Showing papers on "Xanthine published in 1994"
TL;DR: Although mammalian xanthine oxidase exists originally as a dehydrogenase form in freshly prepared samples, it is converted to an oxidase form during purification, either irreversibly by proteolysis or reversibly by sulfhydryl oxidation of the protein molecule, which changes the protein conformation around FAD.
Abstract: Although mammalian xanthine oxidase exists originally as a dehydrogenase form in freshly prepared samples, it is converted to an oxidase form during purification, either irreversibly by proteolysis or reversibly by sulfhydryl oxidation of the protein molecule. However, avoiding proteolysis the mammalian enzyme can be purified as an interconvertible form and thus can be used to compare directly the properties of xanthine dehydrogenase and the oxidase derived from the same enzyme molecule. The cDNAs encoding the enzyme have been cloned from several sources, and structural information is becoming available. The most significant difference between the two forms is the protein conformation around FAD, which changes the redox potential of the flavin and the reactivity of FAD with the electron acceptors, NAD and molecular oxygen. The flavin semiquinone is thermodynamically stable in xanthine dehydrogenase, but is unstable in xanthine oxidase. Detailed analyses by stopped-flow techniques suggest that the flavin semiquinone reacts with oxygen to form superoxide anion while the fully reduced flavin reacts to form hydrogen peroxide. Although xanthine dehydrogenase can produce greater amounts of superoxide anion than xanthine oxidase during xanthine-oxygen turnover, it seems to be physiologically insignificant because NAD inhibits almost completely the formation of superoxide anion. Although the involvement of this enzyme in reperfusion injury has been proposed, this seems to be more complex than originally envisaged and still remains to be established.
194 citations
Journal Article•
TL;DR: Data indicate that AA and ROS act on glial glutamate transport via distinct noninteracting mechanisms, which could independently and additively contribute to the impairment of reuptake function, a phenomenon observed in pathological conditions such as ischemia/reperfusion injury.
Abstract: Reuptake of glutamate in astrocytes, a critical mechanism involved in the maintenance of physiological excitatory amino acid neurotransmission, is inhibited by both arachidonic acid (AA) and reactive oxygen species (ROS), via incompletely defined molecular mechanisms. Because ROS are generated during AA metabolism and AA can be released as a result of ROS-mediated phospholipase A2 activation, it seems likely that their effects on uptake are mediated by a common mechanism. However, here we show that rapid (10-min) uptake inhibitions by AA or by ROS generated by the xanthine plus xanthine oxidase (XO) reaction are selectively abolished by distinct agents; bovine serum albumin (BSA) acts only on AA, whereas the scavenger enzymes superoxide dismutase (SOD) and catalase (CAT) and the disulfide-reducing agent dithiothreitol (DTT) act only on ROS. Moreover, when added together, xanthine/XO and AA decrease uptake in a fully additive manner. In particular, the effect of xanthine/XO is seen also in the presence of maximal AA inhibition. No major signs of cell damage or chemical reaction between AA and radicals accompany their cumulative effects on uptake. Finally, uptake inhibition elicited by AA and xanthine/XO together is attenuated but not blocked by either BSA, DTT, or SOD/CAT individually, whereas it is fully blocked and substantially reversed by a combination of SOD/CAT and BSA or SOD/CAT, DTT, and BSA. Together, these data indicate that AA and ROS act on glial glutamate transport via distinct noninteracting mechanisms. Therefore, they could independently and additively contribute to the impairment of reuptake function, a phenomenon observed in pathological conditions such as ischemia/reperfusion injury.
179 citations
TL;DR: It is concluded that peroxynitrite, but not .NO or superoxide and hydrogen peroxide, in concentrations likely to be encountered in vivo, caused nitrotyrosine formation and decreased the ability of SP-A to aggregate lipids.
