Tumor necrosis factor-alpha inversely regulates prostaglandin D2 and prostaglandin E2 production in murine macrophages. Synergistic action of cyclic AMP on cyclooxygenase-2 expression and prostaglandin E2 synthesis

Increased synthesis of insulin-like growth factor-1 is induced in murine macrophages by prostaglandin E2 (PGE2) and tumor necrosis factor-alpha (TNFalpha). Accordingly, we have investigated mechanisms regulating synthesis of PGE2 that might contribute to autocrine/paracrine effects on insulin-like growth factor-1 production. In response to zymosan, TNFalpha specifically induced a 5-fold increase in PGE2 synthesis, at the same time decreasing PGD2 production in a reciprocal fashion. Activators of cyclic AMP-dependent protein kinase (PKA), such as PGE2 itself or dibutyryl cyclic AMP, did not modify PGE2 production by themselves but potentiated the TNFalpha-induced increase in PGE2; this effect required both RNA and protein synthesis. No significant change in arachidonate release or production of other eicosanoids was observed. The inducible form of cyclooxygenase-2 (COX2) but not of the constitutive form COX1 was implicated in the generation of both PGE2 and PGD2 in these cells by use of specific inhibitors and effects of dexamethasone. Neither COX1 nor COX2 protein levels were affected by TNFalpha or PKA activators used alone, whereas in association, marked up-regulation of COX2 mRNA and protein was observed. Incubations of cells carried out with PGH2 demonstrated that PGE2 synthase activity was increased after a TNFalpha pretreatment. Taken together, our results suggest that TNFalpha induced a switch from the PGD2 to PGE2 synthesis pathway by regulating PGE2 synthase expression and/or activity and that activators of PKA markedly potentiated the TNFalpha-induced increase in PGE2 through up-regulation of COX2 gene expression.     

Rate of vasoconstrictor prostanoids released by endothelial cells depends on cyclooxygenase-2 expression and prostaglandin I synthase activity

This study was undertaken to investigate the enzymatic regulation of the biosynthesis of vasoconstrictor prostanoids by resting and interleukin (IL)-1(beta)stimulated human umbilical vein endothelial cells (HUVECs). Biosynthesis of eicosanoids in response to IL-1beta, exogenous labeled arachidonic acid (AA), or histamine, as well as their spontaneous release, was evaluated by means of HPLC and RIA. HUVECs exposed to IL-1beta produced prostaglandin (PG) I2 for no longer than 30 seconds after the substrate was added irrespective of the cyclooxygenase (COX) activity, whereas the time course of PGE2 and PGD2 formation was parallel to the COX activity. The ratio of PGE2 to PGD2 produced by HUVECs was similar to that obtained by purified COX-1 and COX-2. Production of PGF2alpha from exogenous AA was limited and similar in both resting and IL-1beta-treated cells. PGF2alpha was the main prostanoid released into the medium during exposure to IL-1beta, whereas when HUVECs treated with IL-1beta were stimulated with histamine or exogenous AA, PGE2 was released in a higher quantity than PGF2alpha. PGF2alpha released into the medium during treatment with IL-1beta and the biosynthesis of PGE2 and PGD2 in response to exogenous AA or histamine increased with COX-2 expression, whereas this did not occur in the case of PGI2. We observed that PGI synthase (PGIS) mRNA levels were not modified by the exposure to IL-1beta, but the enzyme was partially inactivated. When SnCl2 was added to the incubation medium, the transformation of exogenous AA-derived PGH2 into PGE2 and PGD2 was totally diverted toward PGF2alpha. Overall, these results support the conclusions that PGE2 and PGD2 (and also probably PGF2alpha) were nonenzymatically derived from PGH2 in HUVECs. The concept that a high ratio of PGH2 was released by the IL-1beta-treated HUVECs and isomerized outside the cell into PGE2 and PGD2 was supported by the biosynthesis of thromboxane B2 by COX-inactivated platelets, indicating the uptake by platelets of HUVEC-derived PGH2. The IL-1beta-induced increase in the release of PGH2 by HUVECs was suppressed by the COX-2-selective inhibitor SC-58125 and correlated with both COX-2 expression and PGIS inactivation. An approach to the mechanism of inactivation of PGIS by the exposure to IL-1beta was performed by using labeled endoperoxides as substrate. The involvement of HO. in the PGIS inactivation was supported by the fact that deferoxamine, pyrrolidinedithiocarbamate, DMSO, mannitol, and captopril antagonized the effect of IL-1beta on PGIS to different degrees. The NO synthase inhibitor NG-monomethyl-L-arginine also antagonized the PGIS inhibitory effect of IL-1beta, indicating that NO. was also involved. NO. reacts with O2-. to form peroxynitrite, which has been reported to inactivate PGIS. Homolytic fission of the O-O bond of peroxynitrite yields NO2. and HO.. The fact that 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy-PTIO), which reacts with NO. to form NO2., dramatically potentiated the IL-1beta effect suggests that NO2. could be a species implicated in the inactivation of PGIS. Cooperation of HO. was supported by the fact that DMSO partially antagonized the effect of carboxy-PTIO. Although our results on the exact mechanism of the inactivation of PGIS caused by IL-1beta were not conclusive, they strongly suggest that both NO. and HO. were involved.     

