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Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) can exert antidepressant, anti-inflammatory and neuroprotective properties, but the exact molecular mechanism underlying their effects is still not fully understood. We conducted both in vitro and clinical investigations to test which EPA or DHA metabolites are involved in these anti-inflammatory, neuroprotective and antidepressant effects. In vitro, we used the human hippocampal progenitor cell line HPC0A07/03C, and pre-treated cells with either EPA or DHA, followed by interleukin 1beta (IL1β), IL6 and interferon-alpha (IFN-α). Both EPA and DHA prevented the reduction in neurogenesis and the increase in apoptosis induced by these cytokines; moreover, these effects were mediated by the lipoxygenase (LOX) and cytochrome P450 (CYP450) EPA/DHA metabolites, 5-hydroxyeicosapentaenoic acid (HEPE), 4-hydroxydocosahexaenoic acid (HDHA), 18-HEPE, 20-HDHA, 17(18)-epoxyeicosatetraenoic acid (EpETE) and 19(20)-epoxydocosapentaenoic acid (EpDPA), detected here for the first time in human hippocampal neurones using mass spectrometry lipidomics of the supernatant. In fact, like EPA/DHA, co-treatment with these metabolites prevented cytokines-induced reduction in neurogenesis and apoptosis. Moreover, co-treatment with 17(18)-EpETE and 19(20)-EpDPA and the soluble epoxide hydroxylase (sEH) inhibitor, TPPU (which prevents their conversion into dihydroxyeicosatetraenoic acid (DiHETE)/ dihydroxydocosapentaenoic acid (DiHDPA) metabolites) further enhanced their neurogenic and anti-apoptotic effects. Interestingly, these findings were replicated in a sample of n = 22 patients with a DSM-IV Major Depressive Disorder, randomly assigned to treatment with either EPA (3.0 g/day) or DHA (1.4 g/day) for 12 weeks, with exactly the same LOX and CYP450 lipid metabolites increased in the plasma of these patients following treatment with their precursor, EPA or DHA, and some evidence that higher levels of these metabolites were correlated with less severe depressive symptoms. Overall, our study provides the first evidence for the relevance of LOX- and CYP450-derived EPA/DHA bioactive lipid metabolites as neuroprotective molecular targets for human hippocampal neurogenesis and depression, and highlights the importance of sEH inhibitors as potential therapeutic strategy for patients suffering from depressive symptoms.

Solar ultraviolet radiation (UVR) exposure of human skin has beneficial and harmful effects on health, including impact on immune function, inflammation and reportedly mood, but these are not fully elucidated. Since the endocannabinoid system is implicated in many activities including mood alteration, our objective was to (i) determine and quantify circulating levels of a wide range of endocannabinoid and N-acyl ethanolamine (NAE) species (ii) evaluate whether these are modulated by cutaneous UVR exposures, as attained through repeated low level summer sunlight exposure. Wearing goggles to prevent eye exposure, 16 healthy volunteers (23–59 y; 10 light skin, phototype II, and 6 dark skin, phototype V) received the same UVR exposures (1.3 SED, 95% UVA/5% UVB) thrice weekly for 6 weeks, whilst casually dressed to expose ∼35% skin surface area. Blood samples were taken at baseline, days 1, 3 and 5 of week one, then at weekly intervals, and analysed by LC-MS/MS. Eleven endocannabinoids and NAEs were detected and quantified at baseline, with N-palmitoyl ethanolamine the most abundant (30% of total). Levels did not vary according to phototype (p > 0.05), except for the NAE docosapentaenoyl ethanolamide, which was higher in phototype II than V (p = 0.0002). Level of the endocannabinoid, 2-AG, was elevated during the UVR exposure course (p < 0.05 vs. baseline for all subjects; p < 0.01 for each phototype group), with maximum levels reached by week 2–3, while NAE species did not significantly alter. These findings suggest differential involvement of the cutaneous endocannabinoid system in low dose solar UVR responses in humans.

While resolution is believed to switch inflammation off and return the affected tissues back to homeostasis, we found that it triggers a prolonged phase of immune suppression. During resolution of acute inflammation (∼72h) CD4+ and CD8+ lymphocytes as well as natural killer (NK) cells migrated into tissues secreting IFNγ from day 6. IFNγ carried out two distinct functions: firstly, through IP-10, IFNγ indirectly triggered monocyte infiltration, which differentiated into macrophages (moMϕs). Secondly, moMϕs were directly acted upon by IFNγ to express microsomal prostaglandin E synthase-1 (mPGES-1) alongside cyclooxygenase (COX 1) resulting in sustained prostaglandin (PG)E2 synthesis peaking at day 14 and lasting for several weeks. PGE2 suppressed innate immune responses to bacterial infection opening a prolonged window of postresolution infectious opportunity within the host. At the same time, it also differentially modulated multiple aspect of the adaptive immune system including suppressing lymphocyte function whilst also generating myeloid-derived suppressor cells, the net effect of which was impaired uptake/presentation of exogenous antigens. Therefore, we have defined a novel sequence of post-resolution events starting with IFNγ and culminating in sustained PGE2, which we propose dampens the propensity to develop autoimmune responses to endogenous antigens at the cost of local tissue infection.

