Sn-2 Position Of Phospholipids Research Articles

Liver X receptor agonist upregulates LPCAT3 in human aortic endothelial cells.

Endothelial cells (ECs) play an important role in tissue homeostasis. Recently, EC lipid metabolism has emerged as a regulator of EC function. The liver X receptors (LXRs) are involved in the transcriptional regulation of genes involved in lipid metabolism and have been identified as a potential target in cardiovascular disease. We aimed to decipher the role of LXRs in the regulation of lipid metabolism in human aortic endothelial cells. Lipid composition analysis of endothelial cells treated with the LXR agonist T0901317 revealed that LXR activation increased the proportion of polyunsaturated fatty acids (PUFAs) and decreased the proportion of saturated fatty acids. The LXR agonist decreased the uptake of fatty acids (FAs) by ECs. This effect was abolished by LXRα silencing. LXR activation increased the activity and the expression of lysophosphatidylcholine acyltransferase, LPCAT3, which is involved in the turnover of FAs at the sn-2 position of phospholipids. Transcriptomic analysis also revealed that LXRs increased the expression of key genes involved in the synthesis of PUFAs, including FA desaturase one and 2, FA elongase 5 and fatty acid synthase. Subsequently, the LXR agonist increased PUFA synthesis and enhanced arachidonic acid, eicosapentaenoic acid, and docosahexaenoic acid content in the EC phospholipids. Modification of the FA composition of ECs by LXRs led to a decrease of arachidonate and linoleate derived prostaglandins synthesis and release. No change on markers of inflammation induced by plasma from sickle cell patient were observed in presence of LXR agonist. These results identify LXR as a key regulator of lipid metabolism in human aortic endothelial cells and a direct effect of LXR agonist on lysophosphatidylacyl transferase (LPCAT3).

Division of the role and physiological impact of multiple lysophosphatidic acid acyltransferase paralogs

BackgroundLysophosphatidic acid acyltransferase (LPAAT) is a phospholipid biosynthesis enzyme that introduces a particular set of fatty acids at the sn-2 position of phospholipids. Many bacteria have multiple LPAAT paralogs, and these enzymes are considered to have different fatty acid selectivities and to produce diverse phospholipids with distinct fatty acid compositions. This feature is advantageous for controlling the physicochemical properties of lipid membranes to maintain membrane integrity in response to the environment. However, it remains unclear how LPAAT paralogs are functionally differentiated and biologically significant.ResultsTo better understand the division of roles of the LPAAT paralogs, we analyzed the functions of two LPAAT paralogs, PlsC4 and PlsC5, from the psychrotrophic bacterium Shewanella livingstonensis Ac10. As for their enzymatic function, lipid analysis of plsC4- and plsC5-inactivated mutants revealed that PlsC4 prefers iso-tridecanoic acid (C12-chain length, methyl-branched), whereas PlsC5 prefers palmitoleic acid (C16-chain length, monounsaturated). Regarding the physiological role, we found that plsC4, not plsC5, contributes to tolerance to cold stress. Using bioinformatics analysis, we demonstrated that orthologs of PlsC4/PlsC5 and their close relatives, constituting a new clade of LPAATs, are present in many γ-proteobacteria. We also found that LPAATs of this clade are phylogenetically distant from principal LPAATs, such as PlsC1 of S. livingstonensis Ac10, which are universally conserved among bacteria, suggesting the presence of functionally differentiated LPAATs in these bacteria.ConclusionsPlsC4 and PlsC5, which are LPAAT paralogs of S. livingstonensis Ac10, play different roles in phospholipid production and bacterial physiology. An enzyme belonging to PlsC4/PlsC5 subfamilies and their close relatives are present, in addition to principal LPAATs, in many γ-proteobacteria, suggesting that the division of roles is more common than previously thought. Thus, both principal LPAATs and PlsC4/PlsC5-related enzymes should be considered to decipher the metabolism and physiology of bacterial cell membranes.

The expanding role of lyso-phosphatidylcholine acyltransferase-3 (LPCAT3), a phospholipid remodeling enzyme, in health and disease.

