Selenium and selenoproteins in prostanoid metabolism and immunity

Abstract

Selenium (Se) is an essential trace element that functions in the form of the 21st amino acid, selenocysteine (Sec) in a defined set of proteins. Se deficiency is associated with pathological conditions in humans and animals, where incorporation of Sec into selenoproteins is reduced along with their expression and catalytic activity. Supplementation of Se-deficient population with Se has shown health benefits suggesting the importance of Se in physiology. An interesting paradigm to explain, in part, the health benefits of Se stems from the observations that selenoprotein-dependent modulation of inflammation and efficient resolution of inflammation relies on mechanisms involving a group of bioactive lipid mediators, prostanoids, which orchestrate a concerted action toward maintenance and restoration of homeostatic immune responses. Such an effect involves the interaction of various immune cells with these lipid mediators where cellular redox gatekeeper functions of selenoproteins further aid in not only dampening inflammation, but also initiating an effective and active resolution process. Here we have summarized the current literature on the multifaceted roles of Se/selenoproteins in the regulation of these bioactive lipid mediators and their immunomodulatory effects.     

Study of diet and drug interactions on prostanoid metabolism

To study the extent to which combinations of different dietary lipids stimulate or inhibit prostanoid synthesis groups of 12 rats were fed diets containing 10% (w/w) of either safflower oil, hydrogenated coconut oil/safflower oil, cod liver oil/safflower oil or cod liver oil/linseed oil for a period of four weeks. All diets, with the exception of the safflower oil feed, contained similar levels of linoleic acid. Two further groups of rats placed on the cod liver oil diets were injected with indomethacin (4 mg/kg, i.p.) every three days to establish the completeness of dietary prostaglandin (PG) inhibition. In spite of a 20 fold difference in dietary linoleic acid content, the safflower oil group had similar PG generating capacities to the saturated fat control group, suggesting tight metabolic control of PGs and their precursors. Although there were prostanoid variations in tissue responses, both of the cod liver oil diets substantially reduced generation of aortic, whole blood and renal prostanoids, and decreased urinary PG excretion. The degree of inhibition of renal PGs was substantially greater in the cod liver oil/linseed oil group, with prostaglandin levels being 35% lower than those observed in the cod liver oil/safflower oil fed animals suggesting that linolenic acid and the marine oil fatty acids act synergistically to inhibit formation of 2-series prostaglandins. Concurrent administration of omega-3 fatty acids and indomethacin reduced PG levels further than those obtainable by diet alone, demonstrating that the diets did not result in maximal inhibition. Awareness of these various effects is important for both physiological or clinical studies in which dietary manipulations are used as a means of modifying prostanoid synthesis.     

Antiplatelet activity of synthetic pyrrolo-benzylisoquinolines

Pyrrolo-benzylisoquinolines were prepared as target compounds and their antiplatelet aggregation activity, adreno-receptor affinity, and cytotoxicity were screened. Compounds 1d-9d showed specific antiplatelet aggregation activity induced by arachidonic acid and collagen. Among them, 8d and 9d exhibited better activity than the reference drug, aspirin and 9d also showed inhibition of platelet aggregation by all four inducers.     

Postprandial hyperlipemia inhibits platelet aggregation without affecting prostanoid metabolism

The objective of this work was to characterize changes in platelet aggregability during postprandial hypertriglyceridemia with special emphasis on arachidonic acid metabolism. Ten healthy young men consumed 100 g fat after a fasting period of 12 hr. In-vitro platelet aggregation induced by ADP and collagen was measured at 0, 3, 5, and 9 hours after the fat intake. The major arachidonic acid metabolites, 12-hydroxyeicosatetraenoic acid (12-HETE), thromboxane A2 (TXA2), prostaglandin F2 alpha (PGF2a), and prostaglandin E2 (PGE2) produced during collagen-induced platelet activation were quantified by gas chromatography/mass spectrometry. A significant decrease in platelet aggregability induced by both ADP and collagen was detected during the postprandial hyperlipemia. No significant changes could be found in the prostanoid pattern of collagen activated platelets. There was no correlation between the degree of the inhibition of platelet aggregation and the relative or absolute increase of triglyceride-levels in the plasma during the postprandial hyperlipemia.     

Methylglyoxal: metabolism and biological activity

The enzymic formation of methylglyoxal from dihydroxyacetone phosphate and aminoacetone (metabolites of carbohydrates and proteins) is considered. Methylglyoxal transformation into lactic and pyruvic acids is related to energy metabolism, catabolism and anabolism dissociation processes in carbohydrates and proteins, and, probably, to maintenance of asymmetrical entropy in vivo on the constant level. Attention is paid to the methylglyoxal inhibition of enzymes, its interaction with glutathione and polyamines affecting the mechanisms regulating protein synthesis and cellular division. The methods for obtaining and quantitative determination of methylglyoxal are described.     

