Prostaglandin-related immune suppression in cattle.

Prostaglandins (PGs) are lipid mediators derived from arachidonic acid by several enzymes including cyclooxygenase (COX)-1 and COX-2. We have previously shown that PGE2 regulates immune responses, such as Th1 cytokine production and T-cell proliferation, in cattle. However, it is still unclear whether other PGs are involved in the regulation of immune responses in cattle. Here, immunosuppressive profiles of PGs (PGA1, PGB2, PGD2, PGE2, PGF1α and PGF2α) were firstly examined using bovine peripheral blood mononuclear cells (PBMCs). In addition to PGE2, PGA1 significantly inhibited Th1 cytokine production from PBMCs in cattle. Further analyses focusing on PGA1 revealed that treatment with PGA1 in the presence of concanavalin A (con A) downregulated CD69, an activation marker, and IFN-γ expression in both CD4+ and CD8+ T cells. Sorted CD3+ T cells stimulated with con A were cultivated with PGA1, and IFN-γ and TNF-α concentrations decreased upon PGA1 treatment. Taken together, these results suggest that the treatment with PGA1in vitro inhibits T-cell activation, especially Th1 cytokine production, in cattle.

Assessment of the effects of prostaglandins on myometrial and leiomyoma cells in vitro through microRNA profiling.

It is well known that prostaglandin (PG) E2 and PGF2α are secreted in copious amounts from the menstruating uterus. The aim of the present study was to determine whether PGs affect the growth of uterine leiomyomas (ULs) to the same extent as estrogen or progesterone (P4). The present study evaluated the expression of eight microRNAs (miRNAs) by reverse transcription­quantitative polymerase chain reaction (RT­qPCR) through treatment with estradiol (E2), P4, PGE2, PGF2α and each antagonist or cyclooxygenase­2 (COX­2) inhibitor of cultured leiomyoma and myometrial cells (LC and MC, respectively). The eight miRNAs were divided into two groups according to their primary biological action, namely apoptosis­regulating miRNAs (let­7a, miR­21, miR­26a and miR­200a) and inflammation­regulating miRNAs (miR­29b, miR­93, miR­106b and miR­100b). PGE2 induced significantly higher expression of the 3 anti­apoptotic miRs, let­7a, miR­16a and miR­200a, in LC when compared with the non­treated control or E2. PGE2 significantly promoted a greater expression of let­7a and miR­26a in LC when compared with P4. Overall, PGE2 exerted the highest anti­apoptotic and anti­inflammatory effect in LC, which was comparable with E2. It was not observed among the inflammation­regulating miRNAs in LC. PGF2α did not exert effects as prominent as those of PGE2. In MC, PGs and sex steroids exerted no similar effects on MC compared with LC. The present study demonstrated that PGE2 levels during menstruation may affect the growth of preexisting ULs without affecting the normal myometrium. Therefore, the control of secretion of PGs from the menstruating uterus or the administration of antagonists may be an alternative therapy for inhibiting the growth of ULs.

New insights into the effects of onion consumption on lipid mediators using a diet-induced model of hypercholesterolemia.

The levels and roles of lipid mediators can be modified in response to nutritional stimuli. The aim of this study was to investigate shifts in oxylipin and sphingolipid profiles stimulated by a hypercholesterolemic (HC) diet along with the modulating effects of onion introduced as an antioxidant functional ingredient characterized in the diet (HCO). Oxylipin and sphingolipid profiles were determined in plasma and tissues from Wistar rats using LC-MS/MS. Plasma ω-3 and ω-6 PUFA-derived oxylipins decreased in rats after 7 weeks of HC feeding, but did not evidence a further shift with HCO diet. Onion ingredient supplementation modulated the hepatic concentrations of prostaglandins and enhanced ω-3 oxylipins in the liver of HCO-fed rats relative to the HC group. The HC diet induced shifts in plasma sphingolipids, increasing sphingoid bases, dihydroceramides and ceramides, whilst the sphingomyelin, hexosylceramide and lactosylceramide families decreased. The HCO diet modified some HC diet-induced changes in sphingolipids in liver and spleen tissue. Onion supplementation effected changes in lipid mediator levels in diet-induced hypercholesterolemic Wistar rats. The potential of onion as regulator of pro-inflammatory mediators, and possible enhancer of pro-resolution pathways, warrants further study of the interaction of functional ingredients with bioactive lipid mediators and their potential impact on inflammation, oxidative stress and organ dysfunction.