The turnover of fatty acids (FAs) at the sn-2 position of phospholipids is mediated by the reciprocal actions of phospholipases A2 and lyso-PL acyltransferases (LPLAT). LPCAT3, a major LPLAT isoform, exhibits a strong specificity for polyunsaturated FAs s (PUFAs). Although the enzyme was originally studied in the context of cardiometabolism, recent investigations have shed light on the role of LPCAT3 in other tissues such as skeletal muscle and in unexpected biological processes such as cell death and oncogenesis. The three-dimensional structure of LPCAT3 has been elucidated allowing further understanding of the mechanism of the acylation reaction as well as the substrate specificity of the enzyme. In skeletal muscle, LPCAT3-mediated phospholipid remodeling modulates membrane domain clustering and insulin signalingLPCAT3 plays an important role in the process of ferroptosis by modulating the PUFA content of phospholipids and possibly of plasmalogens.In tumor-associated macrophages, LPCAT3 can prevent ER stress induced by the tumor microenvironment and may equally modulate antitumor immunity. LPCAT3 is an attractive therapeutic target in the cardiometabolic disorders. Nevertheless, the involvement of LPCAT3 in processes such as cell death and oncogenesis demands caution with respect to the potential deleterious effects of enzyme modulation.

Emerging Role of Phospholipids and Lysophospholipids for Improving Brain Docosahexaenoic Acid as Potential Preventive and Therapeutic Strategies for Neurological Diseases.

Docosahexaenoic acid (DHA, 22:6n-3) is an omega-3 polyunsaturated fatty acid (PUFA) essential for neural development, learning, and vision. Although DHA can be provided to humans through nutrition and synthesized in vivo from its precursor alpha-linolenic acid (ALA, 18:3n-3), deficiencies in cerebral DHA level were associated with neurodegenerative diseases including Parkinson’s and Alzheimer’s diseases. The aim of this review was to develop a complete understanding of previous and current approaches and suggest future approaches to target the brain with DHA in different lipids’ forms for potential prevention and treatment of neurodegenerative diseases. Since glycerophospholipids (GPs) play a crucial role in DHA transport to the brain, we explored their biosynthesis and remodeling pathways with a focus on cerebral PUFA remodeling. Following this, we discussed the brain content and biological properties of phospholipids (PLs) and Lyso-PLs with omega-3 PUFA focusing on DHA’s beneficial effects in healthy conditions and brain disorders. We emphasized the cerebral accretion of DHA when esterified at sn-2 position of PLs and Lyso-PLs. Finally, we highlighted the importance of DHA-rich Lyso-PLs’ development for pharmaceutical applications since most commercially available DHA formulations are in the form of PLs or triglycerides, which are not the preferred transporter of DHA to the brain.

Lipid Droplets, Phospholipase A2, Arachidonic Acid, and Atherosclerosis.

Lipid droplets, classically regarded as static storage organelles, are currently considered as dynamic structures involved in key processes of lipid metabolism, cellular homeostasis and signaling. Studies on the inflammatory state of atherosclerotic plaques suggest that circulating monocytes interact with products released by endothelial cells and may acquire a foamy phenotype before crossing the endothelial barrier and differentiating into macrophages. One such compound released in significant amounts into the bloodstream is arachidonic acid, the common precursor of eicosanoids, and a potent inducer of neutral lipid synthesis and lipid droplet formation in circulating monocytes. Members of the family of phospholipase A2, which hydrolyze the fatty acid present at the sn-2 position of phospholipids, have recently emerged as key controllers of lipid droplet homeostasis, regulating their formation and the availability of fatty acids for lipid mediator production. In this paper we discuss recent findings related to lipid droplet dynamics in immune cells and the ways these organelles are involved in regulating arachidonic acid availability and metabolism in the context of atherosclerosis.

DHA-enriched phosphatidylcholine suppressed angiogenesis by activating PPARγ and modulating the VEGFR2/Ras/ERK pathway in human umbilical vein endothelial cells.

