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The altered levels of phospholipids (PLs) and lysophospholipids (LPLs) in prostate cancer (CaP) and benign tissues in our previous findings prompted us to explore PLs and LPLs as potential biomarkers for CaP. Urinary lipidomics has attracted increasing attention in clinical diagnostics and prognostics for CaP. In this study, 31 prostate tissues obtained from radical prostatectomy were assessed using high‐resolution matrix‐assisted laser desorption/ionization imaging mass spectrometry (HR‐MALDI‐IMS). Urine samples were collected after digital rectal examination (DRE), and urinary lipids were extracted using the acidified Bligh‐Dyer method. The discovery set comprised 75 patients with CaP and 44 with benign prostatic hyperplasia (BPH) at Kyoto University Hospital; the validation set comprised 74 patients with CaP and 59 with BPH at Osaka University Hospital. Urinary lipidomic screening was performed using MALDI time‐of‐flight MS (MALDI‐TOF/MS). The levels of urinary lysophosphatidylcholine (LPC) and phosphatidylcholines (PCs) were compared between the CaP and BPH groups. The (PC [34:2] + PC [34:1])/LPC (16:0) ratio was significantly higher (P, Scatter plots of the urinary phosphatidylcholines/lysophosphatidylcholine (PCs/LPC) ratios and serum prostate‐specific antigen (PSA) levels in the Kyoto University Hospital (KUHP)/discovery and Osaka University Hospital (OUHP)/validation sets.

The lipid mediators platelet-activating factor (PAF) and lysophosphatidylcholine (LPC) play relevant pathophysiological roles in Trypanosoma cruzi infection. Several species of LPC, including C18:1 LPC, which mimics the effects of PAF, are synthesized by T. cruzi. The present study identified a receptor in T. cruzi, which was predicted to bind to PAF, and found it to be homologous to members of the progestin and adiponectin family of receptors (PAQRs). We constructed a three-dimensional model of the T. cruzi PAQR (TcPAQR) and performed molecular docking to predict the interactions of the TcPAQR model with C16:0 PAF and C18:1 LPC. We knocked out T. cruzi PAQR (TcPAQR) gene and confirmed the identity of the expressed protein through immunoblotting and immunofluorescence assays using an anti-human PAQR antibody. Wild-type (WT) and knockout (KO) parasites were also used to investigate the in vitro cell differentiation and interactions with peritoneal mouse macrophages; TcPAQR KO parasites were unable to react to C16:0 PAF or C18:1 LPC. Our data are highly suggestive that PAF and LPC act through TcPAQR in T. cruzi, triggering its cellular differentiation and ability to infect macrophages.

Membrane lipids serve as substrates for the generation of numerous signaling lipids when plants are exposed to environmental stresses, and jasmonic acid, an oxidized product of 18‐carbon unsaturated fatty acids (e.g., linolenic acid), has been recognized as the essential signal in wound‐induced gene expression. Yet, the contribution of individual membrane lipids in linolenic acid generation is ill‐defined. In this work, we performed spatial lipidomic experiments to track lipid changes that occur locally at the sight of leaf injury to better understand the potential origin of linolenic and linoleic acids from individual membrane lipids. The central veins of tomato leaflets were crushed using surgical forceps, leaves were cryosectioned and analyzed by two orthogonal matrix‐assisted laser desorption/ionization mass spectrometry imaging platforms for insight into lipid spatial distribution. Significant changes in lipid composition are only observed 30 min after wounding, while after 60 min lipidome homeostasis has been re‐established. Phosphatidylcholines exhibit a variable pattern of spatial behavior in individual plants. Among lysolipids, lysophosphatidylcholines strongly co‐localize with the injured zone of wounded leaflets, while, for example, lysophosphatidylglycerol (LPG) (16:1) accumulated preferentially toward the apex in the injured zone of wounded leaflets. In contrast, two other LPGs (LPG [18:3] and LPG [18:2]) are depleted in the injured zone. Our high‐resolution co‐localization imaging analyses suggest that linolenic acids are predominantly released from PCs with 16_18 fatty acid composition along the entire leaf, while it seems that in the apex zone PG (16:1_18:3) significantly contributes to the linolenic acid pool. These results also indicate distinct localization and/or substrate preferences of phospholipase isoforms in leaf tissue.

Recently, studies have shown that neuropathy target esterase (NTE) is essential to placental and normal blood vessel development. However, whether it is involved in abnormal placenta angiogenesis of pre-eclampsia remains unknown. Thus, our aim was to observe the expression of NTE in pre-eclamptic placentas and its effects and mechanism of NTE on the migration and the tube formation of human umbilical vein endothelial cells (HUVECs). Immunohistochemical staining showed that the NTE protein was intensely located in blood vessels of the normal pregnant placenta. However, western blot revealed that the expression level of NTE protein was significantly reduced in pre-eclamptic placenta. The results indicated that overexpression of NTE significantly promoted the migration and the tube formation of HUVECs compared with those of the control and scramble short hairpin RNA (shRNA) group. Conversely, NTE shRNA obviously inhibited the migration and the tube formation of HUVECs. Additionally, chromatography assay evidenced that NTE overexpression significantly reduced the level of phosphatidylcholine (PC) of HUVECs, but NTE shRNA obviously increased the level of PC of HUVECs. Furthermore, exogenous PC and lysophosphatidylcholine (LPC) significantly inhibited the tube formation of HUVECs in a dose-dependent manner. Collectively, our results suggest that reduced NTE in placenta may contribute to abnormal placenta angiogenesis of pre-eclampsia via the dysregulation of PC and LPC metabolism.

Multiple sclerosis is a chronic inflammatory demyelinating disease of the central nervous system (CNS), characterized by accumulated motor disability. However, whether remyelination promotes motor recovery following demyelinating injury remains unclear. Damage to the internal capsule (IC) is known to result in motor impairment in multiple sclerosis and stroke. Here, we induced focal IC demyelination in mice by lysophosphatidylcholine (LPC) injection, and examined its effect on motor behavior. We also compared the effect of LPC‐induced IC damage to that produced by endothelin‐1 (ET1), a potent vasoconstrictor used in experimental stroke lesions. We found that LPC or ET1 injections induced asymmetric motor deficit at 7 days post‐lesion (dpl), and that both lesion types displayed increased microglia/macrophage density, myelin loss, and axonal dystrophy. The motor deficit and lesion pathology remained in ET1‐injected mice at 28 dpl. In contrast, LPC‐injected mice regained motor function by 28 dpl, with corresponding reduction in activated microglia/macrophage density, and recovery of myelin staining and axonal integrity in lesions. These results suggest that LPC‐induced IC demyelination results in acute motor deficit and subsequent recovery through remyelination, and may be used to complement future drug screens to identify drugs for promoting remyelination., In this issue, we examined the effect of focal internal capsule (IC) demyelination on motor behavior in mice. We found that lysophosphatidylcholine (LPC) induced demyelination at the IC resulted in acute asymmetric motor deficit, followed by subsequent remyelination‐associated functional recovery. Our results suggest that IC demyelination is a tractable model for characterizing demyelination and remyelination through behavioral measurements, and may be used to complement future drug discovery efforts for promoting repair in inflammatory demyelinating disorders, such as multiple sclerosis (MS). Images generated with BioRender.

