Endogenous mechanisms that orchestrate resolution of acute inflammation are essential in host defense and the return to homeostasis. Resolvin (Rv)D2 is a potent immunoresolvent biosynthesized during ...active resolution that stereoselectively stimulates resolution of acute inflammation. Here, using an unbiased G protein-coupled receptor-β-arrestin-based screening and functional sensing systems, we identified a receptor for RvD2, namely GPR18, that is expressed on human leukocytes, including polymorphonuclear neutrophils (PMN), monocytes, and macrophages (MΦ). In human MΦ, RvD2-stimulated intracellular cyclic AMP was dependent on GPR18. RvD2-stimulated phagocytosis of Escherichia coli and apoptotic PMN (efferocytosis) were enhanced with GPR18 overexpression and significantly reduced by shRNA knockdown. Specific binding of RvD2 to recombinant GPR18 was confirmed using a synthetic (3)H-labeled-RvD2. Scatchard analysis gave a Kd of ∼10 nM consistent with RvD2 bioactive concentration range. In both E. coli and Staphylococcus aureus infections, RvD2 limited PMN infiltration, enhanced phagocyte clearance of bacteria, and accelerated resolution. These actions were lost in GPR18-deficient mice. During PMN-mediated second organ injury, RvD2's protective actions were also significantly diminished in GPR18-deficient mice. Together, these results provide evidence for a novel RvD2-GPR18 resolution axis that stimulates human and mouse phagocyte functions to control bacterial infections and promote organ protection.
Acute inflammatory responses are protective, yet without timely resolution can lead to chronic inflammation and organ fibrosis. A systems approach to investigate self-limited (self-resolving) ...inflammatory exudates in mice and structural elucidation uncovered novel resolution phase mediators in vivo that stimulate endogenous resolution mechanisms in inflammation. Resolving inflammatory exudates and human leukocytes utilize DHA and other n−3 EFA to produce three structurally distinct families of potent di- and trihydroxy-containing products, with several stereospecific potent mediators in each family. Given their potent and stereoselective picogram actions, specific members of these new families of mediators from the DHA metabolome were named D-series resolvins (Resolvin D1 to Resolvin D6), protectins (including protectin D1–neuroprotectin D1), and maresins (MaR1 and MaR2). In this review, we focus on a) biosynthesis of protectins and maresins as anti-inflammatory–pro-resolving mediators; b) their complete stereochemical assignments and actions in vivo in disease models. Each pathway involves the biosynthesis of epoxide-containing intermediates produced from hydroperoxy-containing precursors from human leukocytes and within exudates. Also, aspirin triggers an endogenous DHA metabolome that biosynthesizes potent products in inflammatory exudates and human leukocytes, namely aspirin-triggered Neuroprotectin D1/Protectin D1 AT-(NPD1/PD1). Identification and structural elucidation of these new families of bioactive mediators in resolution has opened the possibility of diverse patho-physiologic actions in several processes including infection, inflammatory pain, tissue regeneration, neuroprotection-neurodegenerative disorders, wound healing, and others. This article is part of a Special Issue entitled “Oxygenated metabolism of PUFA: analysis and biological relevance”.
•Proresolving mediators are biosynthesized during resolution phase of acute inflammation.•Resolution is a biosynthetically active process.•Protectins and maresins are two structurally distinct families of potent local mediators.•NPD1/PD1 reduces neural inflammation, stimulates resolution and reduces pain.•MaR1 stimulates tissue regeneration and resolution mechanisms, is anti-inflammatory and reduces pain.
Uncovering mechanisms that control immune responses in the resolution of bacterial infections is critical for the development of new therapeutic strategies that resolve infectious inflammation ...without unwanted side effects. We found that disruption of the vagal system in mice delayed resolution of Escherichia coli infection. Dissection of the right vagus decreased peritoneal group 3 innate lymphoid cell (ILC3) numbers and altered peritoneal macrophage responses. Vagotomy resulted in an inflammatory peritoneal lipid mediator profile characterized by reduced concentrations of pro-resolving mediators, including the protective immunoresolvent PCTR1, along with elevated inflammation-initiating eicosanoids. We found that acetylcholine upregulated the PCTR biosynthetic pathway in ILC3s. Administration of PCTR1 or ILC3s to vagotomized mice restored tissue resolution tone and host responses to E. coli infections. Together these findings elucidate a host protective mechanism mediated by ILC3-derived pro-resolving circuit, including PCTR1, that is controlled by local neuronal output to regulate tissue resolution tone and myeloid cell responses.