Abstract: The conditions under which nitric oxide (.NO) may modulate or promote lung injury have not been identified. We hypothesized that .NO-induced injury results from peroxynitrite, formed by the reaction of .NO with superoxide. The simultaneous generation of .NO and superoxide by 3-morpholinosydnonimine (SIN-1, 0.1-2 mM) resulted in oxidation of dihydrorhodamine, a marker of peroxynitrite production, and a dose-dependent decrease in the ability of SP-A to enhance lipid aggregation. Western blot analysis of SIN-1 exposed SP-A samples, overlaid with a polyclonal antibody against nitrotyrosine, were consistent with nitration of SP-A tyrosine residues. Superoxide dismutase (100 U/ml), L-cysteine (5 mM), xanthine oxidase (10 mU/ml) and xanthine (500 microM), or urate (100 microM) prevented the SIN-1-induced dihydrorhodamine oxidation and injury to SP-A. .NO alone, generated by S-nitroso-N-acetylpenicillamine plus 100 microM L-cysteine, or superoxide and hydrogen peroxide, generated by pterin and xanthine oxidase in the absence of iron, did not damage SP-A or oxidize dihydrorhodamine. We concluded that peroxynitrite, but not .NO or superoxide and hydrogen peroxide, in concentrations likely to be encountered in vivo, caused nitrotyrosine formation and decreased the ability of SP-A to aggregate lipids.
171 citations
Journal Article•
TL;DR: Caffeic acid was found to be the active principle exhibiting strong inhibition of xanthine oxidase in this study, and esculetin displayed the strongest activity, and induced competitive inhibition of the enzyme with respect to the substrate x anthine.
Abstract: The stems of Bougainvillea spectabillis Wild (Nyctaginaceae) have been used in folk medicine against hepatitis. Spinasterol, 22, 23-dihydrospinasterol and caffeic acid were isolated from the plant stems and characterized. Caffeic acid has not been previously isolated from this plant but spinasterol has been isolated from the leaves. Caffeic acid was found to be the active principle exhibiting strong inhibition of xanthine oxidase in this study (IC50 = 39.21 microM). In order to study the structure-activity relationship of the phenolics as regards xanthine oxidase inhibition, twelve naturally occurring phenolics (esculetin, scopoletin, scoparone, barbaloin, berberine chloride, sinomenine, osthole, paeonol, honokiol, magnolol, methyleugenol and 6-gingerol) were tested for their inhibitory effects on xanthine oxidase. The results showed that esculetin displayed the strongest activity (IC50 = 28.4 microM), and induced competitive inhibition of the enzyme with respect to the substrate xanthine. The apparent inhibition constant (Ki) of esculetin was 2.369 x 10(-6) M. Since xanthine oxidase serum levels are increased in hepatic and brain tumors, caffeic acid and esculetin should be tested as anti-hepatitis or/and anticancer agents.
160 citations
TL;DR: It is concluded that myocardial lipid peroxidation, estimated as MDA formation, is common during intermittent ischemia-reperfusion sequences in coronary bypass surgery, although some patients may be better protected.
Abstract: BACKGROUNDFree radicals and lipid peroxidation have been suggested to play an important role in the pathophysiology of myocardial reperfusion injury. The purpose of the present study was to monitor myocardial malondialdehyde (MDA) production as an index of lipid peroxidation during ischemia-reperfusion sequences in patients undergoing elective coronary bypass grafting. There has been a lot of debate on the role of xanthine oxidase as a potential superoxide anion generator and thus lipid peroxidation in human myocardium. To evaluate the activity of xanthine oxidase pathway, we measured the changes in the transcardiac concentration differences in adenosine, inosine, hypoxanthine, xanthine, and uric acid.METHODS AND RESULTSThe coronary sinus-aortic root differences (CS-Ao) of MDA, oxypurines, and nucleosides were measured by a recently developed ion-pairing high-performance liquid chromatographic (HPLC) method. Fifteen patients were included in the study, and 13 of them demonstrated a more than 10-fold incre...
122 citations
TL;DR: Caffeic acid was the most potent constituent and was an uncompetitive inhibitor of the enzyme with respect to the substrate xanthine in the isolate of Alsophila spinulosa Hook Tryon.
