Release Of Cyclooxygenase Metabolites Research Articles

Traditional Chinese formula, lubricating gut pill, improves loperamide-induced rat constipation involved in enhance of Cl − secretion across distal colonic epithelium

Aim of the study Lubricating gut pill (LGP), a traditional Chinese formula, was widely used for the treatment of chronic constipation, especially in the elderly, in China. However, it is unclear whether LGP-induced laxative and/or lubricating effect is involved in water and electrolytes transport in distal colonic epithelium. Materials and methods The present study was designed to evaluate the effect of LGP on Cl − secretion across rat distal colonic epithelium mounted in Ussing chambers, and on a rat constipation model induced by loperamide, respectively. Results Application of LGP in the apical side elicited a sustained increase in short circuit current ( I SC) response in a concentration-dependent manner. Evidence that LGP-stimulated I SC was due to Cl − secretion is based on inhibition of current by (a) a Na +–K +–2Cl − cotransporter inhibitor bumetanide, (b) removal of Cl − ions in bath solution, and (c) the cystic fibrosis transmembrane conductance regulator (CFTR) Cl − channel blocker DPC, suggesting that a apical cAMP-dependent Cl − channel was activated. LGP-stimulated I SC was also strongly inhibited by pretreatment with clotrimazole, indicating that the basolateral K + channel was also involved in maintaining this cAMP-dependent Cl − secretion. Pretreatment of tissues with indomethacin, but not atropine, tetrodotoxin or hexamethonium, inhibited LGP-induced response. In a rat constipation model, oral administration with LGP was significantly restored number of fecal pellets, water content and mucus secretion compared with loperamide-treated group alone. Conclusions LGP enhances Cl − secretion that is mostly mediated through the release of cyclooxygenase metabolites, by which provided an osmotic force for the subsequent laxative action observed in the rat constipation model.

Barakol Extracted from Cassia siamea Stimulates Chloride Secretion in Rat Colon

Barakol is a purified extract of Cassia siamea, a plant that has been used as a laxative in traditional medicine. In this study, the effect of barakol on anion transport across the rat colon epithelium was investigated. Colonic epithelium was mounted in Ussing chambers and bathed with Ringer's solution. Addition of 1 mM barakol to the basolateral solution produced a slow increase in short-circuit current (Isc) in proximal colon and distal colon by 24.5 +/- 2.2 and 24.2 +/- 1.4 microA/cm(2), respectively. Barakol increased Isc in a concentration-dependent manner with an EC(50) value of 0.4 mM. The barakol-stimulated increase in Isc was inhibited by subsequent treatment with 500 microM diphenylamine-2-carboxylic acid or 400 microM glibenclamide added to the apical solution and 200 microM bumetanide added to the basolateral solution. Pretreatment of the tissues with 200 microM bumetanide, but not 10 microM amiloride, completely abolished the barakol-increased Isc. Ion substitution experiments showed an inhibition of barakol-stimulated Isc in chloride-free solution but not in bicarbonate-free solution. In addition, pretreatment of tissues with 10 microM tetrodotoxin or 10 microM indomethacin, but not 1 microM atropine or 10 microM hexamethonium, partially inhibited the Isc response by barakol. The present results demonstrated the stimulatory effect of barakol on the bumetanide-sensitive chloride secretion in rat colon. The effect of barakol was partly mediated by the stimulation of submucosal nerves and through the release of cyclooxygenase metabolites. These findings thus provide an explanation for the underlying mechanism of barakol as a secretagogue in mammalian colon.

Platelet-activating factor dilates efferent arterioles through glomerulus-derived nitric oxide.

