The tumour necrosis factor superfamily ligand APRIL (TNFSF13) is released upon platelet activation and expressed in atherosclerosis.

Activated platelets release a wide range of inflammatory mediators, including members of the tumour necrosis factor (TNF) superfamily (e.g. CD40 ligand [CD40L] and LIGHT). Such platelet-mediated inflammation could be involved in atherogenesis and plaque destabilisation. In the present study we investigated whether APRIL, another member of the TNF superfamily that has been detected in megakaryocytes, could be released from platelets upon activation. The release of APRIL was studied in thrombin receptor (SFLLRN) activated platelets, and the expression of APRIL was examined in plasma and within the atherosclerotic lesion in patients with carotid and coronary atherosclerosis. Upon SFLLRN activation, there was a gradual release of APRIL, reaching maximum after 90 minutes. While this pattern is similar to that of CD40L and LIGHT, the release of APRIL was quite differently regulated. Thus, prostaglandin E1, but not inhibitors of metal-dependent proteases and actin polymerisation or the lack of GP IIb/IIIa, blocks APRIL release in activated platelets. With relevance to atherogenesis, we found that patients with coronary artery disease (n=80) had raised plasma levels of APRIL as compared with controls (n=20), and APRIL immunoreactivity was detected in aggregated platelets within the ruptured plaque in patients with myocardial infarction and within macrophages in symptomatic carotid plaques. In conclusion, activated platelets release significant amounts of APRIL in a long-lasting manner, differently regulated than the gradual release of other platelet-derived TNF superfamily ligands. The enhanced expression of APRIL in atherosclerotic disorders, both systemically and within the lesion, may suggest a potential involvement of APRIL in atherogenesis.

Prostaglandins A1 and E1 influence gene expression in an established insect cell line (BCIRL-HzAM1 cells).

Prostaglandins (PGs) and other eicosanoids exert important physiological actions in insects and other invertebrates, including influencing ion transport and mediating cellular immune defense functions. Although these actions are very well documented, we have no information on the mechanisms of PGs actions in insect cells. Here we report on the outcomes of experiments designed to test our hypothesis that PGs modulate gene expression in an insect cell line established from pupal ovarian tissue of the moth Helicoverpa zea (BCIRL-HzAM1 cells). We treated cells with either PGA(1) or PGE(1) for 12 or 24h then analyzed cell lysates by 2-D electrophoresis. Analysis of the gels by densitometry revealed substantial changes in protein expression in some of the protein spots we analyzed. These spots were processed for mass spectrometric analysis by MALDI TOF/TOF, which yielded in silico protein identities for all 34 spots. The apparent changes in three of the proteins were confirmed by semi-quantative PCR, showing that the changes in mRNA expression were reflected in changes in protein expression. The 34 proteins were sorted into six categories, protein actions, lipid metabolism, signal transduction, protection, cell functions and metabolism. The findings support the hypothesis that one mechanism of PG action in insect cells is the modulation of gene expression.

Mechanisms of strain-dependent development of mast cells from mouse splenocytes.

Mast cell development from spleen cells was not triggered by prostaglandin E1 (PGE1) or dibutyryl cAMP (db-cAMP) during a 12 day culture when the spleen cells were obtained from C57BL/6N and DBA/1 mice, but mast cells did develop when the spleen cells were obtained from C3H/HeN, BALB/c and ICR mice. A lack of endogenous IFN-gamma in the initial 2 days of the culture period was responsible for the failure. This was confirmed by adding neutralizing anti-IFN-gamma antibody and rIFN-gamma to the cultures and by determining IFN-gamma levels in the spleen cell cultures. Th1 cells in the spleens of C57Bl and DBA/1 mice were much more sensitive to PGE1 and db-cAMP than Th1 cells from other inbred mice strains, and consequently, IFN-gamma production was inhibited in spleen cell cultures of C57BL and DBA/1 mice on addition of PGE1 or db-cAMP. Furthermore, the different sensitivities of Th1 cells to PGE and db-cAMP were dependent on the different levels of IL-12 p40 monomers or homodimers in the spleen cell cultures. As the endogenous specific inhibitors of IL-12 p70 (heterodimers of p40 and p35), large amounts of IL-12 p40 monomers or homodimers in the spleen cell cultures of C57BL and DBA/1 mice enhanced the ability of PGE1 and db-cAMP to inhibit IFN-gamma production by antagonizing the activity of IL-12 heterodimers. These results indicate that the strain-dependent development of mast cells from mouse splenocytes is related to endogenous IFN-gamma levels, which are regulated by PGE, db-cAMP, IL-12 p70 and IL-12 p40.

Prostaglandin E2 is a potent regulator of interleukin-12- and interleukin-18-induced natural killer cell interferon-gamma synthesis.