Abstract: Diploptene(1), β-sitosterol(2), a mixture of 6′-O-(E-P-coumaroyl)-α-glucopyranose and 6′-O-(E-P-coumaroyl)-β-glucopyranose(3), a mixture of 6′-O-(E-P-caffeoyl)-α-glucopyranose and 6′-O-(E-P-caffeoyl)-β-gIucopyranose(4), caffeic acid(5) and astragalin(6) were isolated from an ethanolic extract of the leaves of Alsophila spinulosa Hook Tryon (Cyatheaceae). The plant has been used in folk medicine for hepatitis, gout, rheumatism and tumor and these compounds were tested for their inhibitory effect on xanthine oxidase. Caffeic acid was the most potent constituent (IC50 = 39.21 μM; Ki = 28.2 μM) and was an uncompetitive inhibitor of the enzyme with respect to the substrate xanthine.
101 citations
TL;DR: The inhibitory action of caffeine on InsP3 receptors suggests that the action of caffeine on the intracellular Ca pool must be interpreted with caution when both ryanodine receptors and InsP 3 receptors are present in the cell.
Abstract: Effects of the xanthine drug caffeine on inositol (1,4,5)-trisphosphate (InsP3)-gated calcium (Ca) channels from canine cerebellum were studied using single channels incorporated into planar lipid bilayers. Caffeine, used widely as an agonist of ryanodine receptors, inhibited the activity of InsP3-gated Ca channels in a noncooperative fashion with half-inhibition at 1.64 mM caffeine. The frequency of channel openings was decreased more than threefold after addition of 5 mM caffeine; there was only a small effect on mean open time of the channels, and the single channel conductance was unchanged. Increased InsP3 concentration overcame the inhibitory action of caffeine, but caffeine did not reduce specific [3H]InsP3 binding to the receptor. The inhibitory action of caffeine on InsP3 receptors suggests that the action of caffeine on the intracellular Ca pool must be interpreted with caution when both ryanodine receptors and InsP3 receptors are present in the cell.
90 citations
TL;DR: The present review summarizes information that has become available about Xanthine oxidoreductase and Interpretations of contradictory findings are presented in order to reduce confusion that still exists with respect to the role of this enzyme in physiology and pathology.
Abstract: Xanthine oxidoreductase is an enzyme which has the unusual property that it can exist in a dehydrogenase form which uses NAD+ and an oxidase form which uses oxygen as electron acceptor. Both forms have a high affinity for hypoxanthine and xanthine as substrates. In addition, conversion of one form to the other may occur under different conditions. The exact function of the enzyme is still unknown but it seems to play a role in purine catabolism, detoxification of xenobiotics and antioxidant capacity by producing urate. The oxidase form produces reactive oxygen species and, therefore, the enzyme is thought to be involved in various pathological processes such as tissue injury due to ischaemia followed by reperfusion, but its role is still a matter of debate. The present review summarizes information that has become available about the enzyme. Interpretations of contradictory findings are presented in order to reduce confusion that still exists with respect to the role of this enzyme in physiology and pathology.
80 citations
TL;DR: No reversibly and dose-dependently suppressed XOD activity in both determination methods and the results indicate that NO suppresses Xod activity through reversible alteration of the flavin prosthetic site.
Abstract: The effects of nitric oxide (NO) on xanthine oxidase (XOD) activity and the site(s) of the redox center(s) affected were investigated. XOD activity was determined by superoxide (O2-) generation and uric acid formation. NO reversibly and dose-dependently suppressed XOD activity in both determination methods. The suppression interval also disclosed a dose-dependent prolongation. The suppression occurred irrespective of the presence or absence of xanthine; indicating that the reaction product of NO and O2-, peroxynitrite, is not responsible for the suppression. Application of synthesized peroxynitrite did not affect XOD activity up to 2 microM. Methylene blue, which is an electron acceptor from Fe/S center, prevented the NO-induced inactivation. The results indicate that NO suppresses XOD activity through reversible alteration of the flavin prosthetic site.
79 citations
TL;DR: Carvedilol protects endothelial cells against oxygen radical mediated cell injury and death by scavenging free radicals, and might potentially contribute to the clinical beneficial effects of carvedilol as an antihypertensive agent.