Despite evidence that platelet-activating factor (PAF) is produced by the glomerulus, its direct action on the glomerular microcirculation is poorly understood. It was recently reported that at picomolar concentrations, PAF dilates isolated microperfused afferent arterioles (Af-Art) via nitric oxide (NO). The present study tested the hypothesis that PAF acts on the glomerulus to release NO, which in turn controls the resistance of the efferent arteriole (Ef-Art). Rabbit Ef-Art were perfused from the distal end (retrograde perfusion [RP]) to eliminate the influence of the glomerulus, or through the glomerulus from the end of the Af-Art (orthograde perfusion [OP]) to maintain the influence of the glomerulus. Ef-Art were preconstricted by approximately 40% with norepinephrine and increasing doses of PAF were added to both the arteriolar perfusate and bath. Only with OP did PAF at picomolar concentrations cause significant dilation: at 400 pmol, the diameter increased by 64 +/- 11% from the preconstricted level (N = 6, P < 0.01). This dilation was completely abolished by pretreatment with an NO-synthesis inhibitor. To study its possible constrictor action, PAF was added to nonpreconstricted Ef-Art. At nanomolar concentrations, PAF constricted Ef-Art similarly in both RP and OP: at 40 nM, the diameter decreased by 24 +/- 4% (N = 6, P < 0.01) and 20 +/- 2% (N = 6, P < 0.01), respectively. This constriction was attenuated by pretreatment with indomethacin (Indo) in both RP (14 +/- 2%, N = 7; P < 0.02 versus without Indo) and OP (10 +/- 2%, N = 6; P < 0.02 versus without Indo). (1) at picomolar concentrations, PAF stimulates the glomerulus to release NO, which in turn dilates the Ef-Art; and (2) at nanomolar concentrations, PAF constricts the Ef-Art partly through release of cyclooxygenase metabolites. Thus, PAF may play a role in glomerular hemodynamics under various physiological and pathological conditions.

Characterization of ETB receptors mediating contractions induced by endothelin-1 or IRL 1620 in guinea-pig isolated airways: effects of BQ-123, FR139317 or PD 145065.

1. We have characterized the receptors mediating contractions to endothelin-1 (ET-1) or IRL 1620, an ETB receptor selective agonist, in isolated strips of tissue prepared from different parts of the guinea-pig airways. We used as antagonists BQ-123 and FR139317 (ETA receptor-selective) and PD 145065 (ETA/ETB receptor non-selective). 2. ET-1 and IRL 1620 (10(-10) M to 10(-6) M) caused similar concentration-dependent contractions of strips of guinea-pig trachea and upper bronchus. In the guinea-pig trachea without epithelium or lung parenchyma, IRL 1620 was less potent than ET-1. 3. In the trachea, contraction to ET-1 (< 10(-8) M) was preceded by a transient relaxation which was inhibited by BQ-123 (10(-5) M) or FR 139317 (10(-5) M) or by the removal of the epithelium. The concentration-response curve to ET-1 in the trachea was shifted to the right by PD 145065 (10(-5) M to 10(-4) M). PD 145065 (10(-4) M) also inhibited the response to ET-1 (3 x 10(-7) M) by 55%. Contractions induced by IRL 1620 were not affected by BQ-123 (10(-6) M) or FR139317 (10(-6) M) but were significantly attenuated by 10(-5) M of either antagonist. PD 145065 at 10(-6) M strongly attenuated and at 10(-5) M abolished contractions induced by IRL 1620. 4. In the trachea, removal of the epithelium potentiated the effects of both agonists. BQ-123 (10-5 M)had no effect on contractions of the trachea without epithelium induced by ET-1, but FR139317 (10-5 M)caused a significant inhibition. PD 145065 (10-5 M to 10-4 M) caused a shift to the right of the ET-1 concentration-response curve without affecting the contractile effect at 3 x 10-7 M. All three antagonists inhibited contractions induced by IRL 1620.5. In the upper bronchus, BQ-123 (10-5 M) did not affect contractions induced by ET-1, whileFR139317 (10-5 M) attenuated (20-26%) only contractions induced by 1-3 x 10-7 M ET-1. PD 145065(10-5 M to 10-4 M) caused a shift to the right of the ET-1 concentration-response curve. The contractions induced by IRL 1620 were inhibited by BQ-123 or FR139317 (10-5M to 10-4 M). PD 145065(10-6 M) strongly inhibited contractions induced by IRL 1620 and PD 145065 (10-5 M) totally abolished them.6. The contractile action of ET-1 in the lung parenchyma was significantly and similarly attenuated by BQ-123 (10-5 M) or indomethacin (10-5 M), while FRI39317 (10-5 M) was less effective. PD 145065(10-6 to 10-5 M) inhibited contractions to ET-1. IRL 1620, which is less potent than ET-1 in this preparation, was antagonized by PD 145065 (10-5 to 10-6 M) but unaffected by BQ-123 (10-6 M to10-5M) or FR139317 (10-6 M).7. Thus, ETB receptors mediate contractions to ET-1 in all four guinea-pig airway preparations. In addition, contractions to ET-1 in the trachea and lung parenchyma are mediated in part by ETA receptors. In the latter tissue, these ETA receptors mediate contraction through the release of cyclooxygenase metabolites. Similarly, ETA receptors located on the epithelial cells also mediate the release of prostanoids in the trachea with epithelium but they are responsible for transient relaxations. Interestingly,contractions induced by IRL 1620 were more susceptible to inhibition by the different antagonists,most probably because it binds to the endothelin receptors in a reversible manner. High concentrations(10-5 M) of ETA-selective antagonists also inhibit responses to IRL 1620, most probably by an effect at ETB receptors in both the trachea and the upper bronchus.