Apoptosis of vascular endothelial cells (VECs) is highly important in the occurrence and development of atherosclerosis (AS). HomeboxC6 (HOXC6) is expressed in higher levels in multiple malignant tissues, and it influences the malignant biological behavior of the cancer cells. However, the effects of HOXC6 on AS and the apoptosis of VECs have not been fully elucidated. In this study, we demonstrated that HOXC6 expression was increased in aortic wall of AS rats and peripheral blood monocytes of patients with coronary heart disease. Furthermore, it was uncovered that BAX expression was upregulated, while BCL-2 expression was downregulated in the aortic wall of AS rats. The apoptosis of human VECs (HVECs) cultured normally or treated with oxidized low-density lipoprotein in vitro was decreased after transfection with HOXC6-siRNA. Moreover, the results of Western blot analysis unveiled that the expressions of proapoptotic proteins, such as BAX, caspase-3, cleaved-caspase-3, and caspase-9 were reduced, while the expression of antiapoptotic protein, BCL-2, was elevated. Meanwhile, mRNA and protein expressions of phospholipase C beta (PLCβ) were decreased, the phosphorylation levels of protein kinase C zeta (PKCζ) and nuclear transcription factor-κB-p65 (NF-κBp65) and the membrane translocation of PKCζ were reduced as well. Besides, the expression of interleukin-18 (IL-18) protein was downregulated. However, after overexpression of HOXC6, the opposite trends of the abovementioned indices were observed. Furthermore, the inhibition of apoptosis induced by HOXC6-siRNA was reversed by lysophosphatidylcholine, an activator of PKCζ. Taken together, our results indicated that HOXC6 can promote the apoptosis of HVECs and may be involved in the occurrence and development of AS, which may be partially associated with the activation of PLCβ/PKCζ/NF-κBp65/IL-18 signaling pathway.

Glycerophospholipids (GPs) and sphingolipids (SPs) are important lipid components in the body and play biological functions. Omega-3 polyunsaturated fatty acids (n-3 PUFAs) are important nutrients, and their supplements are commonly used for preventing some diseases. However, the effect of n-3 PUFAs on the human glycerophospholipidome and sphingolipidome is unclear. We used targeted lipidomics to study the GP and SP profile of healthy individuals after supplementation with n-3 PUFAs for 3, 7, 14 and 21 days. Fuzzy c-means clustering was used to cluster the lipid species into six classes reflecting different changed-content patterns after n-3 PUFA supplementation. Among the species with significantly changed content, lysophospholipids were the most sensitive; their content started to increase on day 3. The content of phosphatidylserines increased at a later stage. The content of most of the phosphatidylcholines and alkylphosphatidylcholines decreased on day 21. A correlation network analysis of lipid species suggested that some enzymes involved in the metabolism of lysophospholipids and phosphatidylserines were regulated by n-3 PUFAs. Levels of creatine kinase-MB (CK-MB), urea, glucose, triglycerides and total bilirubin were altered by n-3 PUFA at 21 days. Correlation analysis revealed that the level of CK-MB was negatively correlated with those of species in lysophosphatidic acid, lysophosphatidylcholine, lysophosphatidylethanolamine and phosphatidylserine classes, which were increased by n-3 PUFA supplementation. With the analysis in this work, we demonstrated the regular pattern of n-3 PUFAs on GP and SP metabolism, which provides a pharmacological basis for n-3 PUFAs for clinical application.

Background Pancreaticobiliary maljunction and intrahepatic gallstones are at a high risk for biliary malignancy. Lysophosphatidylcholine (LPC) is increased in the bile of these patients, and we have previously reported that LPC-induced cytotoxicity causes senescence-associated secretory phenotype (SASP) in cholangiocytes. We aimed to determine the protective effect of phosphatidylcholine (PC) on LPC-induced cholangiocyte cytotoxicity. Methods MMNK-1, a human immortalized cholangiocyte cell line was treated with LPC with or without PC. To assess the biological effects of SASP components on cholangiocarcinoma, HuH28 and HuCCT1 (human cholangiocarcinoma cell lines) were cultured in the conditioned media where MMNK-1 cells treated with LPC. Results The presence of PC reduced reactive oxygen species generation and oxidative DNA damage in MMNK-1 treated with LPC. Moreover, SA-β-gal activity was markedly downregulated by PC. The secretion of SASP components, including interleukin (IL)-8, IL-6, and C-C motif chemokine ligand 2 was also substantially reduced in the presence of PC. Cellular proliferation and migration were enhanced in HuCCT1 and HuH28 cells when cultured in the conditioned media, and these observations were suppressed by simultaneous addition of PC. Conclusion PC protects cholangiocytes against LPC-induced cytotoxicity and cellular senescence, suggesting its potential as a target for inhibiting LPC-related carcinogenesis and its promotion.

Obesity alters skeletal muscle lipidome and promotes myopathy, but it is unknown whether aberrant muscle lipidome contributes to the reduction in skeletal muscle contractile force-generating capacity. Comprehensive lipidomic analyses of mouse skeletal muscle revealed a very strong positive correlation between the abundance of lysophosphatidylcholine (lyso-PC), a class of lipids that is known to be downregulated with obesity, with maximal tetanic force production. The level of lyso-PC is regulated primarily by lyso-PC acyltransferase 3 (LPCAT3), which acylates lyso-PC to form phosphatidylcholine. Tamoxifen-inducible skeletal muscle-specific overexpression of LPCAT3 (LPCAT3-MKI) was sufficient to reduce muscle lyso-PC content in both standard chow diet- and high-fat diet (HFD)-fed conditions. Strikingly, the assessment of skeletal muscle force-generating capacity ex vivo revealed that muscles from LPCAT3-MKI mice were weaker regardless of diet. Defects in force production were more apparent in HFD-fed condition, where tetanic force production was 40% lower in muscles from LPCAT3-MKI compared to that of control mice. These observations were partly explained by reductions in the cross-sectional area in type IIa and IIx fibers, and signs of muscle edema in the absence of fibrosis. Future studies will pursue the mechanism by which LPCAT3 may alter protein turnover to promote myopathy.

Aim To assess the relevance of lipoprotein-associated phospholipase A2 activity as a diagnostic and prognostic marker for renal microvascular diseases. Methods We analysed lipoprotein-associated phospholipase A2 activity and lysophosphatidylcholine levels (as a surrogate marker of oxidative stress) in 165 adolescents (aged 17.0 ± 2.3 years) with a history of Type 1 diabetes greater than 10 years. Clinical data were obtained from the German/Austrian nationwide Diabetes-Patients Follow-up (DPV) registry at blood collection and on average 2.4 ± 1.3 years later at follow-up. Relationships between lipoprotein-associated phospholipase A2 activity and clinical, demographic and laboratory variables, lysophosphatidylcholine levels and presence of albuminuria were evaluated by multivariable linear and logistic regression. Results Lipoprotein-associated phospholipase A2 activity was higher in male than female adolescents (P = 0.002). Albuminuria was present in 14% (22/158) of participants at baseline, and 5% (4/86) of participants without albuminuria at baseline developed albuminuria until follow-up. Lipoprotein-associated phospholipase A2 activity was associated neither with present nor with incident albuminuria. Lysophosphatidylcholine did not correlate with lipoprotein-associated phospholipase A2 activity. Cross-sectional bivariate correlation as well as multivariable linear regression analysis revealed a negative correlation of lipoprotein-associated phospholipase A2 activity with HbA1c and HDL-cholesterol. Conclusions Lipoprotein-associated phospholipase activity was not associated with surrogate markers for oxidative stress and early diabetic nephropathy. The association of decreased lipoprotein-associated phospholipase A2 activity with poor glucose control might limit its function as a predictor of micro- and macrovascular diseases in Type 1 diabetes.

Estrogen sulfotransferase (SULT1E) mainly catalyzes the sulfation of estrogens, which are known to prevent the pathogenesis of atherosclerosis. Recently, we found that peptides with a YKDG sequence specifically bind to oxidized low-density lipoprotein (Ox-LDL), which plays a major role in the pathogenesis of atherosclerosis. Here, we investigated the interaction between human SULT1E1 (hSULT1E1), which has a YKEG sequence (residues 61-64) unlike other human SULTs, and Ox-LDL. Results from polyacrylamide gel electrophoresis and western blotting demonstrated that hSULT1E1 specifically binds to Ox-LDL and its major lipid component (lysophosphatidylcholine; LPC), and platelet-activating factor (PAF), which bears a marked resemblance to LPC in terms of structure and activity. Moreover, an N-terminally fluorescein isothiocyanate (FITC)-labeled decapeptide (MIYKEGDVEK; FITC-hSULT1E1-P10) corresponding to residues 59-68 of hSULT1E1 specifically binds to Ox-LDL, LPC, and PAF. Unveiling the specific interaction between hSULT1E1 and Ox-LDL, LPC, and PAF provides important information regarding the mechanisms underlying various diseases caused by Ox-LDL, LPC, and PAF, such as atherosclerosis. In addition, FITC-hSULT1E1-P10 could be used as an efficient fluorescent probe for the detection of Ox-LDL, LPC, and PAF, which could facilitate the mechanistic study, identification, diagnosis, prevention, and treatment of atherosclerosis.

