Sarah K. Orr

The emerging role of group VI calcium-independent phospholipase A2 in releasing docosahexaenoic acid from brain phospholipids

Brain phospholipids are highly enriched in docosahexaenoic acid (DHA; 22:6n-3). Recent advances indicate that 22:6n-3 is released from brain phospholipids via the action of phospholipase A2 (PLA2) in response to several stimuli, including neurotransmission, where it then acts as a secondary messenger. Furthermore, it is now known that released 22:6n-3 is a substrate for several oxygenation enzymes whose products are potent signaling molecules. One emerging candidate PLA2 involved in the release of 22:6n-3 from brain phospholipids is the group VI calcium-independent phospholipase A2 (iPLA2). After a brief review of brain 22:6n-3 metabolism, cell culture and rodent studies facilitating the hypothesis that group VI iPLA2 releases 22:6n-3 from brain phospholipids are discussed. The identification of PLA2s involved in cleaving 22:6n-3 from brain phospholipids could lead to the development of novel therapeutics for brain disorders in which 22:6n-3 signaling is disordered.

Regulation of brain polyunsaturated fatty acid uptake and turnover

The brain is particularly enriched in glycerophospholipids with either arachidonic or docosahexaenoic acid esterified in the stereospecifically numbered-2 position. In this paper, we review how combining a kinetic approach to study the uptake and turnover of arachidonic and docosahexaenoic acids within brain phospholipids of unanesthetized rats, along with chronic administration of antimanic drugs (lithium, valproate and carbamazepine), have advanced our understanding of how polyunsaturated fatty acids (PUFA) enter the brain, and the mechanisms that regulate their turnover within brain phospholipids. The incorporation rates of arachidonic and docosahexaenoic acid from the plasma unesterified pool into brain phospholipids closely approximate independent measures of their consumption rates by the brain, suggesting this is quantitatively the major pool for uptake of these PUFA. Antimanic drugs (lithium and carbamazepine) that downregulate the activity of the calcium-dependent cytosolic phospholipase A(2) (cPLA(2)) transcription factor AP-2, and in turn the expression and activity of cPLA(2,) lead to a selective downregulation in brain arachidonic acid turnover. Furthermore, targeting arachidonoyl-CoA formation via ordered, non-competitive inhibition of an acyl-CoA synthetase with valproate also selectively decreases brain arachidonic acid turnover. Drugs that increase brain cPLA(2) activity (N-methyl-d-aspartic acid and fluoxetine) are correlated with increased turnover of arachidonic acid in brain phospholipids. Altered PUFA metabolism has been implicated in several neurological disorders, including bipolar disorder and Alzheimers disease. Identifying the enzymes that regulated brain PUFA metabolism could lead to new therapeutic approaches for these disorders.

Unesterified docosahexaenoic acid is protective in neuroinflammation

Docosahexaenoic acid (22:6n‐3) is the major brain n‐3 polyunsaturated fatty acid and it is possible that docosahexaenoic acid is anti‐inflammatory in the brain as it is known to be in other tissues. Using a combination of models including the fat‐1 transgenic mouse, chronic dietary n‐3 polyunsaturated fatty acid modulation in transgenic and wild‐type mice, and acute direct brain infusion, we demonstrated that unesterified docosahexaenoic acid attenuates neuroinflammation initiated by intracerebroventricular lipopolysaccharide. Hippocampal neuroinflammation was assessed by gene expression and immunohistochemistry. Furthermore, docosahexaenoic acid protected against lipopolysaccharide‐induced neuronal loss. Acute intracerebroventricular infusion of unesterified docosahexaenoic acid or its 12/15‐lipoxygenase product and precursor to protectins and resolvins, 17S‐hydroperoxy‐docosahexaenoic acid, mimics anti‐neuroinflammatory aspects of chronically increased unesterified docosahexaenoic acid. LC‐MS/MS revealed that neuroprotectin D1 and several other docosahexaenoic acid‐derived specialized pro‐resolving mediators are present in the hippocampus. Acute intracerebroventricular infusion of 17S‐hydroperoxy‐docosahexaenoic acid increases hippocampal neuroprotectin D1 levels concomitant to attenuating neuroinflammation. These results show that unesterified docosahexaenoic acid is protective in a lipopolysaccharide‐initiated mouse model of acute neuroinflammation, at least in part, via its conversion to specialized pro‐resolving mediators; these docosahexaenoic acid stores may provide novel targets for the prevention and treatment(s) of neurological disorders with a neuroinflammatory component.