Antiplatelet activity of N-methoxycarbonyl aporphines from Rollinia mucosa

Bioassay-directed fractionation of the stems of Rollinia mucosa led to the isolation of new N-methoxycarbonyl aporphine alkaloids, romucosine A (1), romucosine B (2), romucosine C (3), and romucosine D (4), along with the known alkaloid, N-methoxylcarbonyl-nornuciferine (5). Alkaloids 1 and 4 exhibited significant inhibition of collagen, arachidonic acid, and platelet activating factor-induced platelet aggregation, and alkaloid 3 also showed an inhibitory effect on arachidonic acid induced platelet aggregation.     

Intracellular measurement of prostaglandin E2: effect of anti-inflammatory drugs on cyclooxygenase activity and prostanoid expression.

Cyclooxygenase (COX) converts arachidonic acid to prostaglandin (PG) H2, which is further metabolized to various prostaglandins, prostacyclin and thromboxane A2. COX exists in at least two different isoforms. COX-1 is constitutively expressed, whereas COX-2 is induced by proinflammatory stimuli. Prostaglandin E2 is a major metabolite of COX activation. In order to compare the activity of target ligands and COX inhibitors on PGE2 synthesis and release, the responsiveness of several cell lines to the calcium ionophore A23187, bacterial lipopolysaccharide (LPS), nonsteroidal anti-inflammatory drugs (NSAIDs), and the glucocorticoid, dexamethasone, were investigated. For intracellular measurements, the culture supernatant was aspirated, and the cells were thoroughly washed and lysed with dodecyltrimethylammonium bromide. Intracellular and secreted PGE2 were measured with an enzyme immunoassay. A23187 and LPS increased intracellular PGE2 in a dose-dependent manner. Kinetic experiments with A23187-stimulated mouse 3T3 fibroblast cells revealed a distinct biphasic response in COX activity. In the presence of NSAIDs or dexamethasone, there was a dose-dependent inhibition in intracellular PGE2 with A23187-stimulated 3T3 cells. Inhibitory studies demonstrated an apparent increased sensitivity of COX activity to the action of inhibitors when measuring intracellular PGE2 compared with using cell culture supernatants. Indeed, intracellular PGE2 levels were comprehensively reduced in the presence of low concentrations of inhibitor. The utilization of cell culture lysates and, in particular, measurement of intracellular PGE2 should prove useful for identifying new COX inhibitors.     

Influence of sulfasalazine, 5-aminosalicylic acid and sulfapyridine on prostanoid synthesis and metabolism in rabbit colonic mucosa

The effects of sulfasalazine (SASP) and its cleavage products 5-aminosalicylic acid (5-ASA) and sulfapyridine (SP) on prostanoid (PG) synthesis and degradation were determined in rabbit colonic mucosa fractions in vitro. When the microsomal fraction was incubated with (14C)arachidonic acid, 10(-3) M SASP and SP did not markedly change the formation of labeled PGE2, PGF2 alpha, TxB2 and 6-keto-PGF1 alpha X 10(-4) M 5-ASA increased synthesis about 2.7-fold; the pattern of PG identified was unaltered. In the presence of the 10-fold higher concentration of 5-ASA, PG synthesis remained elevated at a similar level. When the cytosolic fraction was incubated with (3H)PGE2, 10(-3) M 5-ASA was without influence and 10(-3) M SP decreased slightly PGE2 breakdown. However, SASP showed a pronounced inhibitory effect at 10(-5) M and inhibition of PGE2 degradation was complete at 10(-3) M SASP. The results are compatible with the assumption that stimulation of PG synthesis by 5-ASA is related to therapeutic benefit in the treatment of ulcerative colitis.     

Lipid storage in cultured articular chondrocytes due to prostanoid precursors and a prostanoid synthesis inhibitor

Lapine articular chondrocytes were subcultured in the presence or absence of the prostanoid precursors, arachidonic acid or dihomo-gamma-linolenic acid, and the cyclooxygenase inhibitor indomethacin. Lipid storage was studied microscopically using the Sudan black staining method. Control chondrocyte cultures showed a weakly positive staining reaction until confluence was reached, at which point the intra-cytoplasmic lipid content decreased. Both arachidonic acid and dihomo-gamma-linolenic acid at 100 mumol/l caused a marked increase in lipid storage which continued even after confluence was achieved. 1 mumol/l concentrations were indistinguishable from controls, whereas 10 mumol/l concentrations elicited a slight increase in lipid storage compared with controls. The prostaglandin cyclooxygenase inhibitor indomethacin did not affect chondrocyte lipid storage. However, administration of a prostanoid precursor in the presence of indomethacin caused a massive increase in intra-cytoplasmic storage of lipid, eventually leading to cell death. A possible explanation is that indomethacin may alter chondrocyte lipid metabolism in the presence of substrate molecules by rechanneling lipid synthesis away from the prostaglandin pathway to other lipid synthetic pathways.     

