Role of platelet-activating factor (PAF) in the initiation of the decidual reaction in the rat

Injection of PAF into the left uterine horn induced a dose-dependent decidua-like reaction in the pseudopregnant rat. This reaction was maximal when PAF was injected at Day 5 of pseudopregnancy and was blocked by the specific PAF antagonist, BN 52021. BN 52021 did not interfere with the decidual reaction induced by prostaglandin E-2 or insertion of a cotton thread in the uterine horn. In contrast, a decidua-like reaction was not evoked by the inactive lyso-PAF, demonstrating the specificity of the action of PAF. The decidua-like reaction induced by PAF involves the generation of cyclooxygenase metabolites of arachidonic acid since it was inhibited by indomethacin. The histological alterations induced by PAF were similar to those observed after embryo implantation, strengthening the postulate for a role of the autacoid in the early stages of pregnancy.     

Interference of the PAF-acether antagonist BN 52021 with endotoxin-induced hypotension in the guinea-pig

Because of the potential role of PAF-acether in the pathogenesis of endotoxin shock, we examined the preventive and curative effects of BN 52021, a new PAF-acether antagonist in guinea-pig challenged with S. Typhimurium endotoxin. A biphasic reduction of mean arterial pressure was elicited by i.v. endotoxin (300 micrograms/kg) in control animals, with a rapid drop of blood pressure (maximal decrease within 10 min), partial recovery at 20 min and a second gradual decrease after 30 min. Treatment with BN 52021 injected 15 min prior to endotoxin reduced the initial rapid drop of blood pressure from 38.5 +/- 5 mmHg in vehicle-treated controls (n = 15) to 17 +/- 3 mmHg (p less than 0.01) in animals treated with 1 mg/kg BN 52021(n = 10) and to 9.5 +/- 8 mmHg (p less than 0.01) in guinea-pigs treated with 6 mg/kg BN 52021 (n = 5). The early hypotensive phase was associated with severe thrombocytopenia-leukopenia; only the thrombocytopenia was reduced by BN 52021. The prolonged secondary phase of hypotension was reduced by BN 52021 pretreatment whereas a small increase of hematocrit persisted. The two phases of the arterial pressure profile during endotoxic shock were not observed in animals previously made thrombopenic by rabbit and anti-platelet serum and only the late hypotensive phase persisted. This late hypotension induced by endotoxin in thrombopenic animals was suppressed by BN 52021 pretreatment suggesting that BN 52021 may act via a platelet-independent mechanism. The intravenous injection of BN 52021 during the prolonged secondary phase of shock was followed by an immediate increase of the depressed blood pressure. This increase of blood pressure was dose-dependent, maximum at 6 mg/kg BN 52021, and observed in normal and thrombopenic animals. The interference of BN 52021 with endotoxin shock may be related to its PAF-acether antagonist properties and suggests that PAF-acether is an important participant in endotoxic shock.     

The effect of platelet-activating factor and its antagonist--BN 52021--on immune reactions in vivo in mice and in vitro

The effect produced by the injection of platelet activation factor (PAF) and its antagonist BN 52021 on the intensity of humoral immune response in (CBA x C57BL)F1 mice was studied. PAF was found to stimulate the formation of antibodies to sheep red blood cells. In addition PAF stimulated the phagocytic activity of mouse peritoneal macrophages. The stimulation of immune response under the action of PAF may be attributed to an increase in the phagocytic activity of macrophages. The stimulating effect of PAF on immune response in vivo was abolished by the injection of BN 52021, the antagonist of PAF. At the same time the dose-dependent decrease of immune response was observed after the injection of BN 52021. Indomethacin, an inhibitor of prostaglandin synthesis, when administered to mice treated with BN 52021, abolished the BN 52021-induced suppression of humoral immune response. Mouse peritoneal macrophages, treated in vitro with BN 52021, were found to produce significantly more prostaglandin E than control macrophages. Thus, BN 52021 induced the suppression of humoral immune response in vivo; this suppression was probably due to the action of prostaglandin E2, a messenger of the second order. Besides, the PAF antagonist BN 52021 significantly decreased leukotriene B4 production by macrophages in vitro. BN 52021 may be supposed to switch over the synthesis and/or secretion of arachidonic acid from the lipoxygenase pathway to the cycloxygenase one.     

