial product. This is converted to 7S-hydroperoxy,17S-hydroxy-DHA by the action of a lipoxygenase, and thence via epoxy intermediates to resolvin (RvD1 or 7S,8R,17S‑trihydroxy-docosa-4Z,9E,11E,13Z,15E,19Z-hexaenoic acid) and epimeric resolvin D2 (RvD2 or 7S,16R,17S-trihydroxy-docosa-4Z,8E,10Z,12E,14E,19Z-hexaenoic acid), i.e. all contain a 17S-hydroxyl group.
A further lipoxygenase-generated intermediate from 17S-hydroxy-DHA is transformed via an epoxide 4S,5S-epoxy-17S-hydroxy-DHA to resolvins D3 and D4 (4S,11R,17S- and 4S,5R,17S-trihydroxydocosahexaenoic acids, respectively). 17R- and 17S‑hydroxy-DHA have anti-inflammatory properties of their own, although they have generally been viewed simply as pathway markers and have been found in blood samples. Two further resolvins designated RvD5 and RvD6 (7S,14S- and 4S,17S‑dihydroxydocosahexaenoic acids, respectively), with an isomer in the cornea termed Rv6Di, and other related oxylipins have now been characterized.
Docosapentaenoic acid (22:5(n-3) or DPA) can also be converted to resolvins, first by 17-lipoxygenation to 17-hydroperoxy-8Z,10Z,13Z,15E,19Z-docosapentaenoic acid followed by a 5-lipoxygenase-like reaction to yield three products, of which the most abundant is 7,8,17-trihydroxy-9,11,13,15E,19Z-docosapentaenoic acid (designated RvD1n-3 DPA). Four further metabolites of DPA have a hydroxyl group in position 13 and have been designated as 13‑series resolvins (RvTs). For example, RvT1 is (7,13,20-trihydroxy-8,10,14,16Z,18-docosapentaenoic acid. Biosynthesis of these molecules is initiated in the vascular endothelium when cyclooxygenase-2 converts docosapentaenoic acid to 13-hydroperoxy-7Z,10Z,14E,16Z,19Z-docosapentaenoic acid, which is rapidly hydrolysed to the 13(R) intermediate for donation to neutrophils where it is converted to RvT1.
The marine diatom (microalgae) Cylindrotheca closterium produces appreciable amounts of SPMs, including RvE3 and novel structurally related eicosanoids derived from EPA, including 14S/R,17R,18R-trihydroxy-eicosatetraenoic acid, which displays inflammation-resolving and anti-inflammatory bioactivities in animals.
Catabolism: Resolvin E1 is eventually de-activated in tissues by oxidation by the 15-hydroxyprostaglandin dehydrogenase common to the catabolism of many oxylipins, then proceeds via at least four distinct pathways, including conversion to 18- and 12‑oxo‑RvE1, for example, prior to further oxidation. Similarly, RvD1 is de-activated by oxidation to 8- or 17-oxo-RvD1 as the first step in catabolism, while RvD4 is oxidized to 17‑oxo‑RvD4.
5. Maresins and Related Docosanoids
An alternative single oxygenation is found in human macrophages and platelets in which a mediator termed maresin 1 (7R,14S-dihydroxy-docosa-4Z,8E,10E,12Z,16Z,19Z-hexaenoic acid - ‘macrophage mediator in resolving inflammation’ or ‘MaR1’) is formed. Biosynthesis is initiated in macrophages by 14‑lipoxygenation of DHA by the action of human 12-lipoxygenase (ALOX12 or ALOX15), producing the hydroperoxy intermediate 14S-hydroperoxydocosa-4Z,7Z,10Z,12E,16Z,19Z-hexaenoic acid that is subsequently converted to 13S,14S-epoxy-maresin. This is believed to undergo enzymatic hydrolysis via an acid-catalysed nucleophilic attack by water at carbon-7, resulting in the introduction of a hydroxyl group at that position and double-bond rearrangement to form the stereochemistry of bioactive maresin 1 (the required enzymes have yet to be characterized).
