Resolvin D3 Is Dysregulated in Arthritis and Reduces Arthritic Inflammation.

Uncontrolled inflammation is a unifying component of many chronic inflammatory diseases, such as arthritis. Resolvins (Rvs) are a new family from the endogenous specialized proresolving mediators (SPMs) that actively stimulate resolution of inflammation. In this study, using lipid mediator metabololipidomics with murine joints we found a temporal regulation of endogenous SPMs during self-resolving inflammatory arthritis. The SPMs present in self-resolving arthritic joints include the D-series Rvs, for example, RvD1, RvD2, RvD3, and RvD4. Of note, RvD3 levels were reduced in inflamed joints from mice with delayed-resolving arthritis when compared with self-resolving inflammatory arthritis. RvD3 was also reduced in serum from rheumatoid arthritis patients compared with healthy controls. RvD3 administration reduced joint leukocytes as well as paw joint eicosanoids, clinical scores, and edema. Taken together, these findings provide evidence for dysregulated endogenous RvD3 levels in inflamed paw joints and its potent actions in reducing murine arthritis.

Proresolving and cartilage-protective actions of resolvin D1 in inflammatory arthritis.

Rheumatoid arthritis (RA) is a debilitating disease characterized by persistent accumulation of leukocytes within the articular cavity and synovial tissue. Metabololipidomic profiling of arthritic joints from omega-3 supplemented mice identified elevated levels of specialized proresolving lipid mediators (SPM) including resolvin D1 (RvD1). Profiling of human RA synovial fluid revealed physiological levels of RvD1, which - once applied to human neutrophils - attenuated chemotaxis. These results prompted analyses of the antiarthritic properties of RvD1 in a model of murine inflammatory arthritis. The stable epimer 17R-RvD1 (100 ng/day) significantly attenuated arthritis severity, cachexia, hind-paw edema, and paw leukocyte infiltration and shortened the remission interval. Metabololipidomic profiling in arthritic joints revealed 17R-RvD1 significantly reduced PGE2 biosynthesis, while increasing levels of protective SPM. Molecular analyses indicated that 17R-RvD1 enhanced expression of genes associated with cartilage matrix synthesis, and direct intraarticular treatment induced chondroprotection. Joint protective actions of 17R-RvD1 were abolished in RvD1 receptor-deficient mice termed ALX/fpr2/3-/- . These investigations open new therapeutic avenues for inflammatory joint diseases, providing mechanistic substance for the benefits of omega-3 supplementation in RA.

Maresin 1 Biosynthesis and Proresolving Anti-infective Functions with Human-Localized Aggressive Periodontitis Leukocytes.

Localized aggressive periodontitis (LAP) is a distinct form of early-onset periodontitis linked to periodontal infection with uncontrolled inflammation and leukocyte-mediated tissue destruction. The resolution of inflammation is an active process orchestrated by specialized proresolving lipid mediators (SPMs). Since the level of the Maresin pathway marker 14-hydroxy-docosahexaenoic acid (14-HDHA) was lower in activated peripheral blood from LAP patients, we investigated the Maresin 1 (MaR1) biosynthetic pathway in these subjects and its role in regulating phagocyte functions. Macrophages from LAP patients had a lower level of expression of 12-lipoxygenase (∼30%) and reduced MaR1 (LAP versus healthy controls [HC], 87.8 ± 50 pg/10(6) cells versus 239.1 ± 32 pg/10(6) cells). Phagocytosis by LAP macrophages was reduced ∼40% compared to that of HC, and killing of periodontal pathogens, including Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans, were similarly reduced. LAP neutrophils also displayed slower kinetics (∼30%) and decreased maximal phagocytosis (∼20% lower) with these pathogens than those of HC. The administration of MaR1 at 1 nM enhanced phagocytosis (31 to 65% increase), intracellular antimicrobial reactive oxygen species production (26 to 71% increase), bacterial killing of these periodontal pathogens (22 to 38% reduction of bacterial titers), and restored impairment of LAP phagocytes. Together, these results suggest that therapeutics targeting the Maresin pathway have clinical utility in treating LAP and other oral diseases associated with infection, inflammation, and altered phagocyte functions.