Some lipid digestion pathways in fish deviate from those in mammals, and many differences may also be species dependent. This report describes a pathway for monoacylglycerol (MAG) and lysophospholipid absorption by intestinal enterocytes in brown trout that may be of significance in salmonids. When culturing primary cells in a medium containing 1- and 2-MAG, we observed a massive hydrolysis of unesterified fatty acids. The hydrolysis activity was retained in the medium even after the removal of the cells. To further characterize these activities, both extracellular and isolated membrane proteins were tested for lipase activity toward triacylglycerol (TAG), diacylglycerol (DAG), MAG, phosphatidylcholine (PtdCho), and lysoPtdCho. In both cases, the main hydrolyzing activity was toward MAG followed by lysoPtdCho with very little activity toward DAG, TAG, or PtdCho. The extracellular and membrane proteins were partially purified by fast protein liquid chromatography and identified by proteomics (liquid chromatography-tandem mass spectrometry) focusing on lipase/hydrolase enzymes. In the membrane protein fraction, the data suggested that MAG was produced as an intermediate in the hydrolysis of lysoPtdCho by either lysophospholipase C or lysophospholipase D activity. Both abhydrolase-domain-containing protein 6 and abhydrolase-domain-containing protein 12 were identified in the membrane protein and they could be responsible for the hydrolysis of MAG. In the culture medium, low-peptide matches were found for ABHD6 and phospholipases and further studies are needed. In summary, trout enterocytes are capable of hydrolyzing MAG and lysoPtdCho. The enzymes are both extracellular and membrane bound. The pathways may be of significance during lipid absorption in fish lacking a 1,3 specific pancreatic lipase.

International audience; Fatty acids have many health benefits in a great variety of diseases ranging from cardiovascular to cerebral diseases. For instance, docosahexaenoic acid (DHA), which is highly enriched in brain phospholipids, plays a major role in anti-inflammatory or neuroprotective pathways. Its effects are thought to be due, in part, to its conversion into derived mediators such as protectins. 1-Lyso,2-docosahexaenoyl-glycerophosphocholine (LysoPtdCho-DHA) is one of the physiological carrier of DHA to the brain. We previously synthesized a structured phosphatidylcholine to mimic 1-lyso,2-docosahexaenoyl-glycerophosphocholine, named AceDoPC(A (R)) (1-acetyl,2-docosahexaenoyl-glycerophosphocholine), that is considered as a stabilized form of the physiological LysoPtdCho-DHA and that is neuroprotective in experimental ischemic stroke. Considering these, the current study aimed at enzymatically oxygenate DHA contained within AceDoPC(A (R)) to synthesize a readily structured oxidized phospholipid containing protectin DX (PDX), thereafter named AceDoxyPC (1-acetyl,2-PDX-glycerophosphocholine). Identification of this product was performed using liquid chromatography/tandem mass spectrometry. Such molecule could be used as a bioactive mediator for therapy against neurodegenerative diseases and stroke.

Introduction Zellweger spectrum disorders (ZSD) are a group of genetic metabolic disorders caused by a defect in peroxisome biogenesis. This results in multiple metabolic abnormalities, including elevated very long-chain fatty acid (VLCFA) levels. Elevated levels of C26:0-lysophosphatidylcholine (C26:0-lysoPC) have been shown in dried blood spots (DBS) from ZSD patients. However, little is known about the sensitivity and specificity of this marker and C26:0-carnitine, another VLCFA-marker, in ZSD. We investigated C26:0-lysoPC and C26:0-carnitine as diagnostic markers for ZSD in DBS and fibroblasts. Methods C26:0-lysoPC levels in 91 DBS from 37 different ZSD patients were determined and compared to the levels in 209 control DBS. C26:0-carnitine levels were measured in 41 DBS from 29 ZSD patients and 97 control DBS. We measured C26:0-lysoPC levels in fibroblasts from 24 ZSD patients and 61 control individuals. Results Elevated C26:0-lysoPC levels (> 72 nmol/L) were found in 86/91 ZSD DBS (n=33/37 patients) corresponding to a sensitivity of 89.2%. Median level was 567 nmol/l (range 28-3133 nmol/l). Consistently elevated C26:0-carnitine levels (> 0.077 mu mol/L) in DBS were found in 16 out of 29 ZSD patients corresponding to a sensitivity of 55.2%. C26:0-lysoPC levels were elevated in 21/24 ZSD fibroblast lines. Discussion C26:0-lysoPC in DBS is a sensitive and useful marker for VLCFA accumulation in patients with a ZSD. C26:0-carnitine in DBS is elevated in some ZSD patients, but is less useful as a diagnostic marker. Implementation of C26:0-lysoPC measurement in the diagnostic work-up when suspecting a ZSD is advised. This marker has the potential to be used for newborn screening for ZSD

Lysophophatidylcholine (LysoPC) is an abundant constituent in human plasma. Patients with malignant cancer diseases have attenuated LysoPC plasma levels, and thus LysoPC has been examined as a metabolic biomarker for cancer prediction. Preclinical studies have shown that solid tumor cells drastically degrade LysoPCs by incorporating their free fatty acids into cell membrane phospholipids. In this way, LysoPC C18:0 reduced the metastatic spread of murine melanoma B16.F10 cells in mice. Although membrane rigidification may have a key role in the attenuation of metastasis, evidence for this has yet to be shown. Therefore, the present study aimed to determine how LysoPC reduces the metastatic capacity of B16.F10 cells. Following cellular preincubation with LysoPC C18:0 at increasing concentrations and lengths of time, cell migration was most significantly attenuated with 450 μm LysoPC C18:0 at 72 h. Biosensor measurements suggest that, despite their abundance in B16.F10 cells, LysoPC-sensitive G protein-coupled receptors do not appear to contribute to this effect. Instead, the attenuated migration appears to result from changes in cell membrane properties and their effect on underlying signaling pathways, most likely the formation of focal adhesion complexes. Treatment with 450 μm LysoPC C18:0 activates protein kinase C (PKC)δ to phosphorylate syndecan-4, accompanied by deactivation of PKCα. Subsequently, focal adhesion complex formation was attenuated, as confirmed by the reduced activity of focal adhesion kinase (FAK). Interestingly, 450 μm LysoPC C18:1 did not affect FAK activity, explaining its lower propensity to affect migration and metastasis. Therefore, membrane rigidification by LysoPC C18:0 appears to prevent the formation of focal adhesion complexes, thus affecting integrin activity as a key for metastatic melanoma spread.

In recent years, the number of patients suffering from Lyme Disease (LD) has significantly increased. The most dangerous manifestation of LD is neuroborreliosis associated with invasion of the central nervous system by Borrelia burgdorferi. Phospholipids (PL) and their metabolites are involved in inflammation, which plays a dominant, but still unclear, role in the pathogenesis of neuroborreliosis. We analyzed the plasma PL profiles of neuroborreliosis patients (n = 8) and healthy volunteers (n = 8) using a lipidomic approach. Significant increases in the lysophosphatidylcholines LysoPtdCho 16:0 and LysoPtdCho 18:2 were observed. The plasma of neuroborreliosis patients appeared to have an increased relative abundance of sphingomyelin CerPCho d18:1/24:1 and a decrease in CerPCho d18:0/18:0. Principal components analysis of the relative abundances of all PL class species distinguished between neuroborreliosis patients and healthy subjects. This is the first report comparing PL classes and their molecular species in neuroborreliosis patients and healthy subjects. Electronic supplementary material The online version of this article (doi:10.1007/s11745-016-4212-3) contains supplementary material, which is available to authorized users.