Chronic sinusitis: characterization of cellular influx and inflammatory mediators in sinus lavage fluid

Chronic sinusitis is a recurrent disorder commonly found in atopic individuals, yet few studies have explored the role of inflammatory mediators in sinusitis. Sinus lavage fluid from ten patients with chronic sinusitis obtained during endoscopic surgery was analyzed for total cell counts and then assayed for histamine, immunoreactive leukotriene C4/D4/E4 (LTC4/D4/E4), and prostaglandin D2 (PGD2). All ten patients had been unresponsive to medical treatment, including oral corticosteroids in most cases. High concentrations of histamine, LTC4/D4/E4 and PGD2 were found in sinus fluid and were comparable to levels seen in nasal secretions of allergic rhinitis patients following allergen challenge. In the sinus fluid, inflammatory cells were predominantly neutrophils with only low percentages of mast cells, basophils or eosinophils. On the basis of the histamine and PGD2 concentrations in sinus fluid, we conclude that mast cell/basophil activation does occur in chronic sinusitis and may contribute to the persistent inflammation present in sinusitis.     

Heterogeneous effects of protamine on human mast cells and basophils

To investigate the mechanisms of anaphylactoid reactions to protamine, we have examined the in vitro effects of increasing concentrations of protamine (10(-6)-3 x 10(-4) mol litre-1) on the release of preformed (histamine and tryptase) and de novo synthesized (peptide leukotriene C4 (LTC4) or prostaglandin D2 (PGD2)) mediators from human basophils and mast cells isolated from lung parenchyma, heart, skin and synovial tissues. Protamine 10(-6)-3 x 10(-4) mol litre-1 induced release of histamine, but not de novo synthesis of LTC4 from basophils. At concentrations from 10(-5) to 3 x 10(-4) mol litre-1 it induced histamine release from human heart (mean 6.5 (SEM 1.5)%), skin (17.7 (4.1)%) and to a lesser extent from synovial mast cells, but not from lung mast cells. Protamine also caused the release of tryptase from heart mast cells (12.8 (3.2) micrograms/10(7) cells), but did not induce de novo synthesis of LTC4 and PGD2 from lung and skin mast cells. In these experiments cross-linking of IgE by anti-IgE caused release of LTC4 or PGD2 from human basophils or mast cells. These results demonstrate that protamine acted as an incomplete secretagogue, causing the release of preformed mediators from human basophils and mast cells.     

Reduction by arachidonic acid of prostaglandin I2-induced cyclic AMP formation. Involvement of prostaglandins E2 and F2 alpha