Docosahexaenoic acid-enriched phosphatidylcholine (DHA-PC) is a new generation of omega-3 lipids, which contains an ester bond linking DHA at the sn-2 position of phospholipid. DHA-PC has become the interest recently as its better bioavailability and anti-oxidation capacity. In this study, the anti-angiogenic effect of DHA-PC was evaluated. The capacities of proliferation, migration, tube formation of human umbilical vein endothelial cells were significantly declined after DHA-PC treatment. Furthermore, DHA-PC inhibited the neovascularization of the chick chorioallantoic membrane in vivo. Mechanism results indicated that DHA-PC enhances the expression of peroxisome proliferator-activated receptor γ (PPARγ) at transcriptional and translational level, subsequently down-regulates the VEGFR2 expression and VEGFR2-mediated downstream Ras/ERK pathway, resulting in significant reduction in proliferation and differentiation. Additionally, PPARγ-specific antagonist GW9662 partly reversed the inhibition effects of DHA-PC on tube formation and neovascularization, suggesting that DHA-PC exerts anti-angiogenesis effect through activating PPARγ. These findings indicated that DHA-PC has a great prospect of anti-tumor angiogenesis therapy.

Lysophosphatidic acid acyltransferase from the thermophilic bacterium Thermus thermophilus HB8 displays substrate promiscuity.

ACP: acyl carrier protein; CAPS: N-cyclohexyl-3-aminopropanesulfonic acid; CoA: coenzyme A; CYMAL-6: 6-cyclohexyl-1-hexyl-β-D-maltoside; DDM: n-dodecyl-β-D-maltoside; DTNB: 5,5´-dithiobis(2-nitrobenzoic acid); EPA: eicosapentaenoic acid; G3P: glycerol 3-phosphate; HEPES: N-2-hydroxyethylpiperazine-N´-2-ethanesulfonic acid; LPA: lysophosphatidic acid; MS: mass spectrometry; PA: phosphatidic acid.

Disrupting a phospholipase A2 gene increasing lipid accumulation in the oleaginous yeast Yarrowia lipolytica.

Phospholipase A2 (PLA2 ) is a diverse superfamily that hydrolyzes fatty acyl ester bonds at the sn-2 position of phospholipids. The correlation between phospholipid metabolism and the anabolism of neutral lipids remains unclear in yeasts. This study aims to explore the effects of PLA2 on lipid accumulation in the oleaginous yeast Yarrowia lipolytica. This study identified an actively expressed phospholipase A2 gene (PLA2-3, YAIL0_E16060g) in Y. lipolytica by quantitative PCR analysis. The gene PLA2-3 was disrupted in the strain po1gΔKu70 by homologous recombination and in the strain po1g-G3 by a CRISPR-Cas9 system, which caused an increase in stress sensitivity while the cell growth was not altered under fermentative conditions. Lipid production was performed in both flasks and bioreactors. The results showed that the lipid titre and lipid content were improved over 25% and 8-30%, respectively, in PLA2-3 disrupted strains compared to the controls. Disruption of the phospholipase PLA2-3 gene could effectively improve lipid production in Y. lipolytica. This study presented a strategy on improving the lipid production of oleaginous yeasts and a similar strategy might be used in other oleaginous microbes.

A Novel Lysophosphatidic Acid Acyltransferase of Escherichia coli Produces Membrane Phospholipids with a cis-vaccenoyl Group and Is Related to Flagellar Formation.

Lysophosphatidic acid acyltransferase (LPAAT) introduces fatty acyl groups into the sn-2 position of membrane phospholipids (PLs). Various bacteria produce multiple LPAATs, whereas it is believed that Escherichia coli produces only one essential LPAAT homolog, PlsC—the deletion of which is lethal. However, we found that E. coli possesses another LPAAT homolog named YihG. Here, we show that overexpression of YihG in E. coli carrying a temperature-sensitive mutation in plsC allowed its growth at non-permissive temperatures. Analysis of the fatty acyl composition of PLs from the yihG-deletion mutant (∆yihG) revealed that endogenous YihG introduces the cis-vaccenoyl group into the sn-2 position of PLs. Loss of YihG did not affect cell growth or morphology, but ∆yihG cells swam well in liquid medium in contrast to wild-type cells. Immunoblot analysis showed that FliC was highly expressed in ∆yihG cells, and this phenotype was suppressed by expression of recombinant YihG in ∆yihG cells. Transmission electron microscopy confirmed that the flagellar structure was observed only in ∆yihG cells. These results suggest that YihG has specific functions related to flagellar formation through modulation of the fatty acyl composition of membrane PLs.

Inhibition of Cytosolic Phospholipase A2α Impairs an Early Step of Coronavirus Replication in Cell Culture.