The decrease in skeletal muscle mitochondrial oxidative capacity with age adversely affects muscle strength and physical performance. Factors that are associated with this decrease have not been well characterized. Low plasma lysophosphatidylcholines (LPC), a major class of systemic bioactive lipids, are predictive of aging phenotypes such as cognitive impairment and decline of gait speed in older adults. Therefore, we tested the hypothesis that low plasma LPC are associated with impaired skeletal muscle mitochondrial oxidative capacity. Skeletal muscle mitochondrial oxidative capacity was measured using in vivo phosphorus magnetic resonance spectroscopy (31P‐MRS) in 385 participants (256 women, 129 men), aged 24–97 years (mean 72.5) in the Baltimore Longitudinal Study of Aging. Postexercise recovery rate of phosphocreatine (PCr), k PCr, was used as a biomarker of mitochondrial oxidative capacity. Plasma LPC were measured using liquid chromatography–tandem mass spectrometry. Adults in the highest quartile of k PCr had higher plasma LPC 16:0 (p = 0.04), 16:1 (p = 0.004), 17:0 (p = 0.01), 18:1 (p = 0.0002), 18:2 (p = 0.002), and 20:3 (p = 0.0007), but not 18:0 (p = 0.07), 20:4 (p = 0.09) compared with those in the lower three quartiles in multivariable linear regression models adjusting for age, sex, and height. Multiple machine‐learning algorithms showed an area under the receiver operating characteristic curve of 0.638 (95% confidence interval, 0.554, 0.723) comparing six LPC in adults in the lower three quartiles of k PCr with the highest quartile. Low plasma LPC are associated with impaired mitochondrial oxidative capacity in adults.

As plant seed oils provide animals with essential fatty acids (FAs), genes that regulate plant lipid metabolism have been used in genetic manipulation to improve dietary seed oil composition and benefit human health. Herein, the Arabidopsis thaliana cytosolic acyl‐CoA‐binding proteins (AtACBPs), AtACBP4, AtACBP5, and AtACBP6 were shown to play a role in determining seed oil content by analysis of atacbp (atacbp4, atacbp5, atacbp6, atacbp4atacbp5, atacbp4atacbp6, atacbp5atacbp6, and atacbp4atacbp5atacbp6) seed oil content in comparison with the Col‐0 wild type (WT). Triacylglycerol (TAG) composition in electrospray ionization‐mass spectrometer (ESI‐MS) analysis on atacbp6 seed oil showed a reduction (−50%) of C58‐TAGs in comparison with the WT. Investigations on fatty acid composition of atacbp mutants indicated that 18:2‐FA accumulated in atacbp6 and 18:3‐FA in atacbp4, both at the expense of 20:1‐FA. As TAG composition can be modified by acyl editing through phosphatidylcholines (PC) and lysophosphatidylcholines (LPC), total PC and LPC content in atacbp6 mature seeds was determined and ESI‐MS analysis revealed that LPC had increased (+300%) at the expense of PC. Among all the 14 tested PC species, all (34:1‐, 34:2‐, 34:3‐, 34:4‐, 34:5‐, 34:6‐, 36:2‐, 36:3‐, 36:5‐, 36:6‐, 38:2‐, 38:3‐, and 38:4‐PCs) but 36:4‐PC were lower in atacbp6 than the WT. In contrast, all LPC species (16:0‐, 18:1‐, 18:2‐, 18:3‐, and 20:1‐LPC) examined were elevated in atacbp6. LPC abundance also increased in atacbp4atacbp5, but not atacbp4 and atacbp5. Interestingly, when LPC composition in atacbp4atacbp5 was compared with atacbp4 and atacbp5, significant differences were observed between atacbp4atacbp5 and each single mutant, implying that AtACBP4 and AtACBP5 play combinatory roles by affecting LPC (but not PC) biosynthesis. Furthermore, PC‐related genes such as those encoding acyl‐CoA:lysophphosphatidylcholine acyltransferase (LPCAT1) and phospholipase A2 alpha (PLA2α) were upregulated in atacbp6 developing seeds. A model on the role of AtACBP6 in modulating TAG through regulating LPCAT1 and PLA2α expression is proposed. Taken together, cytosolic AtACBPs appear to affect unsaturated TAG content and are good candidates for engineering oil crops to enhance seed oil composition.

The liver X receptor (LXR) is a cholesterol‐sensing nuclear receptor that has an established function in lipid metabolism; however, its role in inflammation is elusive. In this study, we showed that the LXR agonist GW3965 exhibited potent anti‐inflammatory activity by suppressing the firm adhesion of monocytes to endothelial cells. To further address the mechanisms underlying the inhibition of inflammatory cell infiltration, we evaluated the effects of LXR agonist on interleukin‐8 (IL‐8) secretion and nuclear factor‐kappa B (NF‐κB) activation in human umbilical vein endothelial cells (HUVECs). The LXR agonist significantly inhibited lysophosphatidylcholine (LPC)‐induced IL‐8 production in a dose‐dependent manner without appreciable cytotoxicity. Western blotting and the NF‐κB transcription activity assay showed that the LXR agonist inhibited p65 binding to the IL‐8 promoter in LPC‐stimulated HUVECs. Interestingly, knockdown of the indispensable small ubiquitin‐like modifier (SUMO) ligases Ubc9 and Histone deacetylase 4 (HDAC4) reversed the increase in IL‐8 induced by LPC. Furthermore, the LPC‐induced degradation of inhibitory κBα was delayed under the conditions of deficient SUMOylation or the treatment of LXR agonist. After enhancing SUMOylation by knockdown SUMO‐specific protease Sentrin‐specific protease 1 (SENP1), the inhibition of GW3965 was rescued on LPC‐mediated IL‐8 expression. These findings indicate that LXR‐mediated inflammatory gene repression correlates to the suppression of NF‐κB pathway and SUMOylation. Our results suggest that LXR agonist exerts the anti‐atherosclerotic role by attenuation of the NF‐κB pathway in endothelial cells.

Recent results have suggested that plant lysophosphatidylcholine:acyl-coenzyme A acyltransferases (LPCATs) can operate in reverse in vivo and thereby catalyse an acyl exchange between the acyl-coenzyme A (CoA) pool and the phosphatidylcholine. We have investigated the abilities of Arabidopsis AtLPCAT2, Arabidopsis lysophosphatidylethanolamine acyltransferase (LPEAT2), S. cerevisiae lysophospholipid acyltransferase (Ale1) and S. cerevisiae lysophosphatidic acid acyltransferase (SLC1) to acylate lysoPtdCho, lysoPtdEtn and lysoPtdOH and act reversibly on the products of the acylation; the PtdCho, PtdEtn and PtdOH. The tested LPLATs were expressed in an S. cervisiaeale1 strain and enzyme activities were assessed in assays using microsomal preparations of the different transformants. The results show that, despite high activity towards lysoPtdCho, lysoPtdEtn and lysoPtdOH by the ALE1, its capacities to operate reversibly on the products of the acylation were very low. Slc1 readily acylated lysoPtdOH, lysoPtdCho and lysoPtdEtn but showed no reversibility towards PtdCho, very little reversibility towards PtdEtn and very high reversibility towards PtdOH. LPEAT2 showed the highest levels of reversibility towards PtdCho and PtdEtn of all LPLATs tested but low ability to operate reversibly on PtdOH. AtLPCAT2 showed good reversible activity towards PtdCho and PtdEtn and very low reversibility towards PtdOH. Thus, it appears that some of the LPLATs have developed properties that, to a much higher degree than other LPLATs, promote the reverse reaction during the same assay conditions and with the same phospholipid. The results also show that the capacity of reversibility can be specific for a particular phospholipid, albeit the lysophospholipid derivatives of other phospholipids serve as good acyl acceptors for the forward reaction of the enzyme. Electronic supplementary material The online version of this article (doi:10.1007/s11745-015-4102-0) contains supplementary material, which is available to authorized users.

