The effect of malotilate, a derivative of malotilate and a flavenoid on eicosanoid production in inflammatory bowel disease in rats

Acetic acid induced colitis in rats was used to investigate the effects of malotilate, a drug which has been shown to inhibit 5-1ipoxygenase in human macrophages, the malotilate derivate ZY16268 and the flavenoid ZY16369 on the eicosanoid production and the colonic morphology in inflammatory bowel disease. Acetic acid produced an acute inflammatory response in the colon, associated with a markedly raised inflammation score (15.8 vs. < 0.5), based on a seven-scaled scoring system which includes observation of haemorrhage, submucosal oedema, cellular infiltration, goblet cell depletion, loss of architecture, crypt abscesses and serosal involvement, of which every item was subdivided as mild, moderate and severe. Incubation of colonic mucosa from rats treated with arachidonic acid and stimulated with A23187 showed an increase of the cyclooxygenase product 12-hydroxy-heptadecatrienoic acid (HHT) and the 12-1ipoxygenase product (12-HETE) and a decrease in the formation of 6-keto-prostaglandin F(1alpha)(6kPGF(1alpha)) in comparison with normal rat mucosa. Malotilate, ZY16268 and ZY16369 all resulted in a decrease in HHT, leukotriene B(4) (LTB(4))-like compounds and 12-hydroxyeicosaenoic acid (12-HETE) production. None of the tested compounds significantly reduced the colonic damage by acetic acid although the formation of 12-HETE was proportional to the histologically obtained inflammation score. There were marked differences in eicosanoid formation patterns between rat and human mucosa, both normal and inflamed. In view of the hyperacute nature of the mucosal damage and the marked differences in eicosanoid production, acetic acid induced colitis in rats is probably not a suitable model of ulcerative colitis in humans.     

The effect of Bulgarian propolis against Trypanosoma cruzi and during its interaction with host cells

Propolis has shown activity against pathogenic microorganisms that cause diseases in humans and animals. The ethanol (Et-Blg) and acetone (Ket-Blg) extracts from a Bulgarian propolis, with known chemical compositions, presented similar activity against tissue culture-derived amastigotes. The treatment of Trypanosoma cruzi-infected skeletal muscle cells with Et-Blg led to a decrease of infection and of the intracellular proliferation of amastigotes, while damage to the host cell was observed only at concentration 12.5 times higher than those affecting the parasite. Ultrastructural analysis of the effect of both extracts in epimastigotes revealed that the main targets were the mitochondrion and reservosomes. Et-Blg also affected the mitochondrion-kinetoplast complex in trypomastigotes, offering a potential target for chemotherapeutic agents.     

The effect of essential fatty acid deficiency on eicosanoid production in the inflamed rat colonic mucosa

Eicosanoids are potent mediators of inflammation and are synthesized in increased quantity in active ulcerative colitis. To elucidate the role of prostaglandin E2, thromboxane A2, prostaglandin I2, and leukotriene B2 in acute chemical colitis induced by 4% acetic acid, we utilized an animal model which has a deficiency of arachidonic acid, the precursor of eicosanoids due to an essential fatty acid deficient diet. Forty-eight hours after colitis was induced, mucosal synthesis of the cyclooxygenase products, prostaglandin E2, thromboxane A2, and prostaglandin I2, was significantly decreased in essential fatty acid deficient rats compared to normal controls. However, the 5-lipoxygenase product, leukotriene B4, was not different between groups. The decrease in cyclooxygenase products did not correlate with any change in the severity of colonic inflammation as assessed by gross morphology, histology, or myleoperoxidase activity. Thus inhibition of formation of the cyclooxygenase products of arachidonate metabolism does not appear to improve the degree of inflammation under the experimental conditions employed in this study.     