Effect of cyclosporin A and the platelet-activating factor (PAF) antagonist, BN 52021, on PAF- and antigen- induced bronchoconstriction in the guinea-pig

The effects of the PAF antagonist, BN 52021, and cyclosporin A (CsA), either alone or in combination, on PAF- and antigen- induced bronchoconstriction were investigated in control and passively sensitized guinea-pigs, respectively. Although single administration of CsA alone has no effect on the PAF-induced bronchoconstriction, a marked inhibition of this phenomenon is observed when the drug is given along with an inactive dose of BN 52021. This effect of the association of the two drugs on the bronchoconstriction is also related to an action on the PAF-induced alterations in the number of leukocytes and platelets. In addition, administration of CsA for 48 hrs, which alone does not influence PAF-induced bronchoconstriction, markedly increases the inhibition evoked by BN 52021. Although bolus administration of CsA has no effect on the antigen -induced bronchoconstriction, a marked inhibition of this phenomenon is observed when the drug is given for 2 days. This inhibition by CsA is not further enhanced when the animals are also treated with BN 52021. These results strengthen the hypothesis that PAF and the immune system are involved in the regulation of bronchopulmonary reactions.     

The accumulation of platelet activating factor in the injured cornea may be interrelated with the synthesis of lipoxygenase products

The rabbit cornea accumulates platelet activating factor (PAF) three hrs after alkali burn. PAF was isolated by HPLC and assayed by platelet aggregation. This bioactivity was blocked by the PAF receptor antagonists BN 52021 and alprazolam. Added PAF increases the chemiluminescence response of the cornea in vitro and BN 52021 inhibits this effect. In vivo experiments show that the synthesis of 5-HETE and 12-HETE is inhibited by the PAF antagonist BN 52021. It is concluded that a metabolic interrelationship may exist between the PAF cycle and the lipoxygenation of arachidonic acid, and that drugs that affect these lipid mediators may modulate the inflammatory response of the anterior segment of the eye.     

Effect of platelet activating factor antagonist treatment on gentamicin nephrotoxicity

To assess whether PAF could be involved in the gentamicin-induced nephrotoxicity, we have studied the effect of PAF antagonist BN-52021 on renal function in rats after gentamicin (GENTA) treatment. Experiments were completed in 21 Wistar rats divided into three groups: group GENTA was injected with gentamicin 100 mg kg(-1) body wt/day s.c. for 6 days. Group GENTA + BN received gentamicin and BN-52021 i.p. 5 mg kg(-1) body wt/day. A third group served as control. Rats were placed in meta-bolic cages and plasma creatinine and creatinine clearance were measured daily. GENTA group showed a progressive increase in plasma creatinine, a drop in creatinine clearance and an increase in urinary excretion of N-acetyl-beta-D-glucosaminidase and alkaline phosphatase. GENTA + BN group showed a lesser change in plasma creatinine and a creatinine clearance, but no difference with GENTA group in urinary excretion of NAG and AP were observed. Histological examination revealed a massive cortical tubular necrosis in rats treated with gentamicin, whereas in BN-52021 injected animals tubular damage was markedly attenuated. The present results suggest a role for PAF in the gentamicininduced nephro-toxicity.     