Arachidonic acid reverses the increase in cyclic AMP levels of washed human platelets exposed to prostaglandin (PG)I2, under conditions where the PGH2 analogue U46619 is ineffective. This effect of arachidonic acid was inhibited by aspirin, a cyclooxygenase inhibitor, but not by the thromboxane (Tx) synthase inhibitor Ridogrel, which induces, by inhibiting the conversion of PGH2 into TxA2, an overproduction of PGE2, PGD2 and PGF2 alpha. Addition of PGE2 or PGF2 alpha, which share a receptor with PGI2, to washed human platelets also induced a decrease in cyclic AMP levels, but PGD2, which interacts with a different receptor, had no effect. Thus neither PGD2, PGG2, PGH2, TxA2 nor TxB2 formed from arachidonic acid via the cyclooxygenase pathway is involved in the decrease in cyclic AMP levels. These findings were confirmed using forskolin, a diterpene from the labdane family, which enhanced the formation of cyclic AMP synergistically with the PGs. Also, arachidonic acid, unlike U46619, is able to reverse the inhibition of platelet aggregation by PGI2 after a lag phase of about 4 min. Our data indicate that arachidonic acid decreased cyclic AMP levels through its cyclooxygenase metabolites PGE2 and PGF2 alpha probably interacting competitively with the receptor of PGI2. In addition, intracellular cyclic AMP levels and the degree of aggregation of platelets by arachidonic acid seem to be inversely correlated.     

Evaluation of efficacy of traditional Chinese medicines in the treatment of childhood bronchial asthma: clinical trial, immunological tests and animal study. Taiwan Asthma Study Group

Traditional Chinese medicines (TCM) have been used to treat bronchial asthma for several centuries and a certain degree of clinical benefit has been observed; however, scientific substantiation is lacking. A multicenter, double-blind and placebo-controlled study was therefore conducted to evaluate the clinical efficacy in terms of symptom score, medication score, morning and evening PEFRs, and changes of immunoregulatory function, such as distribution of lymphocyte subsets and in vivo and in vitro production of lymphokines (IFN-gamma and IL-4) and inflammatory mediators (histamine, PGE2 and LTC4). Furthermore, the protective effect of TCM on the late asthmatic reaction (LAR) was evaluated by using asthmatic guinea pigs. Three hundred and three asthmatic children were classified by Chinese doctors, according to a standardized questionnaire designed on the basis of basic logic of Chinese medicine, into three groups of specific constitution (group A, B and C). Group A consisted of 32 herb A-treated patients and 34 placebo-treated; group B, 74 herb B-treated and 64 placebo-treated; and group C, 55 herb C-treated and 44 placebo-treated. The study period was six months. The results were: 1) Both treatment group and placebo group showed an improvement in all clinical parameters, thus demonstrating a placebo effect. However, the improvement was usually greater in the former than the latter, although only the difference in PEFR was significant; 2) Herb A could increase total T cell and decrease B cell; 3) Herb A and B enhanced production of PGE2 but not LTC4, IFN-gamma and IL-4; 4) There was a general tendency for in vivo and in vitro production of histamine to decrease at the end of study in both treatment group and placebo group; however, the decrease was significantly greater in the former than the latter; 5) In asthmatic guinea pigs, 10-day's pretreatment with Chinese herbs could reverse the decrease of sGaw, suppress eosinophilia in bronchoalveolar lavage fluid (BALF), prevent the eosinophil infiltration of airways, increase PGE2 production and decrease LTC4 production in serum and BALF. Thus, traditional Chinese medicines did show a certain degree of clinical efficacy. The decreased production of histamine and LTC4, increased production of PGE2 that were found in both asthmatic children and asthmatic guinea pigs, and prevention of occurrence of LAR by suppressing eosinophil infiltration of airways and preserving airway conductance that were observed in asthmatic guinea pigs after allergen challenge might be used to account partly for the effectiveness.     

Histamine and eicosanoid levels in plasma and peripheral blood leucocyte cultures during experimental infection of cattle with bluetongue virus serotype 11

Three calves were sensitized with three doses of inactivated BTV-11 UC8 strain and then experimentally infected with the homologous virus. In addition, four BTV-seronegative heifers were also experimentally infected with BTV-11. Granulocyte rich fractions of peripheral blood leucocyte (PBL-GRF) cultures from BTV-sensitized/infected calves and from control unexposed cattle were exposed in vitro with BTV-11. Histamine, leukotriene (LT) C4 and prostaglandin (PG) D2 were assayed in supernatant fluids. Plasma histamine levels increased in BTV-infected heifers from 10.1 +/- 2 ng/ml at Day 0 to 23.1 +/- 6.6 ng/ml at Day 12 following virus exposure. In addition, in this experimental group the concentration of PGF2 alpha (mean 551.97 +/- 243.54 pg/ml) increased significantly (P < or = 0.05) compared with control cattle (mean 467.3 +/- 73.9 pg/ml). Bluetongue virus induced histamine and LTC4 release after in vitro infection of PBL-GRF. Release of LTC4 was significantly (P < or = 0.05) higher in PBL-GRF cultures from sensitized and control animals than in unexposed PBL-GRF cultures. In contrast to these results, PGD2 was not released after BTV infection of PBL-GRF in vitro. The histamine release caused by BTV was virus-specific and mainly mediated by an immunological reaction, since the release was significantly reduced by removal of cell surface immunoglobulins.     