Coronavirus replication is associated with intracellular membrane rearrangements in infected cells, resulting in the formation of double-membrane vesicles (DMVs) and other membranous structures that are referred to as replicative organelles (ROs). The latter provide a structural scaffold for viral replication/transcription complexes (RTCs) and help to sequester RTC components from recognition by cellular factors involved in antiviral host responses. There is increasing evidence that plus-strand RNA (+RNA) virus replication, including RO formation and virion morphogenesis, affects cellular lipid metabolism and critically depends on enzymes involved in lipid synthesis and processing. Here, we investigated the role of cytosolic phospholipase A2α (cPLA2α) in coronavirus replication using a low-molecular-weight nonpeptidic inhibitor, pyrrolidine-2 (Py-2). The inhibition of cPLA2α activity, which produces lysophospholipids (LPLs) by cleaving at the sn-2 position of phospholipids, had profound effects on viral RNA and protein accumulation in human coronavirus 229E-infected Huh-7 cells. Transmission electron microscopy revealed that DMV formation in infected cells was significantly reduced in the presence of the inhibitor. Furthermore, we found that (i) viral RTCs colocalized with LPL-containing membranes, (ii) cellular LPL concentrations were increased in coronavirus-infected cells, and (iii) this increase was diminished in the presence of the cPLA2α inhibitor Py-2. Py-2 also displayed antiviral activities against other viruses representing the Coronaviridae and Togaviridae families, while members of the Picornaviridae were not affected. Taken together, the study provides evidence that cPLA2α activity is critically involved in the replication of various +RNA virus families and may thus represent a candidate target for broad-spectrum antiviral drug development.IMPORTANCE Examples of highly conserved RNA virus proteins that qualify as drug targets for broad-spectrum antivirals remain scarce, resulting in increased efforts to identify and specifically inhibit cellular functions that are essential for the replication of RNA viruses belonging to different genera and families. The present study supports and extends previous conclusions that enzymes involved in cellular lipid metabolism may be tractable targets for broad-spectrum antivirals. We obtained evidence to show that a cellular phospholipase, cPLA2α, which releases fatty acid from the sn-2 position of membrane-associated glycerophospholipids, is critically involved in coronavirus replication, most likely by producing lysophospholipids that are required to form the specialized membrane compartments in which viral RNA synthesis takes place. The importance of this enzyme in coronavirus replication and DMV formation is supported by several lines of evidence, including confocal and electron microscopy, viral replication, and lipidomics studies of coronavirus-infected cells treated with a highly specific cPLA2α inhibitor.

A novel calcium-independent phospholipase A2 and its physiological roles in development and immunity of a lepidopteran insect, Spodoptera exigua

Phospholipase A2 (PLA2) catalyzes hydrolysis of ester linkage at sn-2 position of phospholipids. At least 15 groups (I-XV) of PLA2 gene superfamily are associated with various physiological processes such as digestion, secretion, immunity, and maintenance of membrane integrity. This study suggests that various insects encode putative Group VI PLA2s representing intracellular and calcium-independent PLA2s (iPLA2). These insect iPLA2s are separated into at least two subgroups: iPLA2A (Group VIA-like) and iPLA2B (non-Group VIA). Most insects encode genes of iPLA2B type, although their biological functions are currently unknown. This study predicted a novel iPLA2 from Spodoptera exigua (a lepidopteran insect) (SeiPLA2B) and analyzed its physiological functions by RNA interference (RNAi). SeiPLA2B encodes 336 amino acid sequence with a predicted size of about 36.6 kDa and an isoelectric point at pH 8.61. It possesses a lipase catalytic site, but does not have ankyrin repeats in the amino terminal region. Phylogenetic analysis indicated that SeiPLA2B was clustered with other Group VI iPLA2s, in which SeiPLA2B was closely associated with Group VIF gene while SeiPLA2A was closely related to Group VIA gene. SeiPLA2B was expressed in all developmental stages of S. exigua. In larval stage, SeiPLA2B was expressed in fat body, hemocyte, and epidermis, but not in digestive tract. SeiPLA2B RNAi significantly reduced PLA2 enzyme activities and resulted in developmental retardation and immunosuppression. Though RNAi treatment did not significantly change fatty acid composition in fat body lipids, it significantly increased lipid peroxidation. Taken together, our results suggest that SeiPLA2B plays important roles in the development and immunity of S. exigua.

