PGE2 secretion from organ cultured gastric mucosa: Correlation with cyclooxygenase activity and endogenous substrate release

In gastrointestinal research the in vitro release of prostaglandins from incubated or cultured biopsies is a widely used method to estimate prostaglandin synthesis. We therefore investigated the rate limiting mechanisms of PGE₂ release in organ cultured gastric mucosa of the rabbit, determining PGE₂ secretion from organ cultured mucosal biopsies by radioimmunoassay and prostaglandin synthesizing capacity by in vitro incubation of mucosal homogenate or microsomes with [¹⁴C]-arachidonic acid.Freshly taken biopsies secreted PGE₂ at an initial high rate, that decreased during the following 4 hrs of culture. This PGE₂ release was dose dependently reduced by inhibitors of the prostaglandin cyclooxygenase. 5mM acetylsalicylic acid (ASA) maximally suppressed PGE₂ secretion to 7% of controls, and the inhibition by ASA was quantitatively similar at every given culture period. PGE₂ release was markedly increased by carbenoxolone but was only slightly activated by extracellular calcium and the Ca⁺⁺-ionophore A23187. However, Ca⁺⁺/A23187 were unable to maintain PGE₂ secretion at the initial rate.PGE₂ secretion was undisturbed in calcium-free medium but was reduced to 50–60% of controls by excess EDTA. The intracellular calcium chelator 1,2-bis-(2-aminophenoxy)-ethane-N,N,N′,N′,-tetraacetic acid-acetoxymethyl ester (BAPTA-AM) similarly inhibited PGE₂ release to 72% of controls. In contrast, PGE₂ release was unaffected by the intracellular calcium antagonist 3,4,5-trimethylene-bis(4-formylpyridinium bromide) dioxime (TMB-8), the calmodulin antagonists N-(6-aminohexyl)-1-5-chloro-1-naphthalenesulfonamide (W-7) and calmidazolium (compound R24571) or various direct inhibitors of endogenous arachidonic acid release like tetracaine, bromophenacyl bromid, neomycine or low dose quinacrine, indicating that the reduction of PGE₂ release by EDTA or BAPTA may be mediated by mechanisms different from substrate release. In contrast, an inhibition of PGE₂ secretion by quinacrine at high concentrations (≥ 0.8mM) was attributed to a direct inhibition of the prostaglandin cyclooxygenase, similar to ASA. Finally, the reduction of the prostaglandin synthesizing capacity by ASA was strongly correlated with the inhibition of PGE₂ secretion, also at low concentrations and minor degrees of inhibition.From these data we conclude, that the activity of the prostaglandin cyclooxygenase is rate limiting for PGE₂ secretion from organ cultured mucosal biopsies rather than arachidonic acid release by a phospholipase A₂. This should be considered for interpretation of studies based on prostaglandin release from cultured mucosa.     

Questionable role of leukotriene B4 in monosodium urate (MSU)-induced synovitis in the dog

Monosodium urate (MSU)-induced synovitis in the dog's stifle (knee joint) is similar to an acute gouty attack in man in which a loss of function of the joint correlates with massive influx of neutrophils and the release of an assortment of inflammatory mediators (e.g. histamine, bradykinin, lysosomal enzymes, complement and eicosanoids) into the synovial space. We found in the urate-induced inflammatory exudates 3 hr post MSU the following: 88 million leukocytes/ml (95% neutrophils) and eicosanoid concentrations of LTB₄, LTC₄, and PGE₂ of < 0.1, 1.4 and 20 ng/ml, respectively. Isotonic saline injected knee joints at 3 hr contained 5 million leukocytes/ml (95% neutrophils) and concentrations of LTB₄, LTC₄, and PGE₂ of < 0.1, 0.7 and 0.2 ng/ml, respectively. Intrasynovial injections of 1 μg LTB₄, 10 μg PGE₂ or the combination of LTB₄ and PGE₂ produced no reduction of paw pressure for up to 3 hr. Leukocyte concentrations measured at 3 hr in joints injected with these arachidonic acids metabolites were similar to saline controls. These results question the role of LTB₄ as a chemotactic and inflammatory mediator in urate-induced synovitis in the dog but confirm the importance of PGE₂ and possibly LTC₄ in this model.     

