Inhibitory Effect Of Prostaglandin E2 Research Articles

The inhibitory effect of prostaglandin E1 on oxidative stress-induced hepatocyte injury evaluated by calpain-mu activation.

Prostaglandin E1 (PGE1) is known to inhibit ischemia-reperfusion injury of the liver. The calcium-dependent neutral proteinase, calpain-mu, is involved in oxidative stress-induced hepatocyte injury. We investigated the mechanisms of cytoprotection by PGE1, focusing on the elevation of intracellular calcium ([Ca2+]i), activation of calpain-mu, and calpain-mu-mediated activation of protein kinase C-alpha (PKC-alpha). Cultured hepatocytes were treated with various amounts of PGE1 (0, 0.1, 1.0, 10, and 100 ng/ml) for 30 min and subsequently with 0.5 mM tert-butyl hydroperoxide (TBHP). Cell injury was evaluated by the release of lactate dehydrogenase. Plasma membrane bleb formation was examined by phase contrast microscopy. Activation of calpain-mu and limited degradation of PKC-alpha was evaluated by Western blotting using antibodies that specifically recognize the amino-terminal regions of calpain-mu and PKC-alpha. [Ca2+]i was measured by confocal microscopy using Fluo-3AM. LDH release from cells treated with 10 ng/ml PGE1 was significantly lower than from untreated cells (135 +/- 12 vs. 258 +/- 18 IU/L, respectively; P < 0.05). Morphologically, many blebs were observed in untreated cells, but very few were seen in those treated with 10 ng/ml PGE1. Western blotting revealed that the amount of activated calpain-mu and [Ca2+]i increased up to 1,300 nM at 35 min after the addition of TBHP (0.5 mmol/L) in control experiments (without PGE1). PGE1 (10 ng/ml) delayed the rise in [Ca2+]i for about 30 min, but did not suppress it completely. PKC-alpha decreased in experiments using PGE1 (10 ng/ml). PGE1 exerts its cytoprotective effect in TBHP-induced hepatocyte injury partly by inhibiting Ca2+-calpain-mu-mediated mechanisms.

Prostaglandin E2 inhibits spontaneous inhibitory postsynaptic currents in rat supraoptic neurones via presynaptic EP receptors.

Prostaglandin E2 (PGE2) has been implicated in the excitatory regulation of magnocellular neurones in the supraoptic nucleus (SON). We have recently reported that PGE2 excited SON neurones by directly activating postsynaptic PGE2 receptors (EP receptors) of a subclass other than EP1-3, but did not affect excitatory postsynaptic currents (EPSCs). In the present study, we examined presynaptic effects of PGE2 on rat SON neurones by measuring spontaneous inhibitory postsynaptic currents (IPSCs) by a slice patch-clamp technique. PGE2 inhibited spontaneous IPSCs in a dose-dependent and reversible manner. PGE2 selectively suppressed the frequency of IPSCs without affecting the amplitude of IPSCs in the presence of tetrodotoxin, a blocker of Na+ channels, indicating that the effects were presynaptic. The inhibitory effects of PGE2 on the frequency of IPSCs were mimicked by the EP1/EP3 agonists, 17PT-PGE2 and sulprostone, and the EP2/EP3 agonist, misoprostol, whereas the EP2 agonist, butaprost, or the FP agonist, fluprostenol, had little effect. The effects of PGE2 on IPSCs were unaffected by the selective EP1 antagonist, SC-51322. They were unaffected also by antagonists of GABAB and alpha2 adrenergic receptors, which are present at presynaptic terminals of GABA neurones in the SON and cause suppression of spontaneous IPSCs. The inhibitor of PG synthesis, indomethacin, had little effect on spontaneous IPSCs and on the inhibitory effects of PGE2 as well as of the GABAB agonist, baclofen, and noradrenaline. These results suggest that PGE2 inhibits release of GABA from the GABAergic terminals innervating SON neurones by activating presynaptic EP receptors, presumably of the EP3 subclass, and that such a presynaptic mechanism may play a role in the excitatory regulation of SON neurones by PGE2.

EP3 receptors inhibit antidiuretic-hormone-dependent sodium transport across frog skin epithelium.