Effects of vasoactive intestinal polypeptide on antigen-induced bronchoconstriction and thromboxane release in guinea-pig lung.

1. Exogenous vasoactive intestinal polypeptide (VIP) infused into the pulmonary artery of isolated and ventilated lungs of guinea-pigs decreased, in a dose-dependent fashion (1.0-10.0 nmol), airway resistance and thromboxane B2 (TXB2, the stable hydrolysis product of TXA2) release in the perfusion medium. Prostacyclin (PGI2) synthesis, as reflected by the release of its stable hydrolysis product 6-oxo-PGF1 alpha, was unaffected. Pretreatment with the 5-lipoxygenase inhibitor BWA4c (3.5 x 10(-5) M) did not modify the bronchodilatory effect of VIP or its inhibitory action on TXB2 release. 2. Basal release of immunoreactive VIP from perfused lungs decreased from an initial value of 0.96 +/- 0.10 ng min-1 (mean +/- s.e.mean) in the first 2 min to an average of 0.58 +/- 0.10 ng min-1 in the following 15-20 min. 3. Antigen challenge with ovalbumin (0.1%) in sensitized lungs caused an anaphylactic reaction in 45% of tested lungs, concomitant with a 5 fold increase in both VIP and TXB2 release. Tetrodotoxin pretreatment (10(-6) M) reduced basal VIP release by > 80% and abolished the VIP increase observed during anaphylaxis, without modifying TXB2 release or the bronchoconstrictor response. 4. Indomethacin (10(-6) M) inhibited TXB2 synthesis and release by > 90%, delayed the bronchoconstrictor response and blunted the increased VIP release during lung anaphylaxis, without influencing basal VIP release. 5. The 5-lipoxygenase inhibitor BWA4c (3.5 x 10(-5) M) blunted the increase of TXB2 and VIP release from guinea-pig lung and attenuated the bronchoconstrictor response following ovalbumin challenge. 6. The administration of exogenous VIP as a continuous infusion (10-8 M) attenuated the bronchoconstriction and the release of cyclo-oxygenase metabolites following antigen challenge.7. Acetylcholine (10-6-l0-5 M) infused into the pulmonary artery induced a dose-dependent bronchoconstriction not associated with enhanced VIP or TXB2 release.8. The TXA2 mimetic U-46619 (0.1-1.0 nmol) caused dose-dependent increases in airway resistance,concomitant with an up to 10 fold increase in VIP release. VIP inhibited arachidonate-induced in vitro aggregation of washed rabbit platelets in a dose-dependent manner over a dose range 10-8 10-6 M.Despite the antiaggregatory effect of VIP, TXB2 and PGE2 synthesis was reduced only to a minor extent,and there was no redirection of arachidonate metabolism from TXA2 to PGE2, indicating that VIP does not act as a TX synthase inhibitor in vitro.9. We conclude that VIP may play a role in regulating bronchial smooth muscle reactivity in lung anaphylaxis by inhibiting the synthesis and release of TXA2, a potent vasoactive and bronchoconstrictor agent. TXA2, on the other hand, strongly enhances neuronal VIP release.

The Effects of Different Silicas on Arachidonic Acid Metabolism in Alveolar Macrophages

Bovine alveolar macrophages (BAM) prelabeled with 3H-arachidonic acid (AA) were exposed in vitro to different doses of DQ-12, Minusil-5, and Sigma silicas, or carbonyl iron beads. Arachidonic acid metabolites released into the culture medium by BAM were identified and quantitated using high performance liquid chromatography (HPLC). Cytotoxicity was assessed by the release of lactate dehydrogenase (LDH). At doses of 0.1 or 0.25 mg of DQ-12 silica and of 0.25 or 0.5 mg of Minusil-5 and Sigma silica, the release of cyclooxygenase metabolites (TXB2, PGE2, PGF2, and HHT) comprised greater than 95% of the total released AA metabolites. Silica doses above 0.5 mg led to 5-lipoxygenase metabolite release (LTB4, its two nonenzymatic isomers, and 5-HETE). This shift to 5-lipoxygenase metabolite release paralleled increased cellular cytotoxicity and was observed for each of the silicas. In contrast to silica stimulation, carbonyl iron beads elicited only small quantities of cyclooxygenase metabolites, no 5-lipoxygenase metabolites, and showed little cytotoxicity toward BAM. The relative potency of each particulate for stimulating the release of AA metabolites and LDH was calculated with DQ-12 greater than Minusil-5 greater than Sigma much greater than carbonyl iron beads. Our results indicate that the cytotoxic and presumed fibrogenic potential of a silica may be correlated with the potency to stimulate the release of 5-lipoxygenase metabolites from AM.

