Effect of platelet-activating factor on lipoprotein lipase and blood lipids

We investigated the effect of platelet-activating factor (PAF) and of the PAF specific antagonist CV-6209 on plasma lipid metabolism, and particularly on post-heparin plasma lipolytic activity in male Wistar rats. Lipoprotein lipase (LPL) activity was enhanced by intravenous injection of PAF before intravenous injection of heparin when the PAF dose was low (0.2 μg/kg). PAF activated hepatic triacylglycerol lipase (HTGL) activity dose-dependently. Plasma triacylglycerols (TG) significantly decreased with the activation of LPL and/or HTGL. Plasma total cholesterol (TC) and phospholipid (PL) levels decreased at a low dose of PAF (0.2 μg/kg), but increased when higher doses were used. The PAF antagonist CV-6209 partially reversed the PAF induced effects on HTGL, TC and PL. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Characterization of the coronary vascular responses to platelet-activating factor in the isolated perfused heart

Platelet-activating factor (PAF) is a potent phospholipid mediator with diversein vivo andin vitro coronary vascular effects. In the present study, the coronary vascular responses to bolus injections of PAF were compared in rat hearts perfused under constant flow and under constant pressure. Low levels of PAF (1 pmol) produced vasodilatation only, while higher PAF concentrations (100 pmol) produced initial vasodilatation which was followed by a vasoconstriction under both experimental conditions. To determine species differences in PAF action, the effect of PAF was tested on perfused guinea pig hearts. Unlike in perfused rat hearts, only a dosedependent vasoconstrictor response was observed in perfused guinea pig hearts following a bolus injection of 1 fmol to 10 pmol of PAF. The results from repeated injections of PAf indicated that depletion of vasoactive mediators induced by PAF or receptor desensitization may explain a failure of a second injection of PAF to initiate a vasoconstrictor response. After PAF injection, the coronary vascular response to leukotriene was not altered, indicating that the reduced vasoconstrictor effect of a second injection of PAF cannot be due to a reduced ability of the smooth muscle to constrict. The study demon-strates that similar coronary vascular responses to PAF are observed in perfused rat hearts under either constant flow rate or constant pressure and that some of the variable coronary vascular responses reported may be due to the difference between animal species.     

Metabolism of platelet-activating factor by blood platelets and plasma

High performance liquid chromatography in combination with a radioactivity detector was used to study the metabolism of platelet-activating factor (1-0-alkyl-2-acetyl-sn-glycero-3-phosphocholine) by washed platelets, platelet-free plasma and platelet-rich plasma obtained from rabbits and humans. Degradation of platelet-activating factor in plasma was completely inhibited by diisopropylfluorophosphate and was partially inhibited by ethylenediamine tetraacetic acid. Washed platelets metabolized platelet-activating factor not only to the 2-lyso compound but also, by reacylation of this lyso intermediate, to an analogue of platelet-activating factor probably containing a long-chain acyl group at thesn-2 position. These transformations occurred, but to a lesser extent, in platelet-rich plasma.     

Effect of platelet-activating factor antagonist BN 52063 on the nephrotoxicity of cisplatin

Cisplatin (DDP) is an effective anticancer agent that has been successfully applied against various solid tumors. However, DDP commonly causes nephrotoxicity. We observed that DDP led to significant alterations in renal microcirculation when administered to Munich-Wistar rats, with a concomitant decrease in single nephron glomerular filtration rate due to reduction in glomerular plasma flow and transcapillary hydraulic pressure difference. BN 52063, a platelet-activating factor antagonist, caused a striking change in acute renal failure induced by DDP leading toward normalization of all parameters of renal function. The results suggest that BN 52063 could be used as a novel drug to control DDP nephrotoxicity. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Production of platelet-activating factor by human normodense and hypodense eosinophils

Normodense eosinophils and neutrophils from normal donors produced considerable amounts of plateletactivating factor (PAF) when stimulated with ionophore A23187. PAF produced by eosinophils appeared to be degraded more rapidly than PAF formed by neutrophils, suggesting a higher activity of PAF-degrading enzyme in eosinophils. Substantial proportions of PAF newly formed by both eosinophils and neutrophils were shown to be cell-associated. By comparison, hypodense eosinophils obtained from a patient with idiopathic hypereosinophilic syndrome produced an extremely large amount of PAF and released much of it into the incubation medium. The accelerated formation of PAF in hypodense eosinophils may be related to various cardiovascular complications associated with hypereosinophilic syndrome. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Tumor necrosis factor release by human monocytes stimulated with platelet-activating factor