Leukotriene and prostaglandin production in rat brain synaptosomes treated with phospholipase A2 neurotoxins and enzymes

β-Bungarotoxin (β-BuTX) and notexin cause an irreversible blockade of neurotransmitter release through specific and potent effects at the presynaptic nerve terminal, however, the mechanism of action in uncertain. We examined the effects of β-BuTX and notexin on LT and PG production in rat cerebrocortical synaptosomes in order to determine if eicosanoid production might mediate or regulate the pharmacological actions of these phospholipase A₂(PLA₂) neurotoxins. The effects of the PLA₂ enzymes isolated from Naja naja atra and Naja nigricollis snake venoms (which are not presynaptic selective) on LT and PG production were compared with the effects of β-BuTX and notexin. N. n. atra PLA₂, β-BuTX, and notexin (all 50 nM) produced a time dependent rise in free fatty acids as measured in synaptic plasma membranes isolated from treated synaptosomes. Both the PLA₂ neurotoxins and enzymes stimulated LTC₄, LTB₄, and PGE₂ production, as measured by radioimmunoassay. In all cases, the PLA₂ enzymes were more potent than the PLA₂ neurotoxins. This observations correlates with their relative enzymatic potencies, as measured by free fatty acid generation. EDTA and BSA antagonized PLA₂ induced LTB₄ production and BSA also antagonized PLA₂ induced PGE₂ production. These results suggest that stimulation of eicosanoid production does not mediate the potent and specific presynaptic actions of β-BuTX and notexin.     

Tissue differences in vascular permeability induced by leukortriene B4 and prostaglandin E2 in the rat

The activity of synthetic LTB₄ and PGE₂, in increasing vascular permeability was tested simultaneously in seventeen different organs in the rat. Rats were injected in the aortic arch through a cannula in the carotid artery with ¹²⁵-I-albumin, ⁵¹Cr-erythrocytes, and ⁵⁷Co-EDTA. The rats were then injected through the carotid artery cannula with LTB₄, PGE₂ or a combination of LTB₄ and PGE₂. Eight minutes later the rats were killed and the activity of ¹²⁵I, ⁵¹Cr, and ⁵⁷Co measured in different organs. Changes in vascular permeability were infered from changes in the ratios of the isotope activities. LTB₄ (15 μg/kg) induced enhanced permeability in caecum, small bowel, skin, fat pad, stomach, pancreas, and aorta, but not in the heart, brain, colon, testes, diaphragm, forelimb, cremaster muscle, lung, kidney or eye. A lower dose of LTB₄, 3 μg/kg, enhanced vascular permeability in caecum, small bowel, skin, stomach, and aorta. PGE₂ (1 μg/kg) enhanced vascular permeability only in the caecum. A combination of LTB₄ (3 μg/kg) and PGE₂ (1 μg/kg) was more potent than either alone. Rats depleted of neutrophils with anti-neutrophil serum were less sensitive to LTB₄ than intact rats. These findings suggest that the vasculatures of different tissues in the rat vary markedly in their susceptibility to LTB₄ induced increases in permeability.     

Production of leukotriene B4 and prostaglandin E2 by turbot (Scophthalmus maximus) leukocytes

Production of two eicosanoids derived from lipoxygenase and cyclooxygenase activities: leukotriene B₄ (LTB₄) and prostaglandin E₂ (PGE₂), respectively, have been simultaneously determined in turbot (Scophthalmus maximus) blood leucocyte and kidney macrophage supernatants by a reverse phase high performance liquid chromatography (HPLC) system coupled with a Diode–Array detector. Levels of LTB₄ after calcium ionophore challenge were 4.08 ng ml⁻¹ in blood leukocyte supernatants and 0.25 ng ml⁻¹ in kidney macrophage supernatants. The levels found for PGE₂ were 428.23 and 606.67 ng ml⁻¹ in blood leukocytes and kidney macrophage supernatants, respectively. When blood leukocytes were treated with the respective inhibitors for the enzymes implicated on the synthesis of both compounds an inhibition of 90.35% was observed for PGE₂ and 76.44% for LTB₄. The detection limit of the method was 0.15 ng ml⁻¹ for LTB₄ and 50 ng ml⁻¹ for PGE₂.     