Synthesis of interferon (IFN)-gamma by natural killer (NK) cells is an important pro-inflammatory event with interleukin (IL)-12 and IL-18 playing major inductive roles. However, other temporal events are likely to regulate such processes and as prostaglandin E2 (PGE2) is ubiquitous during inflammation this study tested the hypothesis that PGE2 was capable of directly modulating cytokine-induced NK cell IFN-gamma synthesis in the absence of other immune cells. Using homogeneous NK cell lines to establish direct effects, PGE2 (0.1-1 micro m) was found to suppress NK cell IFN-gamma synthesis and antagonized the potent synergistic IFN-gamma-inducing effects of IL-12 and IL-18. The actions of PGE2 were mimicked by synthetic PGE2 analogues including misoprostol and butaprost. The selective EP2 receptor agonist butaprost, but not the EP1/EP3 agonist sulprostone, suppressed IFN-gamma synthesis and exclusively competed with PGE2 for receptor binding on NK cells. Further analysis showed that PGE2 did not modulate IL-12 receptor mRNA expression and the effects of PGE2 could be mimicked by the phosphodiesterase inhibitor 3-iosobutyl-1-methylxanthine. The absence of demonstrable receptor modulation coupled with the observed suppression of IFN-gamma synthesis by both EP2 receptor-selective agonists and IBMX suggest that PGE2 acts directly on NK cells via EP2 receptors with its downstream effects on cAMP metabolism. This conclusion is further supported by findings that PGE2 and its analogues consistently elevated levels of cAMP in NK cells. The ability of PGE2 to antagonize the potent inductive signal provided by the combination of IL-12 and IL-18 supports the concept that PGE2 may play an important role in limiting innate inflammatory processes in vivo through direct suppression of NK cell IFN-gamma synthesis.

Glucocorticoid involvement in suppression of NK activity following surgery in rats.

We studied plasma factors mediating suppression of NK activity (NKA) following surgery. Plasma from operated rats suppressed NKA of splenocytes, leukocytes, and purified natural killer (NK) cells, and charcoal stripping nullified suppression. The glucocorticoid antagonist mifepristone prevented suppression, whereas blockers of reactive oxygen metabolites, opioids, catecholamines, prostaglandin-E2, and histamine did not. NKA dropped as corticosterone levels peaked postoperatively, and administration of relevant doses of corticosterone suppressed NKA. Inhibition of glucocorticoid synthesis prevented plasma from suppressing NKA but merely attenuated NKA suppression in operated rats. Thus, postoperative concentrations of corticosterone can directly suppress NKA but additional factors probably act in vivo.

The modulatory effects of prostaglandin-E on cytokine production by human peripheral blood mononuclear cells are independent of the prostaglandin subtype.

The production of inflammatory mediators, relevant to (auto)immune diseases and chronic inflammatory conditions, can be modulated by dietary intake of n-3 and n-6 long chain polyunsaturated fatty acids (PUFAs). It was suggested that these effects are related to the formation of different series of eicosanoids, in particular prostaglandin-E (PGE). In this study we investigated whether prostaglandin subtypes metabolized from arachidonic acid (PGE2), dihomo-gamma-linolenic acid (PGE1) or eicosapentaenoic acid (PGE3) have different effects on T-cell proliferation and cytokine production in vitro. Freshly isolated human peripheral blood mononuclear cells (PBMC) were stimulated with concanavalin A (ConA) or lipopolysaccharide (LPS) in the presence or absence of exogenous PGE1, PGE2 or PGE3. We found that tumour necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma) and to a lesser extent interleukin (IL)-10 production was inhibited by all PGE-subtypes in ConA-stimulated PBMC concomitant with unaffected IL-2 levels. The modulated cytokine production of ConA stimulated cells was independent of T-cell proliferation. PGE2 and PGE1 moderately stimulated proliferation, while PGE3 inhibited the proliferative response to some extent. In LPS-stimulated PBMC, TNF-alpha production was inhibited by all PGE-subtypes, whereas IL-6 remained unaffected and IL-10 production was increased. Time course experiments on the effects of PGE-subtypes on cytokine production after ConA or LPS stimulation showed these effects to be time dependent, but indifferent of the prostaglandin subtype added. Overall, the modulatory effects of PGE on cytokine production were irrespective of the subtype. This may implicate that the immunomodulatory effects of PUFAs, with respect to cytokine production, are not caused by a shift in the subtype of PGE.

A multi-centre, double-blind, placebo-controlled study of liposomal prostaglandin E1 (TLC C-53) in patients with acute respiratory distress syndrome.