Abstract: Objective: Oxygen radical mediated endothelial injury plays an important role in cardiovascular disease. Carvedilol, a new β blocker and antihypertensive agent, has been shown to have antioxidant activity. The aim of this study was to determine whether carvedilol protects oxygen radical induced endothelial injury. Methods: Cultured bovine pulmonary artery (BPAEC) and human umbilical vein endothelial cells (HUVEC) were used and oxygen radicals were generated by xanthine-xanthine oxidase or phorbol myristate acetate (PMA) activated human neutrophils. Cell injury was assessed by lactate dehydrogenase (LDH) release and cell death, or 51Cr release from prelabelled BPAEC. The electron paramagnetic resonance (EPR) spin trapping technique was used to detect the amount of radical spin adducts formed in cell lipids. Results: Carvedilol dose dependently inhibited xanthine-xanthine oxidase induced LDH release from BPAEC and HUVEC, with IC50 values of 3.8 μM and 2.6 μM, respectively, and significantly reduced cell death by xanthine-xanthine oxidase. Other β blockers tested (propranolol, labetalol, pindolol, and celiprolol) showed a mild effect or no effect at all. Increasing the time of pretreatment with carvedilol enhanced its cell protective effect against oxidative stress. Carvedilol also protected BPAEC dose dependently from PMA activated, neutrophil induced cell injury. Carvedilol had no effect on xanthine oxidase activity. EPR study confirmed that xanthine-xanthine oxidase induced the formation of lipid derived radicals in cell lipids and carvedilol scavenged free radicals, as indicated by the decreased EPR signal. Conclusions: Carvedilol protects endothelial cells against oxygen radical mediated cell injury and death by scavenging free radicals. The prevention of oxidative injury to endothelial cells might potentially contribute to the clinical beneficial effects of carvedilol as an antihypertensive agent.
Cardiovascular Research 1994; 28 :400-406
65 citations
TL;DR: Reduction of free radical formation decreases platelet aggregation and may eliminate cyclic flow variations, whereas promotion of freeradical generation enhances platelet aggregating and may induce cyclicflow variations.
TL;DR: It is suggested that ROS may at low fluxes stimulate PG-synthesis under physiological conditions and alter cartilage metabolism and structure in conditions where they are overproduced, such as in rheumatoid arthritis, and in hemochromatosis and other iron storage diseases.
TL;DR: The use of 6-thioxanthine to detect resistant mutants in the presence of many wild-type parasites provides a simple selection and back-selection for mutations that affect the hypoxanthine-guanine phosphoribosyltransferase gene of T. gondii.
TL;DR: The acute inflammatory process is initiated when an extravascular stimulus provokes capillary dilatation and increased permeability, and recruits circulating neutrophils to the site, and the highly reactive (and highly toxic) hydroxyl radical is formed.
Abstract: The acute inflammatory process is initiated when an extravascular stimulus provokes capillary dilatation and increased permeability, and recruits circulating neutrophils to the site. Damage to vascular structures is seen at these sites and evidence of injury occurs early in the evolution of the lesion. Studies carried out over the past 10 years in a number of laboratories have elucidated many of the biochemical events that lead to endothelial cell damage at sites of inflammation. Activated neutrophils bind tightly to the target cells and this is accompanied by neutrophil generation of superoxide anion and hydrogen peroxide and by release of granule enzymes. Neutrophil-derived hydrogen peroxide gains access to the interior of the target cell where it induces a breakdown of cellular ATP and a build-up of ATP metabolites. Among these are xanthine and hypoxanthine, substrates for xanthine oxidase. Exposure of the target cell to other neutrophil products (specifically, elastase) induces the interconversion of xanthine dehydrogenase to xanthine oxidase. Formation of uric acid from hypoxanthine and xanthine by the oxidase form of the enzyme results concomitantly in the generation of superoxide anion. In addition to providing a source of intracellular reducing equivalents, the target (endothelial) cell is also the source of iron. Target cell iron, maintained in the reduced form by intracellular oxidants, combines with neutrophil-derived hydrogen peroxide to form the highly reactive (and highly toxic) hydroxyl radical. This oxidant is most likely the direct mediator of injury.
