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Highlights: [•] Lipid mediators promote the induction and resolution of inflammation. [•] The complexity of lipid metabolism network necessitates systems biology approaches. [•] Advances in mass spectrometry allow for multiplex quantification of lipid species. [•] In vitro and in vivo lipidomic profiling has generated intriguing insights. [•] Comparative analysis of several infection models revealed common immune mechanisms. [Copyright &y& Elsevier]

Abstract: Inflammation is a beneficial host reaction to tissue damage and has the essential primary purpose of restoring tissue homeostasis. Inflammation plays a major role in containing and resolving infection and may also occur under sterile conditions. The cardinal signs of inflammation dolor, calor, tumor and rubor are intrinsically associated with events including vasodilatation, edema and leukocyte trafficking into the site of inflammation. If uncontrolled or unresolved, inflammation itself can lead to further tissue damage and give rise to chronic inflammatory diseases and autoimmunity with eventual loss of organ function. It is now evident that the resolution of inflammation is an active continuous process that occurs during an acute inflammatory episode. Successful resolution requires activation of endogenous programs with switch from production of pro-inflammatory towards pro-resolving molecules, such as specific lipid mediators and annexin A1, and the non-phlogistic elimination of granulocytes by apoptosis with subsequent removal by surrounding macrophages. These processes ensure rapid restoration of tissue homeostasis. Here, we review recent advances in the understanding of resolution of inflammation, highlighting the pharmacological strategies that may interfere with the molecular pathways which control leukocyte survival and clearance. Such strategies have proved beneficial in several pre-clinical models of inflammatory diseases, suggesting that pharmacological modulation of the resolution process may be useful for the treatment of chronic inflammatory diseases in humans. [Copyright &y& Elsevier]

Abstract: α-Bulnesene is a sesquiterpenoid isolated from the water extract of Pogostemon cablin. It showed a potent and concentration-dependent inhibitory effect on platelet-activating factor (PAF) and arachidonic acid (AA) induced rabbit platelet aggregation. In a radioligand binding assay for the PAF receptor, α-bulnesene competitively inhibited [3H]PAF binding to the PAF receptor with an IC50 value of 17.62±5.68μM. α-Bulnesene also dose-dependently inhibited PAF-induced intracellular Ca2+ increase in fluo-3/AM-loaded platelets (IC50 values of 19.62±1.32μM). Furthermore, α-bulnesene inhibited AA-induced thromboxane B2 (TXB2) formation and prostaglandin E2 (PGE2) formation. These results indicate that the inhibitory effect of α-bulnesene on platelet aggregation was due to a dual activity; specifically the chemical blocked PAF-induced intracellular signal transduction and interfered with cyclooxygenase activity, which resulted in a decrease in thromboxane formation. This study is the first to demonstrate that α-bulnesene is a PAF receptor antagonist as well as an anti-platelet aggregation agent. [Copyright &y& Elsevier]

Abstract: P388D1 cells release free arachidonic acid (AA) and prostaglandin E2 (PGE2) upon stimulation with platelet-activating factor (PAF) and zymosan. The response to PAF is dependent on priming of the cells with bacterial lipopolysaccharide (LPS). In the LPS/PAF pathway, both AA and PGE2 release are dependent on transcription and translation, whereas in the zymosan pathway the release of these compounds appears to be largely independent of these processes. Using quantitative real-time PCR, we analyzed the expression of mRNAs that encode proteins potentially responsible for the dependency of the LPS/PAF pathway on gene expression. These include all the phospholipases A2 (PLA2) that we detected in P388D1 cells, cyclooxygenases (COX), COX-1 and COX-2, the membrane-associated prostaglandin E synthase-1 (mPGES-1), the lipocalin-type prostaglandin D2 synthase (PGDS), hematopoietic PGDS and the subunit Gαi2 of heterotrimeric G-proteins. None of the mRNAs encoding PLA2s, PGDSs, or Gαi2 are substantially altered during LPS priming. However, cyclooxygenase-2 is up-regulated during LPS priming and after stimulation of the cells with zymosan. A modest but significant increase of mPGES-1 mRNA was also detected upon stimulation with zymosan. Thus, the dependency of the LPS/PAF-induced PGE2 production on gene expression can be attributed to the production of cyclooxygenase-2. The dependency of AA release on gene expression is not due to altered expression of any of the PLA2s. We suggest that an accessory regulatory protein affecting the release of AA must be responsible. Using HPLC we separated lipids that are secreted upon stimulation with LPS/PAF and zymosan and found that in both pathways PGD2 is the dominant prostaglandin produced and also detected PGE2, PGF2α and AA besides several unidentified compounds. [Copyright &y& Elsevier]

