Specific Platelet-activating Factor Receptor Antagonist Research Articles

Effect of Platelet-Activating Factor on Barrier Function of ARPE-19 Cells.

AimTo examine the effects of platelet-activating factor (PAF) on the barrier functions of cultured retinal pigment epithelial (RPE) cells.MethodsA human RPE cell line (ARPE-19) was cultured on microporous filter supports and treated with PAF and WEB 2086, a specific PAF-receptor (PAF-R) antagonist. The permeability of the RPE monolayer was measured using transepithelial electrical resistance (TER) and sodium fluorescein flux. The expression of the tight junction protein zonula occludens (ZO)-1 and the adherens junction protein N-cadherin was assessed using immunohistochemistry and Western blotting. We also measured the vascular endothelial growth factor (VEGF) concentrations in PAF-treated cultures and re-measured RPE monolayer permeability in the presence of VEGF-neutralizing antibodies.ResultsPAF significantly decreased the TER and enhanced the sodium fluorescein flux of the RPE monolayer and downregulated the expression of ZO-1 and N-cadherin. These effects were abolished by WEB 2086-mediated blockage of the PAF-R. PAF stimulation increased VEGF expression in RPE cells, and the antibody-mediated neutralization of VEGF caused a partial recovery of the barrier properties.ConclusionThe barrier functions of ARPE-19 cells were altered by PAF, and these effects were partly mediated by an upregulation of VEGF expression in these cells. Our results contribute to the growing body of evidence supporting the role of PAF in choroidal neovascularization. Our findings suggest that PAF is a novel target in the development of therapies for increased permeability of the RPE monolayer.

A histamine-independent itch pathway is required for allergic ocular itch

Non-histaminergic TRPA1 neural pathway is required for the development of allergic ocular itch. Pharmacological inhibition of TRPA1 channel can lead to novel therapeutic strategies to treat ocular itch in severe allergic conjunctivitis.

CSH72—A Specific Antagonist of Platelet‐activating Factor in Rabbit Platelets

CSH72 (N-methyl-1-benzyl-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline HCl) inhibited the aggregation and ATP-release reaction induced by platelet-activating factor (PAF) in a concentration-dependent manner in rabbit washed platelets. The IC50 values of CSH72, kadsurenone and BN52021 on PAF (3·6 nM)-induced platelet aggregation were 61·5 ± 7·2, 18·0 ± 2·8 and 10·6 ± 2·3 μM, respectively. CSH72 also concentration-dependently inhibited [3H]PAF (3·6 nM) binding to washed platelets with an IC50 value of 46·1 ± 1·9 μM. The concentration-response curve of PAF-induced platelet aggregation was shifted to the right by CSH72 with pA2 and pA10 values of 5·47 and 4·68, respectively. CSH72 (300 μM) significantly inhibited PAF (3·6 nM)-induced thromboxane B2 formation but exhibited no effects on those induced by arachidonic acid (100 μM) and thrombin (0·1 units mL−1). CSH72 (300 μM) also completely abolished [3H]inositol monophosphate formation induced by PAF but not that caused by collagen or thrombin in the presence of indomethacin (20 μM). It is suggested that CSH72 is a specific PAF-receptor antagonist as an antiplatelet aggregatory agent.

Trichinella spiralisinfection enhances protein kinase C phosphorylation in guinea pig alveolar macrophages

To learn more about the signalling pathways involved in superoxide anion production in guinea pig alveolar macrophages, triggered by Trichinella spiralis infection, protein level and phosphorylation of mitogen activated protein (MAP) kinases and protein kinase C (PKC) were investigated. Infection with T. spiralis, the nematode having 'lung phase' during colonization of the host, enhances PKC phosphorylation in guinea pig alveolar macrophages. Isoenzymes beta and delta of PKC have been found significantly phosphorylated, although their location was not changed as a consequence of T. spiralis infection. Neither in macrophages from T. spiralis-infected guinea pig nor in platelet-activating factor (PAF)-stimulated macrophages from uninfected animals, participation of MAP kinases in respiratory burst activation was statistically significant. The parasite antigens seem to act through macrophage PAF receptors, transducing a signal for enhanced NADPH oxidase activity, as stimulating effect of newborn larvae homogenate on respiratory burst was abolished by specific PAF receptor antagonist CV 6209. A suppressive action of T. spiralis larvae on host alveolar macrophage innate immunological response was reflected by diminished protein level of ERK2 kinase and suppressed superoxide anion production, in spite of high level of PKC phosphorylation.

