Effect of flunixin meglumine on plasma prostanoid concentrations in horses with colic in the perioperative period

In the present study the significance of eicosanoids in the development of shock in horses on the basis of ileus has been investigated using the prostanoids thromboxane B2 (TXB2) and prostaglandine E2 (PGE2) as indicators. The prostanoid synthesis inhibitor flunixin meglumine was to be examined regarding its efficacy in the effective blockade of the synthesis of these mediators within the peri-operative timeframe as well as its effects on clinical signs and laboratory parameters. 21 horses suffering from ileus and ready for surgical intervention received an intravenous flunixin dosis of 1.1 mg/kg body weight immediately after the initial examination and prior to the surgical procedure. 20 colic horses receiving surgical treatment without application of the drug served as control group. Reference data concerning the approximate standard plasma levels of the prostanoids were determined in 10 healthy horses. Plasma levels of thromboxane B2 and prostaglandine E2 in all colic horses, treatment group as well as controls, initially proved to be significantly higher than the reference values in healthy horses. The untreated control group showed plasma levels highly exceeding the standards within the course of investigation. The application of flunixin meglumine resulted in an effective inhibition of the prostanoid synthesis. Post-operatively as well as within the whole period of investigation the plasma levels of PGE2 and TXB2 of the treated group were considerably lower than those of the control group. Flunixin meglumine had a favorable effect on several cardiovascular parameters. The experimental data concerning the effects of flunixin meglumine thus could be validated in a clinical setting, especially the effective inhibition of the cyclooxygenase enzyme system. The application of the prostanoid synthesis inhibitor flunixin meglumine can be judged as being effective in limiting shock progress in the peri-operative setting given reliable diagnosis.     

Differential sensitivities of human blood monocytes and alveolar macrophages to the inhibition of prostaglandin endoperoxide synthase-2 by interleukin-4

Interleukin-4 (IL-4) is a potent immunomodulatory cytokine synthesized and released by Th2 lymphocytes, mast cells and basophils. It has important effects on monocyte/macrophage cell lines, regulating the secretion of several cytokines, and the production of eicosanoids. In human monocytes and macrophages, IL-4 increases the expression of 15-lipoxygenase and 15-HETE production, but suppresses the inducible isoform of the prostaglandin H synthase (PGHS-2) enzyme and prostanoid synthesis. Prostanoids, in particular prostaglandin E2 (PGE2) have important functions in modulating inflammatory and fibrotic processes. We compared the effect of IL-4 on the expression of PGHS-2 in human alveolar macrophages (AM) and blood monocytes (BM) activated with physiologically distinct stimuli, lipopolysaccharide (LPS) or IL-1 in vitro. The induction of PGHS-2 mRNA and protein, and prostanoid synthesis by all stimuli was inhibited by exogenous IL-4 in both cell types. However, monocytes were more susceptible to this effect of IL-4 than alveolar macrophages.     

Measurements of urinary prostaglandins in young ovulatory women during the menstrual cycle and in postmenopausal women

The purpose of the present work was to study the prostaglandin excretion in young nonpregnant ovulatory women during the menstrual cycle on the one hand and in postmenopausal women on the other hand and to investigate the influence of female sex hormones (estradiol, progesterone) on urinary prostanoid excretion. Urinary excretion rates of prostaglandin E2 (PGE2), 6-keto-PGF1 alpha, thromboxane B2 (TxB2) and their metabolites PGE-M (11 alpha-hydroxy-9, 15-dioxo-2,3,4,5,20-pentanor-19-carboxyprostanoic acid), 2,3-dinor-6-keto-PGF1 alpha, 2,3-dinor-TxB2 and 11-dehydro-TxB2 were determined by gas chromatography-triple stage quadrupole mass spectrometry (GC/MS/MS) in 41 young non-pregnant women during the follicular phase and during the luteal phase and in 23 postmenopausal women. Excretion rates of all urinary prostanoids were not significantly different in the follicular phase when compared with the luteal phase. In contrast to the young ovulatory women, PGE2 and TxB2 were significantly higher in postmenopausal women. Concerning the other prostaglandins significant differences between these groups of women did not exist. Although serum levels of estradiol and progesterone were different in young and postmenopausal women, sex hormones have not been shown to correlate with prostaglandins. Our data do not suggest sex hormones to be responsible for the difference in the prostaglandin excretion in women of reproductive age and in women in the menopause. Further systematic investigations into age dependency of prostaglandin excretion in women are necessary.     