We examined the effect of prostaglandin E2 (PGE2) on antidiuretic hormone (ADH)-dependent Na+ transport and cAMP production in isolated frog skin epithelium. ADH caused an increase in transepithelial Na+ transport and a decrease in cellular potential, indicating an increase in apical Na+ permeability. Subsequent addition of PGE2 decreased Na+ transport and repolarised the cells. The PGE2 receptor EP1/3-selective analogue sulprostone and the PGE2 receptor EP2/3-selective analogue misoprostol were able to mimic the effect of PGE2. ADH increased cellular cAMP levels, whereas PGE2, sulprostone and misoprostol were able to reduce the ADH-dependent cAMP production. Measurements of intracellular Ca2+ concentration ([Ca2+]i) revealed that it was unaffected by both PGE2 and sulprostone. The inhibitory effect of PGE2 on ADH-dependent Na+ transport was also observed in Ca2+-depleted epithelia. We conclude that ADH stimulates transepithelial Na+ transport by increasing cellular cAMP levels, whereas PGE2 inhibits ADH-dependent Na+ transport by activating EP3-type receptors, which decrease cellular cAMP levels. We have found no evidence that [Ca2+]i is involved in the regulation of ADH-dependent Na+ transport by PGE2.

Modulation of noradrenaline release from the sympathetic nerves of human right atrial appendages by presynaptic EP3- and DP-receptors.

Strips of human right atrial appendages were preincubated with [3H]noradrenaline and then superfused with physiological salt solution containing inhibitors of uptake1 and uptake2. Tritium overflow was evoked by transmural electrical stimulation (standard frequency: 2 Hz). Prostaglandin E2 (PGE2) inhibited the electrically evoked tritium overflow; at the highest concentration investigated, tritium overflow was reduced by about 80% and the pIC50% value was 7.14. The effect of PGE2 was not affected by rauwolscine, which, by itself, increased the evoked overflow. Naproxen failed to affect the evoked tritium overflow and its inhibition by PGE2. The inhibitory effect of PGE2 on the electrically evoked tritium overflow was mimicked by prostaglandin E1, the EP1/EP3-receptor agonist sulprostone and the EP2/EP3-receptor agonist misoprostol with the rank order of potency (pEC50%): sulprostone (7.68) > misoprostol (7.10) > PGE1 (6.39). In contrast, PGF2alpha, the IP/EP1-receptor agonist iloprost and the stable thromboxane A2 analogue U46619 (9,11-dideoxy-11alpha,9alpha-epoxy-methanoprostaglandin++ + F2alpha) did not change evoked tritium overflow. PGD2 caused facilitation. These results suggest that the sympathetic nerve fibres innervating human atrial appendages are endowed with presynaptic inhibitory EP3 and facilitatory DP-receptors. The EP3-receptors appear not to be tonically activated and do not interact with the alpha2-autoreceptors.

Prostaglandin E2 suppression of acetylcholine release from parasympathetic nerves innervating guinea-pig trachea by interacting with prostanoid receptors of the EP3-subtype.

1. We have demonstrated recently that exogenous prostaglandin E2 (PGE2) inhibits electrical field stimulation (EFS)-induced acetylcholine (ACh) release from parasympathetic nerve terminals innervating guinea-pig trachea. In the present study, we have attempted to characterize the pre-junctional prostanoid receptor(s) responsible for the inhibitory action of PGE2 and to assess whether other prostanoids modulate, at a prejunctional level, cholinergic neurotransmission in guinea-pig trachea. To this end, we have investigated the effect of a range of both natural and synthetic prostanoid agonists and antagonists on EFS-evoked [3H]-ACh release. 2. In epithelium-denuded tracheal strips pretreated with indomethacin (10 microM), PGE2 (0.1 nM-1 microM) inhibited EFS-evoked [3H]-ACh release in a concentration-dependent manner with an EC50 and maximal effect of 7.62 nM and 74% inhibition, respectively. Cicaprost, an IP-receptor agonist, PGF2alpha and the stable thromboxane mimetic, U46619 (each at 1 microM), also inhibited [3H]-ACh release by 48%, 41% and 35%, respectively. PGD2 (1 microM) had no significant effect on [3H]-ACh release. 3. The selective TP-receptor antagonist, ICI 192,605 (0.1 microM), completely reversed the inhibition of cholinergic neurotransmission induced by U-46619, but had no significant effect on similar responses effected by PGE2 and PGF2alpha. 4. A number of EP-receptor agonists mimicked the ability of PGE2 to inhibit [3H]-ACh release with a rank order of potency: GR63799X (EP3-selective) > PGE2 > M&B 28,767 (EP3 selective) > 17-phenyl-omega-trinor PGE2 (EP1-selective). The EP2-selective agonist, AH 13205 (1 microM), did not affect EFS-induced [3H]-ACh release. 5. AH6809 (10 microM), at a concentration 10 to 100 times greater than its pA2 at DP-, EP1- and EP2-receptors, failed to reverse the inhibitory effect of PGE2 or 17-phenyl-omega-trinor PGE2 on [3H]-ACh release. 6. These results suggest that PGE2 inhibits [3H]-ACh release from parasympathetic nerves supplying guinea-pig trachea via an interaction with prejunctional prostanoid receptors of the EP3-receptor subtype. Evidence for inhibitory prejunctional TP- and, possibly, IP-receptors was also obtained although these receptors may play only a minor role in suppressing [3H]-ACh release when compared to receptors of the EP3-subtype. However, the relative importance of the different receptors will depend not only on the sensitivity of guinea-pig trachea to prostanoids but on the nature of the endogenous ligands released locally that have activity on parasympathetic nerves.