Interactions between platelet activating factor and eicosanoids during endotoxic shock in anaesthetized pigs

The effects of platelet activating factor (PAF) on eicosanoid release during endotoxic shock was investigated in anaesthetized pigs receiving 5 mug kg(-1) Escherichia coli endotoxin (LPS) into the superior mesenteric artery over a 60 min period, by measuring plasma levels of a variety of mediators. Fifteen of the 31 animals infused with LPS and not treated with BN 52021, a PAF receptor antagonist, died within 30 min after the commencement of LPS infusion (non-survivors), while the other 16 survived the experimental period of 3 h, though in a state of shock (survivors). No alterations were observed in plasma concentrations of eicosanoids in the non-survivors. A significant, though transient, increase in eicosanoid concentrations occurred only in the survivors. Treatment with BN 52021 (4 mg kg-1, i.v.) injected 5 min prior to LPS infusion, failed to exert any effect on the survival rate. However, pretreatment with BN 52021 prevented circulatory collapse in the survivors and reduced the concentration of cyclooxygenase enzyme products, without affecting LTB(4) release. Exogenous administration of PAF (0.01 mug kg(-1)) caused hypotension and increased TXB(2) levels although 6-keto PGF(1alpha) and LTB(4) concentrations were unchanged. The data suggest that prostanoid formation may be secondary to PAF release in circulatory collapse evoked by LPS infusion in survivors, and give further support to the suggestion that PAF prostanoid interaction is important during endotoxic shock. However, their role in early death seems to be negligible, indicating the importance of other mediators.     

Effects of platelet activating factor (PAF) and its receptor antagonist BN 52021 on isolated perfused guinea-pig heart

The cardiac effects of PAF and its antagonist BN 52021 have been investigated on the isolated perfused guinea-pig heart maintained at a constant hydrostatic perfusion pressure of 80 cm water. In this model, PAF (1 x 10(-11) to 1 x 10(-7) moles) induced a dose-dependent coronary vasoconstriction, a decrease in heart rate and a fall in contractile force. BN 52021 (1 x 10(-6) to 2 x 10(-4) M) dose-dependently inhibited the vasospasm induced by PAF (1 x 10(-10) moles). BN 52021 also antagonized the decrease in coronary flow and heart rate, but not that of contractile force induced by a high dose of PAF (1 x 10(-7) moles). This dose of PAF also significantly (p less than 0.001) provoked a marked release of TxB2 but did not alter the generation of 6 Keto PGF1 alpha, PGE2 or LTC4. The PAF-induced increase in TxB2 release was completely abolished by BN 52021.     

Inhibition by BN 52021 (ginkgolide B) of the binding of [3H]-platelet-activating factor to human neutrophil granulocytes

The inhibitory effect of BN 52021, a specific antagonist of platelet-activating factor (PAF) on PAF-induced activation of human polymorphonuclear granulocytes (PMNL) and on the binding of [3H]-PAF to neutrophils were examined. BN 52021 over the range of 10(-9)-10(-4) M inhibited PAF-induced degranulation and superoxide production of PMNLs in a dose-dependent manner with Kd values of 0.6 +/- 0.1 x 10(-6) M and 0.4 +/- 0.1 x 10(-6) M, respectively. BN 52021 (up to 1 mM) did not show any agonistic activity and it did not affect neutrophil responses to N-formyl-methionyl-leucyl-phenylalanine or leukotriene B4. The Ki value of BN 52021 for the specific binding of [3H]-PAF to neutrophils was 1.3 +/- 0.5 x 10(-6) M versus a Ki of 1.1 +/- 0.3 x 10(-7) M for PAF itself. BN 52021 did not affect metabolism of PAF by PMNL. These studies indicate that BN 52021 inhibits neutrophil responses to PAF by inhibiting binding of PAF to its specific PMNL receptor.     

