XO Inhibition Research Articles

Long-Term Inhibition of Xanthine Oxidase by Febuxostat Does Not Decrease Blood Pressure in Deoxycorticosterone Acetate (DOCA)-Salt Hypertensive Rats

Xanthine oxidase and its products, uric acid and ROS, have been implicated in the pathogenesis of cardiovascular disease, such as hypertension. We have previously reported that allopurinol inhibition of XO does not alter the progression of deoxycorticosterone acetate (DOCA)-salt hypertension in rats. However other researchers have observed a reduction in blood pressure after allopurinol treatment in the same model. To resolve this controversy, in this study we used the newer and more effective XO inhibitor febuxostat, and hypothesized that a more complete XO blockade might impair hypertension development and its end-organ consequences. We used DOCA-salt hypertensive rats and administered vehicle (salt water) or febuxostat (orally, 5 mg/kg/day in salt water) in a short-term “reversal” experiment (2 weeks of treatment 3 weeks after DOCA-salt beginning) and a long-term “prevention” experiment (treatment throughout 4 weeks of DOCA-salt). We confirmed XO inhibition by febuxostat by measuring circulating and tissue levels of XO metabolites. We found an overall increase in hypoxanthine (XO substrate) and decrease in uric acid (XO product) levels following febuxostat treatment. However, despite a trend for reduced blood pressure in the last week of long-term febuxostat treatment, no statistically significant difference in hemodynamic parameters was observed in either study. Additionally, no change was observed in relative heart and kidney weight. Aortic media/lumen ratio was minimally improved by long-term febuxostat treatment. Additionally, febuxostat incubation in vitro did not modify contraction of aorta or vena cava to norepinephrine, angiotensin II or endothelin-1. We conclude that XO inhibition is insufficient to attenuate hypertension in the rat DOCA-salt model, although beneficial vascular effects are possible.

Antioxidant and Xanthine Oxidase Inhibition Activities of Cynomorium songaricum Extracts

In this study, we evaluated the antioxidant activities and xanthine oxidase inhibition effects of water and ethanol extracts of Cynomorium songaricum. The ethanol extract of C. songaricum (EE) contained more phenolic and flavonoid compounds than the water extract (WE). The antioxidant activities of the extracts were increased as the concentration of the extract increased. The WE has better effectiveness than the EE for DPPH free radical scavenging activity and nitrite scavenging ability. The nitrite scavenging abilities of WE were 90.02% (EC50 653.15 ㎍/mL) at conditions of pH 1.2 and 2,000 ㎍/mL, and 84.34% (EC50 817.17 ㎍/mL) at pH 3.0. The EE has more effective SOD-like activity and XO inhibition than WE. The SOD-like activity of EE was 81.47% at a concentration of 2,000 ㎍/mL, EC50 was 951.70 ㎍/mL. The xanthine oxidase inhibition of the EE, with an EC50 of 112.47 ㎍/mL, is greater than that of ascorbic acid, which was 192.50 ㎍/mL (p＜0.05). These results suggest that the C. songaricum is a potentially useful antioxidant source for the development of nutraceuticals and medicines.

Antioxidants, Free Radicals Scavenging and Xanthine Oxidase Inhibitory Potentials of Ajuga iva L. Extracts.

Introduction: Antioxidant effects of Ajuga iva L extracts (AIE), a traditionally used plant in Algerian folk medicine, were investigated. This plant is widely used to treat diabetes, gastric ulcer, dysuria, painful joints of the limbs and other free radicals related disorders. Methods: In order to determine the antioxidant activities of AIE, the shoot extracts were prepared using solvents of varying polarity. Xanthine oxidoreductase (XOR) was purified from bovine milk. Anti-radical activities were determined by enzymatic and non enzymatic methods: The enzymatic methods were realised by either production of uric acid or reduction of cytochrome c. The non enzymatic methods were conducted using in vitro techniques: NBT test, ß-carotene linoleate acid, 1,1- diphenyl-2 picrylhydrazyl (DPPH) radical-scavenging, ferric reducing /antioxidant power (FRAP) and Ferrous ion chelating activity. Results: All extracts showed inhibitory properties on xanthine oxidase, the IC50 (μM / quercetin equivalen) ranges from 3.878 ± 0.717 to 5.835 ± 0.468), with an additional superoxide scavenging capacity. These extracts showed a potent DPPH radical scavenging activity and a powerful scavenging activity of superoxide anion more than that of gallic acid using NBT test. They possess a similar inhibition ratio of the linoleic acid oxidation to that of the BHT, and gave a reduction power better than rutin in Ferric Reducing Ability of Plasma assay (FRAP). However, they were weaker than EDTA in Fe2+ chelating activity. Conclusion: Ajuga iva L. appears to be a valuable plant and could be used to treat conditions where inhibition and free-radicals scavenging of XO is warranted.