Platelet-activating factor (PAF) induced cytokine production and otitis media with effusion (OME) in the rat middle ear

Objectives We examined the effect of platelet-activating factor (PAF) on IL-8 production in cultured rat middle ear epithelial cells (RMECs), and the concentrations of cytokine, PAF, and PAF-acetylhydrolase (PAF-AH) were also examined in the PAF-induced experimental OME (otitis media with effusion) of rats. Methods Using an enzyme-linked immunospecific assay, we measured the levels of cytokines in the cultured RMECs and the PAF-induced OME of rats. The PAF was quantitated by the platelet-aggregating activity and the PAF-AH was measured by radioimmunoassay. Results Both PAF and C-PAF, which is a stable analogue of PAF, significantly induced production of IL-8 in the RMECs in a dose-dependent manner. The PAF-induced IL-8 production was abolished by co-incubation with WEB2170, a specific PAF receptor antagonist. The concentrations of the cytokines and PAF in the PAF-induced OME of rats were higher on day 1 and the PAF and cytokine levels seemed to correspond well with the persistence of OME. Conclusion PAF may stimulate the local production of cytokines and may induce OME in the middle ear.

Blockades of phospholipase A2 and platelet-activating factor receptors reduce the hemocyte phagocytosis in Rhodnius prolixus: In vitro experiments

The hemocytes phagocytosis in response to microorganisms may play an important role in the cellular immune responses of insects. Here, we have evaluated the effects of the platelet-activating factor (PAF) and eicosanoids in the phagocytosis of hemocyte monolayers of Rhodnius prolixus to the yeast Saccharomyces cerevisiae. Experiments showed that the phagocytosis of yeast cells by Rhodnius hemocytes is very efficient in both controls and cells treated with PAF and arachidonic acid. Phagocytosis of yeast particles is significantly blocked when the specific phopholipase A2 inhibitor, dexamethasone, is applied on the hemocytes. By contrast, dexamethasone-pretreated hemocyte monolayers exhibit a drastic increase in the quantity of yeast cell-hemocyte internalization when the cells are treated by arachidonic acid. In addition, phagocytosis presents significant reduction in hemocyte monolayers treated with a specific PAF receptor antagonist, WEB 2086. Nevertheless, inhibition of phagocytosis with WEB 2086 is counteracted by the treatment of the hemocyte monolayers with PAF. In conclusion, phagocytosis of yeast cells by hemocytes is related to the activation of PAF receptors and eicosanoid pathways in the bloodsucking bug, R. prolixus.

Activation of platelet‐activating factor receptor in SZ95 sebocytes results in inflammatory cytokine and prostaglandin E2 production

Platelet-activating factor (PAF) is a group of phosphocholines with various biological effects mediated by the PAF receptor (PAF-R). Activation of the epidermal PAF-R induces the expression of inflammatory mediators, including cyclooxygenase-2 (COX-2) and prostaglandin E(2) (PGE(2)). The upregulation of COX-2 expression has been shown to be involved in sebocyte proliferation, sebaceous gland inflammation and carcinogenesis. The present study was designed to investigate whether PAF-R activation could induce the expression of COX-2 and production of PGE(2), as well as secretion of the inflammatory cytokine, interleukin-8 (IL-8), in the immortalized sebaceous gland cell line SZ95. Using calcium mobilization studies, we first confirmed that PAF can signal through PAF-R in SZ95 sebocytes. We then found that the production of IL-8 was induced following treatment with PAF-R agonist, however blocked by a specific PAF-R antagonist. Induction of COX-2 expression and increased PGE(2) production were observed in SZ95 sebocytes after PAF-R activation. Finally, it was demonstrated that the production of PGE(2), induced by PAF-R activation and mediated by COX-2 expression, was blocked following PAF-R antagonism in SZ95 sebocytes. These studies suggest that SZ95 sebocytes express functional PAF-Rs and PAF-Rs are involved in regulating the expression of inflammatory mediators, including COX-2, PGE(2) and IL-8.