Regulation of cyclooxygenase-2 and prostaglandin F synthase gene expression by steroid hormones and interferon-tau in bovine endometrial cells

Estradiol (E2) and progesterone are responsible for regulating PG synthesis in the endometrium during the estrous cycle and interferon-tau (IFN-tau) alters PG synthesis during early pregnancy in ruminants. In this study, we examined the effects of these steroid hormones and recombinant bovine IFN-tau (rbIFN-tau) on PG production and on cyclooxygenase-2 (COX-2) and PG F (PGF) synthase (PGFS) gene expression in isolated endometrial cells. E2 decreased both PGF2alpha and PG E2 (PGE2) whereas progesterone increased PGF2alpha secretion in epithelial cells. Steroid hormones had no effect on PG production in stromal cells. rbIFN-tau attenuated both PGF2alpha and PGE2 production in epithelial cells and enhanced their production, and the ratio of PGE2 to PGF2alpha, in stromal cells. Northern blot analysis showed that E2 and rbIFN-tau decreased COX-2 messenger RNA (mRNA) levels in epithelial cells. Conversely, rbIFN-tau increased COX-2 mRNA in stromal cells. Furthermore, rbIFN-tau decreased PGFS mRNA in both cell types and this was associated with the increase in PGE2/PGF2alpha ratio. These results show that the regulation of PG synthesis by steroid hormones is different in endometrial epithelial and stromal cells in vitro. The attenuation of PGF2alpha secretion from epithelial cells and increased PGE2 production in stromal cells by rbIFN-tau are modulated by steroid hormones.     

The emerging role of paclitaxel in breast cancer therapy

Bradykinin and thrombin caused a time- and dose-dependent stimulation of prostanoid biosynthesis in human dental-pulp fibroblasts, as assessed by the release of prostaglandin E2 (PGE2) and 6-keto-prostaglandin F1 alpha (the stable breakdown product of prostacyclin). The stimulatory effect of bradykinin and thrombin on PGE2 biosynthesis was maximal within 5-10 min. The concentration of bradykinin producing half-maximal stimulation (EC50) of PGE2 and prostacyclin formation was 10 nM. EC50 for thrombin-induced formation of PGE2 and prostacyclin were 0.05 and 0.2 U/ml, respectively. Bradykinin analogues with affinity to the bradykinin B2 receptor, but not those with affinity to the B1 receptor, caused a burst of PGE2 formation. The stimulatory action of bradykinin and thrombin on PGE2 biosynthesis was abolished by two structurally different cyclo-oxygenase inhibitors and significantly reduced by two corticosteroids. Thrombin dose-dependently enhanced the incorporation of [3H]-thymidine into DNA in pulpal fibroblasts by a mechanism that was unrelated to the effect on prostanoid biosynthesis. Bradykinin did not affect thymidine incorporation. Thrombin, but not bradykinin, stimulated the biosynthesis of type 1 collagen in the pulpal fibroblasts. The stimulatory effect of thrombin on collagen biosynthesis was not affected by cyclo-oxygenase inhibitors. These data show that human dental-pulp fibroblasts are equipped with receptors for bradykinin and thrombin linked to enhanced prostanoid biosynthesis. Occupancy of the thrombin receptors also leads to a prostaglandin-independent stimulation of cell proliferation and collagen biosynthesis.     

Enhancement of cell proliferation and prostaglandin biosynthesis by 1,8-dihydroxyanthraquinone in the rat large intestine

The effects of 1,8-dihydroxyanthraquinone (DHAQ), a stimulant laxative named danthron, on cell kinetics and prostaglandin (PG) biosynthesis in the gastrointestinal tract were investigated in male 8-week-old F344 rats divided into three groups, each consisting of 10 animals. The animals in groups one, two and three were respectively given diets supplemented with 0%, 0.1% and 0.2% DHAQ for 24 days. PGE2 levels in the colorectal mucosa were significantly (P < 0.05 and 0.001) elevated after DHAQ treatment and showed some evidence of a dependence of DHAQ dose, consistent with the plasma PGE2 levels. BrdU-labeling indices in the large intestinal epithelium were also significantly (P < 0.01) increased, although the other portions of the gut such as the stomach and small intestine were not significantly affected. Excretion of the main urinary metabolite of PGE (PGE-MUM) was significantly (P < 0.001 or 0.01) increased whereas the urinary PGE2 concentration and total PGE2 excretion were not changed. Thus the results of the present study clearly indicate enhancement of cell proliferation by DHAQ in the large intestine epithelia, correlated with increased PGE2 levels in the large intestinal mucosa as well as the plasma, and possible support for the conclusion that quantitative analysis of urinary PGE-MUM, but not PGE2 itself, offer a useful approach for biomonitoring exposure to such stimulant laxatives.     