In addition, 13S,14S-epoxy-maresin (13,14-eMar), which has important biological activities of its own, is the precursor for 13R,14S-dihydroxy-docosahexaenoic acid, designated maresin 2 (MaR2), via the action of the soluble epoxide hydrolase, and this metabolite also displays potent anti-inflammatory and pro-resolving actions. MaR1 can also be produced by circulating neutrophil–platelet aggregates in which DHA is converted to 13S,14S‑eMaR by the platelet 12‑lipoxygenase, before the intermediate is donated to neutrophils for the hydrolysis step. Similar oxygenated compounds with anti-inflammatory properties are formed from 22:5(n-3) and 22:5(n-6) fatty acids with those derived from the former designated MaR2n-3 DPA and MaR3n-3 DPA. MaR1 has been found in primitive invertebrates suggesting that its structure and function have been conserved in evolution.
Maresin-like di-oxygenated metabolites of DHA involving sequential oxidation by 12-LOX and enzymes of the cytochrome P450 family have been shown to occur in macrophages to produce docosanoids such as 14S,21S‑dihydroxy-docosa-4Z,7Z,10Z,12E,16Z,19Z-hexaenoic acid and its epimers. Their synthesis is induced by wounding and they have been shown to promote the healing of wounds. Similar 14,22‑dihydroxy-metabolites are synthesised by a related mechanism in leukocytes and platelets and also promote wound healing. 14S,20R‑dihydroxy-DHA is produced by the action of 12/15-lipoxygenase in eosinophils and has been detected in inflammatory exudates, while 22-hydroxy-MaR1 is produced by neutrophils and 14-oxo-MaR1 by macrophages in inflammation induced by bacterial infection; 22‑hydroxy-MaR1 retains the potent biological actions of its parent.
6. Sulfido-Conjugates of Specialized Pro-Resolving Mediators
Sulfido-peptide conjugated mediators with some structural similarity to the cysteinyl-leukotrienes and with novel biological properties are now known to be produced from SPMs in macrophages, and they have been termed protectin conjugates in tissue regeneration (PCTR), resolvin conjugates in tissue regeneration (RCTR), and maresin conjugates in tissue regeneration (MCTR). For example, while 13S,14S-epoxy-maresin, produced as described above, has biological activity in its own right, it is also the precursor of a family of novel mediators with some structural similarity to the cysteinyl-leukotrienes, i.e. 13-glutathionyl,14-hydroxy-docosahexaenoic (MCTR1), 13‑cysteinylglycinyl,14-hydroxy-docosahexaenoic (MCTR2) and 13‑cysteinyl,14-hydroxy-docosahexaenoic (MCTR3) acids. Indeed, it is now evident that the same enzymes are used for the biosynthesis of these two functionally distinct lipid mediator families. In brief, 13S,14S-epoxy-maresin is acted upon by the leukotriene C4 synthase or by glutathione S-transferase Mu 4 to produce MCTR1, which is converted to first to MCTR2 by a γ-glutamyltransferase and thence to MCTR3 by a dipeptidase.
Similarly, sulfido-conjugates are produced by resolvins (RCT1, RCT2 and RCT3) and protectins, and those of the 17-series are produced by human leukocytes and are highly abundant in lymphatic tissue. RCT3 is 8R-cysteinyl-7S,17S-dihydroxy-4Z,9E,11E,13Z,15E,19Z-docosahexaenoic acid, for example. Macrophages produce many such metabolites including a protectin analogue PCTR1 derived from 16S,17S-epoxy-protectin, and PCTR1 in turn gives rise to PCTR2 and PCTR3. These oxylipins have been detected in lymph nodes, serum and milk in humans, and they regulate the ability of the system to clear bacteria and to repair and regenerate damaged tissues. They have also been detected in mice and primordial organisms such as Planaria.