[Background]Human prostate cancers are highly heterogeneous, indicating a need for various novel biomarkers to predict their prognosis. Lipid metabolism affects numerous cellular processes, including cell growth, proliferation, differentiation, and motility. Direct profiling of lipids in tissue using high-resolution matrix-assisted laser desorption/ionization imaging mass spectrometry (HR-MALDI-IMS) may provide molecular details that supplement tissue morphology. [Methods]Prostate tissue samples were obtained from 31 patients, with localized prostate cancer who underwent radical prostatectomy. The samples were assessed by HR-MALDI-IMS in positive mode, with the molecules identified by tandem mass spectrometry (MS/MS). The effect of identified molecules on prostate specific antigen recurrence free survival after radical prostatectomy was determined by Cox regression analysis and by the Kaplan–Meier method. [Results]Thirteen molecules were found to be highly expressed in prostate tissue, with five being significantly lower in cancer tissue than in benign epithelium. MS/MS showed that these molecules were [lysophosphatidylcholine (LPC)(16:0/OH)+H]+, [LPC(16:0/OH)+Na]+, [LPC(16:0/OH)+K]+, [LPC(16:0/OH)+matrix+H]+, and [sphingomyelin (SM)(d18:1/16:0)+H]+. Reduced expression of LPC(16:0/OH) in cancer tissue was an independent predictor of biochemical recurrence after radical prostatectomy. [Conclusions]HR-MALDI-IMS showed that the expression of LPC(16:0/OH) and SM(d18:1/16:0) was lower in prostate cancer than in benign prostate epithelium. These differences in expression of phospholipids may predict prostate cancer aggressiveness, and provide new insights into lipid metabolism in prostate cancer., Article first published online: 1 SEP 2015

Lysophospholipid acyltransferases (LPLATs) regulate the diversification of fatty acid composition in biological membranes. Lysophosphatidylcholine acyltransferases (LPCATs) are members of the LPLATs that play a role in inflammatory responses. M1 macrophages differentiate in response to lipopolysaccharide (LPS) and are pro-inflammatory, whereas M2 macrophages, which differentiate in response to interleukin-4 (IL-4), are anti-inflammatory and involved in homeostasis and wound healing. In the present study, we showed that LPCATs play an important role in M1/M2-macrophage polarization. LPS changed the shape of PMA-treated U937 cells from rounded to spindle shaped and upregulated the mRNA and protein expression of the M1 macrophage markers CXCL10, TNF-α, and IL-1β. IL-4 had no effect on the shape of PMA-treated U937 cells and upregulated the M2 macrophage markers CD206, IL-1ra, and TGF-β in PMA-treated U937 cells. These results suggest that LPS and IL-4 promote the differentiation of PMA-treated U937 cells into M1- and M2-polarized macrophages, respectively. LPS significantly downregulated the mRNA expression of LPCAT3, one of four LPCAT isoforms, and suppressed its enzymatic activity toward linoleoyl-CoA and arachidonoyl-CoA in PMA-treated U937 cells. LPCAT3 knockdown induced a spindle-shaped morphology typical of M1-polarized macrophages, and increased the secretion of CXCL10 and decreased the levels of CD206 in IL-4-activated U937 cells. This indicates that knockdown of LPCAT3 shifts the differentiation of PMA-treated U937 cells to M1-polarized macrophages. Our findings suggest that LPCAT3 plays an important role in M1/M2-macrophage polarization, providing novel potential therapeutic targets for the regulation of immune and inflammatory disorders.

In acute stage of ischemic stroke, the surrounding zone of fresh infarcts is termed penumbra, where microglia are activated in response to damaged cell-derived proinflammatory mediators. Rescuing penumbra by regulating inflammatory activity would minimize infarct volume, which positively correlates with functional outcome. To elucidate mechanisms by which inflammation occurs in penumbra, we performed immunohistochemical investigations using autopsied human brains affected by acute, subacute and chronic stages of cerebral infarction as well as cell culture experiments using a murine microglia-derived cell line (BV-2). In penumbra of fresh infarcts, immunoreactivity for secretory phospholipase A2 group X (sPLA2 -X), which is responsible for the production and release of the proinflammatory mediator lysophosphatidylcholine (LPC), was intensely detected in neurons and astrocytes. Furthermore, immunoreactivities for the LPC receptors G protein-coupled receptor 132 (G2A) and P2X purinoreceptor 7 (P2X7R), as well as the CC chemokine monocyte chemoattractant protein-1 (MCP-1) and its receptor CCR2, were detectable in activated microglia. Prior to cell culture experiments, it was confirmed that BV-2 cells were immunoreactive for ionized Ca(2+) -binding adaptor molecule 1 (Iba1), G2A, P2X7R, MCP-1 and CCR2. Reverse transcription-quantitative polymerase chain reaction analysis revealed that MCP-1 and CCR2 mRNA expression levels were significantly increased by LPC stimulation. The LPC-driven increase in MCP-1 transcripts was lowered by blockade of G2A or P2X7R or by inhibition of Rho-associated protein kinase (ROCK) or inhibitor of κBα kinase. The LPC-driven increase in CCR2 transcripts was lowered by blockade of G2A or P2X7R or by inhibition of ROCK, phosphatidylinositide 3-kinanse, extracellular signal-regulated kinase kinase, or p38 mitogen-activated protein kinase. The present results provide in vivo and in vitro evidence that in acute stage of ischemic stroke, the sPLA2 -X enzyme product LPC is released from neurons and astrocytes and stimulates penumbra microglia via G2A and P2X7R, thereby exerting the MCP-1/CCR2-mediated neurotoxicity through distinct cell-signaling pathways.

Modeling the phospholipase A1 (PLA1 )-catalyzed partial hydrolysis of soy phosphatidylcholine (PC) in hexane for the production of lysophosphatidylcholine (LPC) and optimizing the reaction conditions using response surface methodology were described. The reaction was performed with 4 g of PC in a stirred batch reactor using a commercial PLA1 (Lecitase Ultra) as the biocatalyst. The effects of temperature, reaction time, water content, and enzyme loading on LPC and glycerylphosphorylcholine (GPC) content in the reaction products were elucidated using the models established. Optimal reaction conditions for maximizing the LPC content while suppressing acyl migration, which causes GPC formation, were as follows: temperature, 60°C; reaction time, 3 h; water content, 10% of PC; and enzyme loading, 1% of PC. When the reaction was conducted with 40 g of PC under these conditions, the reaction products contained 83.7 mol % LPC and were free of GPC. LPC had a higher total unsaturated fatty acid content than original PC had and was mainly composed of linoleic acid (78.0 mol % of the total fatty acids).

Inflammation is essential in defense against infection or injury. It is tightly regulated, as over-response can be detrimental, especially in immune-privileged organs such as the central nervous system (CNS). Microglia constitutes the major source of inflammatory factors, but are also involved in the regulation of the inflammation and in the reparation. Autotaxin (ATX), a phospholipase D, converts lysophosphatidylcholine (LPC) into lysophosphatidic acid (LPA) and is upregulated in several CNS injuries. LPA, a pleiotropic immunomodulatory factor, can induce multiple cellular processes including morphological changes, proliferation, death, and survival. We investigated ATX effects on microglia inflammatory response to lipopolysaccharide (LPS), mimicking gram-negative infection. Murine BV-2 microglia and stable transfected, overexpressing ATX-BV-2 (A +) microglia were treated with LPS. Tumor necrosis factor α (TNFα), interleukin (IL)-6, and IL-10 mRNA and proteins levels were examined by qRT-PCR and ELISA, respectively. Secreted LPA was quantified by a radioenzymatic assay and microglial activation markers (CD11b, CD14, B7.1, and B7.2) were determined by flow cytometry. ATX expression and LPA production were significantly enhanced in LPS treated BV-2 cells. LPS induction of mRNA and protein level for TNFα and IL-6 were inhibited in A+ cells, while IL-10 was increased. CD11b, CD14, and B7.1, and B7.2 expressions were reduced in A+ cells. Our results strongly suggest deactivation of microglia and an IL-10 inhibitory of ATX with LPS induced microglia activation. J. Cell. Biochem. 115: 2123–2132, 2014. © 2014 Wiley Periodicals, Inc.