Mitochondrial disorders

The production of Prostaglandin E2 (PGE2) and Thromboxane B2 (TXB2) by turkey blood monocytes and a chicken mononuclear phagocytic cell line MQ-NCSU after exposure to vitamin E (VE) was examined. Turkey embryos were exposed in ovo to 0 and 10 international units (IU) of VE; blood monocytes were collected at 2 weeks of age and cultured. MQ-NCSU macrophage monolayers were exposed to 0, 0.1, 0.25, and 0.5 IU VE. The monocyte/macrophage cultures were exposed to 1 microgram/mL bacterial lipopolysaccharide (LPS). Non-stimulated parallel cultures were maintained as controls. The PGE2 and TXB2 levels were quantitated in culture supernatants by a competitive ELISA. Blood monocytes from the 10 IU VE poults produced lower PGE2 levels as compared with the 0 IU VE controls. Upon stimulation with LPS, monocytes from the 10 IU VE group exhibited levels of PGE2 that were higher than the 0 IU VE group. Levels of TXB2 were not quantitated in the poult blood monocyte culture supernatants. The PGE2 and TXB2 levels in the supernatant of the VE treated MQ-NCSU macrophage cultures were lower than the 0 IU VE controls. Stimulation with LPS resulted in increased PGE2 and TXB2 production by the VE-exposed macrophages. The results from this study suggest that in ovo or in vitro exposure with VE may either upregulate or downregulate PGE2 and TXB2 production by monocytes/macrophages, and that this production may be dependent upon the exposure to a variety of external stimuli and/or the state of macrophage activation.     

CD40 engagement up-regulates cyclooxygenase-2 expression and prostaglandin E2 production in human lung fibroblasts

A newly emerging view of fibroblasts is that they are vital for initiating inflammation and respond to and direct the activities of leukocytes. Human fibroblasts can express CD40, an activation Ag the ligand of which is displayed by activated leukocytes. We demonstrate here that CD40 engagement on human lung fibroblasts dramatically increases proinflammatory PGE2 synthesis. This up-regulation is mediated through an induction of cyclooxygenase-2 (Cox-2) since Cox-2-selective inhibitors block the up-regulation. Western and Northern blot analyses demonstrated that Cox-2 protein and mRNA are dramatically increased in fibroblasts following CD40 engagement. We conclude that CD40 is a major pathway in human fibroblasts for the induction of Cox-2. There is intense interest in devising strategies for disruption of the CD40-CD40 ligand system to blunt inflammation. Such an intervention would be expected to attenuate the up-regulation of fibroblast Cox-2 and PGE2 production at the site of tissue injury.     

Lipopolysaccharide induces prostaglandin H synthase-2 protein and mRNA in human alveolar macrophages and blood monocytes

We and others have previously demonstrated that human alveolar macrophages produce more PGE2 in response to lipopolysaccharide (LPS) than do blood monocytes. We hypothesized that this observation was due to a greater increase in prostaglandin H synthase-2 (PGHS-2) enzyme mass in the macrophage compared to the monocyte. To evaluate this hypothesis, alveolar macrophages and blood monocytes were obtained from healthy nonsmoking volunteers. The cells were cultured in the presence of 0 to 10 micrograms/ml LPS. LPS induced the synthesis of large amounts of a new 75-kD protein in human alveolar macrophages, and a lesser amount in monocytes. Synthesis of this protein required more than 6 h and peaked in 24 to 48 h; the protein reacted with an anti-PGHS-2 antibody prepared against mouse PGHS-2. Associated with synthesis of the protein was a marked increase in LPS-stimulated and arachidonic acid-stimulated synthesis of PGE2 by alveolar macrophages compared to monocytes. Cells not exposed to LPS contained only PGHS-1 and synthesized very little PGE2 during culture or in response to exogenous arachidonic acid. An LPS-induced mRNA, which hybridized to a human cDNA probe for PGHS-2 mRNA, was produced in parallel with production of this new protein and was produced in much greater amounts by alveolar macrophages compared to blood monocytes. This mRNA was not detectable in cells not exposed to LPS. In contrast, both types of cells contain mRNA, which hybridizes to a cDNA probe for PGHS-1. This mRNA did not increase in response to LPS. LPS also had no effect on PGHS-1 protein. These data demonstrate that PGE2 synthesis in human alveolar macrophages and blood monocytes correlates to the mass of PGHS-2 in the cell. We conclude that the greater ability of the macrophage to synthesize PGE2 in response to LPS is due to greater synthesis of PGHS-2 by the macrophage.     

Induction of cyclooxygenase-2 by parathyroid hormone in human osteoblasts in culture

OBJECTIVE: Parathyroid hormone (PTH) induced bone resorption by osteoclasts depends on the presence of osteoblasts. PTH induced production of prostaglandins by osteoblasts and induction of bone resorption by prostaglandins suggest that these autacoids may be implicated in the effects of PTH on bone. Our objective was to determine if the increase in prostaglandin production induced in human osteoblasts by PTH is due to an increase in cyclooxygenase-2 (COX-2) expression. METHODS: Primary cultures of human osteoblasts were obtained from specimens of trabecular bone. Confluent cells were treated with PTH, dexamethasone or compound NS-398, a specific COX-2 inhibitor. The concentration of prostaglandin E2 (PGE2) in the supernatants was determined by radioimmunoassay and COX-2 mRNA levels evaluated by Northern blot. RESULTS: PTH induced COX-2 mRNA expression and PGE2 production. These effects were time and concentration dependent and were inhibited by dexamethasone. Compound NS-398 reduced PGE2 production to the same extent as dexamethasone, and neither compound had an additive effect on this variable. CONCLUSION: These results show that PTH induces COX-2 expression in human osteoblasts in culture and suggest that this isoenzyme is the main factor in the control of prostaglandin synthesis in these experimental conditions.     