Down-regulation by prostaglandins of type-II phospholipase A2 expression in guinea-pig alveolar macrophages: a possible involvement of cAMP.

We have demonstrated previously that isolated guinea-pig alveolar macrophages (AM) synthesize type-II phospholipase A2 (PLA2-II) through a tumour necrosis factor-alpha (TNF-alpha)-dependent process. This synthesis is enhanced by lipopolysaccharide (LPS) and accompanied by a release of prostaglandin E2 (PGE2) into the medium. Because agents elevating intracellular cAMP, such as PGE2, have been shown to stimulate PLA2-II expression in various cell types, we investigated the modulation of PLA2-II synthesis by cAMP in AM. Surprisingly, incubation of AM with PGE2, dibutyryl-cAMP, cholera toxin or rolipram (an inhibitor of specific cAMP-phosphodiesterase) inhibited both basal and LPS-stimulated PLA2-II expression. The inhibitory effect of PGE2 was observed at concentrations similar to those released by AM. Moreover, treatment of AM with either aspirin or neutralizing PGE2 monoclonal antibody stimulated PLA2-II synthesis. These effects were closely correlated with the ability of these agents to modulate TNF-alpha release, which was decreased by dibutyryl-cAMP and exogenous PGE2, whereas neutralizing PGE2 antibody markedly increased this release. Hence, in contrast to other cell systems, we report that: (i) agents elevating intracellular cAMP levels down-regulate both basal and LPS-induced PLA2-II synthesis, (ii) prostaglandins exert a negative feedback effect on this synthesis, probably through an elevation of intracellular cAMP levels, and (iii) inhibition of TNF-alpha release may account, at least in part, for the down-regulation of PLA2-II expression by endogenously produced prostaglandins and cAMP-elevating agents.

Timing of prostaglandin exposure is critical for the inhibition of LPS- or IFN-gamma-induced macrophage NO synthesis by PGE2.

Macrophage nitric oxide (NO) synthesis is an integral component of the host defense system. We have previously found that NO and prostaglandins interact in a variety of ways. NO modulates Kupffer cell prostaglandin E2 (PGE2) production and we have recently described the inhibitory effects of PGE2 on NO synthesis in both Kupffer cells and hepatocytes. Activated macrophages produce a number of prostaglandins but studies regarding the capacity of prostaglandins to regulate macrophage NO synthesis have yielded conflicting results. We found that exogenous PGE2 decreased lipopolysaccharide (LPS)-induced NO synthesis in murine resident peritoneal macrophages and in the RAW 264.7 murine macrophage cell line. PGE2 also suppressed NO synthesis in response to interferon-gamma (IFN-gamma) alone and a combination of LPS + IFN-gamma. Inhibition of endogenous PGE2 synthesis with indomethacin or ibuprofen had no effect on NO synthesis. PGE2 added with the activating stimulus was most effective. PGE2 lost the capacity to block NO synthesis if added more than 180 min after LPS. PGE2 decreased inducible NO synthesis (iNOS) mRNA and immunoreactive iNOS protein, consistent with the hypothesis that exogenous PGE2 inhibits macrophage iNOS expression but that the inhibition depends on the time and concentration of prostaglandin exposure.