15-Hydroxy-eicosatetraenoic acid (15-Hete) inhibits carragheenan-induced experimental arthritis and reduces synovial fluid leukotriene B4 (LTB4)

15-Hydroxy-eicosatetraenoic acid (15-HETE), a product of arachidonic acid, has no proinflammatory capacity, but can inhibit the formation and the chemotactic response of neutrophils to leukotriene B₄ (LTB₄), a potent mediator of inflammation. The purpose of the present study was to determine whether intraarticular administration of 15-HETE in carragheenan-induced acute arthritis might decrease the levels of LTB₄ in synovial fluid and modify the arthritis. A bilateral acute knee joint arthritis was established in 7 dogs by intraarticular injections of carragheenan every third day. To the right joints, 15-HETE was administered both concomitantly with the carragheenan injections and continously via an osmotic pump. In samples of synovial fluid obtained on day 0, 3 and 10 PGE₂ and LTB₄ were determined using reversed phase high performance liquid chromatography combined with radioimmunoassays and neutrophil chemokinesis. In the presence of 15-HETE the clinical severity of arthritis was significantly reduced and the volume synovial effusate was decreased on an average by 42%. Furthermore, the relative number of neutrophils in histological sections of synovial tissue was decreased by 58%. Intaarticular carragheenan injection induced LTB₄ formation, and maximum levels were obtained on day 3 (279.2 ± 148.2 pg/joint). PGE₂ was also present on a day 3, but maximum levels were detected on day 10 (9.5 ± 4.8 ng/joint). In joints injected with both carragheenan and 15-HETE the levels of LTB₄ on days 3 and 10 were inhibited by 90% and 83%, respectively. For PGE₂ a small but significant decrease was found on both day 3 and on day 10. These results show that LTB₄ may be an important mediator of acute arthritis induced by carragheenan in dogs, and that intraarticular administration of 15- HETE can modify this arthritis by inhibiting LTB₄ formation.     

Delayed-onset synergism between leukotriene B4 and prostaglandin E2 in human skin

The time-course of cutaneous inflammatory responses to LTB₄ and PGE₂ both alone and in combination has been studied in 10 healthy volunteers. LTB₄ induced a transient wheal and flare response in some subjects, maximal at 15 minutes and succeeded by an erythematous, indurated lesion at 2–4 hours. PGE₂ elicited a wheal and erythema response which resolved within 1–2 hours. Combination of LTB₄ and PGE₂ produced acute wheal and erythema responses which did not differ significantly from the summation of responses to the individual constituents of the mixture or from responses to a two-fold increase in the concentration of either component. Wheal and erythema responses persisted, however, with significant potentiation of responses 4 hours after injection. As both leukotrienes and prostaglandins are generated in acute allergic reactions, the effects of these mediators in combination could contribute to persisting and late-onset responses to allergen, in both the skin and lung. In particular, sustained responses to the combination of LTB₄ and PGE₂ might be important in the pathogenesis of inflammatory skin diseases such as psoriasis.     

Opposite effects of adrenalectomy on eicosanoid release in rat peritoneal macrophages and spleen

The effect of adrenalectomy on the formation of cyclo-oxygenase and lipoxygenase products by activated peritoneal rat macrophages was determined and compared with that of the spleen. After isolation, the cells and tissues were incubated with [1-¹⁴C] arachidonic acid and the Ca-ionophore A23187 and the metabolites isolated by HPLC chromatography. The main components formed in the macrophages of the controls are 6-keto-PGF_1α, TxB₂ and 12-HETE. One peak represents 5, 12 di HETE. Smaller amounts of PGF_2α, PGE₂, PGD₂, LTB₄ and 15-HETE are also present. After adrenalectomy, a considerable increase occurs in the amounts of LTB₄, 15-HETE and 12-HETE. The increase in the PG is smaller. The compounds formed from endogenous arachidonic acid are also determined. In the cells of the controls, the formation of LTB₄ is considerably increased after adrenalectomy. In the spleen, PGD₂ and 12-HETE are decreased after adrenalectomy.The effect of the macrophages is most probably related to a diminished amount or inactivation of lipocortin, a glucocorticosteroid induced peptide with PlA₂ inhibitory activity in adrenalectomized animals. In the decrease in formation in the spleen, the absence of the permissive effect of glucocorticosteroids on the hormone-induced lipolysis may play a role.     