TL;DR: Propyl gallate protected hepatocytes against the cascade of oxyradicals produced by xanthine oxidase-hypoxanthine, and approximately halved the amount of lipid conjugated dienes formed in tissues after oxidant damage.
TL;DR: An amperometric xanthine minibiosensor based on a supported biotinylated phospholipid membrane was designed and streptavidin-modified Xanthine oxidase was coupled to provide electrochemical detection of enzymatically generated hydrogen peroxide.
TL;DR: This study demonstrates that the microdialysis technique is suited to monitor metabolically important substances in the renal interstitium.
Abstract: It has been proposed that adenosine, derived from ATP and released into the renal interstitium, mediates a reduction in renal function in ischemic acute renal failure. Because no direct measurements of interstitial adenosine are available, we evaluated an in vivo microdialysis technique to assess the levels of adenosine and its metabolites in the cortex of the normal rat kidney (n = 6). Microdialysis probe implantation did not alter cortical renal blood flow, glomerular filtration rate, or fractional sodium excretion. The interstitial concentration of adenosine was 199 +/- 53 nM, and relative concentrations of inosine, hypoxanthine, xanthine, and uric acid were 99 +/- 47, 182 +/- 29, and 183 +/- 70 nM and 1.8 +/- 0.4 microM, respectively. Infusion of ATP-MgCl2 (n = 5) resulted in a significant increase in the dialysate levels of adenosine (67 +/- 11 to 378 +/- 97 nM), inosine (230 +/- 102 to 803 +/- 219 nM), and uric acid (3.5 +/- 1.3 to 6.9 +/- 1.7 microM). In conclusion, this study demonstrates that the microdialysis technique is suited to monitor metabolically important substances in the renal interstitium.
TL;DR: Superoxide and peroxide uncoupled the hydrolytic activity of the Na+ pump from Rb+ uptake, which would result in dual damage: ion imbalance and continuous hydrolysis of ATP in the cells that are already starved.
Abstract: Free radicals may impair vital functions of several types of tissues including coronary artery smooth muscle. Because the Na+ pump plays a key role in maintaining coronary tone, the effects of superoxide and peroxide on this protein were examined. Ouabain-sensitive Rb+ uptake by denuded coronary artery rings was used in lieu of K+ transport by this pump. It was inhibited by exposing the rings for 90 min either to peroxide [50% inhibitory concentration (IC50) = 0.56 +/- 0.18 mM] or to superoxide generated by xanthine oxidase (XO; 0.3 mM xanthine and xanthine oxidase, IC50 = 0.08 +/- 02 mU/ml). The effect of peroxide was not overcome by superoxide dismutase and that of superoxide was not prevented by catalase. K(+)-activated ouabain-sensitive hydrolysis of p-nitrophenyl phosphate in the plasma membrane-enriched fraction isolated from the coronary artery smooth muscle was monitored as the hydrolytic activity of the Na+ pump. It was inhibited by exposing the membranes only to very high concentrations of peroxide (IC50 = 9.85 +/- 3.5 mM) or XO (IC50 = 5 +/- 2 mU/ml). The exposure to 2.5 mM H2O2 or 0.5 mU/ml XO reduced the Na(+)-dependent acylphosphate levels only by 41 +/- 3 and 30 +/- 4%, respectively even though either inhibited the Rb+ uptake by > 80%. Thus superoxide and peroxide uncoupled the hydrolytic activity of the Na+ pump from Rb+ uptake. We speculate that such an uncoupling in ischemia and reperfusion would result in dual damage: ion imbalance and continuous hydrolysis of ATP in the cells that are already starved.
TL;DR: The release of xanthine oxidoreductase is reported in an animal model that simulates the situation of human thoracic aorta operations and would likely be toxic to vascular endothelium, potentially contributing to multiple organ dysfunction.