Comparison of the Distribution of Unsaturated Fatty Acids at the <i>Sn</i>-2 Position of Phospholipids and Triacylglycerols in Marine Fishes and Mammals

Highly unsaturated fatty acid (HUFA) binding at the sn-2 position of phospholipids (PL) becomes a resource for prostaglandin, leukotriene, resolvin, and protectin synthesis. Both triacylglycerol (TAG) and PL synthesis pathways in vivo are via phosphatidic acid; therefore, the distribution of fatty acid species at the sn-2 position must theoretically be the same for TAG and PL if rearrangement does not occur. However, it is known that little HUFA is located at the sn-2 position of TAG in marine mammals. Therefore, distribution of fatty acid species at the sn-2 position of TAG and PL was compared between marine fishes and mammals in this study. The composition of fatty acids binding at the sn-2 or sn-1,3 position of PL and TAG was analyzed via hydrolysis with enzymes and GC-FID. The results showed that 20:4n-6, 20:5n-3, 22:5n-3, and 22:6n-3 were primarily located at the sn-1,3 positions of TAG in marine mammals. Comparison of the binding positions of HUFA and 16:0 in PL and TAG suggested the existence of Lands' cycle in marine fishes and mammals. In conclusion, both marine fishes and mammals condensed HUFA as a source of eicosanoid at the sn-2 position of PL. Furthermore, abundance ratios for 22:5n-3 or 22:6n-3 at the sn-2 position (sn-2 ratio) in TAG and PL (calculated by the equation: [abundance ratio at sn-2 position of TAG]/[abundance ratio at sn-2 position of PL]) was less than 0.35 in marine mammals; however, it was greater than 0.80 in marine fishes. These differences suggested that the HUFA consisted of 22 carbon atoms and had different roles in marine fishes and mammals.

DHA結合ホスファチジルコリン輸送に関する分子生物学的一考察

DHA levels are not homogeneously distributed throughout an animal. There are two distinct categories, select tissues containing large amounts of DHA and the remaining tissues with normally less than a few mol%. High-DHA tissues, the rod outer segment, sperm, and brain have a large quantity of DHA. DHA is mainly found on in the sn-2 position of phospholipids. DHA is n-3 fatty acid that is essential for normal brain development and cognitive function. It was confirmed a member of the major facilitator superfamily: Mfsd2a as the major transporter for DHA uptake into brain. Mfsd2a is found to be expressed exclusively in endothelium of the blood-brain barrier of micro-vessels. Mfsd2a-KO mice show markedly reduced levels of DHA in brain accompanied by neuronal cell loss in hippocampus and cerebellum. Mfsd2a transports DHA in the form of sn-2 lysophosphatidylcholine, but not unesterified fatty acid, in a sodium ion-system. A remarkable property of MFSD2 is its high-level expression specifically in the placenta, with expression in all other tissues at least 10-fold low―with the expression of the testis in which it is only 4-fold lower than the placenta. DHA is specifically bound to fatty acid binding proteins (Fabp7) in the cytoplasm, and transported to the cell membrane and work as ligands for nuclear receptor transcription factor.

MALDI-TOF MS to monitor the kinetics of phospholipase A2-digestion of oxidized phospholipids

Free fatty acids (FFA) are released through phospholipase A2 (PLA2), which cleaves the fatty acyl residue at the sn-2 position of phospholipids (PL). During inflammatory diseases, reactive oxygen species (such as HOCl) lead to the formation of oxidatively modified PL (e.g., chlorohydrin generation). It is still widely unknown to which extent the oxidation of PL influences their digestibility by PLA2. Additionally, investigations on the impact of the position of the unsaturated fatty acyl residue (sn-1 versus sn-2 position) and modifications of the headgroup (for instance phosphatidylcholine (PC) versus phosphatidylethanolamine (PE)) are also lacking. Therefore, the aim of this study is the investigation of these aspects using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry to elucidate the PL/lysophospholipid (LPL) ratios as measures of the PLA2 digestibility.We will show that oxidative modifications of PL by HOCl have a considerable impact on the PLA2 digestibility, i.e., oxidation of the unsaturated fatty acyl residues leads to a reduced digestibility of both PC and PE. Besides, it will be shown that MALDI MS is a convenient and reliable tool to investigate the related changes.