Allosteric regulation by membranes and hydrophobic subsites in phospholipase A2 enzymes determine their substrate specificity

Lipids play critical roles in several major chronic diseases of our times, including those that involve inflammatory sequelae such as metabolic syndrome including obesity, insulin sensitivity, and cardiovascular diseases. However, defining the substrate specificity of enzymes of lipid metabolism is a challenging task. For example, phospholipase A₂ (PLA₂) enzymes constitute a superfamily of degradative, biosynthetic, and signaling enzymes that all act stereospecifically to hydrolyze and release the fatty acids of membrane phospholipids. This review focuses on how membranes interact allosterically with enzymes to regulate cell signaling and metabolic pathways leading to inflammation and other diseases. Our group has developed “substrate lipidomics” to quantify the substrate phospholipid specificity of each PLA₂ and coupled this with molecular dynamics simulations to reveal that enzyme specificity is linked to specific hydrophobic binding subsites for membrane phospholipid substrates. We have also defined unexpected headgroup and acyl chain specificity for each of the major human PLA₂ enzymes, which explains the observed specificity at a structural level. Finally, we discovered that a unique hydrophobic binding site—and not each enzyme’s catalytic residues or polar headgroup binding site—predominantly determines enzyme specificity. We also discuss how PLA₂s release specific fatty acids after allosteric enzyme association with membranes and extraction of the phospholipid substrate, which can be blocked by stereospecific inhibitors. After decades of work, we can now correlate PLA₂ specificity and inhibition potency with molecular structure and physiological function.     

Effect of exogenous phospholipase A2 and triacylglycerol lipase on the synthesis and release of monoenoic and bisenoic prostaglandins from isolated rat uterus

The possible existence of a selective and independent mechanism subserving the formation of prostaglandin E₁ (PGE₁) and of prostaglandin E₂ (PGE₂) has been reported in previous studies from our group. In the present experiments we have demonstrated that neutral lipid lipases play an important role yielding dihomo-gamma-linolenic acid for the formation of PGE₁. Indeed, exogenous triglyceride lipase added to the incubation bathing solution at a concentration of 150 U/ml increased several fold the production of PGE₁ by isolated uterine strips obtained from spayed rats. Nevertheless the presence of the enzyme did not modify significantly the synthesis and release of bisenoic PGs (PGE₂ and PGF_2α). When triarachidonin was added, as an artificial substrate into the incubating medium in order to detect the presence of endogenous triacylglycerol lipase, we observed a significant increment in the generation of PGE₂ (p < 0.005) and of PGF_2α (p < 0.001) without evident changes in the basal release of PGE₁. On the other hand, the addition of phospholipase A₂ (PLA₂) at 0.2 U/ml, increased significantly the production of PGE₂ (p < 0.001) but failed to alter the concentration of PGE₁ in the incubating solution. Surprisingly, PLA₂ did not enhance the synthesis of PGF_2α in the present experiments, a situation for which we do not have a clear explanation. Exogenous bradykinin (10⁻⁶ M), a well known stimulant of PLA₂ activity in several tissues, also increased significantly (p < 0.001) the production of PGE₂ without altering that of PGE₁. Results presented herein provide strong support to the notion that different enzyme mechanisms and different lipid stores are linked to the formation of PGE₁ and PGE₂ in isolated rat uteri.     

Interaction of human neutrophils with airway epithelial cells: Reduction of leukotriene B4 generation by epithelial cell derived prostaglandin E2