7. Biological Activities of Specialized Pro-Resolving Mediators
The protectins, resolvins and maresins (SPMs) are distinctive lipids with highly stereospecific structures that are endogenous local mediators with strong pro-resolution effects in addition to some immunoregulatory activities at picomolar to nanomolar concentrations. This high degree of stereospecificity is essential for their functions, and unnatural stereo-isomers tend to be much less active. It should be noted that, strictly speaking, pro-resolution is not identical to anti-inflammation. As part of the molecular mechanisms that contribute to removal of inflammatory cells and restoration of tissue integrity once the need for the inflammatory response is over, SPMs actively assist in the resolution of inflammation, once thought to be a passive process. In addition, they are strong attenuators of oxidative stress. In that it controls many aspects of both the innate and adaptive arms of the immune response, the nervous system is a central player in stimulating SPM production with neurotransmitters such acetylcholine being of particular importance. It is noteworthy that SPMs are not immunosuppressive. Several SPMs may be involved in the response to an inflammatory challenge, but each will have its own time-course for biosynthesis and insertion into the process, and the following discussion is necessarily simplistic and can only touch upon what is an increasingly complex and rapidly developing topic.
Although some have yet to be identified, each SPM is believed to interact with its own G protein coupled receptors, which evoke rapid intracellular signalling, and also long-term actions via regulating the expression of specific genes involved in the resolution of inflammation (Table 1). Depending on the cellular context, each receptor is capable of interacting with more than one SPM. For example, RvD1 interacts with DRV1 for homeostatic functions and with ALX/FPR2 for anti-neutrophil actions in resolving inflammation, while for maresin 1, RORα is a nuclear receptor and LGR6 is a plasma membrane G protein coupled receptor. It is noteworthy that SPMs can also antagonize pro-inflammatory receptors, and RvE1, MaR1 and 22-OH-MaR1 are all competitive antagonists of the leukotriene B4 receptor BLT1. In addition, a number of peptides have been identified that bind to some of these receptors with pro-resolving effects.
Table 1. Receptors for SPMs in mice and humans.
|Lipoxin A4 and AT-LXA4, RvD1, AT-RvD1, RVD3||ALX/FPR2 and GPR32|
|RvD1, AT-RvD1, RvD3, LXA4, AT-LXA4, RvD5||DRV1 (GPR32)|
|RvE1 and RvE2||ERV1 (ChemR23) and BLT1|
|DPA resolvins (RvD5n-3PDA)||GPR101|
|Maresin 1||RORα, LGR6|
Resolution of inflammation: Acute inflammation in response to infection or tissue damage is an essential defensive process that is usually characterized by heat, redness, swelling and pain at a simple observational level, and by oedema, accumulation of neutrophils, and then by accumulation of phagocytes such as monocytes and macrophages at a cellular level. The latter are the first line of defense of the innate immune system to defeat pathogens and toxins traveling through the body, and they protect each organ by engulfing and killing invading pathogens and clearing debris from within cells. While the symptoms of chronic inflammation may not appear as serious as those of acute inflammation, they must be controlled as the consequences can include tissue damage and loss of function without proper resolution. Leukotrienes (especially LTB4) and prostaglandins (PGE2 and PGD2) derived from arachidonic acid are important in the early stages of the inflammatory process by stimulating the migration of neutrophils to the affected tissue. However, biosynthesis of these prostaglandins is also critical for resolution of inflammation as they stimulate the induction of the 15‑lipoxygenases that are necessary for the production of lipoxins, resolvins and protectins. Then, efficient resolution of inflammation may have beneficial effects upon atherosclerosis and other chronic autoimmune diseases (see next section).
Early in the acute inflammatory response, the origins are laid for biosynthesis of SPMs, i.e. a change in lipid mediator production in which arachidonic acid metabolism switches from the production of leukotrienes to that of lipoxins. Local mobilization of the precursor DHA occurs before the autacoid protectins, resolvins and maresins are produced at the site of inflammation to help tissues to return to health by promoting resolution of inflammation through recruitment of non-inflammatory monocytes while limiting the recruitment of proinflammatory granulocytes. After this switch in the nature of the mediators formed, macrophages and mast cells are able to remove excess neutrophils together with cellular debris via draining lymph nodes, ideally without leaving remnants of the host defenses or of the invading microorganisms or other inflammatory initiators. It is evident that the presence of aspirin uniquely facilitates the resolution of inflammation. Thus, at local sites of inflammation, aspirin treatment enhances the conversion of the omega-3 fatty acids EPA and DHA to 18R- and 17R-oxygenated products, respectively, i.e. precursors of resolvins of the E and D series, which carry potent anti-inflammatory signals.