Evaluation of the recombinant turkey pancreatic lipase phospholipase activity: A monolayer study

Classical lipases are well known for being enzymes hydrolysing triacylglycérols as substrate, except the porcine pancreatic lipase (PPL) which was able to hydrolyze phosphatidylcholine. Amino acid sequence alignments revealed that Valine 260 residue in PPL lid, postulated to be responsible for the PPL phospholipase activity, was present in the Turkey pancreatic lipase (TPL). The importance of Val 260 in the phospholipase activities expression has been reported. To confirm this fact, Val 260 was mutated to Alanine in the TPL lid. Mutated protein has conserved its phospholipase activity as well as the non mutated TPL. Therefore, Valine 260 residue in the lid is not involved in the pancreatic lipases phospholipase activity. The rTPL phospholipase activity was also studied using monolayer technique. This avian pancreatic lipase has shown phospholipase activity toward differently charged phospholipids. The highest phospholipase activity was found on phosphatidylglycerol (negatively charged substrate) at a surface pressure of 20mN/m, but when a zwitterionic substrate was used (DLPC), a lower activity was found at a surface pressure of 10mN/m. However, it is worth noticing that the TPL phospholipase activity is about 100 fold lower than its lipase activity. GC chromatography analyses of the released fatty acids from the hydrolysis of 1,2-POPC have shown that rTPL hydrolyses esters bonds at the sn-1 as well as the sn-2 position of phospholipids. Hence, rTPL shows a low phospholipase activity in comparison to its activity toward triacylglycerols.

Positional analysis of phosphatidylcholine and phosphatidylethanolamine via LC with a charged aerosol detector

A new method for the positional analysis of egg yolk phospholipids (PLs) (phosphatidylcholine-PC, phosphatidylethanolamine-PE) using liquid chromatography with charge aerosol detector (CAD) is described. The method is based on six-step procedure: 1) extraction of phospholipids from tissue sample, 2) separation of lipid classes by solid phase extraction (SPE), 3) complete regiospecific hydrolysis of phospholipids by phospholipase A2 (PLA2), 4) separation of reaction products (fatty acids from sn-2 position and 1-acyl lysophospholipids) by SPE, 5) chemical hydrolysis of 1-acyl lysophospholipids, and 6) analysis of obtained fatty acids by LC with charge aerosol detection (CAD).Total time of enzymatic hydrolysis of PLs ranged from 10–30min. The reaction products were separated by SPE in three-step gradient elution procedure. Chloroform: methanol mixtures were used as eluents to obtain pure fractions of FAs from sn-2 position of PL and 1-acyl lysoPL (chemically hydrolyzed to FAs).FAs were separated by reversed-phase LC using a gradient elution and detected using CAD detector. This combination enables determination of all fatty acids in a single analysis, and without the sample derivatization. The method was optimized and the response of CAD, linearity, precision and sensitivity of the method were studied.

IPLA2β knockout mouse, a genetic model for progressive human motor disorders, develops age-related neuropathology.

Calcium-independent phospholipase A2 group VIa (iPLA2β) preferentially releases docosahexaenoic acid (DHA) from the sn-2 position of phospholipids. Mutations of its gene, PLA2G6, are found in patients with several progressive motor disorders, including Parkinson disease. At 4months, PLA2G6 knockout mice (iPLA2β(-/-)) show minimal neuropathology but altered brain DHA metabolism. By 1year, they develop motor disturbances, cerebellar neuronal loss, and striatal α-synucleinaccumulation. We hypothesized that older iPLA2β(-/-) mice also would exhibit inflammatory and other neuropathological changes. Real-time polymerase chain reaction and Western blotting were performed on whole brain homogenate from 15 to 20-month old male iPLA2β(-/-) or wild-type (WT) mice. These older iPLA2β(-/-) mice compared with WT showed molecular evidence of microglial (CD-11b, iNOS) and astrocytic (glial fibrillary acidic protein) activation, disturbed expression of enzymes involved in arachidonic acid metabolism, loss of neuroprotective brain derived neurotrophic factor, and accumulation of cytokine TNF-α messenger ribonucleic acid, consistent with neuroinflammatory pathology. There was no evidence of synaptic loss, of reduced expression of dopamine active reuptake transporter, or of accumulation of the Parkinson disease markers Parkin or Pink1. iPLA2γ expression was unchanged. iPLA2β deficient mice show evidence ofneuroinflammation and associated neuropathology with motor dysfunction in later life. These pathological biomarkers could be used to assess efficacy of dietary intervention, antioxidants or other therapies on disease progression in this mouse model of progressive human motor diseases associated with a PLA2G6 mutation.