Airway epithelial cells (AEC) play an active role in the regulation of inflammatory airway disease. In the present study we analyzed the interaction of AEC with polymorphonuclear leukocytes (PMN) in coincubation with respect to their arachidonic acid (AA) metabolism using reversed phase-HPLC and post-HPLC-ELISA. Primary cultures of porcine AEC released predominantly PGE₂, PGF_2a, and 15-hydroxyeicosatetraenoic acid (15-HETE), whereas the major human PMN-derived AA metabolite was the chemotactic factor leukotriene B₄ (LTB₄). In AEC-PMN cocultures stimulated with the calcium ionophore A23187, PMN-related 5-lipoxygenase products were decreased by 45%. This reduction in LTB₄ formation in the presence of AEC was mainly due to PGE₂ generated by the epithelial cells, whereas 15-HETE made a minor contribution. Most of the effect was inhibited by AEC pretreatment with acetylsalicylic acid and restored by addition of equivalent amounts of exogenous PGE₂. LTB₄ degradation was not enhanced in PMN-AEC coincubations. Moreover, reduction of LTB₄ formation in this system did not require an intimate cell-to-cell contact as shown by studies involving filter membranes for PMN-AEC separation. Superoxide anion concentrations were also decreased in PMN-AEC coincubations; this effect, however, was unrelated to PGE₂ for quantitative reasons and was probably due to 2 is the major mediator in the coincubation of porcine AEC and human PMN that downregulates neutrophil responses by activating receptors on the neutrophil. A minor contributor in this course of PMN-AEC interaction may be the 15-HETE transcellular pathway. Overall, airway epithelium appears to play an antiinflammatory role by damping the proinflammatory potential of neutrophils. J. Cell. Physiol. 175:268–275, 1998. © 1998 Wiley-Liss, Inc.     

Role of leukotriene B4 in celecoxib-mediated anticancer effect

Celecoxib, a selective cyclooxygenase-2 (COX-2) inhibitor, has anticancer effect on many cancers associated with chronic inflammation by both COX-2-dependent and COX-2-independent mechanisms. The non-COX-2 targets of celecoxib, however, are still a matter of research. Leukotriene B₄ (LTB₄) has been implicated in prostate and colon carcinogenesis, but little is known about the potential role of LTB₄ in celecoxib-mediated anticancer effect. In this study, we evaluated whether LTB₄ was involved in celecoxib-mediated inhibitory effect on human colon cancer HT-29 cells and human prostate cancer PC-3 cells. Our data showed that survival of both cell lines was obviously suppressed after celecoxib treatment for 72 h in a concentration-dependent manner. However, only in HT-29 cells, this inhibitory effect could be reversed by LTB₄, which promoted survival of HT-29 cells rather than PC-3 cells. Consistent with these results, lioxygenase (LOX) potent inhibitor nordihydroguaiaretic acid (NDGA) had a higher inhibitory effect on HT-29 cells than PC-3 cells. Additionally, ELISA results showed that celecoxib could suppress expression of LTB₄ in both cell lines, whereas, inhibition of PGE₂ was only detected in HT-29 cells. These results indicate that the anticancer effect of celecoxib is COX-2-independent in HT-29 and PC-3 cells and in HT-29 cells primarily via down-regulating LTB₄ production.     

In vitro and in vivo chemotaxis of guinea pig leukocytes toward leukotriene B4 and its w-oxidation products

Leukotriene B₄ (LTB₄), 20-OH-LTB₄, and 20-COOH-LTB₄ were studied for their relative activities towards guinea pig peritoneal eosinophils and neutrophils during in vitro chemotaxis in modified Boyden chambers. The leukotrienes were also injected into guinea pig skin, and the cellular infiltrate in 4 hour biopsies was evaluated histologically. Eosinophils migrated more actively than neutrophils towards LTB₄in vitro, while in vivo, more neutrophils were observed. 20-OH-LTB₄, was markedly less active than LTB₄in vivo and in vitro, and 20-COOH-LTB₄ was barely active at all. Crude ionophore-stimulated neutrophil supernatants (ECF) were more active towards eosinophils than towards neutrophils, both in vivo and in vitro, compared to the pure leukotrienes. The data confirm the potent chemotactic properties of LTB₄ for eosinophils and neutrophils, with less activity of its w-metabolites.     

Antibodies against type II phospholipase A2 prevent renal injury due to ischemia and reperfusion in rats

This study was performed to determine the involvement of type II phospholipase A₂ (PLA₂-II) in renal injury caused by ischemia and reperfusion. Ischemia and reperfusion significantly elevated levels of blood urea nitrogen and serum creatinine in rats. These increases were significantly reduced by i.v. administration of rabbit IgG F(ab′)2 fragments against rat PLA₂-II. Increased levels of acid-stable PLA₂ activity in the kidney were caused by ischemia and reperfusion, and were suppressed by administration of anti-PLA₂-II F(ab′)2. Increased levels of myeloperoxidase activity, a marker of neutrophil infiltration, in the kidney were also reduced after anti-PLA₂-II F(ab′)2 treatment. These results suggest that PLA₂-II plays a pivotal role in pathogenesis of ischemia and reperfusion injury through induction of neutrophil infiltration.     