We have recently identified hyperforin, a lipophilic constituent of the herb Hypericum perforatum (St. John’s wort), as a dual inhibitor of the proinflammatory enzymes cyclooxygenase-1 and 5-lipoxygenase. The aim of the present study was to further elucidate antiinflammatory properties and respective targets of hyperforin. We found that hyperforin inhibited the generation of reactive oxygen species (ROS) as well as the release of leukocyte elastase (degranulation) in human isolated polymorphonuclear leukocytes (PMNL), challenged by the G protein-coupled receptor (GPCR) ligand N-formyl-methionyl-leucyl-phenylalanine (fMLP) with an ic50≈0.3 μM. When PMNL were stimulated with phorbol-12-myristate-13-acetate (PMA) or ionomycin, hyperforin (up to 10 μM) failed to inhibit ROS production and elastase release, respectively. Moreover, hyperforin blocked receptor-mediated Ca2+ mobilization (ic50≈0.4 and 4 μM, respectively) in PMNL and monocytic cells, and caused a rapid decline of the intracellular Ca2+ concentration in resting cells. In contrast, the Ca2+ influx induced by ionomycin or thapsigargin was not suppressed. Comparative studies with the specific phospholipase C inhibitor U-73122 and hyperforin revealed similarities between both compounds. Thus, U-73122 and hyperforin blocked fMLP- and PAF-induced Ca2+ mobilization, ROS formation, and elastase release, but failed to suppress these responses when cells were stimulated by PMA or ionomycin. Also, both compounds rapidly decreased basal Ca2+ levels in resting cells and led to a rapid decline of the Ca2+ elevations evoked by fMLP or PAF. Our data suggest that hyperforin targets component(s) within G protein signaling cascades that regulate Ca2+ homeostasis, coupled to proinflammatory leukocyte functions. [Copyright &y& Elsevier]

Dexamethasone-treated WEHI7.2 mouse thymoma cells readily undergo apoptosis. WEHI7.2 variants that overexpress catalase (CAT38) or Bcl-2 (Hb12) show a delay or lack of apoptosis, respectively, when treated with dexamethasone. This is accompanied by a delay or lack of cytochrome c release from the mitochondria suggesting that alterations in the signaling phase of apoptosis are responsible for the observed resistance. Because membranes are a rich source of signaling molecules, we have used 31P NMR spectroscopy to compare phospholipids and their metabolites in WEHI7.2, CAT38 and Hb12 cells after dexamethasone treatment. Increased lysophosphatidylcholine (lysoPtdC) content accompanied phosphatidylserine (PtdS) externalization in the WEHI7.2 cells. Both changes were delayed in CAT38 cells suggesting phosphatidylcholine (PtdC) metabolites may play a role in steroid-induced apoptotic signaling. The steroid-resistant Hb12 cells showed a dramatic increase in glycerophosphocholine (GPC) content, suggesting increased phospholipid turnover may contribute to the anti-apoptotic mechanism of Bcl-2. [Copyright &y& Elsevier]