Platelet-activating factor (PAF) is a low molecular weight phospholipid which enhances human monocyte cytotoxicity for tumor cells. In the current studies, the capacity of PAF to stimulate release of tumor necrosis factor α (TNFα) by human monocytes was assessed. PAF induced maximal TNFα synthesis 2–3 hr after monocyte stimulation as assessed by dot blotting of cell-associated TNFα using polyclonal anti-TNF antibody. Maximal net release of TNFα occurred 5–16 hr after monocyte stimulation, as assessed by a specific ELISA. Dose-response studies revealed that a maximal two- to three-fold increase in release of TNFα occurs at 10–100 pM PAF. LysoPAF and the optical isomer of PAF did not stimulate release of TNFα, suggesting that stimulation is mediated by specific PAF receptors. Scatchard analysis of [³H]PAF binding to monocyte membranes revealed 651±495 binding sites/monocyte with a Kd of 4.7±4.2×10⁻¹⁰M. PAF is a structurally unique activator of monocytes whose interactions with TNFα and other cytokines may be critical to host defense against tumors. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

川芎嗪对雪旺细胞神经生长因子合成与分泌的影响

目的观察川芎嗪(Tetramethylpyrazine,TMP)对雪旺细胞(Schwann cells,SCs)合成与分泌神经生长因子(Nervegrowth factor,NGF)的影响,探讨其促进周围神经损伤修复的可能机制。方法采用DMSO溶解TMP,将SCs分为正常对照、溶媒对照组和不同浓度TMP组(终浓度分为25、50、100和200μg/ml),作用24 h后,通过MTT法和流式细胞术分析TMP对SCs增殖和凋亡的影响,Real-Time PCR与ELISA法分别检测TMP在mRNA和蛋白质水平对SCs合成与分泌NGF的影响。结果 TMP对SCs的增殖活力及凋亡均无影响。溶媒对照组、25、50μg/ml TMP组NGF基因mRNA水平和细胞培养上清中NGF含量与正常对照组相比,差异均无统计学意义(P>0.05);100、200μg/ml TMP组与正常对照组相比明显增高(P<0.05),两组间差异无统计学意义(P>0.05)。结论 TMP可促进SCs合成与分泌NGF,且该作用具有浓度依赖性,这可能是其促进神经损伤修复的机制之一。     

Cardiovascular effects of platelet-activating factor

Sudden release of platelet-activating factor (PAF) into the circulation can cause hypotension, tachycardia, and circulatory collapse. To further examine this response, we performed detailed studies of cardiovascular function after PAF administration to young domestic pigs and newborn piglets. Our results indicate that circulatory dysfunction after PAF reflects severe constriction of pulmonary resistance vessels and consequent acute right ventricular failure. Although PAF-induced coronary artery constriction and contractile depression may be complicating problems, left ventricular underperfusion and dysfunction after PAF are mainly the result of systemic arterial hypotension and diminished left ventricular filling. The adverse hemodynamic effects of PAF are accompanied by substantial release of thromboxane A₂ (TxA₂). These effects are mimicked by the TxA₂ agonist U-46619 and partially blocked by specific and nonspecific inhibitors of TxA₂ synthesis (OKY-046 and indomethacin). Even more potent blockade of PAF action is exerted by the TxA₂ receptor blocker, SQ 29,548. Taken together, these findings indicate that severe pulmonary vascular constriction and hemodynamic collapse soon after intravenous PAF are at least partially mediated by PAF-induced TxA₂ release. Tachyphylaxis to PAF influence has been observed in studies of leukocyte and platelet function. We hypothesized that tachyphylaxis to PAF might also occur in our studies of constrictor responses in pulmonary vessels. Recently, we have examined the capacity of PAF to produce sustained pulmonary vasoconstriction in openchested, anesthetized newborn piglets. Infusions sufficient to produce 100% increase in mean pulmonary artery pressure after 3 min showed no loss of efficacy when sustained for 30 min. The same was true for infusions of U-46619. Thus, the pulmonary vasoconstrictor influence of PAF or U-46619 is not readily diminished by tachyphylaxis. These findings favor the viewpoint that PAF or TxA₂ release during inflammatory processes could have prolonged adverse actions on pulmonary and systemic circulations. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989. The opinions or assertions contained here are those of the authors. They do not reflect the views of the Department of Defense or the Uniformed Services University of the Health Sciences. The experiments reported here were conducted according to the principles set forth in the “Guide for the Care and Use of Laboratory Animals,” Institute of Laboratory Animal Resources, U.S. Department of Health and Human Services (NIH Publ. 85-23, 1985).     