Human skin has a distinct profile of fatty acids and related bioactive lipid mediators that regulate many aspects of epidermal and dermal homeostasis, including immune and inflammatory reactions. Sebum lipids act as effective antimicrobial agents, shape immune cell communications and contribute to the epidermal lipidome. The essential fatty acid linoleic acid is crucial for the structure of the epidermal barrier, while polyunsaturated fatty acids act as precursors to eicosanoids, octadecanoids and docosanoids through cyclooxygenase, lipoxygenase and cytochrome P450 monooxygenase-mediated reactions, and endocannabinoids and N-acyl ethanolamines. Cross-communication between these families of bioactive lipids suggests that their cutaneous activities should be considered as part of a wider metabolic network that can be targeted to maintain skin health, control inflammation and improve skin pathologies.

Background: A subset of T-cells expresses the B-cell marker CD20 and in rheumatoid arthritis secretes Interleukin (IL)-17. IL-17 secreting T-cells (Th17) have also been implicated in the inflammatory response in the central nervous system in multiple sclerosis (MS) and may be a potential target for elimination by biologic therapeutics. ScFvRit:sFasL comprises of a rituximab-derived antibody fragment scFvRit genetically fused to human soluble FasL that specifically eliminated T-cells.Objective: To determine the presence and phenotype of CD20+T-cells in blood and brain of MS patients. Second, to determine whether scFvRit:sFasL can selectively eliminate CD20 + T-cells. After CD20-selective binding, scFvRit:sFasL is designed to trigger FasL-mediated activation-induced cell death of T-cells, but not B-cells.Methods: Flow cytometry and immunohistochemistry were used to screen for CD20+inflammatory T-cells in MS blood and brain tissue. ScFvRit:sFasL pro-apoptotic activity was evaluated by Annexin-V/PI staining followed by flow cytometry assessment.Results: Peripheral blood (n=11) and chronic but not active lesions of MS patient brains (n=5) contained CD20+inflammatory T-cells. Activated CD20+T-cells were predominantly CD4+and secreted both IL-17 and INF-gamma. ScFvRit:sFasL triggered CD20-restricted FasL-mediated activation-induced cell death in peripheral blood CD20 +T-cells, but not CD20+B-cells.Conclusion: CD20+ inflammatory T-cells are present in blood and chronic brain lesions of MS patients. ScFvRit:sFasL selectively eliminated CD20+T-cells and may eliminate pathogenic T-cells without B-cell depletion. (C) 2014 Elsevier B.V. All rights reserved.

The skin is the primary barrier from the outside environment, protecting the host from injury, infectious pathogens, water loss and solar ultraviolet radiation. In this role, it is supported by a highly organized system comprising elements of innate and adaptive immunity, responsive to inflammatory stimuli. The cutaneous immune system is regulated by mediators such as cytokines and bioactive lipids that can initiate rapid immune responses with controlled inflammation, followed by efficient resolution. However, when immune responses are inadequate or mounted against non-infectious agents, these mediators contribute to skin pathologies involving unresolved or chronic inflammation. Skin is characterized by active lipid metabolism and fatty acids play crucial roles both in terms of structural integrity and functionality, in particular when transformed to bioactive mediators. Eicosanoids, endocannabinoids and sphingolipids are such key bioactive lipids, intimately involved in skin biology, inflammation and immunity. We discuss their origins, role and influence over various cells of the epidermis, dermis and cutaneous immune system and examine their function in examples of inflammatory skin conditions. We focus on psoriasis, atopic and contact dermatitis, acne vulgaris, wound healing and photodermatology that demonstrate dysregulation of bioactive lipid metabolism and examine ways of using this insight to inform novel therapeutics.