Functional platelet-activating factor receptors in immature forms of leukemic blasts

Platelet-activating factor (PAF) is a phospholipid mediator with potent immunoregulatory activities on mature leukocytes. PAF modulates leukocyte cytosolic Ca 2+ concentration ([Ca 2+] i) through a Gq mediated pathway. We highlight, for the first time, Gq transcripts, PAF receptor (PAF-R) transcripts and protein in blast cells of acute myeloid (AML) and lymphoid (ALL) leukemia patients. PAF stimulated [Ca 2+] i in leukemic blast cells; PAF effects being prevented by a specific PAF-R antagonist. In conclusion, functional PAF-R are present in blast cells of patients with acute leukemia; a result that could be of physiologic importance regarding the important effect of PAF on leukocytes maturation and functions.

PAF-receptor is preferentially expressed in a distinct synthetic phenotype of smooth muscle cells cloned from human internal thoracic artery: Functional implications in cell migration

Platelet-activating-Factor (PAF) and its structural analogues formed upon low density lipoprotein oxidation are involved in atherosclerotic plaque formation and may signal through PAF-receptor (PAF-R) expressed in human macrophages and in certain smooth muscle cells (SMCs) in the media, but rarely in the intima of human plaques. Our aim was to determine which SMC phenotype expresses PAF-R and whether this receptor is functional in cell migration. Circulating SMC progenitors and two phenotypically distinct clones of proliferative, epithelioid phenotype vs contractile, spindle-shaped SMCs from the media of adult internal thoracic artery were studied for the presence of PAF-receptor (PAF-R). The levels of specific mRNA were obtained by reverse transcription/real-time PCR, the protein expression was deduced from immunohistochemistry staining, and the functional transmigration assay was performed by Boyden chamber-type chemotaxis assay. Only SMCs of spindle-shape and synthetic phenotype expressed both mRNA and PAF-R protein and in the functional test migrated at low concentrations of PAF. Two unrelated, specific PAF-R antagonists inhibited PAF-induced migration, but did not modify the migration initiated by PDGF. The presence of functional PAF-R in arterial spindle-shaped SMCs of synthetic phenotype may be important for their migration from the media into the intima and atherosclerotic plaques formation.

Platelet-Activating Factor Antagonists Decrease Follicular Dendritic-Cell Stimulation of Human B Lymphocytes

Both B-lymphoblastoid cell lines and tonsillar B lymphocytes express receptors for platelet-activating factor (PAF). In lymph node germinal centres, B lymphocytes interact with follicular dendritic cells (FDCs), which present antigen-containing immune complexes to B lymphocytes. FDCs have phenotypic features that are similar to those of stromal cells and monocytes and may therefore be a source of lipid mediators. In this study, we evaluated the effects of the PAF antagonist WEB 2170 on the activation of tonsillar B lymphocytes by FDCs. FDCs were isolated from tonsils by Bovine Serum Albumin (BSA) gradient centrifugation. After being cultured for 6 to 10 days, they were incubated with freshly isolated B cells in the presence or absence of the specific PAF receptor antagonist WEB 2170. B-lymphocyte proliferation was assessed by [3H]-thymidine incorporation, and immunoglobulin (Ig) G and IgM secretion was assessed by enzyme-linked immunosorbent assay (ELISA). WEB 2170 (10-6 to 10-8 M) inhibited [3H]-thymidine incorporation by up to 35% ± 3%. Moreover, the secretion of IgG and IgM was inhibited by up to 50% by WEB 2170 concentrations ranging from 10-6 to 10-8 M. There was no evidence of toxicity by trypan blue staining, and the addition of WEB 2170 to B cells in the absence of FDCs did not inhibit the spontaneous production of IgG or IgM. The effect of the PAF antagonist is primarily on B lymphocytes, as reverse transcription polymerase chain reaction detected little PAF receptor messenger ribonucleic acid (mRNA) from FDCs. These data suggest that endogenous production of PAF may be important in the interaction of B lymphocytes with FDCs.