Dual inhibitory effect of Glycyrrhiza glabra (GutGard™) on COX and LOX products

Glycyrrhiza glabra and its phytoconstituents have been known to possess widespread pharmacological properties as an anti-inflammatory, anti-viral, antitumour and hepatoprotective drug. In this study, we examined the inhibitory potential of extract of G. glabra (GutGard™) root and its phytoconstituents (glabridin, glycyrrhizin, and isoliquiritigenin) on both cyclooxygenase (COX) and lipoxygenase (LOX) products in order to understand the mechanism of its anti-inflammatory action. Inhibitory effect of GutGard™ and its phytoconstituents on lipopolysaccharide (LPS) induced prostaglandin E₂ (PGE₂), calcimycin (A23187) induced thromboxane (TXB₂), and leukotriene (LTB₄) release was studied using murine macrophages (J774A.1) and human neutrophil (HL-60) cells. Results revealed that, G. glabra and glabridin significantly inhibited PGE₂, TXB₂ (COX) and LTB₄ (LOX), while, isoliquiritigenin exerted inhibitory effect only against COX products but failed to suppress LOX product. However, glycyrrhizin at the tested concentrations failed to exhibit inhibitory effect on both COX and LOX products. Here, we report for the first time that G. glabra (almost devoid of glycyrrhizin) exhibits anti-inflammatory property likely through the inhibition of PGE₂, TXB₂ and LTB₄ in mammalian cell assay system, which could be influenced in part by glabridin and isoliquiritigenin.     

Bio-synthesis of PGD2 and TXB2 by rabbit blood monocytes

A method for the preparation of a highly purified sample of rabbit blood monocytes is described. The metabolism of arachidonic acid (AA) in these cells was studied. Mononuclear cells were prepared by centrifugation on Ficoll-Paque gradients and the monocytes were obtained by further centrifugation and adherence onto plastic culture dishes. These procedures provided a preparation which contained 95% monocytes (non-specific esterase positive). Incubation of [1-¹⁴C]-AA with these cells produced four major metabolites which were separated by TLC; these corresponded to prostaglandin (PG) D₂, thromboxane (TX) B₂, 12-hydroxyheptadecatrienoic acid (HHT) and 12-/15- hydroxyeicosatetraenoic acid (HETE). A minor product which co-migrated with PGE₂ was also detected but neither 6-keto-PGF_1α nor PGF_2α were detected. Also, there was no evidence of the formation of 5-lipoxygenase products (5-HETE and LTB₄) by rabbit monocytes with or without calcium-ionophore A23187-stimulation. The production of PGD₂, TXB₂ and PGE₂ was further confirmed by analyzing [³H]-AA metabolites using high-performance liquid chromatography (HPLC) with tritiated standards as references. The biosynthesis of these compounds from endogenous substrate in A23187-stimulated monocytes was confirmed by specific radioimmunoassays with or without prior HPLC separation. The synthesis of immunoreactive LTB₄ and LTC₄ by A23187-stimulated cells was also monitored and found to be relatively low. The synthesis of PGD₂, TXB₂ and PGE₂ from both exogenous and endogenous substrate was suppressed by treatment of the monocytes with indomethacin (10⁻⁶ M).     

Effect of Marine-Derived n-3 Polyunsaturated Fatty Acids on Major Eicosanoids: A Systematic Review and Meta-Analysis from 18 Randomized Controlled Trials

Background

Marine-derived n-3 polyunsaturated fatty acids (PUFA) may have a beneficial effect on inflammation via lowering pro-inflammatory eicosanoid concentrations. We aimed to assess the effect of marine-derived n-3 PUFA on prostaglandin E₂ (PGE₂), thromboxane B₂ (TXB₂), and leukotriene B₄ (LTB₄) through systematic review and meta-analysis of randomized controlled trials.