It has become apparent that a novel aspect of the resolution process is that SPMs are able to induce changes in the phenotype of macrophages toward a pro-resolution state. During inflammation, polymorphonuclear neutrophils are produced which have generally beneficial effects in countering disease by removing invading pathogens by phagocytosis, but in the longer term or if malfunctioning they may eventually cause trauma and tissue damage through infiltration into tissues. The resolvins, like the lipoxins, appear to have an important role in regulating and indeed inhibiting these harmful effects. In so doing, they oppose the effects of some of the pro-inflammatory prostanoids. A cluster of SPMs, including RvD3, RvD4, and RvD6, are produced in human blood to targets leukocytes at the single-cell level, directly activating extracellular signalling in human neutrophils and monocytes to promote host defense by enhancing phagocytosis and killing bacteria.
Nanomolar concentrations of resolvin E1 derived from EPA dramatically reduce dermal inflammation, peritonitis, dendritic cell migration, lung damage from smoking, and interleukin production. For example, in a model of periodontitis, RvE1 alone of the SPMs was able to reduce local inflammation by suppressing superoxide production directly in primary neutrophils. RvE1 blocks excessive platelet aggregation, it limits the effects of certain human pathogens by enhancing phagocytosis by polymorphonuclear leukocytes, and it is reported to be more effective in reducing inflammatory pain than morphine. RvE2 reduces joint pain in arthritis, and RvD5 has beneficial properties in an experimental model of this disease.
RvD2 and RvD3 derived from DHA have extremely potent regulatory actions on neutrophil trafficking in the picogram range in vivo by stimulating resolution and enhancing innate host defense mechanisms via a specific receptor. Thus in experimental animals, RvD2 interacting with the receptor DRV2 enhances killing and clearance of bacteria and ameliorates the effects of bacterial sepsis. It prolongs survival from severe burns and reduces colitis. In general, RvD3 appears late in resolution suggesting that it has a special role in the last stages of the process. It displays potent tissue protective actions, and for example in uninjured lungs it may regulate tissue tone. RvD4 has novel pro-resolving actions in Staphylococcus aureus infections and is a powerful stimulant for the clearance of apoptotic cells by skin fibroblasts. RvD4 is produced in mouse bone marrow and lung in response to ischemic injury with reductions in the concentrations of pro-inflammatory eicosanoids. It also ameliorates the effects of thrombosis in a mouse model. Similarly, RvD5 controls E. coli and S. aureus infections and lowers the antibiotic requirements for bacterial clearance. Although contradictory results have been obtained in administering resolvins to treat sepsis in some experimental models, this may have been because the appropriate receptors were down regulated.
Protectins appear to operate in the same way as the resolvins in brain tissue. Thus, (N)PD1 has anti-inflammatory effects by promoting resolution of neuroinflammation and nerve regeneration. It protects retinal epithelial cells from apoptosis induced by oxidative stress. In addition, it has protective effects in animal models of stroke and of Alzheimer's disease, and in a mouse model of epilepsy, considerable improvements in the condition were observed after treatment with protectin D1. Amongst its activities in non-neuronal tissues, it promotes apoptosis of T cells, it has beneficial effects towards asthma in nanogram amounts, and it is reported to have therapeutic potential against virus infections by inhibiting viral replication. Protectins and maresins reduce neuropathic pain in experimental animals. PD1 binds to retinal pigment epithelial cells in a highly stereo-selective and specific manner to protect against oxidative stress. Similarly, the elovanoids function to protect these cells and sustain photoreceptor cell integrity in the retina, for example during age-related macular degeneration, together with neural cell integrity in general during such conditions. Protectins synthesised in white adipose tissue have anti-inflammatory effects in obesity and diabetes.