Fatty acids, inflammation, and asthma

Fatty acids and consequently diet play an essential role in the formation of inflammatory mediators involved in the pathogenesis of asthma. Because intake variations of omega-6 (n-6) and omega-3 (n-3) fatty acids ultimately determine cell membrane incorporation, changes in diet have the potential to modify downstream production of inflammatory mediators derived from these compounds. It has long been hypothesized that decreasing the n-6/n-3 ratio could reduce the production of more proinflammatory mediators while increasing the formation of downstream metabolites that can serve to limit or resolve inflammation. In turn, these changes would result in improved asthma outcomes or would lower the risk for asthma incidence. This review will focus on the role of fatty acid inflammatory and resolving mediators and will summarize the clinical and epidemiologic data on how diet and obesity alter fatty acid profiles that can contribute to asthma.

Nanoassemblies containing a fluorouracil/zidovudine glyceryl prodrug with phospholipase A2-triggered drug release for cancer treatment

Secretory phospholipase A2 (sPLA2), which is overexpressed in many tumors, cleaves ester bonds at the sn-2 position of phospholipids. A PLA2-sensitive amphiphilic prodrug, 1-O-octadecyl-2-(5-fluorouracil)-N-acetyl-3-zidovudine-phosphorylglycerol (OFZG), was synthesized and used to prepare nanoassemblies through the injection of a mixture of OFZG/cholesterol/Tween 80 (2:1:0.1, mol:mol:mol) into water. Cholesterol and Tween 80 was incorporated into the OFZG monolayers at the air/water interface to yield nanoassemblies. The resulting nanoassemblies exhibited a narrow size distribution with a mean size of 77.8nm and were stable due to their high surface charges. The in vitro experiments showed that PLA2 degraded OFZG. The nanoassemblies exhibited higher anticancer activity than the parent drug 5-fluorouracil (5-FU) in COLO205, HT-28, and HCT-116 cells. The intravenous (i.v.) administration of the nanoassemblies into mice resulted in the rapid elimination of OFZG from the circulation and its distribution mainly in the liver, lung, spleen, and kidney. After their injection into tumor-bearing mice, the nanoassemblies exhibited anticancer efficiency comparable to that of 5-FU, even though the nanoassemblies contained concentrations of only 1/10 of the molar amount of 5-FU. The lessons learned from the study and methods for the design of PLA2-sensitive amphiphilic prodrugs are also discussed. Enzyme-sensitive amphiphilic combinatorial prodrugs and prodrug-loaded nanoassemblies may represent a new strategy for anticancer drug design.

Structuring of supported hybrid phospholipid bilayers on electrodes with phospholipase A2

The effect of a lipolytic enzyme, pork pancreatic phospholipase A(2), on hybrid bilayer membranes was monitored using voltammetry, impedance spectroscopy and surface plasmon resonance. The hybrid bilayers were prepared by Langmuir-Schaefer transfer of lipid monolayers onto gold electrodes modified with self-assembled alkanethiol monolayers, or by liposome spreading. The electrodes were immersed in the phospholipase aqueous solution to allow adsorption of the enzyme and cleavage of the ester bond in the sn-2 position of phospholipids in the outer leaflet of the hybrid layers. The action of phospholipase A(2) led to perforation of the lipid films. Impedance spectroscopy and surface plasmon resonance were used for monitoring enzyme adsorption, phospholipid hydrolysis and product desorption. The results obtained show that transport efficiency of an electroactive probe, ferrocyanate, and of an electroactive drug, doxorubicin, through the bilayer depends on the action of the enzyme; the state of the lipid layer covering the electrode surface depends on the latter as well. Cyclic voltammetry and electrochemical impedance spectroscopy were used to study this effect. The doxorubicin reduction/oxidation signals appearing at potentials close to those observed using a bare gold electrode indicated facilitated penetration of the drug into the layer. The results obtained were interpreted in terms of pore formation in the lipid matrix; phospholipase A(2) can be considered as a nano-device for high precision perforation of the lipid layer.