Effect of protein depletion on the VLDL triacylglycerol secretion and apoprotein synthesis by the perfused liver from pregnant rats

The effect of protein depletion in the pregnant rat on the polyunsaturated fatty acid incorporation into very low density lipoproteins (VLDL) has been investigated. The apoprotein pattern of these particles was determined. In in vivo experiments the amounts of serum and liver triacylglycerol were determined. VLDL were isolated and their apo C concentration calculated. In in vitro experiments the radioactivity of [³H] leucine incorporated into VLDL apoproteins was measured. The results show that protein depletion during pregnancy promotes a drastic increase in serum and liver triacylglycerol. The VLDL isolated from these animals show an increase in the triacylglycerol/protein ratio and a decrease in their content of apo C. Meanwhile, a significant reduction in the [³H]leucine incorporation into apo C peptides by the perfused liver of protein depleted rats was detected. On the other hand, protein deprivation did not affect labeled linoleic and arachidonic acid incorporation into triacylglycerol of the newly secreted VLDL. Taking these results together, let us deduce that a defective VLDL is secreted by the liver of the protein depleted pregnant rats. The abnormal composition of these particles may influence its normal metabolism through their effects on lipoprotein lipase and this fact could affect the normal supply of polyunsaturated fatty acids to the fetus.     

Biphasic action of platelet-activating factor on isolated guinea-pig ileum

1-0-Hexadecyl-2-0-acetyl-sn-glycero-3-phosphocholine (platelet-activating factor) at 10⁻¹⁰-10⁻⁹ M induced slow contraction of isolated guinea-pig ilcal muscles and the contraction persisted for a long time. At a higher concentration of 10⁻⁷ M, this phospholipid induced more rapid, but not greater, contraction. At higher concentrations (10⁻⁶-10⁻⁵ M), this phospholipid induced a biphasic response: rapid contraction followed by relaxation. At high concentrations, this compound inhibited acetylcholine-induced contractions. The stimulatory effect of this phospholipid was ca. 300 times that of 1-palmitoyl-2-0-acetyl-sn-glycero-3-phosphocholine, while its inhibitory potency on induced contraction was similar to those of 1-palmitoyl-2-0-acetyl-sn-glycero-3-phosphocholine and its lyso derivative. It was suggested that the differences in effects on contraction of different concentrations of 1-0-hexadecyl- and 1-palmitoyl-2-0-acetyl-sn-glycero-3-phosphocholine were due to the dual effects of these compounds on the ileum: a strong stimulatory effect and a moderate inhibitory effect on contraction.     

Significance of platelet-activating factor in mesenteric ischemia-reperfusion

Reperfusion of the ischemic mesenterium is frequently followed by acute circulatory collapse. This review focuses on the possible role of platelet-activating factor (PAF) in ischemia-induced damage. It provides evidence that (i) PAF concentrations are elevated in the mesenteric circulation following temporary ischemia; (ii) administration of exogenous PAF into the superior mesenteric vein mimics many events observed during reperfusion; and (iii) pretreatment of the experimental animals with specific PAF receptor antagonists prevent the circulatory collapse. These findings suggest that PAF may play an important role in the development of circulatory collapse caused by mesenteric ischemia-reperfusion. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

A sensitive and specific radioimmunoassay for platelet-activating factor

A platelet-activating factor (PAF) analog with a reactive ω-aldehyde group at thesn-1 position was synthesized. The hapten-thyroglobulin conjugate was used to immunize rabbits to produce specific antibodies to PAF. The purified immunoglobulin G (IgG) fraction was found to bind stereo-specifically to tritiated PAF and to crossreact minimally with lysoPAF, plasmalogens, and other phospholipids. The radioimmunoassay detected as little as 20 pg of PAF per assay tube and was used to explore agonist-induced synthesis of PAF in rabbit neutrophils. Calcium ionophore A23187 at 1 μM induced PAF synthesis peaking at 2 min and reaching basal levels after 5 min.N-Formyl-Met-Leu-Phe (FMLP) at 0.1 μM also stimulated rapid synthesis and degradation of PAF with a peak at 5 min. Both A23187 and FMLP stimulated PAF synthesis in a dose-dependent manner. The radioimmunoassay should be applicable to the quantitation of PAF in biological samples. Based on a paper originally presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Presence of platelet-activating factor in pyuria in humans