The poxytrin PDX has been found to have distinctive and biological properties that differ from those of PD1. For example, it may have antithrombotic potential as an inhibitor of platelet activation, especially against thromboxane A2-induced aggregation, it may inhibit inflammation associated with cyclooxygenase (COX) activities, and it has been reported to inhibit influenza virus replication by targeting its RNA metabolism.
Maresin 1 is a powerful regulator of resolution of inflammation, tissue regeneration and pain likewise, with effects upon many disease states including lung, vascular and metabolic diseases, and bacterial infections. In general, maresins appear to be especially important for tissue regeneration and wound healing in the later stages of resolution (D-series resolvins are also involved in skin repair after wounding). Mar1 interferes with vanilloid receptor TRPV1 in dorsal root ganglion neurons and blocks capsaicin activity, for example. In planaria, a primordial organism utilized for surgical injury experiments, maresins and RvE1 each markedly improved the rates of tissue regeneration, for example. In addition, the epoxy intermediate in maresin biosynthesis, 13S,14S-epoxy-maresin, affects macrophage activity and function independently by inhibiting the production of inflammatory eicosanoids from arachidonic acid and especially of leukotrienes by direct inactivation of the LTA4 hydrolase. MaR1 is a key mediator of the switch to the macrophage M2 phenotype thereby stimulating macrophage phagocytosis and efferocytosis to enhance the clearance of inflammation without affecting the innate response. It was found to improve the recovery of neurological function after spinal cord injury.
At nanomolar concentrations, sulfido-conjugates derived from epoxy-maresin were shown to resolve E. coli infections, and in general they constitute a novel mechanism of chemical signalling that contains infections, stimulates resolution of inflammation, and promotes the restoration of function in human tissues and those of experimental animals. Similarly, protectin and resolvin sulfido-conjugates stimulated human macrophages and limited the effects of bacterial infections in a dose-dependent manner. For example, RCTR1, RCTR2 and RCTR3 each exert potent anti-inflammatory and proresolving actions with human leukocytes. Similarly, PCRT1 is a potent agonist of monocytes and macrophages and regulates key anti-inflammatory and pro-resolving processes during infection by bacteria. Inflammatory effects of cysteinyl leukotrienes in the lung are countered by cysteinyl maresins.
Although the specialized pro-resolving mediators are produced locally in many different tissues to terminate inflammation, they also reach the circulation and are found in human peripheral blood, suggesting that they may act as anti-inflammatory signals in tissues other than those in which they originate. Proresolving mediators together with lipoxins have been found at bioactive levels in human breast milk and in human and rat placentas, so it is possible that they may have regulatory functions in normal physiological development as well as in pathophysiological processes. It is hope that measurement of the levels of these mediators in infected patients will provide a better understand of their inflammation-resolution status and might predict outcome or whether specific therapeutic treatments are effective.
8. Specialized Pro-Resolving Mediators and Disease
There appears to be little doubt that imbalances between specialized proresolving and proinflammatory mediators leading to impaired resolution are associated with several human diseases. Thus, it is now well established that administration of lipoxins, protectins, resolvins and maresins in vivo and in vitro in animal models can aid the process of recovery from inflammation without compromising host defenses by causing immune suppression, and there is a view that promoting resolution instead of inhibiting inflammation may be a way forward in treating infections. It is evident that SPMs have considerable potential for therapeutic intervention in acute inflammation or chronic inflammatory diseases, and they have been tested in a wide range of experimental models, including peritonitis, periodontitis, colitis, arthritis, psoriasis, dry eye, inflammatory pain, cardiovascular disease (including atherosclerosis), depression, and asthma and other lung ailments. Elovanoids have been reported to counteract the expression of genes for β-amyloid-protein in a model of Alzheimer's disease. These effects are dependent on age, sex and race.