DOI•
01 Jan 1994
TL;DR: As yet, a coherent explanation of the acute and chronic effects of caffeine in terms of blockade of adenosine receptors has not emerged, and interactions between pathways subserved by A1 - and A2a-adcnosine receptors complicate attempts to interpret caffeine pharmacology.
Abstract: The behavioral effects of caffeine appear likely to be due in large measure to antagonism of the action of endogenous adenosine at A1- and A2a-receptors in the central nervous system. Other biochemical mechanisms of action of caffeine, such as release of intracellular calcium, inhibition of phosphodiesterases and blockade of regulatory sites of GABAA-reccptors, would require much higher concentrations than the micromolar concentrations of caffeine associated with behavioral stimulation. However, micromolar concentrations of caffeine also would be expected to cause only a modest blockade of adenosine receptors. Selective adenosine agonists and xanthine antagonists have provided some insights into central roles for adenosine receptor subtypes. Thus, behavioral stimulation by xanthines appears to require blockade of both A1- and A2a-receptors. Chronic blockade of adenosine receptors by caffeine would be expected to result in alterations in the central receptors and pathways that are regulated by adenosine through A1- and A2a-receptors. Indeed, chronic caffeine docs alter the density not only of adenosine receptors, but also of adrenergic, cholinergic, GABAergic and serotonergic receptors. Behavioral responses to agents acting through dopaminergic and cholinergic pathways arc altered. As yet, a coherent explanation of the acute and chronic effects of caffeine in terms of blockade of adenosine receptors has not emerged. Interactions between pathways subserved by A1 - and A2a-adcnosine receptors complicate attempts to interpret caffeine pharmacology, as does the complex control by adenosine receptors of dopamincrgic, cholinergic and other central pathways.
TL;DR: It is demonstrated that spin trapping using DMPO was at least 20-fold more sensitive than the reduction of cytochrome c for the measurement of superoxide anions, and EPR spin trapping may underestimate radical production, probably due to degradation of D MPO radical adducts.
TL;DR: Ca2+ derived from extracellular sources promoted superoxide radical production and renal cell injury by a calmodulin-dependent conversion of xanthine dehydrogenase to xanthin oxidase, a major source of oxygen free radicals during H/R.
Abstract: Hypoxia and reoxygenation (H/R) generate oxygen free radicals that result in renal cell injury. We tested the roles of calcium and calmodulin in mediating xanthine oxidase-derived oxygen free radical production during H/R. Lowering extracellular Ca2+ attenuated lethal cell injury. H/R increased superoxide radical production over basal levels, whereas removing extracellular Ca2+ before hypoxia decreased superoxide radical production to basal levels. Pretreatment with either 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate hydrochloride or thapsigargin, to inhibit release or deplete stores of intracellular Ca2+, did not affect injury following H/R. Ionomycin increased lactate dehydrogenase release during H/R but did not increase superoxide radical to levels greater than that observed for H/R alone. The calmodulin inhibitors trifluoperazine, calmidazolium, or N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide decreased cell injury to varying degrees. Trifluoperazine also decreased superoxide radical production during H/R and was shown to inhibit the conversion of xanthine dehydrogenase to xanthine oxidase. Cell injury and superoxide radical production correlated with cytosolic free Ca2+ during H/R as determined with the Ca(2+)-sensitive fluoroprobe indo 1. Cytosolic free Ca2+ increased slightly during hypoxia and showed a dramatic increase as soon as cells were reoxygenated. Cells incubated in a Ca(2+)-free medium actually showed a small decrease in intracellular Ca2+ despite H/R. In summary, Ca2+ derived from extracellular sources promoted superoxide radical production and renal cell injury by a calmodulin-dependent conversion of xanthine dehydrogenase to xanthine oxidase, a major source of oxygen free radicals during H/R.
TL;DR: Membrane-bound serine- and metalloproteinases were responsible for the degradation of band 3 protein, because the degradation was remarkably inhibited by diisopropyl fluorophosphate and phenylmethylsulfonyl fluoride, and partially by ethylenediaminetetraacetic acid.