Inhibitive effect of purple sweet potato leaf extract and its components on cell adhesion and inflammatory response in human aortic endothelial cells

This study investigated the effects of purple sweet potato leaf extract (PSPLE) and its components, cyanidin and quercetin, on human aortic endothelial cells (HAECs) during the inflammatory process. HAECs were pretreated with 100 μg/mL PSPLE or 10 μM quercetin, cyanidin or aspirin for 18 h followed by TNF-α (2 ng/mL) for 6 h, and U937 cell adhesion was determined. Adhesion molecule expression and CD40 were evaluated; NFκB p65 protein localization and DNA binding were assessed. PSPLE, aspirin, cyanidin and quercetin significantly inhibited TNF-α-induced monocyte-endothelial cell adhesion (p < 0.05). Cyanidin, quercetin and PSPLE also significantly attenuated VCAM-1, IL-8 and CD40 expression, and quercetin significantly attenuated ICAM-1 and E-selectin expression (p < 0.05). Significant reductions in NFκB expression and DNA binding by aspirin, cyanidin and quercetin were also observed in addition to decreased expression of ERK1, ERK2 and p38 MAPK (p < 0.05). Thus, PSPLE and its components, cyanidin and quercetin, have anti-inflammatory effects through modulation of NFκB and MAPK signaling. Further in vivo studies are necessary to explore the possible therapeutic effects of PSPLE on atherosclerosis.     

The effect of turmeric extracts on inflammatory mediator production.

Major compounds of several commonly used botanicals, including turmeric, have been purported to have anti-inflammatory actions. In order to test the anti-inflammatory activity of compounds isolated from rhizomes of Curcuma longa L. (Zingiberaceae), we have established an in vitro test system. HL-60 cells were differentiated and exposed to lipopolysaccharide (LPS) from Escherichia coli (1 microg/ml) in the presence or absence of botanical compounds for 24 h. Supernatants were collected and analyzed for the production of tumor necrosis factor alpha (TNF-alpha) and prostaglandin E2 (PGE2) using standard ELISA assays. Water-soluble extracts were not cytotoxic and did not exhibit biological activity. Organic extracts of turmeric were cytotoxic only at concentrations above 50 microg/ml. Crude organic extracts of turmeric were capable of inhibiting LPS-induced TNF-alpha (IC50 value = 15.2 microg/ml) and PGE2 (IC50 value = 0.92 microg/ml) production. Purified curcumin was more active than either demethoxy- or bisdemethoxycurcumin. Fractions and subfractions of turmeric extracts collected via preparative HPLC had differing biological activity, ranging from no activity to IC50 values of < 1 microg/ml. For some fractions, subfractionation resulted in a loss of activity, indicating interaction of the compounds within the fraction to produce an anti-inflammatory effect. A combination of several of the fractions that contain the turmeric oils was more effective than the curcuminoids at inhibiting PGE2. While curcumin inhibited COX-2 expression, turmeric oils had no effect on levels of COX-2 mRNA.     

The effect of dehydroepiandrosterone on inflammatory response of astroglial cells

An increased interest in neuroinflammation is conditioned by its involvement in various pathological processes in the brain. Astrocytes play an important role in neuroinflammation, participating in its regulation, throwing out a large number of signaling molecules. Steroid compounds, actively produced by astrocytes, are of interest with regards to the regulation of inflammatory processes in the central nervous system. In the present work the effect of dehydroepiandrosterone (DHEA) on astroglial cells (cultured primary rat astrocytes) in a model of inflammation was studied. The inflammatory response was stimulated with lipopolysaccharide (LPS). Expression levels of pro-inflammatory factor TNFα, antinflammatory interleukin IL-10, and both pro- and antiinflammatory protein COX-2 were measured. The expression of IL-10, COX-2, and TNFα mRNA was determined by real-time PCR, COX-2 protein level by immunoblotting method, TNFα and IL-10 release by enzyme immunoassay. The effect of short-term (30 min) and long-term (24 h) exposure to DHEA was evaluated. It was shown that DHEA potentiates LPS-stimulated (1) increase in the IL-10 mRNA level; (2) IL-10 release; (3) does not affect TNFα level, and (4) exerts a weak pulsating bidirectional effect on COX-2. Using trilostane, an inhibitor of 3β-hydroxysteroid dehydrogenase, a key enzyme of DHEA metabolism, it was shown that DHEA metabolites make the main contribution to its effect. Thus, DHEA is of interest as a stimulant of anti-inflammatory processes in the brain.     