Antiplatelet activity of the long-acting thromboxane receptor antagonist BMS 180,291 in monkeys

The effects of the novel TxA₂/prostaglandin endoperoxide (TP) receptor antagonist BMS 180,291 on platelet reactivity was determined ex vivo in conscious African green monkeys. Platelet aggregation responses to U-46,619 were decreased 50% and 100% at 23 to 24 hrs after BMS 180,291 oral doses of 1 and 3 mg/kg, respectively. In addition to inhibiting aggregation, a 3 mg/kg oral dose of BMS 180,291 also produced an 11±3-fold shift to the right in the U-46,619 concentration-response relationship for platelet shape change at 24 hrs after dosing. When the 3 mg/kg oral dose was continued for 11 days, the shift in this concentration-response relationship increased to 26±10- and 93±30-fold at 24 hrs after the 8th and 11th doses, respectively. This progressive inhibition corresponds to 93±3 and 99±1% blockade of platelet TP-receptors responsible for shape change, respectively. Comparable levels of TP-receptor blockade have been previously correlated with antithrombotic and antiischemic activities of TP-receptor antagonists in vivo. Platelet reactivity to U-46,619 had completely recovered on the 7th day after the final dose of BMS 180,291, indicating effective elimination from the circulation over this interval. In separate experiments, a 3-mg/kg i.v. dose of BMS 180,291 produced only marginal and transient hemodynamic effects in anesthetized African green monkeys. Overall, these data demonstrate that BMS 180,291 given orally once a day produces a sustained and therapeutically-relevant level of TP-receptor antagonism.     

Cellular ras activity and phospholipid metabolism

Cellular ras activity has been neutralized in 3T3 cells by microinjection of a specific anti-ras monoclonal antibody. The injected antibody efficiently inhibited proliferation in cells treated with a phorbol ester and a calcium ionophore, or with prostaglandin F2 alpha. These treatments were designed to imitate the action of phospholipase C or of phospholipase A2. In addition, the highly efficient mitogenic potential of phosphatidic acid was inhibited by the injected antibody even more efficiently than was serum-induced proliferation. The close reliance of phospholipid-induced mitogenesis upon ras activity suggests that ras proteins are unlikely to function to control the action of a phospholipase.     

Distribution and function of prostanoid receptors: studies from knockout mice

Recent developments in the molecular biology of the prostanoid receptors has allowed the investigation of the physiological roles of each individual receptor type and subtype. The following article reports the prostanoid receptor distributions deduced from Northern blot and in situ hybridization analyses, summarizes the phenotypes of each receptor knockout mice, and discusses recent studies investigating the effects of each receptor deficiency on the inflammatory response and female reproductive processes. The combination of expression pattern and knockout analyses enabled us to determine which receptor expressed in a particular cell is important for the maintenance of normal and/or pathological physiology. The results from these analyses may be useful in the development of novel therapeutics that can selectively manipulate prostanoid-mediated actions.     

Antiplatelet activity of the long-acting thromboxane receptor antagonist BMS 180,291 in monkeys.

The effects of the novel TxA2/prostaglandin endoperoxide (TP) receptor antagonist BMS 180,291 on platelet reactivity was determined ex vivo in conscious African green monkeys. Platelet aggregation responses to U-46,619 were decreased 50% and 100% at 23 to 24 hrs after BMS 180,291 oral doses of 1 and 3 mg/kg, respectively. In addition to inhibiting aggregation, a 3 mg/kg oral dose of BMS 180,291 also produced an 11 +/- 3-fold shift to the right in the U-46,619 concentration-response relationship for platelet shape change at 24 hrs after dosing. When the 3 mg/kg oral dose was continued for 11 days, the shift in this concentration-response relationship increased to 26 +/- 10- and 93 +/- 30-fold at 24 hrs after the 8th and 11th doses, respectively. This progressive inhibition corresponds to 93 +/- 3 and 99 +/- 1% blockade of platelet TP-receptors responsible for shape change, respectively. Comparable levels of TP-receptor blockade have been previously correlated with antithrombotic and antiischemic activities of TP-receptor antagonists in vivo. Platelet reactivity to U-46,619 had completely recovered on the 7th day after the final dose of BMS 180,291, indicating effective elimination from the circulation over this interval. In separate experiments, a 3-mg/kg i.v. dose of BMS 180,291 produced only marginal and transient hemodynamic effects in anesthetized African green monkeys. Overall, these data demonstrate that BMS 180,291 given orally once a day produces a sustained and therapeutically-relevant level of TP-receptor antagonism.