Studies on terrein as a new class of proteasome inhibitors

The proteasome is an intracellular multicatalytic protease involved in the cell cycle regulation, signaling response, antigen presentation and apoptosis. Since proteasome inhibitors promote cell death by apoptosis, they have been proposed as new anti-tumoral drugs. Terrein, a secondary metabolite secreted by the fungus Aspergillus terreus, was firstly described in 1935. In the present work we report that terrein isolated through the screening for inhibitors of the 20S proteasome showed inhibitory effect upon both chymotrypsin- and trypsin-like activities of the multicatalytic core particle, the 20S proteasome. Despite of the high inhibitory concentration determined in vitro, that verified by incubating cells (fibroblasts and a pulmonary tumor cell line) in the presence of terrein was 4-fold lower indicating the proteasome as a selective intracellular target. Moreover, terrein promoted apoptotic cell death on both fibroblasts and pulmonary tumor cell line tested. Although terrein concentrations (mM range) necessary to elicit apoptosis in the cellular models herein tried were high when compared to those (μM and nM range) of other inhibitors recently described, its chemical structure is not correlated to any other inhibitor reported thus far. Therefore, the present results point out for the possibility of exploring terrein as a new molecular fragment for the development of synthetic proteasome inhibitors.

O proteassomo é uma protease intracelular multicatalítica envolvida na regulação do ciclo e sinalização celular, apresentação antigênica e apoptose. Uma vez que inibidores do proteassomo promovem morte celular por apoptose, esses têm sido propostos como novas drogas anti-tumorais. A terreína, um metabólito secundário secretado pelo fungo Aspergillus terreus, foi primeiramente descrita em 1935. Neste trabalho demonstramos que a terreína, isolada através da bioprospecção de inibidores do proteassomo, mostrou efeito inibitório das atividades do tipo quimiotripsina e tripsina da unidade catalítica do proteassomo 20S. Apesar da alta concentração inibitória determinada in vitro, aquela verificada após incubação de células em cultura na presença de terreína (fibroblasto e tumor pulmonar humano) foi 4 vezes menor, o que sugere que o proteassomo seja um alvo intracelular específico. A terreína promoveu morte celular por apoptose nas duas linhagens ensaiadas. Embora as concentrações de terreína necessárias para desencadear apoptose nos modelos celulares aqui testados tenham sido altas (ordem de mM) quando comparadas com doses utilizadas de outros inibidores descritos recentemente (ordem de μM e nM), sua estrutura química não está relacionada a nenhum outro inibidor conhecido até o momento. Concluímos que estes resultados apontam para a possibilidade de explorar a terreína como um novo fragmento molecular para o desenvolvimento de inibidores sintéticos do proteassomo.

Brain prostaglandin formation is increased by alpha-synuclein gene-ablation during global ischemia.

We have previously demonstrated that alpha-synuclein (Snca) gene ablation reduces brain arachidonic acid (20:4n-6) turnover rate in phospholipids through modulation of endoplasmic reticulum-localized acyl-CoA synthetase activity. Although 20:4n-6 is a precursor for prostaglandin (PG), Snca effect on PG levels is unknown. In the present study, we examined the effect of Snca ablation on brain PG level at basal conditions and following 30s of global ischemia. Brain PG were extracted with methanol, purified on C(18) cartridges, and analyzed by LC-MS/MS. We demonstrate, for the first time, that Snca gene ablation did not affect brain PG mass under normal physiological conditions. However, total PG mass and masses of individual PG were elevated approximately 2-fold upon global ischemia in the absence of Snca. These data are consistent with our previously observed reduction in 20:4n-6 recycling through endoplasmic reticulum-localized acyl-CoA synthetase in the absence of Snca, which may result in the increased 20:4n-6 availability for PG production in the absence of Snca during global ischemia and suggest a role for Snca in brain inflammatory response.

Simultaneous quantification of seven prostanoids using liquid chromatography/tandem mass spectrometry: the effects of arachidonic acid on prostanoid production in mouse bone marrow-derived mast cells.