OBJECTIVE: To evaluate the safety of liposomal PGE1 (TLC C-53) in patients with acute respiratory distress syndrome (ARDS), and determine its efficacy in improving oxygenation and reducing ventilator dependency. DESIGN: A multi-centre, randomized, double-blind, placebo-controlled clinical study. SETTING: Thirty-one hospitals in six European countries. PATIENTS: One hundred two patients with ARDS. INTERVENTIONS: Patients were randomized in a 2:1 ratio to receive infusions of either the study drug TLC C-53 or placebo. Infusions were given over 60 min every 6 h for 7 days. The dose of study drug started at 0.6 microg/kg per h, rising over 24 h to a maximum dose of 1.8 microg/kg per h. MEASUREMENTS AND MAIN RESULTS: Seventy patients received the study drug and 32 placebo. Sixty-nine patients (47 treatment, 22 placebo) completed the study protocol. Patients were monitored for changes in the PaO2/FIO2 ratio, changes in lung compliance, time to off-ventilator and 28-day mortality, in addition to basic haematological and haemodynamic parameters. There were no significant differences in demographics and baseline characteristics between the two groups. There were no differences in the time to off-ventilation (16 days with treatment, 16.6 days with placebo, p=0.94) or in 28-day mortality (30% with treatment, 28% with placebo, p=0.78). There was a difference in the time to achieve a PaO2/FIO2 ratio above 300 in favour of TLC C-53 (10.3 versus 26.5 days) but this was not statistically significant (p=0.23). CONCLUSIONS: TLC C-53 was generally well-tolerated but failed to reduce mortality or duration of mechanical ventilation.

Histamine and prostaglandin interaction in the central regulation of ACTH secretion.

Since prostaglandins (PGs) and histamine (HA) seem to be involved in the activation of the hypothalamic-pituitary-adrenal axis following immunochallenges with lipopolysaccharide (LPS) endotoxin and cytokines, we investigated whether the histaminergic and eicosanoid systems in the male rat brain interact in their regulation of ACTH secretion. The PG synthesis inhibitor indomethacin (10 mg/kg i.p.) attenuated the ACTH response to LPS (10 micrograms/kg i.p.) and HA (30 micrograms i.c.v.). Infusion of PGE1, PGE2 or PGF2 alpha (1 and/or 5 micrograms infused intracerebroventricularly) stimulated ACTH secretion. The ACTH response to PGE1 was inhibited by the HA synthesis inhibitor alpha-fluoromethyl-histidine and the H1 receptor antagonist mepyramine (MEP) but not the H2 receptor antagonist cimetidine (CIM). Neither MEP nor CIM affected the ACTH response to PGE2. MEP and CIM attenuated the ACTH response induced by PGF2 alpha. The findings indicate that HA and some PGs in the brain interact in their stimulatory regulation of ACTH secretion. Such an interaction may also be involved in their mediation of the ACTH response to immunochallenges.

Prostaglandin E1 protects against ischemia-reperfusion injury of the liver by inhibition of neutrophil adherence to endothelial cells.

BACKGROUND: This study investigates the protective mechanism of prostaglandin E1 (PGE1) against hepatic ischemia-reperfusion injury in vivo. It has been demonstrated that activated leukocytes contribute to ischemia-reperfusion injury, and that administration of the monoclonal antibody (mAb) for adhesion molecules reduces the injury by inhibiting leukocyte-endothelial cell adhesion. We therefore attempted to find out whether PGE1 has an effect on the inhibition of neutrophil adherence to endothelial cells after reperfusion. METHODS: We administered anti-intercellular adhesion molecule 1 (ICAM-1) mAb, antiserum against rat polymorphonuclear leukocytes, or PGE1 to a rat model of left lobar ischemia for 60 min followed by reperfusion. Leukocyte adherence was observed by intravital fluorescence microscopy. The effect of PGE1 on the expression of adhesion molecules was analyzed by immunohistochemistry and flow cytometry. RESULTS: Ischemia-reperfusion caused endothelial dysfunction and hepatocellular injury with leukostasis in postsinusoidal venules. Anti-ICAM-1 mAb administration or leukopenia ameliorated both the hepatocellular injury and endothelial dysfunction. Although PGE1 administration did not affect the serum interleukin-8 level, it significantly decreased hepatic injury and leukostasis in the reperfused liver. Immunohistochemical findings showed that PGE1 decreased ICAM-1 expression on endothelial cells, but did not affect lymphocyte function-associated antigen 1, and membrane attack complex 1 on neutrophils in flow cytometric analysis. CONCLUSIONS: We conclude that PGE1 protects the liver against ischemia-reperfusion injury by reducing leukocyte-endothelial cell adhesion via down-modulation of ICAM-1 expression on the endothelium.

[Nonspecific anaphylactoid reaction in low IgE patients].

We reported two cases of anaphylactoid reaction caused by ranitidine and alprostadil. We suggest that these anaphylactoid reactions are nonspecific reactions which are observed in low IgE patients, because plasma IgE was still low 6 weeks after allergic reaction and these drugs have not been given to them before and eosinophilic and basophilic cells showed no change at all. We have to consider that there is a high risk of allergic reaction during general anesthesia, because many drugs are injected intravenously during general anesthesia.