•Vagotomy reduces tissue PCTR1 amounts and ILC3 numbers, delaying infection resolution•ILC3 depletion alters peritoneal macrophage phenotype and reduces peritoneal PCTR1•Acetylcholine upregulates the PCTR pathway in mouse and human ILC3s•ILC3 or PCTR1 restores peritoneal macrophage responses and resolution of infections
Resolution of the inflammatory response to infections is important to prevent damage to the host. Dalli et al. find that the vagus nerve promotes bacterial clearance by regulating tissue ILC3 numbers and upregulating the protective mediator PCTR1, which conditions peritoneal macrophage responses and promotes resolution of infection.
Resolution of acute inflammation is an active process locally controlled by a novel genus of specialized pro-resolving mediators (SPM) that orchestrate key resolution responses. Hence, it is of ...general interest to identify individual bioactive mediators and profile their biosynthetic pathways with related isomers as well as their relation(s) to classic eicosanoids in mammalian tissues. Lipid mediator (LM)-SPM levels and signature profiles of their biosynthetic pathways were investigated using liquid chromatography-tandem mass spectrometry (LC-MS-MS)-based LM metabololipidomics. LM and SPM were identified using ≥6 diagnostic ions and chromatographic behavior matching with both authentic and synthetic materials. This approach was validated using the composite reference plasma (SRM1950) of 100 healthy individuals. Using targeted LM metabololipidomics, we profiled LM and SPM pathways in human peripheral blood (plasma and serum) and lymphoid organs. In these, we identified endogenous SPM metabolomes, namely, the potent lipoxins (LX), resolvins (Rv), protectins (PD), and maresins (MaR). These included RvD1, RvD2, RvD3, MaR1, and NPD1/PD1, which were identified in amounts within their bioactive ranges. In plasma and serum, principal component analysis (PCA) identified signature profiles of eicosanoids and SPM clusters. Plasma-SPM increased with omega-3 and acetylsalicylic acid intake that correlated with increased phagocytosis of Escherichia coli in whole blood. These findings demonstrate an approach for identification of SPM pathways (e.g., resolvins, protectins, and maresins) in human blood and lymphoid tissues that were in amounts commensurate with their pro-resolving, organ protective, and tissue regeneration functions. LM metabololipidomics coupled with calibration tissues and physiological changes documented herein provide a tool for functional phenotypic profiling.
The role of inflammation in obesity-related pathologies is well established. We investigated the therapeutic potential of LipoxinA4 (LXA4:5(S),6(R),15(S)-trihydroxy-7E,9E,11Z,13E,-eicosatetraenoic ...acid) and a synthetic 15(R)-Benzo-LXA4-analog as interventions in a 3-month high-fat diet (HFD; 60% fat)-induced obesity model. Obesity caused distinct pathologies, including impaired glucose tolerance, adipose inflammation, fatty liver, and chronic kidney disease (CKD). Lipoxins (LXs) attenuated obesity-induced CKD, reducing glomerular expansion, mesangial matrix, and urinary H2O2. Furthermore, LXA4 reduced liver weight, serum alanine-aminotransferase, and hepatic triglycerides. LXA4 decreased obesity-induced adipose inflammation, attenuating TNF-α and CD11c+ M1-macrophages (MΦs), while restoring CD206+ M2-MΦs and increasing Annexin-A1. LXs did not affect renal or hepatic MΦs, suggesting protection occurred via attenuation of adipose inflammation. LXs restored adipose expression of autophagy markers LC3-II and p62. LX-mediated protection was demonstrable in adiponectin−/− mice, suggesting that the mechanism was adiponectin independent. In conclusion, LXs protect against obesity-induced systemic disease, and these data support a novel therapeutic paradigm for treating obesity and associated pathologies.
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•Lipoxins attenuated high-fat diet-induced liver and kidney disease•LXA4 attenuated adipose inflammation, promoting a macrophage M1-to-M2 switch•LXA4 restored obesity-induced attenuation of autophagy markers LC3-II and p62•LXA4-mediated protection was adiponectin independent, but restored Annexin-A1
Börgeson et al. investigated the role of anti-inflammatory lipid mediators in obesity. LipoxinA4 and a synthetic lipoxin analog protected against obesity-induced kidney and liver disease. Lipoxins mediated protection by decreasing adipose inflammation and promoting a macrophage M1-to-M2 switch. Lipoxin-mediated protection was adiponectin independent, but correlated with restored adipose Annexin-A1 levels.
Chronic unresolved inflammation plays a causal role in the development of advanced atherosclerosis, but the mechanisms that prevent resolution in atherosclerosis remain unclear. Here, we use targeted ...mass spectrometry to identify specialized pro-resolving lipid mediators (SPM) in histologically-defined stable and vulnerable regions of human carotid atherosclerotic plaques. The levels of SPMs, particularly resolvin D1 (RvD1), and the ratio of SPMs to pro-inflammatory leukotriene B
(LTB
), are significantly decreased in the vulnerable regions. SPMs are also decreased in advanced plaques of fat-fed Ldlr
mice. Administration of RvD1 to these mice during plaque progression restores the RvD1:LTB
ratio to that of less advanced lesions and promotes plaque stability, including decreased lesional oxidative stress and necrosis, improved lesional efferocytosis, and thicker fibrous caps. These findings provide molecular support for the concept that defective inflammation resolution contributes to the formation of clinically dangerous plaques and offer a mechanistic rationale for SPM therapy to promote plaque stability.