Improved method for harvesting human Schwann cells from mature peripheral nerve and expansion in vitro

The pharmacological profile of a novel dual inhibitor, tepoxalin and of its carboxylic acid metabolite on cyclooxygenase and lipoxygenase pathways was evaluated by in vitro incubation with synovial tissue. Tissue specimens obtained at surgery in rheumatoid arthritis (RA, n = 10) or osteoarthritis (OA, n = 11) patients were incubated. Tepoxalin (10(-7), 10(-6), 10(-5) M) decreased eicosanoid release calculated in % of tyrode control for OA: LTC4 to 71-33%, 6-keto-PGF1a to 37-20%, PGE2 to 29-6%. For RA: LTC4 to 56-22%, 6-keto-PGF1a to 43-22%, PGE2 to 57-32%. Similarly, its metabolite (10(-7), 10(-5)M) decreased release in OA: LTC4 to 99 and 60%, PGE2 to 42 and 20%, 6-keto-PGF1a to 54 and 25%. In RA:LTC4 to 81 and 45%, PGE2 to 61 and 30%, 6-keto-PGF1a to 46 and 18%. Significance (P < 0.05) was achieved for all but 1 group (LTC4 metabolite at 10(-7)M vs tyrode). In summary a marked and dose dependent decrease of LT and PG release was obtained when incubating the dual inhibitor tepoxalin and its active carboxylic acid metabolite with synovial tissue at doses expected to be reached in the joint during therapy.     

Prostaglandin D2 measurement in nasal secretions is not a reliable marker for mast cell activation in atopic patients

BACKGROUND: Prostaglandin D2 (PGD2) is a very important mast cell product during the early-phase nasal allergic reaction. However, the quantification of PGD2 in nasal secretions has not yet been well established. OBJECTIVE: Quantitative determination of PGD2 in nasal secretions of atopic patients (n = 17) after nasal allergen challenge (NAC) and in non-allergic healthy volunteers (n = 10). METHODS: The nasal microsuction sampling technique was used to obtain the nasal secretions with an exactly known and minimally diluted volume. A sensitive and specific enzyme immunoassay was chosen to measure the more stable 11-methoxime derivative of PGD2, which was obtained after extraction in acetone/ethanol and conversion using methoxamine-HCl. The concentrations of PGD2 in nasal secretions obtained from 10 non-allergic healthy volunteers were used as reference values. RESULTS: There was no significant difference in the concentrations of PGD2 between men (median: 569 pg/mL) and women (median: 407 pg/mL), nor between the baseline concentrations from atopic patients (median: 410 pg/mL) and non-allergic controls (median: 477 pg/mL). In the atopic patient group, PGD2 did not significantly increase during the entire sampling period after NAC. The absence of PGD2 response contrasted with the nasal symptoms manifested by sneezing, increased nasal airway resistance, and the significant increases of the concentrations of histamine, tryptase, and leukotriene C4 (LTC4) 5 min after NAC. CONCLUSION: This observation suggests that the measurement of PGD2 alone in the nasal secretions does not give reliable information on mast cell activation during a nasal allergic reaction.     

Biogenic amines in silage, apparent postruminal passage, and the relationship between biogenic amines and digestive function and intake by steers