Granulomatous hypersensitivity to Schistosoma mansoni egg antigens in human schistosomiasis. III. In vitro granuloma modulation induced by immune complexes.

Granulomatous hypersensitivity to parasite eggs of Schistosoma mansoni is an important factor in the development of morbidity in chronic schistosomiasis. It has been demonstrated previously that the chronic, well-tolerated, intestinal form of schistosomiasis is associated with the establishment and maintenance of a variety of immunoregulatory mechanisms. We have used an in vitro model of granuloma formation for the purpose of studying the regulation of granulomatous hypersensitivity to S. mansoni egg antigens, mediated by immune complexes (IC). Our results show that the peripheral blood mononuclear cells (PBMCs) from patients with active schistosome infections, when treated with sera from chronic schistosomiasis patients, were able to induce an inhibitory activity on in vitro granuloma formation. Significant modulation of the in vitro granuloma reaction remained after treatment of PBMCs with isolated IC or manufactured IC with soluble egg antigen (SEA) and purified IgG from pooled chronic schistosomiasis sera. In contrast to granuloma modulation stimulated with whole molecule IgG-SEA IC, the incubation of PBMCs with F(ab')2 IgG-SEA IC did not induce any suppression of the granulomatous hypersensitivity to SEA. It appears in this model system that IC may inhibit the activity of granuloma formation by stimulating macrophages to release suppressive mediators. We have demonstrated this possibility by inhibition of prostaglandin activity using indomethacin. The addition of indomethacin to the granuloma culture significantly reduced in vitro granulomatous hypersensitivity to S. mansoni eggs in patients with chronic intestinal schistosomiasis and do so by inducing macrophages to secrete prostaglandins.

Evidence that circulating 6keto prostaglandin E1 causes the platelet defect of Bartter's syndrome.

Bartter's syndrome is associated with activation of prostaglandin metabolism. In the present study we provide several lines of evidence that a circulating metabolite of prostacyclin, 6ketoPGE1 is responsible for a defect in platelet function present in patients with Bartter's syndrome. In platelet aggregometry studies, plasma from patients contained platelet inhibitory activity which was fully neutralized by coincubation with antibody directed against 6ketoPGE1. Fractionation of lipophilic extracts of plasma by high pressure liquid chromatography yielded a platelet inhibitory fraction which comigrated with authentic 6ketoPGE1 and was neutralized by anti 6ketoPGE1 antibody. Lastly, direct measure of the plasma concentration of 6ketoPGE1 by specific radioimmunoassay indicates a 2-fold increase in patients with Bartter's syndrome (133 +/- 9.1 vs 60.7 +/- 12.3 picograms/ml; p less than 025). These studies provide firm evidence that the platelet dysfunction present in patients with Bartter's syndrome is attributable to an increase in the plasma concentration of 6ketoPGE1. In addition, these data provide further evidence in support of the centrality of activation of prostaglandin metabolism in the pathophysiology of Bartter's syndrome.

Identification of 6-keto-prostaglandin E1 obtained from isolated perfused kidney of the rabbit.

We studied the release of 6-keto-prostaglandin (PG) E1-like material into urinary and venous effluents of the isolated perfused kidney of the rabbit. After high performance liquid chromatographic (HPLC) separation, material coeluting with authentic 6-keto-PGE1 was measured by radioimmunoassay, platelet antiaggregation bioassay and characterized further by gas chromatography/mass spectrometry-single ion monitoring (GC/MS-SIM). Injection of either arachidonic acid or ATP into the renal artery stimulated the release of prostacyclin [measured as immunoreactive (i)-6-keto-PGF1 alpha] and i-6-keto-PGF1. HPLC fractions containing i-6-keto-PGE1, and coeluting with 6-keto-PGE1 standard, exhibited potent inhibition of platelet aggregation. The presence of authentic 6-keto-PGE1 was verified by the GC/MS-SIM spectra in HPLC zones from which radioimmunoassayable and bioassayable 6-keto-PGE1-like material was recovered.

Effect of Rumex nepalensis extracts on histamine, acetylcholine, carbachol, bradykinin, and PGs evoked skin reactions in rabbits.

The antihistaminic, anticholinergic, antibradykinin, and/or antiprostaglandin activity of aqueous and the alcoholic leaf extracts of the plant Rumex nepalensis were tested on the prepared site on the back of rabbits, using a control and a test site. Aqueous extract was found to reduce the mean size of the wheal produced by histamine, acetylcholine, carbachol, and bradykinin. The alcoholic extract reduced the size of the wheal produced by histamine, acetylcholine and carbachol. The results indicate that Rumex nepalensis may have antihistaminic, anticholinergic, and/or antibradykinin activity.