Inducible nitric oxide synthase expression in activated rat microglial cultures is downregulated by exogenous prostaglandin E2 and by cyclooxygenase inhibitors

Prostaglandins and nitric oxide (NO) are among the numerous substances released by activated microglial cells, the brain resident macrophages, and they mediate several important microglial functions. We have previously shown that cyclooxygenase-2 (COX-2) and inducible NO synthase (iNOS), the two key enzymes in prostaglandin and NO synthesis, respectively, are rapidly co-induced in rat neonatal microglial cultures activated by bacterial endotoxin (lipopolysaccharide [LPS]) and that COX-2 expression appears to be under the negative control of endogenous as well as exogenous NO. In this study we show that exogenous prostaglandin E2 (PGE2), which is known to increase cyclic adenosine monophosphate (cAMP) levels in microglial cells, downregulates LPS-induced iNOS expression in a dose-dependent manner. The involvement of cAMP in the PGE2-dependent inhibition of iNOS is supported by several pieces of evidence. First, iNOS expression was also inhibited by agents such as isoproterenol and forskolin, which cause an elevation of cAMP levels, and by dibutyryl cAMP (dbcAMP), a cAMP stable analogue. Second, the inhibitory effect of PGE2 was mimicked by 11-deoxy-16,16-dm PGE2, a selective agonist at the PGE2 receptor subtype EP2, coupled to the activation of adenylyl cyclase, but not by sulprostone, a potent agonist at receptor subtypes EP3 and EP1, associated with an inhibition of adenylyl cyclase activity and intracellular Ca2+ elevation, respectively. Third, the inhibitory effect of PGE2 on NO synthesis was blocked by SQ 22,536, a specific inhibitor of adenylyl cyclase. Interestingly, the abrogation of endogenous prostanoid production by several COX inhibitors caused a reduction of iNOS expression, suggesting a positive modulatory effect of endogenous prostanoids of iNOS expression, as opposed to the inhibitory effect of exogenous PGE2.

The inhibitory effect of prostaglandin E2 on rat neutrophil aggregation.

Rat peritoneal neutrophils stimulated by N-formyl-methionyl-leucyl-phenylalanine (fMLP) produce an aggregation response that can be inhibited by prostaglandin E2 (PGE2) with an IC50 value of 2.6 x 10(-9) M. Although PGE2 can stimulate [3H]cAMP production in neutrophils (EC50 4.3 x 10(-8) M), the anti-aggregation response cannot be significantly attenuated by inhibitors of adenylate cyclase or protein kinase A, neither can it be potentiated by inhibition of phosphodiesterase activity. Despite these observations, it still remains possible that PGE2-mediated inhibition of rat neutrophil aggregation is a cAMP-dependent response mediated by highly localized changes in neutrophil cAMP. The inhibitory effect of PGE2 does not appear to depend on effects on intracellular calcium or K(ATP) channels. Similarities exist between PGE2 and the profile of activity of phosphatidylinositol 3-kinase (PI 3-kinase) inhibitors, suggesting that PI 3-kinase is a possible target for PGE2 action in rat neutrophils.

Prejunctional inhibition of cholinergic responses by prostaglandin E2 in human bronchi.

The effect of prostaglandin E2 (PGE2) on cholinergic responses elicited by electrical field stimulation (EFS) (8 Hz, 1 ms, 8 V for 10 s every 100 s) of human isolated human bronchi was studied. Indomethacin (1 microM) was found to produce approximately a 100% increase in the cholinergic response. In the presence of indomethacin, PGE2 (1-300 nM) was found to concentration-dependently inhibit the cholinergic responses to EFS. A maximal inhibition was obtained with 100 nM PGE2 which produced an 80% inhibition of the cholinergic response. By contrast 100 nM PGE2 was without significant effect on the concentration response curve to exogenously added acetylcholine. The inhibitory effect of PGE2 on the EFS-induced cholinergic responses was not modified by the large conductance Ca(2+)-activated K+ channel inhibitor charybdotoxin (100 nM). These results suggest that endogenously released prostaglandins, presumably PGE2, act to inhibit cholinergic responses by a prejunctional mechanism in human isolated bronchi. In contrast to other autocoids, this inhibition does not involve the activation of large-conductance Ca(2+)-activated K+ channels.