The relationship between the occurrence of platelet-activating factor (PAF) and neutrophils in urine from patients with urinary tract infection was examined. PAF was detected in human pyuria, when leukocyte levels reached at least 300 cells/μL (n=45), but not in normal urine (n=12). The amount of PAF found in pyuria, measured by platelet aggregation assay, was 0.01 to 13.3 pmol/mL. A close correlation was seen between the amount of PAF present and the number of urinary leukocytes (p<0.01, r=0.70). The leukocytes in pyuria consisted almost entirely of neutrophils (96±4%, mean ±S.D.). Our findings suggest that the occurrence of PAF is associated with the accumulation of neutrophils in urine. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Involvement of platelet-activating factor in zymosan-induced rat pleurisy

The role of platelet-activating factor (PAF) in inflammatory reactions was studied in zymosan-induced rat pleurisy. Pleurisy was induced by injection of a 2% zymosan suspension into the pleural cavity of rats. The time course of pleural exudate accumulation, the exudation rate, and exudate leukocyte numbers were followed then for 96 hr. Peak pleural exudate accumulation was about 3 mL at 24 hr, whereas the exudation rate increased biphasically with peaks at 0.5 hr and 5 hr. The migration of leukocytes into the pleural cavity increased with time up to 48 hr. The polymorphonuclear leukocytes were the dominant white cells in the exudate between 5 and 16 hr, but mononuclear leukocytes started to outnumber them around 24 hr. Pretreatment with cyproheptadine (5 mg/kg), an inhibitor of both histamine and seotonin, significantly suppressed pleural fluid accumulation and the exudation rate at 0.5 hr. The PAF antagonist CV-6209 (1 mg/kg) significantly suppressed pleural fluid accumulation and the exudation rate at both 0.5 and 5 hr. At either time point, the parameters were not suppressed by indomethacin. We detected PAF activity in the high-performance liquid chromatography (HPLC) fraction (with a retention time corresponding to that of authentic PAF) of the exudates at 0.5 hr, 5 hr, and 16 hr using an aggregation bioassay with washed rabbit platelets. The results suggest that in zymosan-induced rat pleurisy, histamine and/or serotonin are the main mediators of exudation at 0.5 hr and that PAF may be partly responsible for exudation at 0.5 hr and later at 5 hr to 16 hr. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Bioactions of 5-hydroxyicosatetraenoate and its interaction with platelet-activating factor

In a variety of stimulated cells, platelet-activating factor (PAF) and numerous arachidonate derivatives are coproducts that form as a consequence of receptor-mediated phospholipid mobilization. These lipid co-products produce a plethora of biological effects in a wide variety of cell systems. Furthermore, they often have a fascinating, although less widely appreciated, interaction. 5-HETE, at submicromolar concentrations, exerts relatively few direct bioactions. It does, however, potently (16–160 nM) raise cytosolic free calcium [Ca²⁺]_i and augment PAF-induced responses in human polymorphonuclear neutrophils (PMN) by as much as 100- to 1000-fold. 5-HETE acts on PMN by a structurally specific, stereospecific and pertussis toxin-inhibitable mechanism. In addition, PMN exposed to 5-HETE exhibit homologous but not heterologous desensitization. These findings suggest that 5-HETE, like PAF, may bind to its own specific plasmalemmal receptors to exert its unique set of bioactions. However, further investigation is required to demonstrate any putative 5-HETE receptors. Other potential mechanisms of 5-HETE-induced bioactions together with the possible effects of 5-HETE on PAF transduction mechanisms are also discussed. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Studies on the role of platelet-activating factor in blood pressure regulation

Circulating levels of 1-O-hexadecyl-2-acetyl-sn-glycero-3-phosphocholine (C₁₆PAF) in human subjects were measured by gas chromatography/mass spectrometry using negative ion chemical ionization. The mean (±S.D.) circulating C₁₆PAF levels in patients with essential hypertension (18.1±5.3 pg/mL, n=16) were not significantly different from those in normotensive subjects (17.2±7.2 pg/mL, n=14). During a salt balance study, high salt intake (20 g/day) significantly increased the circulating level of C₁₆PAF, and changes in circulating C₁₆PAF significantly and positively correlated with changes in mean arterial blood pressure (r=0.47, p<0.05). Changes in C₁₆PAF also correlated with changes in creatinine clearance (r=0.55, p<0.05), but did not correlate with changes in plasma sodium concentration, plasma chloride concentration and plasma volume. An intravenous injection of 50 μg of human atrial natriuretic peptide (hANP) decreased circulating C₁₆PAF levels from 20.0±2.7 to 13.9±2.4 pg/mL of blood (n=10, p<0.01) in healthy subjects. The data appear to indicate that C₁₆PAF levels are changed by salt intake-induced mild increase in blood pressure, and that hANP may be an endogenous factor which lowers circulating C₁₆PAF. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Acylation of lyso platelet-activating factor by splenocytes of the rainbow trout,Oncorhyncus mykiss