Novel effect of Y-24180, a presumed specific platelet-activating factor receptor antagonist, on Ca 2+ levels and growth of human osteosarcoma cells

In human osteosarcoma MG63 cells, the effect of Y-24180, a presumed specific platelet-activating factor (PAF) receptor antagonist, on intracellular Ca 2+ concentration ([Ca 2+] i) and proliferation was measured by using fura-2 and tetrazolium as fluorescent dyes, respectively. Y-24180 (1–5 μM) caused a rapid and sustained [Ca 2+] i rise in a concentration-dependent manner. The [Ca 2+] i rise was inhibited by 35% by dihydropyridines or removal of extracellular Ca 2+, but was not altered by verapamil and diltiazem. In Ca 2+-free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca 2+-ATPase, caused a monophasic [Ca 2+] i rise, after which 5 μM Y-24180 failed to increase [Ca 2+] i; conversely, depletion of Ca 2+ stores with 5 μM Y-24180 abolished thapsigargin-induced [Ca 2+] i rise. U73122, an inhibitor of phoispholipase C, inhibited histamine-induced, but not 5 μM Y-24180-induced [Ca 2+] i rise. Overnight treatment with 0.1–5 μM Y-24180 inhibited cell proliferation in a concentration-dependent manner. Together, these findings suggest that Y-24180 acts as a potent and cytotoxic Ca 2+ mobilizer in human osteosarcoma cells, by inducing both extracellular Ca 2+ influx and intracellular Ca 2+ release. Alterations in cytosolic Ca 2+ regulation may lead to interferences of various cellular functions; thus, attention should be exercised in using Y-24180 as a selective PAF receptor antagonist.

HIV drug nevirapine (Viramune): risk of severe hepatotoxicity.

Platelet-activating factor (PAF) has recently been demonstrated to be metabolized by B lymphocytes and to cause enhancement of Ig synthesis by Ig-secreting B lymphoblastoid cell lines. We have now examined some of the early activation events triggered by PAF binding to three Ig-secreting B cell lines, LA350 (IgM secreting), HSCE- (IgG secreting), and U266 (IgE secreting). After addition of 10(-7) to 10(-11) M PAF, but not equimolar concentrations of the inactive metabolite lyso-PAF, all three cell lines demonstrated rapid dose-dependent increases in free cytosolic Ca2+ concentrations ([Ca2+]i). The increases in [Ca2+]i resulted from both the release of Ca2+ from internal stores as well as transmembrane Ca2+ uptake. Addition of PAF triggered the rapid hydrolysis of phosphatidylinositol bisphosphate and accumulation of inositol phosphates. PAF also increased expression of the cell cycle-active genes c-fos and EGR2 in a dose-dependent fashion. The stimulated increases in [Ca2+]i and phosphatidylinositol bisphosphate hydrolysis and the increases in gene expression were all inhibited by the specific PAF receptor antagonist Web 2086. The LA350 cell line (which expresses surface IgM) was also shown to increase [Ca2+]i after addition of anti-IgM antibodies. Sequential addition of PAF or anti-IgM antibody in either order failed to reveal any evidence for heterologous desensitization. Furthermore, the PAF receptor antagonist did not affect anti-IgM induced changes in [Ca2+]i. These data provide evidence for the presence of functional PAF receptors on B lymphoblastoid cells and indicate a potential role for PAF in the regulation of B cell activation.

Platelet activating factor receptor binding plays a critical role in jet fuel-induced immune suppression