ALOX5 gene variants affect eicosanoid production and response to fish oil supplementation

The objective of this study was to determine whether 5-lipoxygenase (ALOX5) gene variants associated with cardiovascular disease affect eicosanoid production by monocytes. The study was a randomized, double-masked, parallel intervention trial with fish oil (5.0 g of fish oil daily, containing 2.0 g of eicosapentaenoic acid [EPA] and 1.0 g of docosahexaenoic acid [DHA]) or placebo oil (5.0 g of corn/soy mixture). A total of 116 subjects (68% female, 20-59 years old) of African American ancestry enrolled, and 98 subjects completed the study. Neither ALOX5 protein nor arachidonic acid-derived LTB4, LTD4, and LTE4 varied by genotype, but 5-hydroxyeicosatetraenoate (5-HETE), 6-trans-LTB4, 5-oxo-ETE, 15-HETE, and 5,15-diHETE levels were higher in subjects homozygous for the ALOX5 promoter allele containing five Sp1 element tandem repeats ("55" genotype) than in subjects with one deletion (d) (three or four repeats) and one common ("d5" genotype) allele or with two deletion ("dd") alleles. The EPA-derived metabolites 5-HEPE and 15-HEPE and the DHA-derived metabolite 17-HDoHE had similar associations with genotype and increased with supplementation; 5-HEPE and 15-HEPE increased, and 5-oxo-ETE decreased to a greater degree in the 55 than in the other genotypes. This differential eicosanoid response is consistent with the previously observed interaction of these variants with dietary intake of omega-3 fatty acids in predicting cardiovascular disease risk.     

Garlic and onions: their effect on eicosanoid metabolism and its clinical relevance

Garlic (Allium sativum) and onion (Allium cepa) are among the oldest of all cultivated plants. Additionally, both plants have been used as medicinal agents for thousands of years. Both garlic and onion have been shown to have applications as antimicrobial, antithrombotic, antitumor, hypolipidaemic, antiarthritic and hypoglycemic agents. In recent years, extensive research has focussed on the beneficial and medicinal properties of garlic and onions. In particular, the use of these agents in the treatment and prevention of cardiovascular disease and cancer is an area of considerable investigation and interest.     

The effects of alcoholism and smoking on platelet eicosanoid production in vitro

Ethanol induces changes in eicosanoid synthesis in blood platelets and brain tissue. Cigarette smoking also causes alterations in eicosanoid formation. This preliminary report examined in vitro platelet sonicate eicosanoid production using 14C-arachidonic acid (14C-AA) and in separate experiments, 14C-PGH2, as substrates. Radiometric thin layer chromatography (TLC) was used to identify the products formed. Eicosanoid product formation in platelet sonicates collected from 28 abstinent male alcoholics were compared to those from 11 male control subjects. All but one of the alcoholics were chronic smokers and all control subjects were non-smokers. All smokers abstained from smoking for 12 h prior to the blood collection to control for any acute effects of cigarette smoke on eicosanoid production. Significant reductions in platelet sonicate production of PGD2 and PGE2 in vitro were observed in alcoholic smokers when 14C-PGH2, but not 14C-AA, was the substrate. These reductions were predicted equally well by two variables, smoking and alcoholism, using several statistical models. This is the first investigation that controlled for the acute effects of smoking and accounted for the potential effects of cigarette smoking on platelet eicosanoid production in alcoholics. Because cigarette smoking is prevalent among alcoholics, future studies on the role of eicosanoids in alcoholism should control for smoking.     

The effect of extracts from ginger rhizome on inflammatory mediator production

Compounds from rhizomes of Zingiber officinale, commonly called ginger, have been purported to have anti-inflammatory actions. We have used an in vitro test system to test the anti-inflammatory activity of compounds isolated from ginger rhizome. U937 cells were differentiated and exposed to lipopolysaccharide (LPS) from Escherichia coli (1 microg/ml) in the presence or absence of organic extracts or standard compounds found in ginger (6-, 8-, 10-gingerol or 6-shogaol) for 24 h. Supernatants were collected and analyzed for the production of prostaglandin E(2) (PGE(2)) and tumor necrosis factor alpha (TNF-alpha) by standard ELISA assays. Predominant compounds in the organic extracts were identified as 6-, 8- 10-gingerols and 6-, 8-, 10-shogaols. Organic extracts or standards containing gingerols were not cytotoxic, while extracts or standards containing predominantly shogaols were cytotoxic at concentrations above 20 microg/ml. Crude organic extracts of ginger were capable of inhibiting LPS induced PGE(2) (IC(50)<0.1 microg/ml) production. However, extracts were not nearly as effective at inhibiting TNF-alpha (IC(50)>30 microg/ml). Thirty three fractions and subfractions, prepared by column chromatography, were analyzed for bioactivity. Extracts containing either predominantly gingerols or shogaols (identified by HPLC) were both highly active at inhibiting LPS-induced PGE(2) production (IC(50)<0.1 microg/ml), while extracts that contained unknown compounds were less effective (IC(50)<3.2 microg/ml). Extracts or standards containing predominantly gingerols were capable of inhibiting LPS-induced COX-2 expression while shogaol containing extracts had no effect on COX-2 expression. These data demonstrate that compounds found in ginger are capable of inhibiting PGE(2) production and that the compounds may act at several sites.     