n-3 Polyunsaturated fatty acids in animal models with neuroinflammation

Neuroinflammation is present in the majority of acute and chronic neurological disorders. Excess or prolonged inflammation in the brain is thought to exacerbate neuronal damage and loss. Identifying modulators of neuroinflammation is an active area of study since it may lead to novel therapies. Omega-3 polyunsaturated fatty acids (n-3 PUFA) are anti-inflammatory in many non-neural tissues; their role in neuroinflammation is less studied. This review summarizes the relationship between n-3 PUFA and brain inflammation in animal models of brain injury and aging. Evidence by and large shows protective effects of n-3 PUFA in models of sickness behavior, stroke, aging, depression, Parkinsons disease, diabetes, and cytokine- and irradiation-induced cognitive impairments. However, rigorous studies that test the direct effects of n-3 PUFA in neuroinflammation in vivo are lacking. Future research in this area is necessary to determine if, and if so which, n-3 PUFA directly target brain inflammatory pathways. n-3 PUFA bioactive metabolites may provide novel therapeutic targets for neurological disorders with a neuroinflammatory component.

Human milk proresolving mediators stimulate resolution of acute inflammation

Human milk contains nutrients and bioactive products relevant to infant development and immunological protection. Here, we investigated the proresolving properties of milk using human milk lipid mediator isolates (HLMIs) and determined their impact on resolution programs in vivo and with human macrophages. HLMIs reduced the maximum neutrophil numbers (14.6±1.2 × 106–11.0±1.0 × 106 cells per exudate) and shortened the resolution interval (Ri; 50% neutrophil reduction) by 54% compared with peritonitis. Using rigorous liquid-chromatography tandem-mass spectrometry (LC-MS-MS)-based lipid mediator (LM) metabololipidomics, we demonstrated that human milk possesses a proresolving LM-specialized proresolving mediator (LM-SPM) signature profile, containing SPMs (e.g. resolvins (Rv), protectins (PDs), maresins (MaRs), and lipoxins (LXs)) at bioactive levels (pico-nanomolar concentrations) that enhanced human macrophage efferocytosis and bacterial containment. SPMs identified in human milk included D-series Rvs (e.g., RvD1, RvD2, RvD3, AT-RvD3, and RvD4), PD1, MaR1, E-series Rvs (e.g. RvE1, RvE2, and RvE3), and LXs (LXA4 and LXB4). Of the SPMs identified in human milk, RvD2 and MaR1 (50 ng per mouse) individually shortened Ri by ∼75%. Milk from mastitis gave higher leukotriene B4 and prostanoids and lower SPM levels. Taken together, these findings provide evidence that human milk has proresolving actions via comprehensive LM-SPM profiling, describing a potentially novel mechanism in maternal–infant biochemical imprinting.

N-3 polyunsaturated fatty acids in animal models with neuroinflammation: An update.

Neuroinflammation is a characteristic of a multitude of neurological and psychiatric disorders. Modulating inflammatory pathways offers a potential therapeutic target in these disorders. Omega-3 polyunsaturated fatty acids have anti-inflammatory and pro-resolving properties in the periphery, however, their effect on neuroinflammation is less studied. This review summarizes 61 animal studies that tested the effect of omega-3 polyunsaturated fatty acids on neuroinflammatory outcomes in vivo in various models including stroke, spinal cord injury, aging, Alzheimers disease, Parkinsons disease, lipopolysaccharide and IL-1β injections, diabetes, neuropathic pain, traumatic brain injury, depression, surgically induced cognitive decline, whole body irradiation, amyotrophic lateral sclerosis, N-methyl-D-aspartate-induced excitotoxicity and lupus. The evidence presented in this review suggests anti-neuroinflammatory properties of omega-3 polyunsaturated fatty acids, however, it is not clear by which mechanism omega-3 polyunsaturated fatty acids exert their effect. Future research should aim to isolate the effect of omega-3 polyunsaturated fatty acids on neuroinflammatory signaling in vivo and elucidate the mechanisms underlying these effects.

The Fat-1 Mouse has Brain Docosahexaenoic Acid Levels Achievable Through Fish Oil Feeding

Fat-1 transgenic mice endogenously convert n-6 to n-3 polyunsaturated fatty acids (PUFA). The aims of this study were to test whether a) fish oil feeding can attain similar brain n-3 PUFA levels as the fat-1 mouse, and b) fat-1 mouse brain docosahexaenoic acid (22:6n-3; DHA) levels can be potentiated by fish oil feeding. Fat-1 mice and their wildtype littermates consumed either a 10% safflower oil (SO) or a 2% fish oil and 8% safflower oil chow (FO). Brain total lipid and phospholipid fraction fatty acids were analyzed using GC-FID. Wildtype mice fed FO chow had similar brain levels of DHA as fat-1 mice fed SO chow. Fat-1 mice fed FO chow had similar brain n-3 PUFA levels as fat-1 mice fed SO chow. In conclusion, brain levels of DHA in the fat-1 mouse can be obtained by and were not further augmented with fish oil feeding.