Applying military jet fuel (JP-8) or commercial jet fuel (Jet-A) to the skin of mice suppresses the immune response in a dose-dependant manner. The release of biological response modifiers, particularly prostaglandin E 2 (PGE 2), is a critical step in activating immune suppression. Previous studies have shown that injecting selective cyclooxygenase-2 inhibitors into jet fuel-treated mice blocks immune suppression. Because the inflammatory phospholipid mediator, platelet-activating factor (PAF), up-regulates cyclooxygenase-2 production and PGE 2 synthesis by keratinocytes, we tested the hypothesis that PAF-receptor binding plays a role in jet fuel-induced immune suppression. Treating keratinocyte cultures with PAF and/or jet fuel (JP-8 and Jet-A) stimulates PGE 2 secretion. Jet fuel-induced PGE 2 production was suppressed by treating the keratinocytes with specific PAF-receptor antagonists. Injecting mice with PAF, or treating the skin of the mice with JP-8, or Jet-A, induced immune suppression. Jet fuel-induced immune suppression was blocked when the jet fuel-treated mice were injected with PAF-receptor antagonists before treatment. Jet fuel treatment has been reported to activate oxidative stress and treating the mice with anti-oxidants (Vitamins C, or E or beta-hydroxy toluene), before jet fuel application, interfered with immune suppression. These findings confirm previous studies showing that PAF-receptor binding can modulate immune function. Furthermore, they suggest that PAF-receptor binding may be an early event in the induction of immune suppression by immunotoxic environmental agents that target the skin.

Platelet-activating factor inhibits the secretion of platelet-activating factor acetylhydrolase by human decidual macrophages.

To clarify the role of platelet-activating factor (PAF) in parturition, the effects of PAF on the secretion of PAF-acetylhydrolase (PAF-AH), a PAF-inactivating enzyme, by decidual macrophage populations were examined. The cells were isolated from human decidual tissue by enzymatic digestion, Ficoll-Paque centrifugation, or flow cytometric sorting. The nonhydrolyzable agonist of PAF, carbamyl-PAF (C-PAF), inhibited the secretion of PAF-AH by either decidual cells or flow cytometrically purified decidual macrophages. A specific PAF receptor antagonist, WEB 2086, blocked the C-PAF-induced inhibition. Lyso-PAF, a metabolite of PAF, had no effect on the enzyme secretion. An intracellular calcium channel blocker, bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid, tetra(acetoxymethyl)-ester, partially blocked the inhibition by C-PAF, whereas extracellular calcium channel blockers, nifedipine and verapamil, were without effect. The inhibitory effect of C-PAF was also partially blocked by protein kinase C (PKC) inhibitors, sphingosine and H-7. A PKC activator, 12-O-tetradecanoylphobol 13-acetate, decreased the secretion of PAF-AH. The decrease was abolished by the addition of sphingosine and H-7. It is suggested that PAF inhibits the PAF-AH secretion by decidual macrophages and that the inhibitory action is mediated by a signal transduction mechanism involving intracellular calcium and PKC.

Induction of IL-4 by platelet-activating factor.

Platelet-activating factor (PAF) is a phospholipid inflammatory mediator which is synthesized by a variety of cells, including monocytes, endothelial cells, mast cells and neutrophils. PAF acts via a recently cloned PAF receptor, present on monocytes and endothelial cells, but not on non-activated lymphocytes. IL-4 is mainly produced by T lymphocytes, and belongs to the Th2 subset of T helper cells. IL-6 is mainly a monocyte/macrophage-derived cytokine with multiple proinflammatory effects. We here report that PAF induces IL-4 production, as determined by ELISPOT. Antibodies to MHC class II inhibited the IL-4 stimulatory effects of PAF. PAF also had the capacity to induce IgA production, as determined by ELISPOT, and IL-6 production in peripheral blood mononuclear cells (PBMC) as determined by ELISA. These PAF-mediated effects were completely inhibited by a specific PAF-receptor antagonist, WEB 2170. Taken together, our data indicate that PAF activates T lymphocytes to IL-4 production by an indirect, monocyte-dependent mechanism dependent on MHC class II. PAF also enhances antibody formation and IL-6 production from PBMC. These findings indicate that PAF activates immune-competent cells, which may be of importance in inflammatory diseases such as asthma, vasculitis and atherosclerosis.