The present study was designed to assess the effect of supplementation with dietary cytidine (5′)-diphosphocholine (CDP-choline), a source of cytidine and choline, on memory in young and older rats. Although the hippocampal-dependent memory deficits in aged rats are well documented, cognitive functioning in early aging has not been as thoroughly evaluated. Female Sprague–Dawley rats (3 or 15 months of age) consumed either a control diet or a diet supplemented with CDP-choline (approximately 500 mg/kg/day) for 8 weeks, after which they were trained to perform spatial and cued versions of the Morris water maze. Compared with young rats, aged rats exhibited a selective deficit in spatial memory tasks that required rats to retain information for 24 h or longer. CDP-choline supplementation protected against the development of this deficit, but had no memory-enhancing effect in normal young rats. These findings suggest that early-aged rats display a selective impairment in hippocampal-dependent long-term memory, and that dietary CDP-choline supplementation can protect against this deficit. [Copyright &y& Elsevier]

1,2-Diacylglycerols (DAGs) can prime polymorphonuclear leukocytes (PMNL) for enhanced release of arachidonic acid (AA) and generation of 5-lipoxygenase (5-LO) products upon subsequent agonist stimulation. Here, we demonstrate that in isolated human PMNL, 1-oleoyl-2-acetylglycerol (OAG) functions as a direct agonist stimulating 5-LO product formation (up to 42-fold). OAG caused no release of endogenous AA, but in the presence of exogenous AA, the magnitude of 5-LO product synthesis induced by OAG was comparable to that obtained with the Ca2+-ionophore A23187. Interestingly, OAG-induced 5-LO product synthesis was not connected with increased 5-LO nuclear membrane association. Examination of diverse glycerides revealed that the sn-2-acetyl-group is important, thus, also 1-O-hexadecyl-2-acetylglycerol (EAG) stimulated 5-LO product formation (up to 8-fold).Treatment of PMNL with OAG did not alter the mobilization of Ca2+ but removal of intracellular Ca2+ abolished the upregulatory OAG effects. Notably, the PKC activator phorbol-myristate-acetate hardly increased 5-LO product synthesis and PKC inhibitors failed to suppress the effects of OAG. Although OAG rapidly activated p38 MAPK and p42/44MAPK, which can stimulate 5-LO for product synthesis, specific inhibitors of these kinases could not prevent 5-LO activation by OAG. Together, OAG acts as a direct agonist for 5-LO product synthesis in PMNL stimulating 5-LO by novel undefined mechanisms. [Copyright &y& Elsevier]

Phospolipase A2 (PLA2) is the esterase activity that cleaves the sn-2 ester bond in glycerophospholipids, releasing free fatty acids and lysophospholipids. The PLA2 activity is found in a variety of enzymes which can be divided in several types based on their Ca2+ dependence for their activity; Ca2+-dependent secretory phosholipases (sPLA2s) and cytosolic phospholipases (cPLA2s), and Ca2+-independent phospholipase A2s (iPLA2s). These enzymes also show diverse size and substrate specificity (i.e., in the fatty acid chain length and extent of saturation). Among the fatty acids released by PLA2, arachidonic acid (AA) is of particular biological importance, because it is subsequently converted to prostanoids and leukotrienes by cyclooxygenases (COX) and lipoxygenases (LOX), respectively. Free AA may also stimulate apoptosis through activation of sphingomyelinase. Alternatively, it is suggested that oxidized metabolites generated from AA by LOX induce apoptosis. Although the precise mechanisms remain to be elucidated, changes are observed in glycerolipid metabolism during apoptotic processes. In some cells induced to undergo apoptosis, AA is released concomitant with loss of cell viability, caspase activation and DNA fragmentation. Such AA releases appear to be mediated by activation of cPLA2 and/or iPLA2. For example, tumor necrosis factor-α (TNF-α)-induced cell death is mediated by cPLA2, whereas Fas-induced apoptosis appears to be mediated by iPLA2. Some discrepancies among early experimental results were probably caused by differences in the experimental conditions such as the serum concentration, inhibitors used that are not necessarily specific to a single-type enzyme, or differential expression of each PLA2 in cells employed in the experiments. Recent studies eliminated such problems, by carefully defining the experimental conditions, and using multiple inhibitors that show different specificities. Accordingly, more convincing data are available that demonstrate involvement of some PLA2s in the apoptotic processes. In addition to cPLA2 and iPLA2, sPLA2s were recently found to play roles in apoptosis. Moreover, new proteins that appear to control PLA2s are being discovered. Here, the roles of PLA2s in apoptosis are discussed by reviewing recent reports. [Copyright &y& Elsevier]