Inhibitory effect of prostaglandin E1 on gastric secretion during general anesthesia in humans

The present study was undertaken to clarify the effects of prostaglandin E1 (PGE1) on gastric secretion during general anesthesia. Thirty-three patients, 16 with (PGE1 group) and 17 without (control group) PGE1 administration, scheduled for selective surgery were studied during general anesthesia with nitrous oxide (67%) and enflurane (1%-2% inspired). PGE1 was administered at a rate of 50-200 ng·kg(-1)·min(-1) when hypotensive medication was required. In the PGE1 group, gastric juice was collected serially three times before and during administration and 60 min after discontinuation of PGE1. In the control group, it was collected three times corresponding to those in the PGE1 group. The pH of gastric juice increased significantly, and the acidity and pepsin activity decreased after the beginning of the administration of PGE1, and these changes were observed even 1h after discontinuation. There was significant differences in the pH, acidity, and pepsin activity between the two groups after administration of PGE1. The results indicate that PGE1 inhibits gastric secretion at doses that produce a sufficient hypotensive effect under general anesthesia.

Investigation of the inhibitory effects of PGE2 and selective EP agonists on chemotaxis of human neutrophils.

1. The aims of this study were to investigate the inhibitory effects of prostaglandin E2 (PGE2) on chemotaxis of N-formyl-methionyl-leucine-phenylalanine (FMLP)-stimulated human neutrophils, and to test the hypothesis that cyclic AMP is the second messenger involved. For this purpose, the inhibitory effect of selective EP agonists, and the modulatory effects of the adenylate cyclase inhibitor, SQ 22536, the protein kinase A (PKA) inhibitors H-89 and Rp-cAMPs, and the type IV phosphodiesterase (PDE) inhibitors, rolipram and Ro20-1724 have been examined. 2. Chemotaxis has been measured using blindwell chambers. When human neutrophils were stimulated with FMLP (100 nM), PGE2 inhibited chemotaxis in a concentration-dependent manner (0.01-10 microM), with an EC50 of 90 +/- 24.5 nM, a maximum effect ranging from 45-75% and a mean inhibition of 64.5 +/- 2.4%. 3. The EP2-receptor agonists, 11-deoxy PGE1, butaprost and AH 13205 also inhibited chemotaxis. The order of potency of these agonists was PGE2 > butaprost (EC50 = 106.4 +/- 63 nM) > 11-deoxy PGE1 (EC50 = 140.9 +/- 64.7 nM) > AH 13205 (EC50 = 1.58 +/- 0.73 microM). Correlation of the ability of EP2 agonists to increase cyclic AMP and to inhibit chemotaxis was poor (r = 0.38). 4. The IP agonist, cicaprost gave similar increases in cyclic AMP to those achieved with PGE2, yet produced 50% of the maximum inhibition of chemotaxis observed with PGE2. 5. Slight potentiation of the inhibitory effects of PGE2 after type IV PDE block was observed with rolipram (EC50 for PGE2 = 57.2 +/- 5.9; 35.2 +/- 6.8 nM) but not Ro20-1724 (EC50 for PGE2 = 216.0 +/- 59.7; 97.8 +/- 50.6 nM). Type IV PDE inhibitors are themselves potent inhibitors of chemotaxis with EC50 values of 23.0 +/- 2.3 and 73.6 +/- 10.3 nM for rolipram and Ro20-1724, respectively. 6. Inhibition of cyclic AMP production with the adenylate cyclase inhibitor SQ 22,536 (0.1 mM) failed to antagonize inhibition of chemotaxis by PGE2 (EC50s for PGE2 of 57.2 +/- 5.9 and 56.8 +/- 27.3 nM, in the absence and presence of SQ 22,536, respectively) despite a reduction in the increase in cyclic AMP induced by PGE2. 7. Inhibition of PKA with either H-89 (10 microM) or Rp cyclic AMPS (10 microM) similarly failed to antagonize inhibition of chemotaxis by PGE2; EC50 for PGE2 of 90 +/- 40 and PGE2 + H-89 60 +/- 17 nM; PGE2 216.0 +/- 58.7 and PGE2 + Rp cyclic AMP 76.9 +/- 14.7 nM. 8. Of the two PKA inhibitors tested, H-89 (10 microM) and Rp cyclic AMPS (10 microM), the more effective inhibitor of PGE2-induced inhibition of neutrophil superoxide anion generation was H-89 (EC50s for PGE2 were 0.36 +/- 0.1 and > 10 microM, respectively). We have previously shown this to be a cyclic AMP-dependent effect of PGE2. 9. Confirmation of block of PKA by H-89 was suggested by the finding that H-89 blocked inhibition of superoxide anion generation observed with the type IV PDE inhibitors rolipram and Ro20-1724; EC50s of 12.9 +/- 8.9 nM for rolipram alone and rolipram+H-89 > 1 microM; Ro20-1724 alone 59.5 +/- 28.1 nM and Ro20-1724 + H-89 > 1 microM. 10. The results suggest that inhibition of chemotaxis by PGE2 and EP2 agonists is not mediated by increased neutrophil cyclic AMP levels.