We have developed a method for the simultaneous estimation of the levels of the prostanoids 6-keto prostaglandin (PG) Flalpha, PGB2, PGD2, PGE2, PGF2(alpha), PGJ2, and thromboxane (TX) B2 in blood- or serum-containing medium using liquid chromatography-tandem mass spectrometry. These prostanoids and their deuterium derivatives, which were used as internal standards, were subjected to solid-phase extraction using Empore C18 HD disk cartridges and analyzed in the selected reaction-monitoring mode. A linear response curve starting at 10 pg of prostanoid/tube was observed for each prostanoid. The accuracy of the method was demonstrated with samples containing known amounts of the prostanoids. Furthermore, we used this method to analyze the prostanoids produced in mouse bone marrow-derived mast cells stimulated with arachidonic acid, which resulted in the production of PGD2, PGE2, PGF2alpha, and TXB2. The results suggest that this simultaneous quantification method is useful for the analysis of the production of biomedically important prostanoids.

Differentiation of cyclooxygenase 1- and 2-derived prostanoids in mouse kidney and aorta.

Accumulating evidence indicates cyclooxygenase (COX) 1 and COX2 differentially regulate cardiovascular and renal function. We have demonstrated previously in mice that COX2 inhibition enhances angiotensin II-induced hypertension, and COX1 inhibition attenuates the pressor effect of angiotensin II. To further elucidate the mechanism underlying the functional difference of COX1 versus COX2 inhibition, the present studies examined the prostaglandin (PG) profiles derived in COX1- or COX2-inhibited mouse kidney and aorta using gas chromatographic/mass spectrometric assays. PGE2 is the most abundant prostanoid in both renal cortex and medulla in normal C57BL/6J mice, followed by PGI2, PGF2alpha and thromboxane A2. In contrast PGI2 was most abundant in aorta followed by thromboxane A2, PGE2, and PGF2alpha. PGD2 was undetectable in control kidney or aorta. At baseline, inhibition of COX1 decreased total prostaglandins in renal cortex, medulla, and aorta, whereas COX2 inhibition decreased total prostaglandins only in renal medulla. Angiotensin II infusion significantly increased COX2-dependent/COX1-independent PGE2 and PGI2 in renal cortex and medulla. Angiotensin II also significantly increased renal PGF2alpha in cortex, but not in medulla, through both COX1- and COX2-dependent mechanisms. These studies demonstrate that although COX1 primarily contributes to basal prostanoid production in the kidney and aorta, angiotensin II increases renal vasodilator prostanoids predominately via COX2 activity. These effects may contribute to the specific effect of COX2 inhibitors to increase blood pressure.

Acetylcholine-induced endothelium-dependent contractions in the SHR aorta: the Janus face of prostacyclin.

In the spontaneously hypertensive rat (SHR) and aging Wistar-Kyoto rats (WKY), acetylcholine releases an endothelium-derived contracting factor (EDCF) produced by endothelial cyclooxygenase-1, which stimulates thromboxane A2 receptors (TP receptors) on vascular smooth muscle. The purpose of the present study was to identify this EDCF by measuring changes in isometric tension and the release of various prostaglandins by acetylcholine. In isolated aortic rings of SHR, U 46619, prostaglandin (PG) H2, PGF2alpha, PGE2, PGD2, prostacyclin (PGI2) and 8-isoprostane, all activate TP receptors of the vascular smooth muscle to produce a contraction (U 46619>>8-isoprostane=PGF2alpha=PGH2>PGE2=PGD2>PGI2). The contractions produced by PGH2 and PGI2 were fast and transient, mimicking endothelium-dependent contractions. PGI2 did not relax isolated aortic rings of WKY and SHR. Acetylcholine evoked the endothelium-dependent release of thromboxane A2, PGF2alpha, PGE2, PGI2 and most likely PGH2 (PGI2>>PGF2alpha>or=PGE2>TXA2>8-isoprostane, PGD2). Dazoxiben abolished the production of thromboxane A2, but did not influence the endothelium-dependent contractions to acetylcholine. The release of PGI2 was significantly larger in the aorta of SHR than in WKY, and the former was more sensitive to the contractile effect of PGI2 than the latter. The inhibition of PGI-synthase was associated with an increase in PGH2 spillover and the enhancement of acetylcholine-induced endothelium-dependent contractions. Thus, in the aorta of SHR and aging WKY, the endothelium-dependent contractions elicited by acetylcholine most likely involve the release of PGI2 with a concomitant contribution of PGH2.