2,3,5,4′-Tetrahydroxystilbene-2-O-β-d-glucoside (TSG) has been shown to protect human umbilical vein endothelial cells (HUVECs) from lysophosphatidylcholine (LPC)-induced injury; however, the underlying molecular mechanism remains to be determined. The aim of this study was to investigate the protective mechanism of TSG against LPC-induced injury in HUVECs. We established a stable LPC-induced cell model by treating HUVECs with various concentrations of LPC and found 10.0 µg/mL of LPC to be optimal for inducing HUVECs injury. The effects of TSG on LPC-induced cell injury were assessed by cell counting kit-8, apoptosis assay, transmission electron microscope, and measurement of malondialdehyde (MDA), the antioxidant enzymes superoxide dismutase (SOD), reactive oxygen species (ROS), glutathione peroxidase, and mitochondrial membrane potential. The mRNA and protein levels of caspase-3, Bax, Bcl-2, PARP-1, and cytochrome C were assayed by real-time reverse transcriptase-polymerase chain reaction and immunoblotting, respectively. TSG pretreatment was able to prevent LPC-induced HUVECs injury and restore cell viability in a concentration-dependent manner. LPC treated cells showed typical apoptotic morphological changes including cytoplasmic vacuolation, swollen mitochondria, and characteristic biochemical hallmarks of apoptosis including loss of mitochondrial membrane potential, activation of caspase-3, decrease of Bcl-2, increase of PARP-1, upregulation of Bax, and release of cytochrome C, all of which were apparently inhibited by TSG pretreatment. Treatment of HUVECs with LPC led to decrease of SOD and glutathione peroxidase in addition to rapid increase of MDA and ROS levels. Pretreatment with TSG restored SOD and glutathione peroxidase levels to that of normal levels, and significantly decreased ROS and MDA levels. Our data indicate that TSG inhibits apoptosis of HUVECs mediated by LPC through blocking the mitochondrial apoptotic pathway and suggest that the mechanisms underlying the protective effects of TSG are related to the activation of SOD and glutathione peroxidase, the clearance of intracellular ROS, and reduction of lipid peroxidation. © 2014 IUBMB Life, 66(10):711–722, 2014

Rationale Sake is made from fermented rice and has been drunk in Japan for more than 1000 years. The rice must be polished prior to fermentation to obtain high-quality sake. It is traditionally recognized that the quality of sake is improved as the rice polishing ratio (percentage removed in the polishing process) increases. However, the underlying chemistry of the rice polishing process is incompletely understood. Herein, we analyzed the distribution of lysophosphatidylcholine (LPC) molecular species with unsaturated fatty acids in rice, as their presence is thought to exert a negative effect on the flavor of sake. Methods The distribution of LPC molecular species in rice was visualized via matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). Results LPC (16:0) is ubiquitously present in the endosperm of rice while LPC (18:0) is localized in the core of the endosperm. In contrast, LPC (18:2) and LPC (18:1) are present in the outer region of the endosperm. Conclusions The enhancement of the quality of sake as the polishing ratio of the rice increases might be explained in terms of the distribution of LPC with unsaturated fatty acids in the rice.

Excess hepatic fat accumulation leads to nonalcoholic steatohepatitis (NASH), a serious threat to health for which no effective treatment is available. However, the mechanism responsible for fatty acid uptake by hepatocytes remains unclear. Using the human hepatocyte-derived tumor cell line HepG2, we found that fatty acid influx is mediated by a heterotetrameric plasma membrane protein complex consisting of plasma membrane fatty acid-binding protein, caveolin-1, CD36, and calcium-independent membrane phospholipase A2 (iPLA2β). Blocking iPLA2β with the bile acid-phospholipid conjugate ursodeoxycholate-lysophosphatidylethanolamide (UDCA-LPE) caused the dissociation of the complex, thereby inhibiting fatty acid influx (IC50 47 μM), and suppressed the synthesis of all subunits through a reduction in lysophosphatidylcholine from 8.0 to 3.5 μmol/mg of protein and corresponding depletion of phosphorylated c-Jun N-terminal kinase. These findings were substantiated by an observed 56.5% decrease in fatty acid influx in isolated hepatocytes derived from iPLA2β-knockout mice. Moreover, steatosis and inflammation were abrogated by UDCA-LPE treatment in a cellular model of NASH. Thus, iPLA2β acts as an upstream checkpoint for mechanisms that regulate fatty acid uptake, and its inhibition by UDCA-LPE qualifies this nontoxic compound as a therapeutic candidate for the treatment of NASH.-Stremmel, W., Staffer, S., Wannhoff, A., Pathil, A., Chamulitrat, W. Plasma membrane phospholipase A2 controls hepatocellular fatty acid uptake and is responsive to pharmacological modulation: implications for nonalcoholic steatohepatitis.

Objective Novel therapeutic interventions aimed at myelin repair are now under development for neuroprotection as well as functional recovery of patients with multiple sclerosis. However, development of myelin repair therapy necessitates a noninvasive approach for measuring changes in myelin content in vivo in a quantitative fashion not yet possible using magnetic resonance imaging. For this reason, we developed a novel positron emission tomography (PET) probe, termed [11C]MeDAS, that is capable of longitudinally imaging central nervous system myelin content. Methods The binding properties of [11C]MeDAS for myelin were systematically evaluated by in vitro and in situ fluorescent staining of the spinal cord and the brain, and by in vivo competitive blocking studies. Longitudinal PET studies were conducted in 3 rat models involving acute focal neuroinflammation in the brain, lysophosphatidylcholine (LPC)-induced focal demyelination in the spinal cord, and experimental autoimmune encephalomyelitis (EAE). Image-guided myelin repair therapy was conducted in an LPC rat model using a mesenchymal stem cell-based hepatocyte growth factor (HGF). Biodistribution and acute toxicity studies of [11C]MeDAS were also conducted. Results MeDAS selectively stains myelin in the spinal cord and brain. Neuroinflammation did not affect [11C]MeDAS uptake in the brain as long as the myelin sheaths remained intact. Longitudinal PET studies in LPC and EAE rat models demonstrate that [11C]MeDAS uptake changes correlate with associated myelin loss in the spinal cord. Furthermore, using [11C]MeDAS-PET, the efficacy of myelin repair therapy with HGF was longitudinally monitored in vivo. Interpretation [11C]MeDAS-PET is a promising imaging marker for monitoring myelin pathology in vivo, future applications of which in humans should be achievable. Ann Neurol 2013;74:688–698

SummaryReasons for performing study Equine summer eczema, also known as insect bite hypersensitivity, affects horses recurrently during summer months. The treatment of this allergic pruritus is difficult and therefore there is a need for efficacious treatments. Autoserum therapy, based on the use of autogenous serum that is specifically prepared for oral administration and given when the animal shows clinical signs has been introduced recently. Lipids are thought to be responsible for the effect of this therapy. Objectives The main aim of this study was to analyse the phospholipid content of autogenous serum preparations and to further assess whether these preparations have different lipid profiles depending on the clinical status of the horse. The hypothesis is that the major serum phospholipids typical of the horse are present in the autoserum preparation. Study design Descriptive controlled clinical study. Methods Sera were collected from 10 affected and 6 healthy horses, prepared in a similar fashion and the lipids contained in the resulting autoserum preparations were analysed by electrospray ionisation mass spectrometry. Results The major phospholipid classes detected were phosphatidylcholine, sphingomyelin, phosphatidic acid and traces of lysophosphatidylcholine. Horses with summer eczema had significantly abundant concentrations of phosphatidylcholine (P = 0.042) and sphingomyelin (P = 0.0017) in comparison with healthy horses, while the concentration of phosphatidic acid was significantly higher in healthy horses (P = 0.0075). Conclusions The autoserum preparation contains minute amounts of the main serum phospholipids in differing concentrations in healthy horses and horses with an allergic skin disease.