Sildenafil protects human mammary epithelial cells against ROS production induced by estradiol.

Several studies suggest that xanthine dehydrogenase (XDH) and its oxidase form (XO) play an important role in various types of ischemic and vascular injuries. Recently, we have demonstrated that estradiol (E2) induces a significant decrease of the expression and activity of XDH and of its conversion to XO in human mammary epithelial cells. E2 is known to induce upregulation of eNOS gene expression in aortic endothelial cells. Because the XO-derived O2·- combines with ·NO to yield ONOO-, and considering that ONOO- converts XDH to XO, the resulting increase of XO activity and reactive oxygen species production would eventually lead to a further increase of ONOO- production, thus creating a vicious cycle of oxidative stress. Our previous study has indicated that sildenafil has a protective effect on human mammary epithelial cells as a consequence of XO inhibition and of the resulting decrease of free oxygen radicals that can impair the expression of NADPH oxidase and type 5 phosphodiesterase (PDE-5). In the present study, we report that the dual inhibitory effect exerted by sildenafil on both XO and PDE-5 is a consequence of a structural modification induced by O2·-, also consisting of the release of a piperazine group that could in turn inhibit the XO enzyme.

Xanthine oxidase inhibitor allopurinol attenuates the development of diabetic cardiomyopathy

In this study we investigated the effect of the xanthine oxidase inhibitor allopurinol on cardiac dysfunction, oxidative‐nitrosative stress, apoptosis, poly(ADP‐ribose) polymerase (PARP) activity, and fibrosis associated with diabetic cardiomyopathy in mice. Diabetes was characterized by increased myocardial, liver and serum XO activity, increased myocardial ROS generation, p22phox, p40phox, p47phox, p91phox mRNA expression, iNOS expression, NT generation, caspase 3/7 and PARP activity/expression, chromatin fragmentation and fibrosis, and declined systolic and diastolic myocardial performance. Allopurinol attenuated the diabetes‐induced increased myocardial, liver and serum XO activities, myocardial ROS, NT generation, iNOS expression, apoptosis, PARP activity and fibrosis, which were accompanied by improved systolic and diastolic performance of the hearts of treated diabetic animals. Thus, XO inhibition with allopurinol improves type I diabetes‐induced cardiac dysfunction by decreasing oxidative/nitrosative stress and fibrosis, which may have important clinical implications for the treatment and prevention of diabetic cardiomyopathy and vascular dysfunction.

Isolation, purification and identification of ellagic acid derivatives, catechins, and procyanidins from the root bark of Anisophyllea dichostyla R. Br.

The root bark of Anisophyllea dichostyla R. Br. is traditionally used in the Democratic Republic Congo for the treatment of several conditions such as anorexia, fatigue and intestinal infections. We have identified and quantitated several polyphenol antioxidants in the methanol extract of the root bark (120 g). The polyphenol content (3.32 g/kg) was predominantly ellagitannins (25%) and polyhydroxyflavan-3-ols (catechins and procyanidins, 75%) with 3′- O-methyl-3,4-methylenedioxo ellagic acid 4′- O-β- d-glucopyranoside and (–)-epicatechin as the major species in each class. These two compounds and the following species were identified unequivocally by NMR spectroscopy: (+)-catechin, (−)-epicatechin 3- O-gallate, 3- O-methyl ellagic acid, 3,3′-di- O-methyl ellagic acid, 3′- O-methyl-3,4-methylenedioxo ellagic acid, 3′- O-methyl-3,4-methylenedioxo ellagic acid 4′- O-β- d-glucopyranoside, and 3′- O-methyl ellagic acid 4- O-β- d-xylopyranoside. The following additional compounds were purified by semi-preparative HPLC and tentatively identified on the basis of UV spectra, HPLC–ESI-MS and nano-ESI-MS–MS: (+)-catechin-3- O-β- d-glucopyranoside, epicatechin-(4β → 8)-catechin (procyanidin B 1), epicatechin-(4β → 8)-epicatechin (procyanidin B 2), an (epi)catechin trimer, 3- O-methyl ellagic acid 4- O-β- d-glucopyranoside, (–)-epicatechin 3- O-vanillate, 3,4-methylenedioxo ellagic acid 4′- O- β- d-glucopyranoside, and 3,3′-di- O-methyl ellagic acid 4- O-β- d-xylopyranoside. Fractionation of the raw extract by column chromatography on silicic acid yielded 10 fractions. In the hypoxanthine/xanthine oxidase antioxidant assay system, CC-9 which contained a range of polyphenols dominated by (−)-epicatechin- O-gallate proved to be the most potent antioxidant fraction (IC 50 = 52 μg/mL) in terms of ROS scavenging. In terms of XO inhibition CC-8, dominated by (epi)catechin trimer and which also contained appreciable amounts of 3′- O-methyl ellagic acid 4′- O-β- d-xylopyranoside, as well as the catechins (+)-catechin-3- O-β- d-glucopyranoside, epicatechin-(4β → 8)-catechin (procyanidin B 1), and (−)-epicatechin 3- O-gallate, proved to be the most potent (IC 50 = 36 μg/mL).