Resolution of inflammation is now held to be an active process where autacoids promote homeostasis. Using functional-metabololipidomics and in vivo systems, herein we report that endogenous n-3 ...docosapentaenoic (DPA) acid is converted during inflammation-resolution in mice and by human leukocytes to novel n-3 products congenerous to D-series resolvins (Rv), protectins (PD) and maresins (MaR), termed specialized pro-resolving mediators (SPM). The new n-3 DPA structures include 7,8,17-trihydroxy-9,11,13,15E,19Z-docosapentaenoic acid (RvD1(n-3 DPA)), 7,14-dihydroxy-8,10,12,16Z,19Z-docosapentaenoic acid (MaR1(n-3 DPA)) and related bioactive products. Each n-3 DPA-SPM displayed protective actions from second organ injury and reduced systemic inflammation in ischemia-reperfusion. The n-3 DPA-SPM, including RvD1(n-3 DPA) and MaR1(n-3 DPA), each exerted potent leukocyte directed actions in vivo. With human leukocytes each n-3 DPA-SPM reduced neutrophil chemotaxis, adhesion and enhanced macrophage phagocytosis. Together, these findings demonstrate that n-3 DPA is converted to novel immunoresolvents with actions comparable to resolvins and are likely produced in humans when n-3 DPA is elevated.
We aimed to characterize metabolites during tuberculosis (TB) disease and identify new pathophysiologic pathways involved in infection as well as biomarkers of TB onset, progression and resolution. ...Such data may inform development of new anti-tuberculosis drugs. Plasma samples from adults with newly diagnosed pulmonary TB disease and their matched, asymptomatic, sputum culture-negative household contacts were analyzed using liquid chromatography high-resolution mass spectrometry (LC-MS) to identify metabolites. Statistical and bioinformatics methods were used to select accurate mass/charge (m/z) ions that were significantly different between the two groups at a false discovery rate (FDR) of q<0.05. Two-way hierarchical cluster analysis (HCA) was used to identify clusters of ions contributing to separation of cases and controls, and metabolomics databases were used to match these ions to known metabolites. Identity of specific D-series resolvins, glutamate and Mycobacterium tuberculosis (Mtb)-derived trehalose-6-mycolate was confirmed using LC-MS/MS analysis. Over 23,000 metabolites were detected in untargeted metabolomic analysis and 61 metabolites were significantly different between the two groups. HCA revealed 8 metabolite clusters containing metabolites largely upregulated in patients with TB disease, including anti-TB drugs, glutamate, choline derivatives, Mycobacterium tuberculosis-derived cell wall glycolipids (trehalose-6-mycolate and phosphatidylinositol) and pro-resolving lipid mediators of inflammation, known to stimulate resolution, efferocytosis and microbial killing. The resolvins were confirmed to be RvD1, aspirin-triggered RvD1, and RvD2. This study shows that high-resolution metabolomic analysis can differentiate patients with active TB disease from their asymptomatic household contacts. Specific metabolites upregulated in the plasma of patients with active TB disease, including Mtb-derived glycolipids and resolvins, have potential as biomarkers and may reveal pathways involved in TB disease pathogenesis and resolution.
Biomarkers are needed for predicting the effectiveness of disease modifying antirheumatic drugs (DMARDs). Here, using functional lipid mediator profiling and deeply phenotyped patients with early ...rheumatoid arthritis (RA), we observe that peripheral blood specialized pro-resolving mediator (SPM) concentrations are linked with both DMARD responsiveness and disease pathotype. Machine learning analysis demonstrates that baseline plasma concentrations of resolvin D4, 10S, 17S-dihydroxy-docosapentaenoic acid, 15R-Lipoxin (LX)A
and n-3 docosapentaenoic-derived Maresin 1 are predictive of DMARD responsiveness at 6 months. Assessment of circulating SPM concentrations 6-months after treatment initiation establishes that differences between responders and non-responders are maintained, with a decrease in SPM concentrations in patients resistant to DMARD therapy. These findings elucidate the potential utility of plasma SPM concentrations as biomarkers of DMARD responsiveness in RA.
Solid-phase extraction coupled with liquid chromatography tandem mass spectrometry provides a robust and sensitive approach for the identification and quantitation of specialized pro-resolving ...mediators (lipoxins, resolvins, protectins, and maresins), their pathway markers and the classic eicosanoids. Here, we provide a detailed description of the methodologies employed for the extraction of these mediators from biological systems, setup of the instrumentation, sample processing, and then the procedures followed for their identification and quantitation.
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