Endothelin-induced constriction of the ductus venosus in fetal sheep: developmental aspects and possible interaction with vasodilatory prostaglandin.

1. The ductus venosus is actively regulated in the fetus, but questions remain on the presence of a functional sphincter at its inlet. Using fetal sheep (0.6-0.7 gestation onwards), we have examined the morphology of the vessel and have also determined whether endothelin-1 (ET-1) qualifies as a natural constrictor being modulated by prostaglandins (PGs). 2. Masson's staining and alpha-actin immunohistochemistry showed a muscular, sphincter-like formation at the ductus inlet and a muscle layer within the wall of the vessel proper. This muscle cell component increased with age. 3. ET-1 contracted dose-dependently isolated sphincter and extrasphincter preparations of the ductus from term fetus. This ET-1 effect also occurred in the premature, but its threshold was higher. 4. BQ123 (1 microm) caused a rightward shift in the ET-1 dose-response curve, while indomethacin at a threshold concentration (28 nm) tended to have an opposite effect. 5. Big ET-1 also contracted the ductus sphincter but differed from ET-1 for its lesser potency and inhibition by phosphoramidon (50 microm). 6. The ductus sphincter (term and preterm) and extrasphincter (term) released 6-keto-PGF(1alpha) (hence PGI(2)) and, to a lesser degree, PGE(2) at rest and their release increased dose-dependently upon ET-1 treatment. Both basal and stimulated release was curtailed by endothelium removal. 7. BQ123 and phosphoramidon reduced slightly the contraction of ductus sphincter to indomethacin (2.8 microm). 8. We conclude that the ductus contains a contractile mechanism in the sphincter and extrasphincter regions. ET-1 lends itself to a role in the generation of contractile tone and its action may be modulated by prostaglandins.

The efficacy and safety of dose escalation of dorzolamide used in combination with other topical antiglaucoma agents.

We investigated the dose-escalation profile of dorzolamide used in combination with other antiglaucoma agents in patients with primary glaucoma and ocular hypertension. In a prospective, open-label study, 78 patients received dorzolamide 0.5% in addition to other topical antiglaucoma agents for > or =4 weeks. The concentration of dorzolamide was then escalated to 1.0% and intraocular pressure (IOP) measured every 4 weeks for 12 weeks. Dose escalation of dorzolamide from 0.5% to 1.0% resulted in a significant reduction in IOP throughout the 12 weeks of treatment at the higher dose. Mean baseline IOP was 19.7 mmHg. At 4, 8, and 12 weeks after dose escalation, mean IOP had decreased to 17.8 (-9.4%), 17.6 (-10.8%), and 17.5 (-10.7%) mmHg. No serious drug-related adverse effects were reported. These results indicate that dose escalation of dorzolamide from 0.5% to 1.0% is effective and well tolerated as adjunctive therapy for patients in whom IOP is insufficiently controlled by combination therapy.

Quantitative high-performance liquid chromatography/electrospray ionization tandem mass spectrometric analysis of 2- and 3-series prostaglandins in cultured tumor cells.

This paper describes a rapid and simple technique for the simultaneous quantitative analysis of PGE(2), PGE(3), and other closely related prostaglandins from cultured cells using liquid chromatography/electrospray ionization tandem mass spectrometry. This method permits quantification of selected individual prostaglandins derived either from arachidonic acid (AA) or eicosapentaenoic acid (EPA) from cell extracts without tedious derivatization, lengthy sample preparation, and separation required by GC-MS- or HPLC-UV-based methods. The validation assessment showed that the quantitative determination is linear (r(2)>0.999) for both PGE(2) and PGE(3) in the range tested (1-500 ng/ml, 0.0028-1.4 microM) and a coefficient of variation lower than 10% was obtained for samples analyzed on 3 separate days. The detection limit was 2.5 pg for both PGE(2) and PGE(3). Extraction efficiency of PGE(2) and PGE(3) from cell suspensions ranged from 89.4 to 98.2%. As an application of the method, prostaglandins formed by EPA in human lung cancer A549 cells were determined. A 62% reduction of PGE(2) formation was noted when A549 cells were treated with 10 microM of EPA. Concomitantly, EPA increased formation of PGE(3) by 10-fold in A549 cells. This is the first report that unequivocally demonstrates that EPA can be converted to PGE(3) by cyclooxygenase in human cancer cells.