Characterization of specific binding sites for PAF in the iris and ciliary body of rabbit

The protective effect exerted by BN 52021 a specific PAF-receptor antagonist in experimentally induced ocular inflammatory disorders led us to investigate the possible presence of specific receptors for PAF in rabbit iris and ciliary body. Two classes of PAF binding sites were found in isolated iris and ciliary process of pigmented rabbit eyes: a high affinity site Kd1 congruent to 4.9 +/- 0.47 nM, Bmax1 congruent to 3.17 +/- 0.50 pmoles/mg protein, a low affinity sites Kd2 congruent to 11.6 +/- 0.33 nM, Bmax2 congruent to 12.46 +/- 2.3 pmoles/mg protein for iris. The specific binding was not affected by lyso-PAF the biologically inactive precursor and metabolite of PAF, up to 10(-6) M; inhibition by unlabelled PAF demonstrated a biphasic curve partially antagonized by BN 52021. The present results demonstrate the presence of specific binding sites for PAF in rabbit eyes which could mediate the action of this mediator in eye inflammatory processes and explain the protective effect observed with BN 52021.     

A study with BN 52021 demonstrates the involvement of PAF-acether in IgE-dependent anaphylactic bronchoconstriction

The involvement of platelet activating factor (PAF) in anaphylaxis was examined by recording the pulmonary responses of anesthetized passively sensitized guinea-pigs to the aerosolization of ovalbumin. Animals were tested with and without BN 52021 (a ginkgolide B, PAF receptor antagonist) pretreatment. Aerosolization of ovalbumin produced a bronchoconstriction (BC) which could be made refractory to additional challenges with the antigen. In animals desensitized to ovalbumin, aerosolization of PAF produced an unattenuated BC. Guinea pigs desensitized by repeated aerosolizations of PAF subsequently showed reduced responses to aerosolized antigen suggesting that PAF may be involved in the BC. Animals pretreated with BN 52021, were protected against the effects of systemically administered PAF and also showed reduced responses to aerosolized antigen. Aerosolization of the leukotriene antagonist, FPL 55712, was ineffective against anaphylactic BC under conditions where catecholamine and histamine release were blocked.     

Antagonism of PAF-induced death in mice

The ability of three platelet activating factor (PAF) antagonists, BN52021, L652, 731 and 48740RP, and the leukotriene antagonist FPL55712 to block iv PAF-induced death was tested in mice. PAF-induced sudden death was been previously characterized as a model of systemic anaphylaxis and circulatory shock related its hypotensive actions. Of the drugs, BN52021 and L652, 731 provided dose-dependent protection against PAF toxicity, whereas the others had no effect. 48740RP was, however active against PAF-induced rabbit platelet aggregation. BN52021 was inactive in three other mouse sudden death models in which arachidonic acid, U46619 or collagen combined with epinephrine is injected iv to provoke a thrombotic/ischemic sudden death. In contrast, the TXA₂ antagonist SQ29548 inhibited the acute toxicity of two of these latter challenges (arachidonic acid and thromboxane agonist U46619), but was inactive against PAF lethality.These results suggest that PAF toxicity in mice is a specific model for PAF agonism, and is not mediated by TXA₂ or peptido-leukotrienes. Further, PAF-induced mortality should be a simple and useful technique for testing potential PAF antagonists for activity by various routes of administration.     

Inhibition of human lymphocyte proliferation and interleukin 2 production by platelet activating factor (PAF-acether): reversal by a specific antagonist, BN 52021

When added to a 72 h culture of human peripheral blood mononuclear leukocytes stimulated with phytohemagglutinin, PAF-acether caused a significant inhibition (40-65%) of proliferation at concentrations of 10(-8) to 10(-6) M. This inhibition was reversed by the specific PAF antagonist, BN 52021. It was also reversed by indomethacin, suggesting that PAF-acether mediated this suppression via cyclooxygenase metabolites of arachidonic acid. IL-2 production, measured at 24 h of lymphocyte proliferation, was similarly impaired (50-66%) by 10(-8)-10(-6) M PAF-acether. IL-2 production was brought up to 90% of control values when both PAF-acether and BN 52021 (10(-4) M) were added together to the lymphocyte cultures. These studies suggest a significant immunoregulatory role for PAF-acether and a potential use of BN 52021 as a biological response modifier.     