Intra-plaque production of platelet-activating factor correlates with neoangiogenesis in human carotid atherosclerotic lesions

Platelet-activating factor (PAF) is a phospholipid mediator synthesized by activated inflammatory and endothelial cells. Recently PAF has been shown to contribute to neoangiogenesis in several experimental models. Here we evaluated the presence of PAF and its potential role in neovascularization within human atherosclerotic plaques. The amount of PAF extracted from 18 carotid plaques (266.65+/-40.07 pg/100 mg dry tissue; mean +/- SE) was significantly higher than that extracted from 18 normal arterial specimens (6 from carotid artery and 12 from aorta) (4.72+/-2.31 pg/100 mg dry tissue; mean +/- SE). The levels of PAF significantly correlated with the infiltration of CD68-positive monocytes and the extent of neovascularization, detected as von Willebrand Factor-positive cells. The amount of PAF also correlated with the area occupied by TNF-alpha-expressing cells. The absence of enhanced level of PAF in the circulation of atherosclerotic patients suggests a local production of this mediator within the plaque. The lipid extracts of atherosclerotic plaques containing high levels of PAF-bioactivity, but not those of control arteries, were angiogenic in a murine Matrigel model. WEB 2170, a specific PAF receptor antagonist, significantly prevented angiogenesis induced by the lipid extracts of atherosclerotic plaques. Our results indicate a local production of PAF within the atherosclerotic plaques and suggest that it may contribute to intra-plaque neoangiogenesis.

Platelet-activating factor, a critical mediator in the pathogenesis of dextran sulfate sodium-induced colitis in rats.

Disorder of mucosal immunity based on an imbalance between proinflammatory and anti-inflammatory cytokines is believed to be a major factor in the pathogenesis of ulcerative colitis. Platelet-activating factor potentially stimulates the production of proinflammatory cytokines and recruits inflammatory cells. The aim of this study was to determine whether and to what extent platelet-activating factor plays a role in the pathogenesis of ulcerative colitis. Using dextran sulfate sodium-induced colitis in rats as a model of ulcerative colitis, we analyzed the composition of cellular infiltrates and the local tissue expression of messenger ribonucleic acid for cytokine-induced neutrophil chemoattractant and tumor necrosis factor-alpha. To directly assess the impact of platelet-activating factor on the development of colitis, we also determined the efficacy of a specific platelet-activating factor receptor antagonist for preventing dextran sulfate sodium-induced colitis. The activity of colitis was well correlated with the upregulation of cytokine-induced neutrophil chemoattractant and tumor necrosis factor-alpha messenger ribonucleic acid in local tissues and infiltration of cytokine-induced neutrophil chemoattractant-positive neutrophils and ED1-positive macrophages. The platelet-activating factor receptor antagonist effectively ameliorated colitis, along with causing a decrease in the tissue cytokine-induced neutrophil chemoattractant messenger ribonucleic acid level and a decline in neutrophil and macrophage infiltration. However, the antagonist did not alter tissue levels of tumor necrosis factor-alpha messenger ribonucleic acid. Platelet-activating factor plays a critical role in the pathogenesis of dextran sulfate sodium-induced colitis through recruitment of cytokine-induced neutrophil chemoattractant-positive neutrophils and macrophages and/or stimulation of cytokine-induced neutrophil chemoattractant release from activated neutrophils. The tissue level of tumor necrosis factor-alpha messenger ribonucleic acid does not closely reflect the activity of colitis.

Involvement of platelet-activating factor in hepatic apoptosis and necrosis in chronic ethanol-fed rats given endotoxin.

Platelet-activating factor (PAF)-a potent activator of neutrophils-plays an important role in the pathogenesis of endotoxin-induced tissue injury. However, the role of PAF in hepatic damage during alcoholic hepatitis remains unclear. The aims of the present study were to test whether PAF contributes to hepatic injury in an animal model of alcoholic hepatitis and to investigate the involvement of the Fas-receptor/Fas-ligand system in this process. Male Sprague-Dawley rats were pair-fed with Lieber-DeCarli ethanol liquid diet or isocaloric control diet for 6 weeks. Liver injury was induced by the intravenous (i.v.) injection of lipopolysaccharide (LPS) (1 mg/kg). Rats were pretreated with a specific PAF receptor antagonist (TCV-309; 100 mg/kg i.v.) or vehicle 1 h before LPS treatment. Chronic ethanol administration remarkably sensitized the rats to the effects of LPS, with resultant severe hepatocellular injury, accompanied by significant increases in serum levels of alanine aminotransferase (ALT), tumour necrosis factor (TNF)-alpha and interleukin (IL)-8 (CINC/gro). Histological examination of the damaged livers showed hepatocyte apoptosis and necrosis with extensive infiltration by neutrophils, whereas immunohistochemical studies revealed enhanced Fas-receptor expression on hepatocytes and hepatic accumulation of neutrophils expressing Fas-ligand. Pretreatment with the PAF receptor antagonist protected against hepatic injury, suppressing hepatocyte apoptosis and necrosis, infiltration of neutrophils, expression of Fas-receptor and Fas-ligand, and serum TNF-alpha levels. Our study suggests that PAF is an important mediator of hepatic injury in the ethanol/endotoxin model of alcoholic hepatitis.