Inflammation in the tumour microenvironment is a defining property of cancers that is exacerbated by carcinogens. By stimulating the resolution of inflammation, SPMs have been important anti-cancer activities in that they suppress tumour growth and enhance cancer therapy by stimulating clearance of tumour debris by macrophages. They also inhibit the eicosanoid/cytokine storm produced by pro-inflammatory mediators. In particular, the aspirin-triggered specialized proresolving mediators (AT-SPMs), such as aspirin-triggered resolvins (AT-RvDs) and lipoxins (AT‑LXs),have pronounced anti-tumour activity, and may provide an explanation for the broad efficacy of aspirin in the treatment of many cancers (in addition to its inhibition of prostaglandin biosynthesis). SPMs may re-programme tumour-associated macrophages to an anti-tumour phenotype. As SPMs are active at much lower concentrations than aspirin, it is hoped that they may be a non-toxic route to harnessing the anticancer activity of aspirin so reducing carcinogen-induced morbidity and mortality.
In relation to cardiovascular disease, it has been reported that the deleterious effects of leukotriene LTB4 resulting from an excessive inflammatory response are countered by the presence of specialized proresolving mediators, especially RvD1, in experimental models of atherosclerosis and suggest a new therapeutic approach to limiting the progression of atherosclerotic plaques and promoting plaque stability. Similarly, RvE1 controls vascular inflammation and protects against atherosclerosis by modifying the uptake of oxidized LDL, while enhancing phagocytosis of macrophages. During blood coagulation, a specific SPM cluster has been detected that consists of resolvin El (RvE1), RvD1, RvD5, lipoxin B-4, and maresin 1 at bioactive concentrations (0.1-1 nM), together with eicosanoids, such as prostaglandins, thromboxanes and leukotrienes. Similarly, studies with animal models suggest that supplementation with RvD2 may decrease the brain damage caused by myocardial infarction and reverse the resultant neurological dysfunction.
In addition, RvD5n-3 DPA and PD1n-3 DPA are protective against intestinal inflammation and intestinal diseases including inflammatory bowel disease; the latter also promotes resolution of neuroinflammation and may be beneficial in epilepsy. RvD1 produced by goblet cells of the eye interacts with its receptors ALX/FPR2 and GPR32 to set off a chain of reactions that result in mucin secretion into the tear film to protect the ocular surface and maintain a healthy interface between the eye and the environment. As examples of the therapeutic potential of SPMs, a phase III clinical trial of a RvE1 analogue against dry eye syndrome is underway, and NPD1/PD1 is in clinical development for neurodegenerative diseases. There is hope that SPMs may prove efficacious in treating the low-level inflammatory conditions that occurs in obesity, including insulin resistance, type 2 diabetes and non-alcoholic fatty liver disease, and they are considered to be candidate drugs for treating chronic inflammation and infection in patients with cystic fibrosis.
In general, specialized proresolving mediators appear to be beneficial towards attack by bacteria, fungi, viruses and parasites, including such debilitating diseases as influenza and malaria. For example, the biological actions of resolvin RvD1 are additive to those of antibiotics, so reducing the doses required to clear both Gram-positive and Gram-negative infections. Similarly, protectin PDX has therapeutic potential in that it suppresses replication of the influenza virus, but by inhibiting the nuclear export of viral mRNA rather than by regulating the resolution of inflammation. Resolvin E1 and lipoxins applied topically were found to have benefits in the treatment of experimental periodontitis. Although SPMs tend to be relatively unstable, they are being used as templates for novel designer drugs or mimetics to treat inflammatory diseases and hopefully will reduce the requirement for antibiotics. For example, the addition of a methyl group to carbon 15 of lipoxin A4 to produce 15(R/S)-methyl-LXA4 prevents metabolism by dehydrogenation, but retains the biological activity of native LXA4 and has shown efficacy and safety in early clinical trials of patients with infantile eczema.
From a nutritional or health standpoint, it has been shown that dietary supplements of the precursor omega-3 fatty acids, taken together with aspirin, stimulate the synthesis of SPMs and may ameliorate the clinical symptoms of many inflammatory disorders by regulating the time course of resolution via the production of resolvins and protectins amongst other effects. Of course, omega-3 fatty acids may have beneficial functions in many normal physiological processes, including ensuring maternal/fetal health.
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|Credits/disclaimer||Updated: September 14th, 2021||Author: William W. Christie|