Aging delays resolution of acute inflammation in mice: reprogramming the host response with novel nano-proresolving medicines.

Aging is associated with an overt inflammatory phenotype and physiological decline. Specialized proresolving lipid mediators (SPMs) are endogenous autacoids that actively promote resolution of inflammation. In this study, we investigated resolution of acute inflammation in aging and the roles of SPMs. Using a self-resolving peritonitis and resolution indices coupled with lipid mediator metabololipidomics, we found that aged mice had both delayed resolution and reduced SPMs. The SPM precursor docosahexaenoic acid accelerated resolution via increased SPMs and promoted human monocyte reprogramming. In aged mice, novel nano-proresolving medicines carrying aspirin-triggered resolvins D1 and D3 reduced inflammation by promoting efferocytosis. These findings provide evidence for age-dependent resolution pathways in acute inflammation and novel means to activate resolution.

Maresin biosynthesis and identification of maresin 2, a new anti-inflammatory and pro-resolving mediator from human macrophages.

Maresins are a new family of anti-inflammatory and pro-resolving lipid mediators biosynthesized from docosahexaenoic acid (DHA) by macrophages. Here we identified a novel pro-resolving product, 13R,14S-dihydroxy-docosahexaenoic acid (13R,14S-diHDHA), produced by human macrophages. PCR mapping of 12-lipoxygenase (12-LOX) mRNA sequence in human macrophages and platelet showed that they are identical. This human 12-LOX mRNA and enzyme are expressed in monocyte-derived cell lineage, and enzyme expression levels increase with maturation to macrophages or dendritic cells. Recombinant human 12-LOX gave essentially equivalent catalytic efficiency (kcat/KM) with arachidonic acid (AA) and DHA as substrates. Lipid mediator metabololipidomics demonstrated that human macrophages produce a novel bioactive product 13,14-dihydroxy-docosahexaenoic acid in addition to maresin-1, 7R,14S-dihydroxy-4Z,8E,10E,12Z,16Z,19Z-docosahexaenoic acid (MaR1). Co-incubations with human recombinant 12-LOX and soluble epoxide hydrolase (sEH) demonstrated that biosynthesis of 13,14-dihydroxy-docosahexaenoic acid (13,14-diHDHA) involves the 13S,14S-epoxy-maresin intermediate produced from DHA by 12-LOX, followed by conversion via soluble epoxide hydrolase (sEH). This new 13,14-diHDHA displayed potent anti-inflammatory and pro-resolving actions, and at 1 ng reduced neutrophil infiltration in mouse peritonitis by ∼40% and at 10 pM enhanced human macrophage phagocytosis of zymosan by ∼90%. However, MaR1 proved more potent than the 13R,14S-diHDHA at enhancing efferocytosis with human macrophages. Taken together, the present findings demonstrate that macrophages produced a novel bioactive product identified in the maresin metabolome as 13R,14S-dihydroxy-docosahexaenoic acid, from DHA via conversion by human 12-LOX followed by sEH. Given its potent bioactions, we coined 13R,14S-diHDHA maresin 2 (MaR2).

Dietary fish oil increases the proportion of a specific neutrophil subpopulation in blood and total neutrophils in peritoneum of mice following endotoxin-induced inflammation.