Both prostaglandin E2 and nitric oxide sequentially mediate the tumor necrosis factor alpha-induced inhibition of surfactant synthesis by human type II pneumocytes.

Tumor necrosis factor alpha (TNF-alpha)-induced inhibition of surfactant synthesis seems to participate in the pathogenesis of the adult respiratory distress syndrome. To examine the ability of human type II pneumocytes to produce nitric oxide (NO) in the presence of TNF-alpha and, in addition, to explore the role of this radical in the transduction of the cytokine signal. Multiple organ donors were the source of lung tissue specimens. Type II pneumocytes were isolated by enzymatic digestion, adherence separation of macrophages, and gradient purification. After 24-hour preculture, cells were cultured for 24 hours in the presence or absence of TNF-alpha (100 ng/mL), sodium nitroprusside (100 mumol/L), N omega-nitro-L-arginine methyl ester (NAME) (1 mmol/L), methylene blue (10 mumol/L),8-bromo-3',5'-cyclic guanosine monophosphate (8-Br-cGMP) (1 mmol/L), prostaglandin E2 (PGE2) (0.1 mumol/L), indomethacin (30 mumol/L), and combinations. The NO release to the medium and cGMP and PGE2 contents of the cells were measured. The incorporation of 14C-labeled glucose (D-[U-14C]glucose) into phosphatidylcholine and phosphatidylglycerol was selectively inhibited either by 8-Br-cGMP or in the presence of TNF-alpha, PGE2, or nitroprusside, all of which caused an increase in the intracellular levels of cGMP. The inhibitory effect of TNF-alpha was partially reverted by indomethacin, NAME, N-monomethyl arginine, or methylene blue. The inhibitory effect of PGE2 was partially reverted by NAME, while that of nitroprusside was reverted by methylene blue, but not by indomethacin. Tumor necrosis factor alpha induced an increase in PGE2 (4.31 +/- 0.27 vs 1.65 +/- 0.17-pg/microgram protein, n = 10, P < .01) and cGMP (0.238 +/- 0.012 vs 0.109 +/- 0.014-pmol/microgram protein, n = 10, P < .01) cell content and in the NO release to the medium (3.10 +/- 0.14 vs 1.19 +/- 0.11-nmol/microgram protein, n = 10, P < .01). The basal NO release to the medium was also increased in the presence of PGE2. The NAME, which blocked NO generation and cGMP increase, did not affect PGE2 production in response to TNF-alpha. However, indomethacin, which blocked PGE2 production, also blunted NO generation and cGMP increase. The NO generation, secondary to PGE2 production, seems responsible for the TNF-alpha-induced inhibition of phosphatidylcholine synthesis by human type II pneumocytes. Nitric oxide seems to exert this effect through activation of guanylyl cyclase.

Inhibitory effect of prostaglandin E1 on gastric secretion during general anesthesia in humans

Inhibitory effect of prostaglandin E1 on gastric secretion during general anesthesia in humans

Prostaglandin-E2 is a potent inhibitor of human interleukin 12 production.

During human immunodeficiency virus infection and allergic diseases, characterized by a dominant T helper (Th) 2 response, overproduction of prostaglandin E2 (PGE2) is observed. In this paper we studied the effect of PGE2 on interleukin (IL)-12 synthesis, because this cytokine has been described to be essential in induction of Th1 responses. IL-12 synthesis was induced in monocytes that were stimulated with Neisseria meningitidis-derived lipopolysaccharide in whole blood cultures. PGE2 almost completely inhibited lipopolysaccharide induced IL-12 production, whereas IL-6 production was only partially inhibited by PGE2. In contrast, the production of IL-10 was approximately twofold enhanced at these conditions. The effects of PGE2 were due to its cAMP-inducing capacity, since they could be mimicked by other cAMP inducers. Recombinant human IL-10 also inhibited IL-12 and IL-6 production. However, the inhibitory effect of PGE2 on IL-12 production was independent of IL-10 since neutralizing anti-IL-10 antibodies were unable to reverse this inhibition. These results suggest that the capacity of an antigen to induce PGE2 synthesis may play a crucial role in the development of either a Th1 or Th2 response.