Schistosoma mansoni: cercarial eicosanoid production and penetration response inhibited by esculetin and ibuprofen

Cercariae of Schistosoma mansoni produce a wide variety of eicosanoids when stimulated by 3.3 mM linoleate. High-performance liquid chromatography indicated that 10(-5) M esculetin dramatically decreased eicosanoid production by cercariae. Ibuprofen (10(-4) M) also decreased eicosanoid production, but to a lesser extent. These results were confirmed by radioimmunoassay using time-dose curves for esculetin and time curves for ibuprofen. The results reported here indicated that, for cercariae, ibuprofen was neither a specific cyclo-oxygenase inhibitor, as has been reported for platelet and endothelial cells, nor was esculetin a specific inhibitor of lipoxygenase, as has been reported for platelets and mastocytoma cells. Rather, both drugs inhibited cyclo-oxygenase and lipoxygenase enzyme systems. Further, the data indicated that two forms of cyclo-oxygenase exist in cercariae (isozymes?), one initiating the conversion of gamma-dihomolinolenate into the 1-series prostaglandins and another acting on arachidonate forming the 2-series prostaglandins. The cyclo-oxygenase acting on arachidonate has a greater sensitivity to both ibuprofen and esculetin than the enzyme acting on gamma-dihomolinolenate. Cercarial lipoxygenases also varied greatly in their sensitivity to ibuprofen.     

Role of oxygen-derived free radicals on rat splanchnic eicosanoid production during acute hemorrhage.

The effect of superoxide dismutase (SOD), an oxygen-derived free radical scavenger, on rat splanchnic eicosanoid synthesis was examined following hemorrhagic shock. Anesthetized male rats were hemorrhaged to 30 mm Hg for 30 minutes (Shock), killed, or treated with the shed blood (Shock plus reperfusion). The Shock plus reperfusion group was treated with saline vehicle or SOD (2500, 5000, 7500, 10,000 or 15,000 U/Kg, i.v.) 15 minutes prior to the reperfusion of the shed blood. The superior mesenteric artery was removed in continuity with the end organ intestine (SV+SI) and perfused in vitro with oxygenated Krebs-Henseleit buffer (3 ml/min at 37 degrees C). Venous effluent was measured for basal release of 6-keto-PGF1 alpha, PGE2 and thromboxane B2 at 15, 30, 60 and 90 minutes of perfusion. The SV+SI compensated for acute shock by increased release of 6-keto-PGF1 alpha (300%) (and not PGE2 or thromboxane B2) which was abolished by reperfusion of the shed blood following shock. Prior treatment of the Shock plus reperfusion group with 7500 U/Kg or more of SOD restored the increased release of SV+SI 6-keto-PGF1 alpha found following shock alone (p less than 0.05). These data provided indirect evidence that ODFRs contributed to endogenous SV+SI regulation of PGI2 synthesis and release during hemorrhagic shock and reperfusion of shed blood.     