A study with BN 52021 demonstrates the involvement of PAF-acether in IgE-dependent anaphylactic bronchoconstriction

The involvement of platelet activating factor (PAF) in anaphylaxis was examined by recording the pulmonary responses of anesthetized passively sensitized guinea-pigs to the aerosolization of ovalbumin. Animals were tested with and without BN 52021 (a ginkgolide B, PAF receptor antagonist) pretreatment. Aerosolization of ovalbumin produced a bronchoconstriction (BC) which could be made refractory to additional challenges with the antigen. In animals desensitized to ovalbumin, aerosolization of PAF produced an unattenuated BC. Guinea pigs desensitized by repeated aerosolizations of PAF subsequently showed reduced responses to aerosolized antigen suggesting that PAF may be involved in the BC. Animals pretreated with BN 52021, were protected against the effects of systematically administered PAF and also showed reduced responses to aerosolized antigen. Aerosolization of the leukotriene antagonist, FPL 55712, was ineffective against anaphylactic BC under conditions where catecholamine and histamine release were blocked.     

Platelet-activating factor: an inflammatory mediator in the acute phase of allergic conjunctivitis in a guinea-pig model

The role of platelet-activating factor (PAF) as a mediator of increased conjunctival vascular permeability was investigated in a guinea-pig model of immediate hypersensitivity. Vascular permeability of the conjunctiva was determined by measuring the albumin content in lavage fluid (LF) after topical challenge with either PAF or ovalbumin. PAF produced a dose-dependent increase of the vascular permeability within minutes. Topical pretreatment with levocabastine, a potent histamine H(1)-antagonist demonstrated no effect towards the vascular permeability in response to PAF provocation. Pretreatment with eyedrops containing the specific PAF antagonist BN 52021 (1%) showed a significant inhibition of the vascular permeability (60.2%) and the clinical score (27.5%) after PAF challenge. In sensitized guinea-pigs, levocabastine showed a marked inhibition of both the vascular permeability (80.5%) and the clinical score (70%) after topical challenge with ovalbumin. BN 2021, although to a lesser extent, showed a similar effect towards the vascular permeability (26.8%) and the clinical score (28%) after antigen provocation. When BN 52021 and levocabastine were administered in combination, the vascular permeability was significantly decreased after antigen challenge in comparison with eyes pretreated with levocabastine alone. These results indicate that PAF plays a role in the acute phase of allergic conjunctivitis in the guinea-pig.     

Effects of a platelet activating factor antagonist (BN 52021) on free fatty acids, diacylglycerols, polyphosphoinositides and blood flow in the gerbil brain: inhibition of ischemia-reperfusion induced cerebral injury

The effects of a platelet activating factor antagonist (BN 52021) on cerebral ischemia-reperfusion was studied in the gerbil. Following ten minutes of bilateral carotid artery ligation, gerbils were reperfused and injected intraperitoneally with either BN 52021 or vehicle-dimethylsulfoxide. Cerebral blood flow and systemic arterial pressure were monitored until 90 minutes of reperfusion. Free fatty acids, diacylglycerols and polyphosphoinositides were then analyzed in forebrains and midbrains. BN 52021 inhibited the maturation of ischemic injury. Cerebral blood flow increased following 60 to 90 minutes of reperfusion. Free fatty acid levels were reduced likely by inhibition of phospholipase A. Phospholipase activity may likely be decreased since there was a tendency to increase phosphatidylinositol-4',5'-bisphosphate and diacylglycerols in BN 52021-treated animals.     