In mammalian systems, platelet-activating factor, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine, (PAF) is rapidly inactivated by a deacetylation/reacylation system that produces 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine which is highly enriched in arachidonic acid. There is some evidence that n−3 fatty acids may have an impact on this system in humans but the nature of this impact is unclear. In rainbow trout, n−3 fatty acids are known to be essential dietary components which are derived through the food chain. Substantial quantities of n−3 fatty acids are found in trout membrane phospholipids. We show here that in sharp contrast to mammalian cells, trout cells acylate lyso platelet-activating factor, alkyl-GPC, 1-O-alkyl-2-lyso-sn-glycero-3-phosphocholine, (lyso-PAF) with a high degree of specificity for n−3 fatty acids. When [³H]lysoPAF was incubated with these cells, only three molecular species of alkylacylglycerophosphocholine were produced, and 92% contained n−3 fatty acids. Since isolated membranes yielded similar results, it appears that the acylation proceedsvia a coenzyme A-independent transacylase as found in mammalian systems.     

Nephrotoxicity of cyclosporine: The role of platelet-activating factor and thromboxane

Cyclosporine (CsA), an immunosuppressive agent, is potentially nephrotoxic. We had previously observed that acute administration of CsA to Munich-Wistar rats induced a decrease in single nephron glomerular filtration rate, due to a decline in glomerular plasma flow, and in the glomerular ultrafiltration coefficient. Moreover, these alterations were prevented when an antagonist of platelet-activating factor (PAF) was administered. In the present study we examined whether the protective effect of the PAF blocker in CsA nephrotoxicity could have been mediated by thromboxane (TxA₂). Our data show that the PAF effects were not mediated by TxA₂, since administration of dazmegrel, a thromboxane synthetase inhibitor, did not ameliorate the acute renal failure caused by CsA. Thus, PAF appears to be a direct mediator of acute CsA nephrotoxicity, while TxA₂ is not significantly involved in this process. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Pooling of blood in postischemic shock is modulated by platelet-activating factor (PAF)

In experiments on dogs,i.v. administration of platelet-activating factor (PAF) (500 ng/kg) was shown to induce hypotension which, apart from decreased myocardial contractility, was characterized by blood pooling in veins (82.6±6.8 mL/kg). This was accompanied by restriction of venous return to the heart and reduction of cardiac output (CO). During postischemic shock the cardio- and hemodynamic disturbances were similar to those induced byi.v. administration of PAF. In the postischemic shock model, preliminary blockage of PAF receptors with the PAF receptor antagonist BN 52021 (6 mg/kg,i.v.) significantly decreased the amount of blood pooled in shock from 38.7±5 to 18.3±2 mL/kg (p<0.01). Simultaneously, the reduction of CO and blood pressure, induced by reperfusion of the continuously ischemized tissues of a rear limb, was less significant in pretreatedvs. the nontreated group. The data suggest that PAF may be involved in postischemic blood pooling and that PAF antagonists could be used to correct postischemic cardio- and hemodynamic disturbances. Based on a paper presented at the Third International Conference on Platelet-Activating Factor and Structurally Related Alkyl Ether Lipids, Tokyo, Japan, May 1989.     

Role of platelet-activating factor (PAF) in superoxide production by human polymorphonuclear leukocytes

The effect of platelet-activating factor (PAF) in superoxide production by human polymorphonuclear leukocytes (PMN) was studied. Cypridina luciferin analog (CLA) dependent chemiluminescence was used to detect superoxide anion radicals. PAF induced superoxide generation in human PMN in a dose-dependent manner. Preincubation with a small amount of PAF (5 x 10⁻⁹ M) enhanced PMN superoxide release induced by various stimuli, such as phorbol myristate acetate (PMA), opsonized zymosan (OZ), calcium ionophore (A23187) andN-formyl-methionyl-leucyl-phenylalanine (FMLP). The PAF antagonist, CV-6209, inhibited superoxide production induced by PAF, but not that induced by other stimuli. These findings would indicate that PAF may play an important role at inflammatory reaction sites and that CV-6209 may inhibit excessive inflammatory reaction.