Copper modulation of macrophage cyclooxygenase metabolite synthesis

The effect of copper on the release of cyclooxygenase metabolites from starch elicited, rat, peritoneal macrophages was investigated. Copper sulphate, in the range 10 −6−10 −5M, inhibited the formation of prostaglandin (PG) E 2 and thromboxane (Tx) B 2, the stable metabolite of TxA 2, in a dose dependent manner but had no effect on the production of 6ketoPGF 1α, the stable product of prostacyclin. At higher concentrations (5×10 −5 and 10 −4M) the synthesis of all three metabolited of arachidonic acid (AA) was stimulated as was the release of radioactivity from macrophages prelabelled with 14C AA. Copper had no effect on the metabolims of exogenous AA however. At 10 −4M copper also stimulated secretion of the lysosomal enzyme, beta-glucuronidase (GUR). Copper nitrate (10 −4M), but not zinc sulphate, also stimulated eicosanoid formation and lysosomal enzyme release. Our results are consistent with the idea that copper stimulated eicosanoid formation via an effect on PL activity.

Dexamethasone inhibits the antigen-induced contractile activity and release of inflammatory mediators in isolated guinea pig lung tissue.

Treatment for 24 h in vitro with dexamethasone inhibited the antigen-induced contractile response in guinea pig tracheal rings and parenchymal strips without inhibiting the contractile response of the tissues to either methacholine or histamine, respectively. Antigen-induced histamine release was inhibited by approximately 50% in both tissues by prior treatment with dexamethasone. Dexamethasone treatment also inhibited the release of immunoreactive sulfidopeptide leukotriene from parenchymal strips. In tracheal rings, dexamethasone treatment reduced spontaneous release of all cyclooxygenase metabolites (PGE2, PGF2 alpha, TXB2, PGD2, and 6-k-PGF1 alpha were tested), with the exception of PGD2, and also inhibited the antigen-induced release of all cyclooxygenase metabolites studied. Dexamethasone-treatment did not inhibit the spontaneous release of cyclooxygenase metabolites in the guinea pig lung strips, and only modestly inhibited the antigen-induced release of PGE2, PGF2 alpha, and PGD2. The results suggest that the inhibition of contractile response of guinea pig lung strips and airway tissue to antigen by dexamethasone is the result of a reduced release of inflammatory mediators. The inhibition by dexamethasone of antigen-induced release of mast cell mediators from guinea pig lung parenchyma contrasts with results previously obtained with human parenchymal lung tissue.

Infectious-inflammatory changes in cyclic AMP levels and in their regulation by prostaglandins in human peritoneal macrophages

Peritoneal macrophages of renal patients on continuous ambulatory peritoneal dialysis (CAPD) have been collected when CAPD was without complications, during an intercurrent infectious peritoneal inflammation and after recovery. Levels of cyclic AMP, release of cyclo-oxygenase metabolites, and responsiveness, in terms of cyclic AMP elevation, to either PGE 2 or to DC-PGI 2 (a stable analogue of PGI2) were examined. Peritoneal. inflammation was associated with a sharp drop in cyclic AMP, which was restored after recovery. Production of TxA 2, PGI 2 and PGE 2 parallelled the direction of changes in cyclic AMP levels, except, that release of PGE 2 entirely failed to recover. Macrophages during the uncomplicated stage of CAPD proved more responsive to DC-PGI 2 than to PGE 2.During inflammation the cells displayed a marked increase in sensitivity towards PG stimulation. Improved sensitivity was more pronounced with PGE 2 than with DC-PGI 2 and so the original difference between responsiveness of the cells to the PGs was abolished. Several findings are compatible with the view that endogenous PGI 2 governs the cyclic AMP levels in human non-inflammatory peritoneal macrophages. However, during infectious-inflammation the cells undergo changes which render a reduced production of PGI2 insufficient to explain the drop in cyclic AMP.