Resolvin D4 stereoassignment and its novel actions in host protection and bacterial clearance.

Resolvins of the D-series are specialized pro-resolving lipid mediators that regulate cellular response by orchestrating resolution networks involved in host responses to injury and infection. Here, endogenous resolvin D4 was identified in human tissues and found to persist late into the resolution phase of acute murine Staphylococcus aureus infections. Completion of the first total synthesis of resolvin D4 established the absolute stereochemical configuration of RvD4 confirmed by matching with endogenous RvD4 from resolving exudates in dorsal pouch S. aureus infections. In vivo, RvD4 (ng/mouse) reduced neutrophilic infiltration (~40%) and enhanced uptake of apoptotic PMN (51%) by human dermal fibroblasts at concentrations as low as 0.1 nM. These results establish the complete stereochemistry of RvD4 as 4S,5R,17S-trihydroxydocosa-6E,8E,10Z,13Z,15E,19Z-hexaenoic acid and its novel pro-resolving actions in S. aureus infections as well as its potent ability to stimulate clearance of apoptotic cells by skin fibroblasts.

The very low density lipoprotein receptor is not necessary for maintaining brain polyunsaturated fatty acid concentrations

Polyunsaturated fatty acids (PUFA), especially docosahexaenoic and arachidonic acids, as well as cholesterol are important for neural development and maintaining brain function. However, in contrast to cholesterol, the brain is unable to synthesize the required amounts of these PUFA de novo and requires a constant supply from plasma. Suggested pools of uptake include plasma unesterified PUFA or the uptake of PUFA-containing lipoproteins via lipoprotein receptors into endothelial cells of the blood brain barrier. Our study tested whether the very low density lipoprotein receptor (VLDLr) is necessary for maintaining brain PUFA and cholesterol concentrations. Moreover, since VLDLr knockout (VLDLr(-/-)) mice have been reported to have behavioural deficits, this study asked the question whether altered brain PUFA and cholesterol concentrations might be related to these deficits. VLDLr(-/-) and wild-type mice had ad libitum access to chow. At 7 weeks of age the mice were sacrificed, and the cortex, cerebellum, hippocampus, and the remainder of the brain were isolated for total fatty acid and cholesterol analyses. There were no differences in total lipid PUFA or cholesterol concentrations in any of the four brain regions between VLDLr(-/-) and wild-type mice. These findings demonstrate that the VLDLr is not necessary for maintaining brain PUFA concentrations and suggest that other mechanisms to transport PUFA into the brain must exist.

Proresolving actions of a new resolvin D1 analog mimetic qualifies as an immunoresolvent

Resolution of inflammation is an active process driven by several new families of endogenous lipid mediators collectively coined specialized proresolving mediators (SPM). Here, we report a synthetic analog of resolvin D1 (RvD1) and aspirin-triggered RvD1, benzo-diacetylenic-17R-RvD1-methyl ester (BDA-RvD1), which was prepared using fewer steps than required for total organic synthesis of natural SPM. BDA-RvD1 was resistant to further metabolism by human recombinant 15-prostaglandin dehydrogenase, a major inactivation pathway for RvD1. In ischemia-reperfusion-initiated second organ injury, BDA-RvD1 intravenously (1 μg) reduced neutrophil infiltration into the lungs by 58 ± 9% and was significantly more potent than native RvD1. BDA-RvD1 at 100 ng/mouse also shortened the resolution interval, Ri, of Escherichia coli peritonitis with a similar potency as RvD1, by ~57%, from Ri 10.5 h to 4.5 h. With isolated human phagocytes, BDA-RvD1 at picomolar concentrations (10(-12) M) stimulated phagocytosis of zymosan A particles. BDA-RvD1 activated human recombinant G protein-coupled receptor 32/DRV1, an RvD1 receptor, in a dose-dependent manner. These results indicate that, both in vivo in mice and with isolated human cells, BDA-RvD1 shares defining proresolving actions of RvD1, including inhibiting leukocyte infiltration and stimulating phagocytosis. Moreover, they provide evidence for a new analog mimetic and example of an immunoresolvent, namely an agent that stimulates active resolution of inflammation, for a potential new therapeutic class.