Omega-3 polyunsaturated fatty acids may have beneficial effects in inflammation, where neutrophil migration and activation are of importance. The effects of dietary fish oil on neutrophil numbers and subpopulations in healthy mice and mice with endotoxin-induced inflammation were determined. Mice were fed a control diet with or without 2.8% fish oil, and half of them were injected intraperitoneally with endotoxin. Blood, peritoneal lavage, bone marrow and spleen were collected. Expression of cell surface molecules was analyzed by flow cytometry, and chemokine concentrations were determined by enzyme-linked immunosorbent assay. Dietary fish oil did not alter the proportion of total neutrophils in blood but increased the proportion of a specific subpopulation of neutrophils 48 h following endotoxin administration. This subpopulation of neutrophils expressed higher levels of CD11b, Ly6G and major histocompatibility complex-II, suggesting a different role for these neutrophils in the inflammatory response. Dietary fish oil did not affect neutrophil numbers in the peritoneum of healthy mice, but 12 h after endotoxin administration, there were fewer neutrophils in the peritoneum of mice fed the fish oil diet than in mice fed the control diet. However, 48 h after endotoxin administration, mice fed the fish oil diet had more neutrophils in peritoneum than mice fed the control diet. These results indicate that, although dietary fish oil may delay recruitment of neutrophils from blood to the peritoneum early in inflammation, it has the potential to increase the number of peritoneal neutrophils later, which may be of benefit as impaired neutrophil migration and activation have been associated with immunosuppression late in inflammation.

Two circulating neutrophil populations in acute inflammation in mice.

OBJECTIVE AND DESIGN: Recent studies indicate that neutrophils are heterogeneous and may have an immunosuppressive role in addition to their well-known phagocytic and bactericidal function. This study examined neutrophil subpopulations in the circulation, peritoneum, spleen and bone marrow from mice at various time points after induction of acute inflammation. MATERIAL, TREATMENT AND METHODS: Female C57BL/6 mice were injected intraperitoneally with lipopolysaccharide (LPS). Blood, peritoneal, spleen and bone marrow cells were collected and counted and expression of surface molecules and chemokine receptors analyzed with flow cytometry. Chemokine and cytokine concentrations in serum and peritoneal fluid were determined by ELISA. RESULTS: Neutrophil numbers in the circulation decreased following administration of LPS but reached similar numbers to those prior to inflammation at 8 h. At that time point, two distinct neutrophil populations were present in the circulation. These two neutrophil populations differed in size, granularity and expression of CD11b and Ly6G. Few neutrophils were recruited into the peritoneum until 24 h after administration of LPS at a time when the neutrophils in the circulation had increased their expression of the chemokine receptor CXCR2. CONCLUSIONS: Induction of acute inflammation leads to the appearance of two circulating neutrophil subpopulations, which may differ in their activation state and function.

Dietary fish oil decreases the proportion of classical monocytes in blood in healthy mice but increases their proportion upon induction of inflammation.

Fish oil can have beneficial effects in health and disease. In healthy individuals, reduction of the inflammatory status may be of benefit, whereas in patients with systemic inflammation, such as sepsis, it is important to diminish the immunosuppression that is thought to contribute to poor outcome. The objective of this study was to determine the effects of dietary fish oil on monocytes/macrophages in blood, bone marrow, spleen, and peritoneum and chemokine concentrations in blood and peritoneum in healthy mice and mice with endotoxin-induced inflammation. Mice were fed a Western-type diet without fish oil (C) or with 2.8% fish oil (FO) for 6 wk and then either killed (healthy mice) or injected i.p. with endotoxin (LPS) and killed after 3, 8, 12, 24, or 48 h. Blood, bone marrow, spleen, and peritoneal lavage were collected. Expression of cell surface molecules and chemokine receptors was analyzed by flow cytometry and chemokine concentrations measured by ELISA. Healthy mice in the FO group had lower proportions of classical monocytes in blood than healthy mice in the C group. LPS administration increased the proportion of classical monocytes in blood in mice in the FO group but not in those in the C group. Healthy mice in the FO group had lower serum concentrations of CCL2 than mice in the C group, but in inflamed mice, CCL2 concentrations were higher in the FO group than in the C group. These results indicate that dietary fish oil can attenuate the inflammatory status in homeostasis but intensify the immune response upon inflammation.