Method and Findings

A structured search strategy on PubMed, Web of Science and Cochrane up to November 2015 was undertaken in this meta-analysis. Standard mean difference was used to calculate the effect size of marine-derived n-3 PUFA on PGE₂, TXB₂ and LTB₄ in a random-effect model. A total of 18 RCTs with 826 subjects were included in this systematic review and meta-analysis. Supplementation of marine-derived n-3 PUFA significantly decreased concentrations of TXB₂ in serum/plasma in subjects with high risk of cardiovascular diseases (SMD:-1.26; 95% CI: -1.65, -0.86) and LTB₄ in neutrophils in unhealthy subjects (subjects with non-autoimmune chronic diseases or auto-immune diseases) (SMD:-0.59: 95% CI: -1.02, -0.16). Subgroup analyses showed a significant reduction of LTB₄ in subjects with rheumatoid arthritis (SMD: -0.83; 95% CI: -1.37, -0.29), but not in non-autoimmune chronic disease patients (SMD: -0.33; 95% CI: -0.97, 0.31). No significant publication bias was shown in the meta-analysis.

Conclusions

Marine-derived n-3 PUFA had a beneficial effect on reducing the concentration of TXB₂ in blood of subjects with high risk of CVD as well as LTB₄ in neutrophils in unhealthy subjects, and that subjects with RA showed lower LTB₄ content with supplementation of marine-derived n-3 PUFA.     

In vitro secretion of digestive phospholipase A2 by midguts isolated from tobacco hornworms,Manduca sexta

We report on secretion of phospholipase A₂ (PLA₂) by in vitro preparations of midguts isolated from tobacco hornworms, Manduca sexta. This enzyme is responsible for hydrolysis of fatty acids from the sn‐2 position of phospholipids, a necessary step in fatty acid absorption. The in vitro midgut preparations are competent to secrete PLA₂ into incubation buffer. Secretion began within the first 30 min of incubation and increased to a maximum at 8 h. We selected 2 h incubations because substantial loss of tissue integrity was observed after 8 h incubations. Using 2 h incubations, we recorded increased secretion of digestive PLA₂ from midguts incubated in buffer amended with diet or with yeast as a component of the diet. We also recorded small increases in secretion of PLA₂ from midguts incubated in buffer amended with a specific phospholipid, phosphatidylcholine. Midguts incubated in buffer amended with increased concentrations of phospholipid did not yield higher levels of PLA₂ activity. Lepidopteran midguts can be divided into three regions, and we recorded the highest secretion of PLA₂ from the middle region and lowest secretion from the anterior region. Because isolated midguts responded to food chemicals with increased secretion of digestive PLA₂, we suggest that secretion of digestive enzymes in tobacco hornworms is regulated by a prandial and/or paracrine mechanism, as suggested for digestive proteases in other insect species. Arch. Insect Biochem. Physiol. 42:179–187, 1999 .© 1999 Wiley‐Liss, Inc.     

Inflammation and pain sensitivity: Effects of leukotrienes D4, B4 and prostaglandin E1 in the rat paw

Leukotrienes (LT's) and prostaglandins (PG's) have been proposed as mediators of vascular permeability change in inflammatory reactions. Also, prostaglandins, especially of the E-type, have been shown to enhance pain responses. In the present studies in rat, the effects of LTB₄ and LTD₄ on edema and pain thresholds were examined in combination with PGE₁ and/or brewer's yeast. Subplantar injections of LTD₄ or LTB₄ induced small increases in paw thickness which were potentiated by the co-administration of PGE₁. LTD₄ alone had no significant effect on the development of the yeast paw edema. LTB₄ was found to reduce significantly the yeast edema and this reduction could be reserved by administration PGE₁. A small but significant decrease in pain threshold was caused by PGE₁ and this was significant enhanced in the presence of LTD₄. LTB₄, like PGE₁, was found to cause slight hyperalgesia but no synergy between the two agents was observed. LTD₄ was found to have no effect on the initial hypoalgesia or subsequent development of hyperalgesia caused by brewer's yeast. Both LTB₄ and PGE₁, however, prevented the initial hypoalgesia and significantly reduced tha latency for development of yeast induced hyperalgesia. These effects of LTB₄ are discussed in terms of possible release of cyclooxygenase products.