The flavo-enzyme xanthine oxidase is under circadian control in the marine alga Gonyaulax.

The activity of xanthine oxidoreductases (xanthine oxidase, XO, EC 1.2.3.2 and xanthine dehydrogenase, XDH, EC 1.1.1.204) in partially purified extracts of Gonyaulax polyedra was measured over 24 h both in a light:dark cycle and in constant light. This is the first demonstration of xanthine oxidoreductase in a unicellular alga. The activity of the O2-dependent form (XO) was found to be 15 times higher in light than in darkness. The same time-of-day specific differences persisted in constant light, demonstrating a control of XO by the circadian clock. In contrast, the activity of the NAD-dependent form (XDH) is not under circadian control. Because pharmacological inhibition of XO also blocks the effect of blue light on the Gonyaulax circadian clock, the possible relationship between XO and light reception in this unicellular alga will be discussed.

Effect of allopurinol on the formation of reactive oxygen species during intense exercise in the horse

Allopurinol was administered to six horses in a cross-over study to determine the relative contribution of xanthine oxidase (xo) activity to the formation of reactive oxygen species (ROS) in the horse during intense exercise. Exercise increased the mean (SEM) plasma lipid hydroperioxide concentration to a maximum of 492·7 (33·4) μM within one minute of exercise completion and maximum levels of both oxidised glutathione (GSSG) in haemolysates of red blood cells and the glutathione redox ratio (GRR) occurred 20 minutes after exercise (87·2 [12·2] μM and 8·9 [0·9] per cent, respectively). Allopurinol significantly reduced lipid hydroperoxides, GSSG and the GRR at the corresponding maximal times after exercise measured during control exercise (2175 [32·1] μM, 63·8 [8·6] μM and 6·8 [0·7] per cent, respectively). Significantly higher levels of hypoxanthine and xanthine were measured after exercise in the plasma of horses that received allopurinol than in control horses, although uric acid levels remained constant. In control horses, plasma uric acid concentrations increased after exercise to a maximum 20 minutes after exercise of 28·1 (2·6) μM, significantly higher than in horses given allopurinol (9·6 [1 ·3] μM). The results show that the inhibition of xO by allopurinol leads to a decrease in the formation of ROs during exercise, and thus a reduction in oxidative stress.

Effect of nitric oxide and cell redox status on the regulation of endothelial cell xanthine dehydrogenase.

We have previously reported that endothelial cell (EC) xanthine dehydrogenase/xanthine oxidase (XD/XO) activity correlates inversely with the O2 tension to which the cells are exposed. Whether this effect is related to the production of reactive O2 species is unclear. We exposed bovine pulmonary artery EC to various conditions that altered the redox status of the cells: 1) hypoxia (3% O2) and normoxia (20% O2); 2) menadione (MEN), known to generate O2 radicals; 3) catalase (CAT) and reduced glutathione (GSH), which detoxify H2O2; and 4) various NO-generating systems. Changes in intracellular XO and XO + XD activities were correlated with rates of extracellular H2O2 release from the same cells. Conditions that decreased extracellular H2O2 release (hypoxia, CAT, and GSH) produced significant and parallel increases in intracellular XO and XO + XD activities in a time-dependent fashion. MEN treatment increased extracellular release of H2O2 and subsequently reduced intracellular XO and XO + XD activities. NO-generating agents did not change extracellular release of H2O2 but significantly reduced XO and XO + XD activities. The latter effect was prevented by reduced hemoglobin. Scavengers of hydroxyl radicals reversed the inhibition of XO and XO + XD activities produced by MEN but not that produced by NO. While NO significantly inhibited XD/XO activity from rat epididymal fat pad, it did not affect XD/XO mRNA expression in these cells. We conclude that intracellular XD/XO activity is sensitive to changes in oxidant-generating and protective systems. Inhibition of XD/XO activity by NO may be mediated through direct binding of NO to the enzyme iron-sulfur moiety or to its sulfhydryl groups.(ABSTRACT TRUNCATED AT 250 WORDS)