Failure of platelet-activating factor (PAF-acether) to induce decidualization in mice and failure of antagonists of PAF to inhibit implantation

The possible role of platelet-activating factor (PAF) in the uterine responses associated with implantation was investigated. Attempts to trigger a decidual cell response in the uteri of hormonally sensitized, ovariectomized mice by instilling PAF-acether (1-1000 ng) intraluminally were unsuccessful. The effect of PAF antagonists on implantation was investigated in females ovariectomized on Day 3 of pregnancy and treated with progesterone. Implantation was induced in these females by injection of 10 ng oestradiol-17 beta on Day 8. Hourly intraperitoneal injections of three PAF antagonists (WEB 2086, CV 3988 and BN 52021 at doses of 1.2-1.4 mg/kg) given over a 24-h period starting 1 h before the injection of oestradiol-17 beta had no significant effect on the occurrence of implantation sites. Intraluminal injection of WEB 2086 (15 micrograms) or BN 52021 (5 micrograms) either 3 h before or 6 h after the nidatory oestradiol also had no significant inhibitory effect on implantation. SRI 63-441 given once daily over the first 4 days of pregnancy at a dose of 40 micrograms/30 g body weight had no inhibitory effect on the establishment of pregnancy. These results are not consistent with a critical role for PAF in implantation in mice.     

Role of platelet-activating factor (PAF) in the bronchopulmonary alterations and beta-adrenoceptor function induced by endotoxin

The possibility that PAF is implicated in the alterations of the beta-adrenoceptor function observed during endotoxemia was investigated. Lung parenchymal strips (LPS) from endotoxin-treated guinea-pig demonstrated specific desensitization to low doses of PAF whereas the contractions induced by histamine and leukotriene D4 were slightly affected. In addition, histamine-contracted LPS from endotoxin-injected animals exhibited decreased responsiveness to isoproterenol, a phenomenon not observed with guinea-pigs also treated with the specific PAF antagonist, BN 52021. No alteration of the sensitivity to isoproterenol of LPS preincubated with PAF was noted, suggesting an indirect effect of the autacoid on beta-adrenoceptor function.     

Effects of platelet activating factor antagonists on arachidonic acid-induced platelet aggregation and TXA2 formation

The effects of the PAF receptor antagonists WEB 2086, WEB 2170, BN 50739 and BN 52021 on AA-induced platelet aggregation (PA) and TXA2 formation were investigated in comparison with the TXA2 synthetase inhibitor HOE 944 and the TXA2 receptor antagonist BM 13.177. All PAF antagonists tested were weak inhibitors of AA-induced PA and TXA2 formation (IC50 values between 80 and 2,737 mumol/l). HOE 944 was effective in concentrations 2-3 orders of magnitude lower than PAF antagonists in inhibiting TXA2 generation. These results imply that the inhibition of TXA2 formation is of minor relevance for the actions of the investigated PAF antagonists in AA-induced PA.     

Involvement of platelet-activating factor (PAF) in endotoxin-induced intestinal motor disturbances in rats

Intestinal myoelectrical activity was investigated in conscious fasted rats chronically implanted with Nichrome electrodes in the duodeno-jejunum. Motility of the small intestine was characterized by the presence of migrating myoelectric complex (MMC) occurring regularly at 16.2 +/- 5.8 minute intervals. Intravenous administration of endotoxin (E. coli S.0111:B4) at a dose of 50 micrograms/kg increased the interval between MMC to 112.6 +/- 26.8 min, the duration of these effects being dose-related between 10 to 100 micrograms/kg. Such a typical myoelectrical alteration, corresponding to rapidly propagated groups of spike bursts, was mimicked by the IP administration of PAF at doses of 10 to 50 micrograms/kg. Previous administration of BN 52021, a specific PAF antagonist at a dose of 50 mg/kg abolished the motor alterations induced by IP injection of PAF (25 micrograms/kg) and significantly (p less than 0.01) reduced by 61.2% those induced by IV endotoxin (50 micrograms/kg). Indomethacin (10 mg/kg IP) as well as SC 19220 (5 mg/kg IV), a PGE2 antagonist, injected prior to endotoxin (50 micrograms/kg IV) or PAF (25 micrograms/kg IP) also reduced significantly (p less than 0.01) the duration of MMC inhibition. It is concluded that endogenous release of PAF is partly responsible for the intestinal motor alterations induced by endotoxin; these effects, strongly reduced after treatment with BN 52021, are also mediated through the release of prostaglandins.