TL;DR: It is demonstrated that in the absence of free metal ions, active oxygen species, generated by activated macrophages or xanthine/xanthine oxidase (XOD), carry out oxidative degradation of collagen fibrils type I in conjunction with proteases.
TL;DR: Fibre-optic biosensors were constructed and the H2O2 generated by the reaction of hypoxanthine and xanthine oxidase was measured by chemiluminescence (CL) detection using luminol and peroxidase.
TL;DR: Adenosine is generated continuously in cultured HUVEC under normal conditions, but is immediately recycled via adenosine kinase after inhibition of ATP synthesis, and in ATP depleted cells, nucleotide catabolism proceeds predominantly via intracellular dephosphorylation of AMP.
Abstract: Adenosine is a key metabolite in myocardial autoregulation and interactions between different types of cells in its production, metabolism and action are vital. In this study, adenosine and other purine catabolite production were analyzed in cultured umbilical vein endothelial cells under control conditions (normal O2 and substrate supply) and with inhibitors of ATP formation (iodoacetate and oligomycin -1 + O). Innhhibitors of adenosine deaminase - erythro-9 (2-hydroxy-3-nonyl) adenine (EHNA) and adenosine kinase 5´ iodotubercidin (ITu) were included where indicated. Nucleotides and catabolites were evaluated by HPLC in the cells and in the medium throughout 45 min of incubation at 370C. Total amounts or increments per culture flask (n = 4-5, ± SEM) were as follows: In controls with EHNA, cellular ATP, ADP and AMP contents were 9.0 ± 0.1, 0.83 ± 0.14 and 0.15 ± 0.03 nmol, while in the medium increases in hypoxanthine, xanthine and uric acid content were 1.0 ± 0.2, 0.4 ± 0.1 nmol and 0.5 ± 0.2 nmol. Adenosine increased by -/006 ± 0.04 nmol. With additional presence of ITu, adenosine rose by 0.9 ± 0.2 nmol (p<0.005). After 45 min incubation with I+O and EHNA, ATP was totally depleted, cellular AMP rose to 3.0 ± 1.2 nmol and no IMP increase was found. Medium adenosine rose by 5.6 ± 0.2 nmol and the increase in sum of other purine catabolites was 2.8 ± 03 nmol. In conclusion, adenosine formed in the endothelial cells under normal O2 and substrate supply is recycled via adenosine kinase. In ATP depleted cells, nucleotide catabolism proceeds predominantly via adenosine.
TL;DR: The in vitro pharmacological profile of putative A2 adenosine antagonists provided a basis to reduce, by further chemical modifications, the affinity at A1 receptor and therefore enhance A2 receptor selectivity.
Abstract: 1. We have characterized the in vitro pharmacological profile of putative A2 adenosine antagonists, two non-xanthine compounds, 5-amino-8-(4-fluorobenzyl)-2-(2-furyl)-pyrazolo [4,3-e]-1,2,4-triazolo[1,5-c] pyrimidine (8FB-PTP) and 5-amino-9-chloro-2-(2-furyl 1,2,4-triazolo [1,5-c] quinazoline (CGS 15943), and the xanthine derivative (E)7-methyl-8-(3,4-dimethoxystyryl)-1,3-dipropyl- xanthine (KF 17837). 2. In binding studies on bovine brain, 8FB-PTP was the most potent (Ki = 0.074 nM) and selective (28 fold) drug on A2 receptors, whereas CGS 15943 and KF 17837 exhibited affinity in the low and high nanomolar range, respectively, and showed little selectivity. 3. In functional studies, 8FB-PTP antagonized 5'-N-ethyl-carboxamidoadenosine (NECA)-induced vasorelaxation of bovine coronary artery (pA2 = 7.98) and NECA-induced inhibition of rabbit platelet aggregation (pA2 = 8.20). CGS 15943 showed weak activity in the platelet aggregation model (pA2 = 7.43) and failed to antagonize NECA-induced vasodilatation. KF 17837 was ineffective in both models up to micromolar concentrations. 4. Antagonism of A1-mediated responses was tested versus 2-chloro-N6-cyclopentyladenosine (CCPA) in rat atria. 8FB-PTP and CGS 15943 also antagonized competitively the negative chronotropic response induced by CCPA. Conversely, KF 17837 was unable to reverse A1-mediated responses. 5. 8FB-PTP is a potent and competitive antagonist of responses mediated by A2 adenosine receptors. The data provided a basis to reduce, by further chemical modifications, the affinity at A1 receptor and therefore enhance A2 receptor selectivity.