Prostanoid receptors of the EP3 subtype mediate the inhibitory effect of prostaglandin E2 on noradrenaline release in the mouse brain cortex.

Mouse or rat brain cortex slices were preincubated with 3H-noradrenaline and superfused with physiological salt solution containing desipramine. We studied the effects of prostaglandin E2 (PGE2), prostaglandin D2 (PGD2) and related drugs on the electrically evoked (50 mA, 2 ms, 0.3 Hz) tritium overflow. PGE2 inhibited the electrically evoked tritium overflow from mouse brain cortex slices; the maximum effect of PGE2 (79%) was attenuated by the alpha 2-adrenoceptor agonist talipexole (to 52%) and enhanced by the alpha 2-adrenoceptor antagonist rauwolscine (to 92%). Rauwolscine was added to the superfusion medium in all subsequent experiments. The effect of PGE2 was readily reversible upon withdrawal from the medium and remained constant upon prolonged exposure of the tissue to the prostanoid. Studies with EP receptor agonists, mimicking the inhibitory effect of PGE2, showed the following potencies (pIC50); sulprostone (8.22); misoprostol (8.00); PGE2 (7.74); PGE1 (7.61); iloprost (5.86). The concentration-response curve of PGE2 was marginally shifted to the right by the EP1 receptor antagonist AH 6809 (6-isopropoxy-9-oxoxanthene-2- carboxylic acid; apparent pA2 3.97) and by the TP receptor antagonist vapiprost (4.50). AH 6809, by itself, did not affect the evoked overflow whereas vapiprost increased it. PGD2 inhibited the evoked overflow at high concentrations (pIC50 4.90); this effect was not altered by the DP receptor antagonist BW A868C (3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl-2- hydroxyethylamino)hydantoin), which, by itself, did not affect the evoked overflow. Indometacin slightly increased the evoked overflow and tended to increase the inhibitory effect of PGE2. PGE2 inhibited the electrically evoked tritium overflow also in rat brain cortex slices.(ABSTRACT TRUNCATED AT 250 WORDS)

Cyclooxygenase inhibitors suppress inhibitory effect of PGE2 on Na-K-ATPase in MDCK cells

In a previous work, we showed that picomolar concentrations of prostaglandin E2 (PGE2) inhibit Na-K-ATPase activity and ouabain binding in a clone of Madin-Darby canine kidney (MDCK) cells. In the present study, we demonstrate that the inhibitory effects of PGE2 on Na-K-ATPase activity, ouabain-sensitive Rb+ uptake, and ouabain binding in MDCK cells were diminished by treatment of the cells with nonsteroidal anti-inflammatory drugs. These results suggested that products of arachidonic acid synthesized through the cyclooxygenase pathway are involved in the inhibitory mechanism of PGE2. Treatment of the cells with arachidonic acid resulted in inhibition of ouabain binding, and the inhibition was eliminated by cyclooxygenase inhibitors. These observations further support the involvement of cyclooxygenase products in the PGE2-induced inhibitory process. Finally, we demonstrated that dopamine inhibits Rb+ influx and ouabain binding in MDCK cells similarly to PGE2. Cyclooxygenase inhibitors suppressed the inhibition of ouabain binding by dopamine, thus also suggesting the involvement of cyclooxygenase products in the inhibitory effect of dopamine.

Inhibition of the transendothelial migration of human T lymphocytes by prostaglandin E2.

To determine whether part of the anti-inflammatory effects of prostaglandin E2 (PGE2) was related to inhibition of T cell interactions with endothelial cells (EC), the effects of PGE2 and other cAMP-elevating agents on the transendothelial migration of human T cells was examined. Although PGE2 did not effect T cell binding to EC, concentration-dependent inhibition of the transendothelial migration of T cells through unstimulated or IL-1-activated EC was observed. PGE2 inhibited the function of both T cells and EC, with maximal inhibition observed when both T cells and EC were treated with PGE2. However, the inhibitory action of PGE2 could not be ascribed to an effect on the adhesion receptor pair, CD11a/CD18-CD54. The inhibitory effect of PGE2 seemed to relate to its capacity to elevate cellular cAMP levels, because 3-isobutyl-1-methylxanthine enhanced PGE2 activity and dibutyryl cAMP and forskolin also inhibited transendothelial migration. The inhibitory effect of PGE2 and the other cAMP-elevating agents on the function of T cells related in part to suppression of their intrinsic locomotory behavior as random migration in the absence of EC was blocked. In EC, PGE2 and the other cAMP-elevating agents increased the barrier function of EC as evidenced by a decrease in the diffusion of [3H]mannitol through the endothelium. These results indicate that part of the anti-inflammatory action of PGE2 relates to its capacity to suppress the transendothelial migration of T cells by cAMP-mediated alterations in the function of both T cells and EC.