Objective Membrane phospholipid species contribute to boundary lubrication that is provided by synovial fluid (SF). Altered levels of lubricants can be associated with increased friction, leading to articular cartilage damage. This study was undertaken to determine whether the composition of phospholipid species is altered in diseases of human knee joints. Methods The study was performed using SF from unaffected controls and patients with early osteoarthritis (OA), late OA, or rheumatoid arthritis (RA). Lipids were extracted from cell- and vesicle-free SF from 9 control donors postmortem and from 17 patients with early OA, 13 patients with late OA, and 18 patients with RA. Phospholipid species were quantified by electrospray ionization tandem mass spectrometry. Results We conducted lipidomic analysis to provide the first detailed overview of phospholipid species in human SF. We identified 130 lipid species belonging to 8 lipid classes (phosphatidylcholine, lysophosphatidylcholine, phosphatidylethanolamine, plasmalogens, phosphatidylserine, phosphatidylglycerol, sphingomyelin, and ceramide). Compared to SF from controls, SF from patients with early OA and those with late OA had higher levels of most phospholipid species. Moreover, the concentrations of 64 and 27 phospholipids differed between RA and early OA SF and between RA and late OA SF, respectively. Also, the levels of 66 phospholipid species were altered in early OA versus late OA. Conclusion Our data indicate disease- and stage-dependent differences in the relative composition and levels of phospholipid species in human SF. Such alterations might affect articular joint lubrication. Because certain phospholipids scavenge reactive oxygen species (ROS) and are pro- or antiinflammatory, any altered phospholipid level might influence ROS-scavenging activity of SF and the inflammatory status of joints. Thus, phospholipids may be associated with the pathogenesis of OA.

The activity of breast milk BSDL was assayed with or without phospholipids as extra-intestinal effector candidates. Phosphatidic acid, lysophosphatidic acid and platelet activating factor but not phosphatidylcholine and lysophosphatidylcholine stimulated BSDL activity at least as efficiently as taurocholate. The apparent dissociation constants of PA and LPA at saturating concentrations of three different substrates were between 0.1 and 13.4 μM and that of PAF was below or equal to 200 pM. Kinetic data suggested the existence of at least one binding site for each of these effectors. PA, LPA and PAF are likely extra-intestinal modulators of BSDL activity.

Acyl-CoA:lysophosphatidylcholine acyltransferase (LPCAT, EC 2.3.1.23) catalyzes acylation of lysophosphatidylcholine (lysoPtdCho) to produce phosphatidylcholine (PtdCho), the main phospholipid in cellular membranes. This reaction is a key component of the acyl-editing process, involving recycling of the fatty acids (FA) mainly at the sn-2 position of PtdCho. Growing evidences indicate that the LPCAT reaction controls the direct entry of newly synthesized FA into PtdCho and, at least in some plant species, it has an important impact on the synthesis and composition of triacylglycerols. Here we describe the molecular characterization of the single LPCAT gene found in the genome of Ricinus communis (RcLPCAT) that is homologous to LPCAT genes of the MBOAT family previously described in Arabidopsis and Brassica. RcLPCAT is ubiquitously expressed in all organs of the castor plant. Biochemical properties have been studied by heterologous expression of RcLPCAT in the ale1 yeast mutant, defective in lysophospholipid acyltransferase activity. RcLPCAT preferentially acylates lysoPtdCho against other lysophospholipids (lysoPL) and does not discriminates the acyl chain in the acceptor, displaying a strong activity with alkyl lysoPL. Regarding the acyl-CoA donor, RcLPCAT uses monounsaturated fatty acid thioesters, such as oleoyl-CoA (18:1-CoA), as preferred donors, while it has a low activity with saturated fatty acids and shows a poor utilization of ricinoleoyl-CoA (18:1-OH-CoA). These characteristics are discussed in terms of a possible role of RcLPCAT in regulating the entry of FA into PtdCho and the exclusion from the membranes of the hydroxylated FA.

Lysophosphatidylcholine (LPC), a major phospholipid component of oxidized low-density lipoprotein (ox-LDL), is implicated in numerous inflammatory diseases, including atherosclerosis. Here, to clarify the relationship between bioactive endothelins (ETs) (which are considered to be potent proinflammatory mediators) and LPC/ox-LDL, we investigated the interaction between ETs and LPC/ox-LDL by fluorescence spectroscopy and western blotting. Tryptophan fluorescence measurements revealed ETs specifically interacted with LPC at concentrations that exceeded the critical micelle concentration (CMC). The tryptophan residue in ETs was not likely to be involved directly in the interaction between ETs and LPC micelles. Tryptophan fluorescence quenching revealed tryptophan residue in ETs where LPC concentrations were below the CMC may be buried deeply in the peptide or may interact with other amino acid residues, whereas tryptophan residue in ETs in the presence of LPC at concentrations exceeding the CMC was exposed outside of the peptide. Furthermore, ETs bind to ox-LDL in a concentration-dependent manner. These results strongly suggest that ox-LDL contains micelle-rich LPCs and that ETs specifically interact with the bioactive LPC micelles. Further study of the interaction between ETs and LPC micelles contained in ox-LDL will provide important information on the development and progression of many inflammatory diseases, including atherosclerosis. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.

Background Production of lysophosphatidylcholine (LPC) via enzyme-catalysed hydrolysis was studied. Three enzymes were employed to conduct the reactions at different temperatures. The starting material, phosphatidylcholine (PC), was dispersed in water (system A) and in ethanol (system B) to define the reaction mixture to carry out the trials. Results It was found that the media employed and the type of biocatalyst (free or immobilized), clearly determine the kinetics of the reactions. PC was well dissolved in ethanol but an opaque emulsion was obtained when it was dissolved in water. Immobilized PLA1 and Novozym 435 were able to convert PC into LPC in ethanol, with yields of 50% and 58.51%, respectively, after 48 h at 50 °C. The highest degree of hydrolysis (70%) was reached with Lipase PS after 48 h at 60 °C in water. Conclusions Both reaction media enabled fairly good yields but water was better. The present work proposes a simple reaction scheme compared with other reports in the literature where different substrates, additives and polar solvents have been employed. LPC has interesting properties as a bioemulsifier, which led us to develop new methods for its production. © 2013 Society of Chemical Industry

Background Lysophosphatidylcholine (LPC) has been implicated in the onset of transfusion-related acute lung injury (TRALI). In plasma, LPC is converted to lysophosphatidic acid (LPA) by autotaxin (ATX). The effect of leukoreduction in the accumulation of these bioactive lipids and ATX in human autologous blood has not been fully investigated. Study Design and Methods The accumulation of choline-containing phospholipids (LPC, sphingomyelin [SM], and phosphatidylcholine [PC]), LPA, and ATX during the storage of autologous blood and the changes caused by leukoreduction were investigated. A total of 26 orthopedic patients were enrolled. Autologous blood was collected as whole blood and, after leukoreduction, preserved refrigerated until use. Prestorage leukoreduced (LR) and non-LR autologous blood samples were analyzed. The time-dependent changes and the effect of the filtration were compared. Results A time-dependent and significant increase in the levels of LPA was observed in both non-LR and LR samples. The concentration of LPA was significantly reduced in LR compared to non-LR samples. The concentration of LPC was higher in LR compared to non-LR samples. The levels of PC, SM, and ATX were not affected by either the storage period or the leukoreduction. Conclusions Leukoreduction of autologous whole blood effectively reduced the accumulation of LPA. On the other hand, prestorage leukoreduction resulted in an increased concentration of LPC, without significantly affecting ATX. Further studies are necessary to confirm the role of LPA in the pathogenesis of adverse effects of blood transfusion, especially TRALI.