The mucosal surface of the digestive tract is a critical barrier between a broad spectrum of noxious and immunogenic substances present in the gastrointestinal lumen and the underlying mucosal immune system. Its preservation following various forms of injury or physiological damage is essential to prevent the invasion of harmful luminal factors into the host, which subsequently may lead to inflammation, uncontrolled immune response, and a disequilibrium of the homeostasis of the host. The preservation of this barrier following injuries is regulated by a broad spectrum of structurally distinct regulatory molecules, including phospholipids. Phospholipids play a pivotal role in the modulation of intestinal inflammation. They have been demonstrated to both promote and inhibit inflammation, and their overall impact in an individual setting seems to be dependent on several factors, including the level of immune cell activation and the presence of other mediators. Modulation of lipid mediators through administration of lysophosphatidic acid (LPA) or lisofylline (LSF), inhibitors of phospholipase A2 (PLA2) biosynthesis or monoclonal antibodies against thromboxane (TBX) or platelet-activating factor (PAF) as a therapeutic approach have been used in several models of inflammation; however, beneficial effects were not always convincing and further studies are warranted. [Copyright &y& Elsevier]

Phospholipase A2 (PLA2) activation generates the release of arachidonic acid (AA) and platelet-activating factor (PAF), two compounds which may be involved in neuroplasticity. In previous studies, we found that PLA2 activation is involved in the development of stimulant sensitization. In the present study, we have examined the roles of AA and PAF in the development of stimulant sensitization using agonists and antagonists selective for PAF receptors or the induction of various AA cascade-mediated eicosanoids. Sprague–Dawley rats were treated for 5 days with cocaine (30 mg/kg) or d-amphetamine (1 mg/kg) preceded 15 min earlier by various antagonists, and then tested following a 10-day withdrawal period for cocaine (15 mg/kg) or d-amphetamine (0.5 mg/kg)-induced locomotion. Consistent with our earlier work, pretreatment with the PLA2 inhibitor quinacrine (25 mg/kg) blocked the development of cocaine and amphetamine sensitization. The lipoxygenase (LOX) inhibitors nordihydroguaiaretic acid (NDGA) (5–10 mg/kg) and MK-886 (1 mg/kg) had no effect on cocaine sensitization. The PAF receptor antagonist WEB 2086 (5–10 mg/kg) reduced the development of cocaine sensitization. The cyclooxygenase (COX) inhibitors indomethacin (1–2 mg/kg), piroxicam (0.5–1 mg/kg), 6-methoxy-2-napthylacetic acid (6-MNA; 0.5–1 mg/kg), and NS-398 (0.5–1 mg/kg) blocked the development of cocaine sensitization. The COX inhibitors indomethacin (2 mg/kg) and 6-MNA (1 mg/kg) also reduced the development of amphetamine sensitization. Rats were administered bilateral intraventral tegmental area (VTA) injections of d-amphetamine (5 μg/side) or saline coadministered with indomethacin (0.5 μg/side) or vehicle three times over 5 days and were then tested after a 10-day withdrawal for d-amphetamine (0.5 mg/kg ip)-induced locomotion. Intra-VTA amphetamine induced a robust form of amphetamine sensitization, which was blocked by coadministration of indomethacin. Unilateral intra-VTA injections of PAF (1 μg) did not significantly alter cocaine (15 mg/kg ip)-induced locomotion when tested after a 3-day withdrawal. These findings suggest that COX, and possibly PAF, activity is involved in the development of stimulant sensitization. Neuroanatomical studies demonstrate that this may occur at the level of the VTA. [Copyright &y& Elsevier]