Prostaglandin biology.

PGs are important mediators of normal physiology, response to injury, and pathologic processes. Dissecting these biochemical and molecular pathways allows development of therapeutic agents that can be [figure: see text] applied to specific clinical situations, while preserving PGs that play a role in normal physiology.

The isoprostanes: novel prostaglandin-like products of the free radical-catalyzed peroxidation of arachidonic acid.

The isoprostanes (IsoPs) are a unique series of prostaglandin-like compounds formed in vivo from the free radical-catalyzed peroxidation of arachidonic acid. This review summarizes our current knowledge regarding these compounds. Novel aspects of the biochemistry and bioactivity of IsoPs are detailed and methods by which these compounds are analyzed are discussed. A considerable portion of this review deals with the utility of measuring IsoPs as markers of oxidant injury in human diseases particularly in association with risk factors that predispose to atherosclerosis, a condition in which excessive oxidative stress has been causally implicated.

Effects of dietary gamma-linolenic acid-rich borage oil combined with marine fish oils on tissue phospholipid fatty acid composition and production of prostaglandins E and F of the 1-, 2- and 3-series in a marine fish deficient in delta5 fatty acyl desaturase.

The effects of gamma-linolenic acid-rich borage oil (BO), in combination with different marine oils, namely an eicosapentaenoic acid (EPA) rich oil (MO) or a DHA-rich oil (TO), on tissue fatty acid composition and prostaglandin production were investigated in turbot, a species which lacks appreciable delta5 fatty acyl desaturase activity. The juvenile turbot grew well on the experimental diets and there were no significant differences in final weights between dietary treatments. Irrespective of the marine oil component, both the BO-containing diets increased tissue phospholipid levels of 18:2n-6 and 18:3n-6, and their respective elongation products, 20:2n-6 and 20:3n-6, compared to fish fed a control diet containing a standard Northern hemisphere fish oil. Both the BO-containing diets increased the production of 1-series prostaglandins (PG), this being observed across all tissues investigated with PGF and especially PGE. The BO/MO diet also reduced 20:4n-6 in tissue phospholipids without affecting 20:5n-3, whereas the BO/TO combination decreased 20:5n-3 but increased 20:4n-6. The production of 2-series and 3-series PGs was also altered by the dietary treatments but the changes were less dependent upon the tissue levels of their respective precursor fatty acids, 20:4n-6 and 20:5n-3. The BO-containing diets had very significant effects on gross fatty acid compositions of the phospholipids including increased proportions of saturated fatty acids and n-6 polyunsaturated fatty acids (PUFA) and decreased proportions of monounsaturated fatty acids and n-3 PUFA. Overall, this study shows that eicosanoid production in turbot tissues can be influenced by dietary fatty acids, not only by changes in the absolute and relative levels of specific eicosanoid precursor PUFA in tissue phospholipids, but also by general effects on membrane composition, structure and function induced by gross fatty acid compositional changes.

A nomenclature system for the isoprostanes.

In 1990, prostaglandin (PG) F2-like compounds were discovered to be produced in abundance in vivo by a free radical mechanism independent of the cyclooxygenase enzyme. Because these compounds are isomeric to cyclooxygenase-derived PGF2 alpha, they were termed F2-isoprostanes (F2-IsoP's). Subsequently, it was also demonstrated that PGD2-like compounds (D2-IsoP's) and PGE2-like compounds (E2-IsoP's) are also produced in vivo as products of this pathway. Four different regioisomers of each of these classes of IsoP's are formed, each of which can be comprised of eight racemic diastereomers. Thus, 64 different F2-IsoP's, E2-IsoP's, and D2-IsoP's can be formed. Interest in these molecules stems not only from the fact that quantification of IsoP's can provide a valuable index of free radical-induced lipid peroxidation in vivo but also from the fact that it has been shown that these compounds are capable of exerting potent biological activity. Because of this potential for exerting biological activity, the chemical syntheses of various IsoP compounds for biological testing has been initiated. As a result, a need for a systematic nomenclature for these compounds has evolved. A facile nomenclature that will allow rational differentiation and designation of each of the isomeric structures comprising the family of IsoP's is presented.