Lysophosphatidylcholine (LPC) induces proinflammatory cytokines by a platelet-activating factor (PAF) receptor-dependent mechanism.

Oxidized low-density lipoprotein (oxLDL) consists of both lipid components and apoprotein B100. OxLDL has both proinflammatory and cytotoxic properties. The present study was undertaken to investigate the effects of components in the lipid moiety of oxLDL on immune activation as determined by cytokine and immunoglobulin secretion. LPC induced interferon-gamma (IFN-gamma) secretion in peripheral blood mononuclear leucocytes from healthy blood donors. The effect varied between individuals, and there were both responders and non-responders. Furthermore, LPC induced enhanced antibody production, indicating B cell activation. None of eight oxysterols, arachidonic acid (AA), or 15-lipoxygenase products of AA tested had immune stimulatory properties. We recently demonstrated that PAF and oxLDL induce IFN-gamma secretion by a common mechanism. LPC-induced IFN-gamma secretion was inhibited by a specific PAF receptor antagonist, WEB 2170, indicating that the PAF receptor is involved in LPC-induced immune activation. Both oxLDL- and LPC-induced antibody formation was inhibited by WEB 2170. Furthermore LPC also induced tumour necrosis factor-alpha secretion, and this effect was inhibited by WEB 2170. LPC is produced during lipid oxidation (as in oxLDL), but also by enzymes such as phospholipase A2. The findings indicate that LPC may play an important role in inflammatory reactions, including atherosclerosis.

Lysophosphatidylcholine and lyso-PAF display PAF-like activity derived from contaminating phospholipids

Lysophosphatidylcholine is an abundant component of plasma and oxidized LDL that displays several biological activities, some of which may occur through the platelet-activating factor (PAF) receptor. We find that commercial lysophosphatidylcholine, its alkyl homolog (lyso-PAF), and PAF all induce inflammation in a murine model of pleurisy. Hydrolysis of PAF to lyso-PAF by recombinant PAF acetylhydrolase abolished this eosinophilic infiltration, implying that lyso-PAF should not have displayed inflammatory activity. Saponification of lyso-PAF or PAF acetylhydrolase treatment of lyso-PAF or lysophosphatidylcholine abolished activity; neither lysolipid should contain susceptible sn-2 residues, suggesting contaminants account for the bioactivity. Lyso-PAF and to a lesser extent lysophosphatidylcholine stimulated Ca2+ accumulation in 293 cells stably transfected with the human PAF receptor, and this was inhibited by specific PAF receptor antagonists. Again, treatment of lyso-PAF or lysophosphatidylcholine with recombinant PAF acetylhydrolase, a nonselective phospholipase A2, or saponification of lyso-PAF destroyed the PAF-like activity, a result incompatible with lyso-PAF or lysophosphatidylcholine being the actual agonist. We conclude that neither lyso-PAF nor lysophosphatidylcholine is a PAF receptor agonist, nor are they inflammatory by themselves. We suggest that PAF or a PAF-like mimetic accounts for inflammatory effects of lysophosphatidylcholine and lyso-PAF. —Marathe, G. K., A. R. Silva, H. C. de Castro Faria Neto, L. W. Tjoelker, S. M. Prescott, G. A. Zimmerman, and T. M. McIntyre. Lysophosphatidylcholine and lyso-PAF display PAF-like activity derived from contaminating phospholipids.