A 4 x 4 Latin square experiment was conducted to examine abomasal passage of biogenic amines in steers fed silage and their related effects on intake, digestibility, and digestive function. Thirty percent of the dry matter (DM) in the diets consisted of alfalfa forage, which was fed as either hay or silage. The DM from alfalfa silage DM was substituted at 0, 33, 67, and 100% for DM from alfalfa hay and was fed to four ruminally and abomasally cannulated steers. The roughage component of the diet constituted 50% of the DM and consisted of 60% alfalfa silage or hay and 40% tropical corn silage. The concentrate was composed mainly of ground corn. The concentrations of putrescine and cadaverine in abomasal digesta increased as alfalfa silage in the diet increased. Abomasal recovery of biogenic amines, a product of their concentration in abomasal digesta and the passage of DM through the abomasum, was negatively correlated with intake. Abomasal recovery of most amines was 5 to 20% of intake. Abomasal recovery of cadaverine was correlated with depressed intake. Total DM intake was reduced 8.3 to 25.8% as the proportion of alfalfa silage in the diet increased. Frequency of reticular contractions, intake, ruminal DM digestibility, ruminal outflow, volatile fatty acids, and total tract DM digestibility decreased in steers fed diets that contained more alfalfa silage. Ruminal fluid pH and NH3 concentration increased in steers fed more alfalfa silage; however, mass and the DM percentage of ruminal contents decreased linearly. Postprandial insulin concentrations were quadratically related to the proportion of alfalfa hay or silage in the diet. Intraruminal metabolism of biogenic amines is extensive based on the relatively low quantities recovered in abomasal digesta; however, the amounts recovered in abomasal digesta were related to intake depression and associated physiological effects.     

Inhibition of nociceptin-induced allodynia in conscious mice by prostaglandin D2

1. We recently showed that intrathecal administration of nociceptin induced allodynia by innocuous tactile stimuli and hyperalgesia by noxious thermal stimuli in conscious mice. In the present study, we examined the effect of prostaglandins on nociceptin-induced allodynia and hyperalgesia. 2. Prostaglandin D2 (PGD2) blocked the allodynia induced by nociceptin in a dose-dependent manner with an IC50 of 26 ng kg(-1), but did not affect the nociceptin-induced hyperalgesia at doses up to 500 ng kg(-1). BW 245C (an agonist for PGD (DP) receptor) blocked the allodynia with an IC50 of 83 ng kg(-1). 3. The blockade of nociceptin-induced allodynia by PGD2 was reversed by the potent and selective DP-receptor antagonist BW A868C in a dose-dependent manner with an ED50 of 42.8 ng kg(-1). 4. Glycine (500 ng kg[-1]) almost completely blocked the nociceptin-induced allodynia. A synergistic effect on the inhibition of nociceptin-evoked allodynia was observed between glycine and PGD2 at below effective doses. 5. Dibutyryl cyclic AMP, but not dibutyryl cyclic GMP, blocked the nociceptin-induced allodynia with an IC50 of 2.9 microg kg(-1). 6. PGE2, PGF2alpha, butaprost (an EP2 agonist) and cicaprost (a PGI receptor agonist) did not affect the nociceptin-induced allodynia. 7. These results demonstrate that PGD2 inhibits the nociceptin-evoked allodynia through DP receptors in the spinal cord and that glycine may be involved in this inhibition.     

Influence of ruminal or abomasal starch hydrolysate infusion on pancreatic exocrine secretion and blood glucose and insulin concentrations in steers

Seven Holstein steers (234 +/- 6.4 kg) surgically fitted with pancreatic cannulas, ruminal and abomasal infusion cannulas, and hepatic-portal vein catheters were used in a 4 x 7 incomplete Latin square design experiment to examine the influence of ruminal and abomasal carbohydrate infusion on enzyme secretion and composition of pancreatic juice. Four treatments were arranged in a 2 x 2 factorial: 1) ruminal starch hydrolysate (SH; 34.2 g/[h.site]) or abomasal water and 2) abomasal SH or ruminal water. Starch hydrolysate is raw cornstarch that has been partially digested by a heat-stable alpha-amylase. Experimental periods were 14 d with 9 to 10 d of adaptation, 4 d of pancreatic collection, and blood collection on d 14. Abomasal SH infusion tended (P < .10) to increase pancreatic fluid secretion. The pH of pancreatic juice was higher (P < .01) for ruminal SH infusion, and abomasal SH infusion tended (P < .10) to result in lower pH of pancreatic juice. alpha-Amylase concentrations (units/milliliter and units/milligram of protein) and secretion (units/hour) were less (P < .002) for abomasal SH infusion. Chymotrypsin concentration (units/liter; P < .01) and secretion (units/hour; P < .10) were less for abomasal SH infusion; however, a rumen x abomasal interaction was found (P < .05). Chloride concentration (milligrams/deciliter) and secretion (milligrams/hour) were increased (P < .01) for abomasal SH infusion. Abomasal SH infusion resulted in increased (P < .01) portal blood glucose concentrations; however, portal plasma insulin concentration was not affected (P > .10). Abomasal SH infusion altered alpha-amylase secretion in steers, but ruminal SH infusion had minimal effect on alpha-amylase secretion. These changes suggest abosamal infusion of SH may negatively impact secretion of pancreatic alpha-amylase.     