Hyperbaric oxygen (HBO) therapy involves the inhalation of 100% oxygen, whilst inside a chamber at greater than atmospheric pressure. It is an effective treatment for chronic diabetic wounds, although the molecular mechanisms involved remain unclear. We hypothesised that HBO could alter inflammatory gene expression in human endothelial cells via a reactive oxygen/nitrogen species-mediated pathway. Endothelial cells were exposed to a chronic wound model comprising hypoxia (2% O(2) at 1 atmosphere absolute (ATA); PO(2) ~2 kPa) in the presence of lipopolysaccharide and TNF-α for 24h, then treated with HBO for 90 min (97.5% O(2) at 2.4 ATA; PO(2) ~237 kPa). 5h post-HBO, 19 genes involved in adhesion, angiogenesis, inflammation and oxidative stress were downregulated. Notably, only angiogenin gene expression, which promotes both angiogenesis and nitric oxide production (reflected by increased eNOS protein expression in this study), was upregulated. This led to a decrease in endothelial IL-8 mRNA and protein, which could help alleviate inflammatory processes during chronic wound healing. This was no longer evident 22.5h post-HBO, demonstrating the importance of daily exposures in HBO treatment protocols. These studies indicate that elevated oxygen transiently regulates inflammatory gene expression in endothelial cells, which may enhance chronic wound healing.

Oxidation of polyunsaturated fatty acids (PUFA) through enzymatic or non-enzymatic free radical-mediated reactions can yield an array of lipid metabolites including eicosanoids, octadecanoids, docosanoids and related species. In mammals, these oxygenated PUFA mediators play prominent roles in the physiological and pathological regulation of many key biological processes in the cardiovascular, renal, reproductive and other systems including their pivotal contribution to inflammation. Mass spectrometry-based technology platforms have revolutionized our ability to analyze the complex mixture of lipid mediators found in biological samples, with increased numbers of metabolites that can be simultaneously quantified from a single sample in few analytical steps. The recent development of high-sensitivity and high-throughput analytical tools for lipid mediators affords a broader view of these oxygenated PUFA species, and facilitates research into their role in health and disease. In this review, we illustrate current analytical approaches for a high-throughput lipidomic analysis of eicosanoids and related mediators in biological samples. This article is part of a Special Issue entitled "Oxygenated metabolism of PUFA: analysis and biological relevance."

The skin produces bioactive lipids that participate in physiological and pathological states, including homeostasis, induction, propagation, and resolution of inflammation. However, comprehension of the cutaneous lipid complement, and contribution to differing roles of the epidermal and dermal compartments, remains incomplete. We assessed the profiles of eicosanoids, endocannabinoids, N-acyl ethanolamides, and sphingolipids, in human dermis, epidermis, and suction blister fluid. We identified 18 prostanoids, 12 hydroxy-fatty acids, 9 endocannabinoids and N-acyl ethanolamides, and 21 non-hydroxylated ceramides and sphingoid bases, several demonstrating significantly different expression in the tissues assayed. The array of dermal and epidermal fatty acids was reflected in the lipid mediators produced, whereas similarities between lipid profiles in blister fluid and epidermis indicated a primarily epidermal origin of suction blister fluid. Supplementation with omega-3 fatty acids ex vivo showed that their action is mediated through perturbation of existing species and formation of other anti-inflammatory lipids. These findings demonstrate the diversity of lipid mediators involved in maintaining tissue homeostasis in resting skin and hint at their contribution to signaling, cross-support, and functions of different skin compartments. Profiling lipid mediators in biopsies and suction blister fluid can support studies investigating cutaneous inflammatory responses, dietary manipulation, and skin diseases lacking biomarkers and therapeutic targets.

Introduction: Rheumatoid arthritis (RA) is considered a T cell driven autoimmune disease, therefore, the ability of B cell depleting biologics, e.g., anti-CD20 antibodies, to alleviate RA is unclear. This study examined the proportions of IL-17-secreting lymphocytes in the blood of healthy subjects and RA patients and determined if Th17 cells belong to a CD20+ subset of T cells.Methods: Fluorescence-activated cell sorting and confocal microscopy verified CD3, CD4/CD8 and CD20-staining of T cells. IL-17 secretion was determined using a commercial assay.Results: In healthy subjects and RA patients blood, the median percentage of total CD20+ lymphocytes was similar (7.5%; n = 6 and 10.3%; n = 9, respectively) and comprised predominantly of B cells (similar to 86%). However, 2-4% of CD3+ T cells from both healthy subjects (n = 7) and RA (n = 8) individuals co-expressed CD20. The peripheral blood of healthy subjects contained few IL-17-secreting CD20+ T cells (Conclusions: In the blood of RA patients, a greater proportion of Th17 cells are of a CD20+ phenotype compared to healthy individuals. These cells may represent an additional target for anti-CD20 therapies.