Lysophosphatidic acid (lysoPtdOH), a lysophospholipid mediator, exerts diverse physiological effects, including angiogenesis, through its specific G-protein-coupled receptors. Previously, we showed that unfertilized hen egg white contains polyunsaturated fatty acid-rich lysoPtdOH and lysophospholipase D (lysoPLD). Here, we examined whether lysoPtdOH was produced by lysoPLD in the presence and absence of a hen fertilized ovum and what the physiological role of lysoPtdOH in hen egg white is. Mass spectrometry showed that fertilized hen egg white contained about 8 μM lysoPtdOH before incubation with an ovum, mainly comprised of 18:1- (12.6 %), 18:2- (37.8 %) and 20:4-molecular species (41.5 %). In an early gestation period, the lysoPtdOH was increased up to 9.6 μM, concomitant with a decrease in the level of polyunsaturated lysophosphatidylcholine (lysoPtdCho). Moreover, lysoPtdOH-degrading activities were found in egg white and the vitelline membrane, showing that these enzymes control lysoPtdOH levels in egg white. In an egg yolk angiogenesis assay, two lysoPtdOH receptor antagonists, Ki16425 and N-palmitoyl serine phosphoric acid (NASP), inhibited blood vessel formation induced by exogenously added 18:1-lysoPtdOH and its precursor lysoPtdCho on the hen yolk sac. Ki16425 and NASP also inhibited blood vessel formation in the chorioallantoic membrane (CAM). Furthermore, the relatively higher levels of LPA₁, LPA₂, LPA₄ and LPA₆ mRNA were present in the yolk sac and CAM. These results suggest that lysoPtdOH produced from lysoPtdCho by the action of lysoPLD in hen egg white is involved in the formation of blood vessel networks through several lysoPtdOH receptors on various extraembryonic membranes, including the yolk sac membrane and CAM.

BACKGROUND: Stored platelet concentrates (PLCs) for transfusion develop a platelet storage lesion (PSL), resulting in decreased platelet (PLT) viability and function. The processes leading to PSL have not been described in detail and no data describe molecular changes occurring in all three components of stored PLCs: PLTs, PLC extracellular vesicles (PLC-EVs), and plasma. STUDY DESIGN AND METHODS: Fifty PLCs from healthy individuals were stored under standard blood banking conditions for 5 days. Changes in cholesterol, glycerophospholipid, and sphingolipid species were analyzed in PLTs, PLC-EVs, and plasma by mass spectrometry and metabolic labeling. Immunoblots were performed to compare PLT and PLC-EV protein expression. RESULTS: During 5 days, PLTs transferred glycerophospholipids, cholesterol, and sphingolipids to newly formed PLC-EVs, which increased corresponding lipids by 30%. Stored PLTs significantly increased ceramide (Cer; +53%) and decreased sphingosine-1-phosphate (−53%), shifting sphingolipid metabolism toward Cer. In contrast, plasma accumulated minor sphingolipids. Compared to PLTs, fresh PLC-EVs were enriched in lysophosphatidic acid (60-fold) and during storage showed significant increases in cholesterol, sphingomyelin, dihydrosphingomyelin, plasmalogen, and lysophosphatidylcholine species, as well as accumulation of apolipoproteins A-I, E, and J/clusterin. CONCLUSION: This is the first detailed analysis of lipid species in all PLC components during PLC storage, which might reflect mechanisms active during in vivo PLT senescence. Stored PLTs reduce minor sphingolipids and shift sphingolipid metabolism toward Cer, whereas in the plasma fraction minor sphingolipids increase. The composition of PLC-EVs resembles that of lipid rafts and confirms their role as carriers of bioactive molecules and master regulators in vascular disease.

Lysophosphatidylcholine, a major phospholipid component of oxidized low-density lipoprotein, is implicated in many inflammatory diseases, including atherosclerosis. We previously reported that Asp-hemolysin-related synthetic peptide (P21) composed of 21 amino acid residues markedly inhibits the bioactivities of oxidized low-density lipoprotein and lysophosphatidylcholine, by directly binding to oxidized low-density lipoprotein and lysophosphatidylcholine. Here, to clarify whether P21 specifically binds to lysophosphatidylcholine and what forms of lysophosphatidylcholine with which P21 interact, we investigated the interaction between P21 containing two tryptophan residues and lysophosphatidylcholine by using fluorescence spectroscopy, polyacrylamide gel electrophoresis, and surface plasmon resonance. From tryptophan fluorescence measurements, N-terminally biotinylated P21 specifically interacted with lysophosphatidylcholine, at concentrations exceeding the critical micelle concentration. From tryptophan fluorescence quenching, the tryptophan residues in biotinylated P21 in the presence of lysophosphatidylcholine were mostly exposed on the outer side of the peptide. From polyacrylamide gel electrophoresis and surface plasmon resonance, bound to 1-palmitoyl-lysophosphatidylcholine at concentrations higher than 100 μm, ensuring stable micelles. These results indicate that biotinylated P21 specifically recognizes lysophosphatidylcholine micelles. Further study of the interaction between biotinylated P21 and lysophosphatidylcholine micelles may provide important information for the prevention and treatment for many inflammatory diseases caused by lysophosphatidylcholine micelles.

Acrosin is an important proteolytic enzyme that is capable of hydrolysing the zona pellucida in bovine oocyte. Lysophosphatydic acid (LPA) derivated from lysophosphatidylcholine (LPC) is known to trigger the acrosome exocytosis. The present study was aimed at examining the acrosin activity variations in LPC-induced acrosome exocytosis and its regulation by tyrosine kinase, protein kinase C (PKC) and voltage-dependent calcium channels (VDCC) in spermatozoa previously capacitated with heparin or quercetin. The enzyme activities were spectrophotometrically measured using N-α-benzoyl-DL-arginine p-nitroanilide as an acrosin-specific substrate. The capacitation and acrosomal reaction were evaluated by chlorotetracycline assay, and the viability and acrosome integrity were evaluated by the trypan blue stain/differential interference contrast. It was observed that LPC induced acrosome exocytosis and increased the activity of acrosin in spermatozoa previously capacitated with heparin. In heparin/LPC-treated samples, it was observed that the inhibition of tyrosine kinase and PKC blocked the acrosome exocytosis and the acrosin activity (p < 0.05). Under these conditions, in heparin-capacitated spermatozoa, the LPC provokes an acrosin activity increase that is independent of calcium influx through VDCC Type L. In cryopreserved bovine spermatozoa, LPC might require modulation, mainly tyrosine kinase participation with respect to PKC activity to induce acrosome exocytosis and increase acrosin activity.

Lysophosphatidylcholine (lysoPC) with polyunsaturated fatty acyl chains has been known to be anti-inflammatory in vivo. In the present study, we examined the effect of docosahexaenoyl-lysophosphatidylcholine (DHE-lysoPC) and 17-hydroxydocosahexaenoyl-lysophosphatidylcholine (17-HDHE-lysoPC) on spleen weight and cytokine level in spleen of mice treated with lipopolysaccharide (LPS). For this purpose, mice were administrated i.p. with DHE-lysoPC or 17-HDHE-lysoPC 1 h before i.p. injection of LPS. First, DHE-lysoPC (50–400 µg/kg) was found to suppress the LPS-induced increase of spleen weight dose-dependently, and such a suppressive effect was greater for 17-HDHE-lysoPC, compared to DHE-lysoPC. Next, in an attempt to see the effect of DHE-lysoPC on cytokine levels in spleen of mice treated with LPS, DHE-lysoPC was found to suppress LPS-induced increase in the levels of cytokines such as TNF-α, IL-1β, or IL-6 in a dose dependent manner (50–400 µg/kg), in contrast to DHA showing a significant action at a high dose (400 µg/kg) only. The greater suppressive effect of 17-HDHE-lysoPC (15–150 µg/kg) than DHE-lysoPC suggested that action of DHE-lysoPC may be enhanced through lipoxygenation process. Presumably in support of this, when the interval time between 17-HDHE-lysoPC administration and LPS challenge was varied, the cytokine-suppressing effect was found to be augmented in a time-dependent manner. Taken all together, it is suggested that DHE-lysoPC and 17-HDHE-lysoPC may be beneficial in suppressing the inflammation in spleen tissue.