Effect of a non-steroidal anti-inflammatory drug on tissue levels of immunoreactive prostaglandin E2, immunoreactive leukotriene, and pain after periodontal surgery

The aim of this study was to measure tissue levels of immunoreactive prostaglandin E2 (iPGE2), immunoreactive leukotriene B4 (iLTB4), and pain after periodontal surgery and to evaluate the effect of the non-steroidal anti-inflammatory drug (NSAID), ibuprofen, on these levels. Two contralateral quadrants in each of nine patients were selected to undergo separate surgical procedures, one with ibuprofen (800 mg 1 hour presurgery and 400 mg postsurgery) and one with a placebo. Intra-operatively, a custom-made microdialysis probe, with a 3,000 dalton molecular weight cut-off, was inserted beneath the soft tissue flap and a dialysate collected every 20 minutes for 4 hours after surgery. Pain perception was measured at the same time intervals using two pain scales. Dialysate samples were assayed using two enzyme immunoassays. Mean tissue levels of iPGE2 in the placebo group increased from 74 nM at 40 minutes to a peak of 261 nM at 200 minutes. Mean tissue levels of iLTB4 in the placebo group fluctuated between 0.2 and 0.6 nM. Pain levels in this group increased continuously with time, peaking at 4 hours. Mean tissue levels of iPGE2 in the ibuprofen group were significantly suppressed, exhibiting more than a 95% reduction. This was accompanied by a significant reduction in pain. Ibuprofen had no detectable effect on tissue levels of iLTB4. These data indicate that iPGE2 and iLTB4 are present at relatively high concentrations in the periodontal tissues after surgery. Since these concentrations exceed the Kd values for binding to their respective receptors, PGE2 and LTB4 may be associated with the development of postsurgical pain and inflammation. These data also indicate that ibuprofen can successfully inhibit iPGE2 production in the periodontal tissues and in this way may help reduce postoperative pain and inflammation.     

Human hepatocyte nuclear factor-4 (hHNF-4) gene maps to 20q12-q13.1 between PLCG1 and D20S17

Male Hartley guinea pigs (480-610 g; n=5) were treated intratracheally with saline, cadmium (Cd, 0.3 mg) as cadmium chloride, selenium (Se, 0.3 or 0.06 mg) as sodium selenite or Se (0.06 mg) and Cd (0.3 mg). After 24 hours, lungs were collected and analyzed for prostaglandin (PGE2), thromboxane (TXB2) and leukotriene (LTC4) levels. Results indicated that, 0.3 mg Se and 0.06 mg Se in combination with 0.3 mg Cd increased PGE2 significantly. Selenium and Cd alone or in combination, decreased LTC4 and TXB2 significantly.     

Effects of prostaglandin E2 and F2 alpha on the skeleton of osteopenic ovariectomized rats