Impact of three different types of exercise on components of the inflammatory response

It was hypothesized that muscle injury would be greater with eccentric than with all-out or prolonged exercise, and that immune changes might provide an indication that supplements the information provided by traditional markers such as creatine kinase (CK) or delayed-onset muscle soreness. Eight healthy males [mean (SE): age = 24.9 (2.3) years, maximum oxygen consumption (VO2(max)) = 43.0 (3.1) ml x kg(-1) x min(-1)] were each assigned to four experimental conditions, one at a time, using a randomized-block design: 5 min of cycle ergometer exercise at 90% VO2(max) (AO), a standard circuit-training routine (CT), 2 h cycle ergometer exercise at 60% VO2(max) (Long), or remained seated for 5 h. Blood samples were analyzed for CK, natural killer (NK) cell counts (CD3(-)/CD16(+)56(+)), cytolytic activity and plasma levels of the cytokines interleukin (IL)-6, IL-10, and tissue necrosis factor alpha (TNF-alpha). CK levels were only elevated significantly 72 h following CT. NK cell counts increased significantly during all three types of exercise, but returned to pre-exercise baseline values within 3 h of recovery. Cytolytic activity per NK cell was not significantly modified by any type of exercise. Prolonged exercise induced significant increases in plasma IL-6 and TNF-alpha. We conclude that the lack of correlation between traditional markers of muscle injury (plasma CK concentrations and muscle soreness rankings) and immune markers of the inflammatory response suggests that, for the types and intensities of exercise examined in this study, the exercise-induced inflammatory response is modified by humoral and cardiovascular correlates of exercise.     

Reduction of eicosanoid production by essential fatty acid depletion does not attenuate the inflammatory response induced by Pseudomonas aeruginosa pneumonia in rat lung.

Sipid mediators of inflammation have been implicated in the pathogenesis of Pseudomonas aeruginosa (PA) related pulmonary damage in patients with cystic fibrosis. We studied the role of these mediators in a rat model of PA endobronchitis using essential fatty acid deficient (EFAD) animals. Whole blood from EFAD animals produced significantly less leukotriene B4 (LTB4) and hydroxyheptadecatrienoic acid when stimulated ex vivo than did whole blood from control animals (p less than 0.005). Similarly, lung lavage fluid from EFAD animals infected with PA contained less LTB4 and thromboxane B2 (TXB2) than that from control animals. Despite these differences, cellular infiltration of airways in response to PA infection was virtually identical in animals from the regular diet and the EFAD groups. Both EFAD and control animals had a significant increase in white blood cells (WBC) in lung lavage fluid at 1, 3 and 6 days following infection with PA when compared to animals receiving sterile beads. Localized areas of consolidation and nodularity were grossly evident in the lungs of all PA infected animals irrespective of their ability to generate the lipid inflammatory mediators. Microscopic examination of lung sections demonstrated similar changes in all infected animals. We conclude that LTB4 and TXB2 production occurs early in the course of PA pulmonary infection in rats. This early rise in lipid mediators is temporally associated with an influx of WBC into the airways. However, attenuation of eicosanoid production by use of an EFAD diet does not lead to a reduction in the inflammatory response to PA infection.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of conjugated linoleic acid on arachidonic acid metabolism and eicosanoid production in human saphenous vein endothelial cells

The effects of a conjugated linoleic acid (CLA) mixture of single isomers (50:50, w/w, cis9,trans11:trans10,cis12) and the individual isomers on (a) the production of resting and calcium ionophore stimulated (14)C-eicosanoids and (b) the incorporation of (14)C-arachidonic acid (AA) into membrane phospholipids of human saphenous vein endothelial cells were investigated. The CLA mixture and the individual isomers were found to inhibit resting production of (14)C-prostaglandin F(2a) by 50, 43 and 40%, respectively. A dose dependent inhibition of stimulated (14)C-prostaglandins was observed with the CLA mixture (IC(50) 100 microM). The cis9,trans11 and trans10,cis12 (50 microM) isomers individually inhibited the overall production of stimulated (14)C-prostaglandins (between 35 and 55% and 23 and 42%, respectively). When tested at a high concentration (100 microM), cis9,trans11 was found to inhibit eicosanoid production in contrast to trans10,cis12 that caused stimulation. The overall degree of (14)C-AA incorporation into membrane phospholipids of the CLA (mixture and individual isomers) treated cells was found to be lower than that of control cells and the cis9,trans11 isomer was found to increase the incorporation of (14)C-AA into phosphatidylcholine. Docosahexaenoic acid, eicosapentaenoic acid and linoleic acid did not alter the overall degree of incorporation of (14)C-AA. The results of this study suggest that both isomers inhibit eicosanoid production, and although trans10,cis12 exhibits pro-inflammatory activity at high concentrations, the CLA mixture maintains its beneficial anti-inflammatory action that contributes to its anti-carcinogenic and anti-atherogenic properties.