The proportion of xanthine oxidase activity of total xanthine oxidoreductase activityin situ remains constant in rat liver under various (patho)physiological conditions

Abstract

Xanthine oxidoreductase is a rate-limiting enzyme of the Activity of xanthine oxidoreductase (total xanthine catabolism of purines (reviewed by Kooij 1 ). The enzyme exists dehydrogenase plus xanthine oxidase) and xanthine oxiin two functionally distinct forms: an oxidized nicotinamide dase was determined cytophotometrically in periportal adenine dinucleotide-dependent dehydrogenase form (Dand pericentral areas of livers of rats under various form; E.C. 1.1.1.204) that produces reduced nicotinamide ad-(patho)physiological conditions that are known to affect enine dinucleotide and urate and an oxygen-dependent oxithe content of reduced glutathione. For this purpose, dase form (O-form; E.C. 1.2.3.2) that produces superoxide rats were either normally fed or fasted for 24 hours, anion radicals and/or hydrogen peroxide and urate. Converfasted for 24 hours, and treated with diethylmaleate that sion of xanthine dehydrogenase into xanthine oxidase may depleted glutathione or treated by in vivo ischemia for occur reversibly via oxidation of sulfhydryl groups and irre-2 hours in the livers. Xanthine oxidoreductase activity versibly by proteolysis. 2-7 These findings were mainly based was shown histochemically with the use of a tetrazolium on biochemical studies performed on homogenates. It was salt procedure, and xanthine oxidase activity was localassumed for a long time that different pathological conditions ized with a cerium-diaminobenzidine-cobalt-hydrogen such as ischemia lead to in vivo conversion of the D-form into peroxide technique in unfixed cryostat sections of the the O-form. 8-13 However, Kooij et al. 14 showed recently that livers. Cytophotometric measurements showed that toconversion of the D-form into the O-form did not occur to any tal xanthine oxidoreductase activity was decreased after significant extent in vivo in livers from fed or fasted rats fasting and ischemia, whereas only ischemia caused reeither or not after in vivo ischemia. However, conversion ocduced xanthine oxidase activity. Moreover, the percentcurred rapidly in vitro in the supernatants of homogenates age of xanthine oxidase of total xanthine oxidoreductase of these livers, and then the conversion rate was increased activity was constant in both periportal and pericentral by fasting and in vivo ischemia. When homogenization proceareas at the level of approximately 4% in normally fed dures were performed under strictly controlled conditions, to and 24-hour fasted and diethylmaleate-treated rats.avoid in vitro conversion of the D-form into the O-form, a Ischemia reduced this percentage in both areas of the constant percentage of 15%-20% oxidase activity of total xanliver to 2%. It was concluded that the amount of endogethine oxidoreductase activity in livers of rats was found under nous reduced glutathione did not affect the percentage all conditions studied. To test the hypothesis that conversion of xanthine oxidase. The low percentage of xanthine oxiof the D-form into the O-form does not occur in vivo to any dase as determined in the pre...

Reperfusion injury and reactive oxygen species: The evolution of a concept

“…The issue of XDH to XO conversion during ischemia has been more extensively evaluated in liver. However, disparate findings have been reported for this tissue, with some reports describing significant conversion during ischemia, while others describe little or no conversion following prolonged ischemia [135] , [136] . There appears to be a growing consensus that the conversion of XDH to XO is not a rate-limiting determinant of ROS production upon reperfusion of ischemic tissue, particularly in liver [123] , [137] .…”

How Biology Handles Nitrite

“…Second, in the course of ischaemia, the mitochondrial electron transfer chain would be disrupted and the ATP synthesis hindered; the subsequent ATP catabolism leads to an accumulation of hypoxanthine and NADH in tissues. − This increase in the concentration of two reducing substrates can “fuel” the enzymes with reducing equivalents to reduce nitrite. Third, as the ATP concentration decreases, the transmembranar ion gradients are dissipated, causing elevated cytoplasmatic calcium concentrations, which, in turn, activate calcium-dependent proteases that would convert the XD into the XO form. ,− In summary, during ischaemia: (i) the pH values are lower enough to provide the acidic conditions required for the nitrite reaction; (ii) reducing substrates are available to supply the necessary electrons; (iii) the formerly prevailing XD form (that reacts with NAD + ) can be converted into the “dioxygen-user” XO, by proteolysis; (iv) the concentration of the competitive dioxygen is very low; and (v) NAD + (regardless of its high concentration) would be no longer a competitive substrate of the nitrite reduction, because XO and AO do not react with it. Therefore, all of the conditions seem to be gathered for nitrite to be reduced by XO and AO during in vivo ischaemia.

12 Mechanism proposed for XO-dependent nitrite reduction to NO under ischaemia.

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“…Another point against the feasibility of these pathways in vivo is related to the conversion of the in vivo-predominant XD into XO. The extent and rate of this conversion are a matter of great controversy: from no conversion at all (with XO being considered as an experimental artifact), to a small (20%) and slow conversion ,,− ,− ,, and a conversion that is enhanced by hypoxic conditions and in vivo ischaemia. , The issue here is the competition between nitrite and NAD + to react with reduced XD. The NAD + concentration (∼0.5–1 mM ,,− ), 2–3 orders of magnitude higher than that of NADH, is not significantly decreased by the NADH accumulation during ischaemia. − As a result, if the conversion of XD into XO is not efficient (or does not occur at all), the NAD + reaction (with a k cat / K m 2–3 orders of magnitude higher) would prevail over the nitrite reduction, and the NO formation by this protein would be seriously compromised.…”

CHAPTER 1. Molybdenum and Tungsten-Containing Enzymes: An Overview

“…rate of conversion of Xd into Xo in vivo have been a matter of great controversy. 237,240,241,[258][259][260][261][262][263][264][265][266] physiologically, mammalian Xd (Xo) is a key enzyme in purine catabolism, where it catalyzes the hydroxylation of both hypoxanthine and xanthine to the terminal metabolite, urate (eqn (1.6)). [154][155][156][157][158][159] it has also been suggested to be involved in the xenobiotic compounds metabolism and in ros-mediated signalling pathways and ros-mediated diseases.…”