This article contains the histomorphometric evaluation of the effects of prostaglandin F2 alpha (PGF2 alpha) on cancellous bone from the lumbar vertebra and cortical bone from the tibial shaft of ovariectomized, osteopenic rats. These effects were then compared with those of prostaglandin E2 (PGE2). Three-month-old rats were either ovariectomized (ovx) or sham-ovx. Then, either PGF2 alpha or PGE2 in doses of 1 and 3 mg/kg/day was given subcutaneously for 21 days at 150 days post ovx. Histomorphometric analysis was performed separately on both the primary and secondary spongiosae of the fourth lumbar vertebral bodies (LVB) and on tibial shafts. The ovx rats exhibited osteopenia in both primary (-23% to -37%) and secondary (-20%) spongiosae of the LVB, but not in the tibial shafts at 150 and 171 days post ovx. In the LVB, PGE2 in doses of 1 or 3 mg/kg/day for 21 days restored trabecular bone volume to the levels of sham-ovx controls in the primary spongiosa. However, in the secondary spongiosa, the treatments only thickened the trabeculae. The effects of the PGF2 alpha treatment were similar to those of the PGE2 in both the primary and the secondary spongiosae. While both PGF2 alpha and PGE2 treatments stimulated bone formation in the LVB as indicated by the increases in labeled perimeter, tissue and bone area-based bone formation rates, PGE2 is about 10 times more potent than PGF2 alpha in these effects. The PGE2 treatment also elevated activation frequency in the LVB, while the PGF2 alpha treatment did not. The treatments differed in that PGE2 at these dose levels did not alter the eroded surface in the LVB while PGF2 alpha decreased it significantly. Thus, the increase of the ratio of labeled to eroded perimeter in the LVB in PGF2 alpha-treated animals was much more than that in PGE2-treated animals. In the tibial shafts, PGE2 in doses of 1 and 3 mg/kg/day produced new marrow trabeculae in 2 of 6 and 3 of 6 of the ovx rats. However, no new trabecula was found in PGF2 alpha-treated tibial shafts. Higher doses of PGE2 also increased periosteal labeled perimeter, MAR, and BFR/BS, while PGF2 alpha did not produce any significant change in these parameters. Both PGE2 and PGF2 alpha in doses of 1 and 3 mg/kg/day increased the labeled perimeter, MAR and BFR/BS and decreased the eroded perimeter in the endocortical surface. We concluded that both PGF2 alpha and PGE2 in doses of 1 and 3 mg/kg/day for 21 days exhibited anabolic bone effects. The effects were mostly confined to an increase in trabecular volume in the primary spongiosa of the LVB and in the endocortical surface of tibial shafts. The tissue level mechanism behind this appears to be that PGE2 and PGF2 alpha can both stimulate osteoblast recruitment and activity. Overall, we found PGE2 to be more potent than PGF2 alpha at the same dose level at the endocortical surface. Furthermore, new marrow trabecular bone formed only after PGE2 treatment. PGF2 alpha differed from PGE2 by significantly reducing the trabecular eroded surface in ovx rats.     

Cyclooxygenase-2-dependent delayed prostaglandin D2 generation is initiated by nerve growth factor in rat peritoneal mast cells: its augmentation by extracellular type II secretory phospholipase A2

When rat serosal connective tissue mast cells (CTMC) were stimulated with nerve growth factor (NGF), the immediate prostaglandin D2 (PGD2) generation was followed by delayed PGD2 generation that occurred between 2 and 24 h, reaching levels as high as 50 ng and 260 ng/10(6) cells in the absence or presence of lysophosphatidylserine (lysoPS), respectively. This delayed PGD2 generation was accompanied by de novo induction of cyclooxygenase (COX)-2, with NGF and lysoPS acting as inducer and enhancer, respectively. COX-2 induction and the attendant delayed PGD2 generation in CTMC were modestly induced by c-kit ligand, but not by Fc epsilonRI cross-linking. This indicated that the stimulus specificity differed from that observed in the immediate phase, in which NGF, c-kit ligand, and Fc epsilonRI cross-linking, either in combination with each other or with lysoPS as a cofactor, elicited comparable levels of PGD2 generation within 10 min, reaching 10 to 20 ng/10(6) cells. Addition of type II secretory phospholipase A2 (sPLA2), a PLA2 isoform that is detected in microg/ml levels in inflammatory exudates, to NGF-stimulated CTMC significantly augmented delayed, but not immediate, PGD2 generation, and this augmentative effect was mediated in part by the enhancement of COX-2 expression by sPLA2. These results suggest that CTMC have the capacity to produce PGD2 over a prolonged period in the presence of tissue-derived cytokines and sPLA2 in a COX-2-dependent manner.     