Nomenclature of isoprostanes: a proposal.

We have proposed a nomenclature system for the isoprostanes, a new class of natural products formed in vivo by the free-radical peroxidation of polyunsaturated fatty acids. Our proposed nomenclature is based on the assignment of four isoprostanes, 1, 9, 17, and 25, as representatives of the four classes of isoprostanes derived from arachidonic acid (AA). We have attempted as much as possible to retain elements from the familiar prostaglandin nomenclature. In this proposal, we have used the abbreviation i.p. for isoprostane. We have classified isoprostane classes or types based on omega-carbon as being the starting reference. Roman numerals I-VI refer the six types of isoprostanes derived from eicosapentaenoic acid (EPA) and III-VI refer to the four types derived from AA. This nomenclature can be applied to isoprostanes derived from other PUFAs also.

Bradykinin and FMLP stimulate prostanoid production by adult rabbit colonocytes in culture.

Normal colonocytes in culture produce prostaglandins both constitutively and in response to inflammatory stimuli. These highly purified proliferative cell populations were isolated from normal adult rabbit proximal and distal colon. Basal prostaglandin production ranged from 3.4 to 11.7 ng.15 min-1 x 10(6) cells-1. Cultures were incubated at 37 degrees C in the presence or absence of bradykinin or N-formyl-methionine-leucine-phenylalanine (FMLP) over concentrations from 10(-9) to 10(-5) M. In both distal and proximal colonocytes bradykinin stimulated a dose-dependent increase in prostaglandin E2 (PGE2) and 6-keto-PGF1 alpha, the stable metabolite of prostacyclin; production peaked at 10(-6) M. Proximal colonocytes responded to FMLP with a bell-shaped curve, with maximal stimulation of both PGE2 and 6-keto-PGF1 alpha occurring at 10(-8) M. Distal colonocytes responded variably to FMLP. Arachidonic acid also stimulated prostanoid production in a concentration-dependent manner, with maximal stimulation occurring at 100 microM. The full synthetic profile of prostanoid production was determined by labeling with [14C]arachidonic acid and by analyzing metabolites using radiochromatography on reverse-phase high-pressure liquid chromatography. Only PGE2 and 6-keto-PGF1 alpha were detected. A similar profile of labeled metabolites occurred when colonocytes were prelabeled with [14C]arachidonic acid and stimulated with bradykinin or FMLP. The degradative capacity of the colonocytes appeared very low. Colonocyte production of protective prostaglandins in response to luminal or other inflammatory stimuli may serve as a mucosal defense mechanism. Prostanoids so produced may also modulate the functions of colonocytes, surrounding cells, or both.

Prostanglandinas. Antiinflamatorios no esteroides. Dolor-inflamación y fiebre / Prostaglandines. Non steroidal antiinflamatory agents. Pain inflamation and fever

Cuando se habla de inflamación, dolor o fiebre, necesariamente hay que mencionar o hablar de prostaglandinas, leucotrienos, prostenoides o epóxidos, sustancias que en buena medida son responsables de aquellas manifestaciones histoclínicas y de sus consecuencias. La mención de los anteriores compuestos implica a sí mismo traer a la mente el grupo de fármacos conocido como antiinflamatorios no esteroides o simplemente "IANE" como se les conoce en términos científicos y cuyo mecanismo de acción influye directamente en aquellas sustancias,a la vez que es responsable de no pocas de sus reacciones secundarias.

[Therapeutic assessment of the evening primrose]. / Ocena zdrowotna nasion z wiesiolka.

In the light of a survey of literature data the chemical composition of described of evening primrose seeds and of the oil obtained from them which has a high content of unsaturated fatty acids, mainly linolic and gamma-linolenic acid. The therapeutic value of the preparations of evening primrose and their effect on the metabolism are discussed. The criteria of the therapeutic quality of the seeds are given.