Elevated prostaglandin E2 production by monocytes is responsible for the depressed levels of natural killer and lymphokine-activated killer cell function in patients with breast cancer

Human natural killer (NK) cells mediate spontaneous cytotoxicity against tumor cells and represent the main precursors of lymphokine-activated killer (LAK) cell activity. A comparison of some aspects of NK and LAK cell activity was undertaken in 85 preoperative patients with breast cancer and 75 healthy donors. NK cell activity (tested in 18-hour cultures of effector peripheral blood mononuclear cells [PBMC] with K562 or MOLT-4 tumor target cells) was significantly diminished in these patients as it was the fully mature LAK cell activity (i.e., interleukin-2 (IL-2)-induced cytotoxicity in PBMC) against NK resistant target cells. Using immunoenzymatic methods we showed that the reduced NK cell activity was due to abnormally high levels of prostaglandin E2 (PGE2) produced by monocytes in culture. PGE2 was found to suppress the production of IL-2 in these cultures. Removal of monocytes from PBMC restored to almost normal levels the deficient NK and LAK cell activity in patients with breast cancer and was also associated with a normalization in the levels of PGE2 and IL-2. Indomethacin and gamma-interferon (IFN-gamma) increased the NK and LAK cell activity in these patients up to the levels of healthy donors. When highly purified CD56+ cells (obtained by an immunomagnetic isolation technique) were used as effector cells, no differences in LAK cell activity could be noticed between healthy donors and patients with cancer. FACS and northern blot analyses demonstrated a PGE2-mediated down-regulation of IL-2 receptor (IL-2R) expression on CD56+ cells that correlated with reduced LAK cell activity. This inhibitory effect of PGE2 was noticeable in long-term LAK cultures and was abrogated in the presence of IFN-gamma or indomethacin. This study may have important implications in the potentiation of NK and LAK cell activity for immunotherapeutic protocols in patients with breast cancer.

Glucocorticoid effects on natural and humoral immunity in mallards

Two studies were conducted to determine the effects of dexamethasone (DEX) on immune function in mallard ducks. Each day ducks were injected intramuscularly with DEX at doses ranging from 0.2–4.0 mg/kg for 28–30 days. Physiologic effects consistent with high dose glucocorticoid (GC) treatment were observed at the 4 mg/kg dose, and included significant body weight loss, lowered hematocrit, and elevated alanine aminotransferase (ALT) activity. At all doses, effects of DEX on the immune system were observed. When DEX was given at 0.2 mg/kg/day, significant suppression of primary IgG antibody titers to sheep erythrocytes (SRBC) was observed. At 1 mg/kg/day, primary IgM and secondary IgM and IgG titers were suppressed as well. These doses of DEX also produced significant elevation in natural killer cell (NKC) activity of peripheral blood mononuclear cells (PBMNC). Removal of adherent cells from the PBMNC prior to NKC assay eliminated the enhancement in NKC activity. Based on these results, it was postulated that the elevation in NKC activity may be due to suppression by DEX of monocyte production of prostaglandin-E2 (PGE-2) resulting in the release of NKC activity from the inhibitory effects of PGE-2. This hypothesis was supported by a measured decrease in PGE-2 production during the NKC assay by cells from DEX-treated birds. Furthermore, an enhanced NKC activity could be reproduced in vitro with the addition of indomethacin or DEX to NKC cultures containing adherent cells from PBMNC. Direct effects of DEX on nonadherent cell NKC activity and lymphocyte viability were only observed at high concentrations (10 −4 M) of DEX, while the phagocytic activity of adhered blood monocytes was inhibited at 10 −6 M DEX. The suppressed phagocytic activity may contribute to the suppressed antibody responses observed in DEX-treated birds. Together, these results support an indirect immunomodulatory effect of DEX on NKC activity and perhaps antibody responses in vivo via altered monocyte function in mallard ducks.