TL;DR: Additional studies are necessary to define precisely the metabolic derangement in idiopathic uric acid overproduction and to assess fully the consequences of increased purine nucleotide degradation, such as free-radical formation, increased adenosine synthesis, and reduced synthesis of signal transducers.
Abstract: Purine nucleotides are synthesized and degraded through a regulated series of reactions which end in the formation of uric acid. Increased uric acid synthesis may be the result of two major pathophysiological disorders: increasedde novo purine synthesis and enhanced purine nucleotide degradation, both of which may be the result of an increased or decreased enzyme activity. In addition, some conditions and disorders associated with uric acid overproduction have been recognized as the result of increased ATP degradation or decreased synthesis of ATP. The clinical manifestations of the diseases leading to excess uric acid synthesis are heterogenous, but symptoms related to uric acid overproduction are always secondary to the precipitation of crystals in soft tissues, joints, and the kidney excretory system. In clinical practice, serum urate concentration and urinary uric acid excretion arc used to assess uric acid synthesis, taking into account that a purine-rich diet can be a confounding variable. Quantification of uric acid precursors, such as adenosine, inosine, guanosine, hypoxanthine, and xanthine, in biological fluids and intracellular nucleotides has provided further insight into the metabolic disturbances underlying disorders associated with uric acid overproduction. Additional studies are necessary to define precisely the metabolic derangement in idiopathic uric acid overproduction and to assess fully the consequences of increased purine nucleotide degradation, such as free-radical formation, increased adenosine synthesis, and reduced synthesis of signal transducers.
TL;DR: In this article, it was shown that β-carotene (BC) does not undergo free radical formation during the first hour of stirring and the use of aldehydes as substrate.
Abstract: The enzymic cooxidation of β-carotene (BC) by xanthine oxidase (XO) in aqueous solutions leads to β-ionone (BI) and derivatives: epoxy-β-ionone (EPBI), dihydroactinidiolide, β-cyclocitral, pseudoionone, etc. We demonstrate, in this article, that this is due to free radical (R . ) formation during the first hour of stirring and the use of aldehydes as substrate. The bleaching of BC does not occur when the common substrate of XO, xanthine, is used; this proves that the superoxide anion O 2 .- alone is not active on BC. BI formation in this case is not observed
TL;DR: Adenine nucleotides of tissue or effluent and nicotinamide adenine dinucleotide, oxidized form (NAD+) of the tissue before and after graft reperfusion and of the effluent were analyzed and ATP, ADP, total AN, total An+total PC (T) and NAD+, in post-transplant tissues were significantly associated with primary graft nonfunction by logistic regression analysis.
Abstract: Sixty-eight primary liver grafts were analyzed to see whether adenine nucleotides (AN: ATP, ADP, and AMP) or purine catabolites (PC: adenosine, inosine, hypoxanthine, and xanthine) of tissue or effluent can predict primary graft nonfunction. AN, PC, and nicotinamide adenine dinucleotide, oxidized form (NAD+) of the tissue before (pretransplant) and after graft reperfusion (post-transplant) and of the effluent were analyzed. The graft outcome was classified into two groups (group A: successful, n = 64; group B: primary nonfunctioning, n = 4). No significant differences were observed in pretransplant measurements between groups A and B, whereas ATP, ADP, total AN, total AN + total PC (T) and NAD+, in post-transplant tissues, were significantly higher in group A. Xanthine in the effluent was significantly higher in group B than in group A. ATP, ADP, total AN, T, and NAD+ in post-transplant tissue were significantly associated with primary graft nonfunction by logistic regression analysis.