Unilateral nasal allergen challenge leads to bilateral release of prostaglandin D2

BACKGROUND: Multiple mediators including prostaglandin D2 and leukotriene B4 have been shown to increase in nasal secretions during the early response to nasal challenge with antigen. OBJECTIVE: Our objective was to investigate the time course of prostanoid and leukotriene B4 release into nasal secretions on both the ipsilateral and contralateral side after a unilateral nasal allergen challenge. METHODS: We performed a controlled, randomized trial. Six volunteers were challenged unilaterally with antigen or diluent in a randomized order and discs were used to collect nasal secretions from both nostrils at 2 min intervals for 20 min after the challenge. Prostanoids and leukotriene B4 (LTB4) in recovered nasal secretions were measured by combined capillary gas chromatography-negative ion chemical ionization mass spectrometry (GC/MS). RESULTS: Nasal allergen challenge resulted in a significant and immediate increase in symptoms and sneezing. PGD2 was significantly elevated above diluent values (0.6 +/- 0.6 pg) 30 s after removal of the allergen disc (P < 0.05), reached its peak (423.2 +/- 182.4 pg) at 2 min and then slowly decreased. PGD2 also increased on the contralateral side after unilateral allergen challenge, reaching peak values about six times lower than on the ipsilateral side (70.8 +/- 21.7 pg at 6 min). Levels of 9a, 11b-PGF2 after antigen provocation became significantly higher than after diluent (0 +/- 0 pg) on the ipsilateral side at 2 min (17.2 +/- 5.9 pg), and reached peak levels at 4 min (25.1 +/- 8.0 pg). LTB4 also increased significantly on the side of challenge. For the other prostanoids measured (PGF2, PGF2 alpha, TxB2, 6kPGF1 alpha), no significant changes in either ipsilateral or contralateral secretions were observed after allergen challenge. CONCLUSIONS: Our study described the kinetics of PGD2 and LTB4 release as well as the contralateral release of PGD2.     

Neurohormonal regulation of histamine release from isolated rabbit fundic mucosal cells

Histamine-containing cells isolated from rabbit fundic mucosa were found in a small cell elutriation fraction (cells with diameter about 9-12 microns) enriched in mucus and endocrine cells and containing less than 1% mast cells (F1 cells). Gastrin (HG-17), pentagastrin and CCK-8 (C-terminal octapeptide of cholecystokinin) dose-dependently stimulated histamine release (EC50, respectively, 0.126 +/- 0.03, 0.92 +/- 0.15 and 0.211 +/- 0.025 nM) and somatostatin inhibited this release. PGE1, PGE2 and PGD2 alone were unable to enhance histamine release even at high concentrations but, when used in combination with gastrin of CCK-8, the release of histamine caused by these peptides was potentiated (about 1.5- to 2-fold). Carbachol also enhanced the liberation of histamine but with a weaker potency and efficacy than the gastrointestinal peptides (EC50: 1.50 +/- 0.06 microM). The use of specific muscarinic antagonists for M1-, M2- and M3-type receptors led us to conclude that an M1 receptor might be involved in the muscarinic-induced stimulation of histamine release. Activators of protein kinase C, 12-O-tetradecanoylphorbol-13-acetate (TPA) and 1-oleyl-2-acetyl-glycerol (OAG) as well as the calcium ionophore, A23187, induced histamine release, whereas agents which increased intracellular cAMP content were devoid of effect.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lack of tactile pain (allodynia) in lipocalin-type prostaglandin D synthase-deficient mice

Prostaglandin (PG) D2 is the most abundant prostanoid produced in the central nervous system of mammals and has been implicated in the modulation of neural functions such as sleep induction, nociception, regulation of body temperature, and odor responses. We generated gene-knockout mice for lipocalin-type PGD2 synthase (L-PGDS) and found that the intrathecal administration of PGE2, an endogenous pain-producing substance, failed to elicit allodynia (touch-evoked pain), which is one typical phenomenon of neuropathic pain, whereas it evoked thermal hyperalgesia, in L-PGDS-/- mice. We also found that the allodynic response induced by the gamma-aminobutyric acid (GABA)A receptor antagonist bicuculline was selectively abolished in the L-PGDS-/- mice, among excitatory and inhibitory agents that induced allodynia in wild-type mice. Interestingly, simultaneous injection of a femtogram amount of PGD2 with PGE2 or bicuculline induced allodynia in L-PGDS-/- mice to the same extent as in wild-type mice. The PGE2- or bicuculline-evoked allodynia in wild-type and in PGD2-supplemented L-PGDS-/- mice was blocked by a PGD2 receptor antagonist given in a femtogram amount. These results reveal that endogenous PGD2 is essential for both PGE2- and bicuculline-induced allodynia.