No deterioration in insulin sensitivity, but impairment of both pancreatic beta-cell function and glucose sensitivity, in Japanese women with former gestational diabetes mellitus

Recent studies have demonstrated a strong correlation between infection and preterm labor. Preterm delivery is also associated with high levels of cytokines and prostaglandins in amniotic fluid. The purpose of this study was to investigate the effect of tumor necrosis factor-alpha (TNF-alpha) on the levels of cyclooxygenase, prostaglandin E2 production (PGE2), and expression of the PGE2 receptor subtype EP1 in amnion WISH cell culture. Amnion WISH cell cultures were incubated in increasing concentrations of TNF-alpha (0-50 ng/ml). Changes in cyclooxygenase and EP1 receptor proteins were evaluated by Western blot analysis. Changes in EP1 mRNA were evaluated by Northern blot, and culture fluid concentrations of PGE2 were estimated by enzyme immunoassay (EIA). EP1 protein (p<0.01), EP1 mRNA (p<0.05), cyclooxygenase-2 (COX-2) protein (p<0.001), and PGE2 concentrations (p<0.01) all increased with increasing concentrations of TNF-alpha. Changes in COX-1 protein were not observed following TNF-alpha-incubation. The results suggest that TNF-alpha may play a role in infection-induced preterm labor by its pleiotropic ability to simultaneously stimulate COX-2 activity, PGE2 concentrations, and PGE2 EP1 receptor levels in human amnion.     

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

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSION: This study may have important implications in the potentiation of NK and LAK cell activity for immunotherapeutic protocols in patients with breast cancer.     

Identification of hepatitis B surface antigen variants with alterations outside the "a" determinant in immunized Singapore infants

Although histological studies have suggested that endothelial cells in bone (BDECs) are associated with some osteolytic bone diseases, it is still unclear how BDECs contribute to bone remodeling. Here we examined the response of BDECs to basic fibroblast growth factor (bFGF, FGF-2) using primary and cloned murine BDECs isolated from the femurs of BALB/c mice. Treatment of primary and cloned BDECs with bFGF induced cyclooxygenase-2 (COX-2) mRNA and protein expression. Furthermore, bFGF promotes the production of prostaglandin E2 (PGE2), which is known to be a potent stimulator of bone resorption and to induce osteoclast formation. Because the secretion of PGE2 was suppressed by COX-2 specific inhibitor NS-398 and by COX-2 antisense oligodeoxynucleotides, bFGF promotes the synthesis of PGE2 in a COX-2-dependent manner. Therefore, endothelial cells in bone are associated with bone remodeling by controlling COX-2 expression and consequently PGE2 production.     

Regulation of cyclooxygenase-2 expression in bovine chondrocytes in culture by interleukin 1alpha, tumor necrosis factor-alpha, glucocorticoids, and 17beta-estradiol

OBJECTIVE: To determine the effects of interleukin 1alpha (IL-1alpha), tumor necrosis factor-alpha (TNF-alpha), dexamethasone, and 17beta-estradiol on the expression of cyclooxygenase-1 (COX-1) and COX-2 in bovine chondrocytes. METHODS: Northern blot analysis was used to quantify COX-1 and COX-2 mRNA expression in primary cultures of bovine chondrocytes and prostaglandin production to evaluate COX activity. RESULTS: IL-1alpha and TNF-alpha increased the expression of COX-2. This effect was independent of de novo protein synthesis and dependent on increased mRNA stability in the case of IL-1alpha. Dexamethasone inhibited the effects of both cytokines. 17beta-estradiol inhibited COX-2 mRNA expression in basal conditions, but had no effect on COX-2 expression induced by cytokines. The specific COX-2 inhibitor compound NS 398 prevented the increase in prostaglandin E2 (PGE2) production induced by the cytokines. COX-1 levels remained stable with all treatments. CONCLUSION: Increase in mRNA stability is a mechanism implicated in the induction of COX-2 by some cytokines. The effects of IL-1alpha and TNF-alpha on PGE2 production are mainly due to an increase in COX-2 activity as shown by the effect of compound NS 398. 17beta-estradiol inhibits COX-2 mRNA expression in basal conditions, suggesting that estrogens could be implicated in the control of cartilage metabolism.     

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.