A new paradigm for XOR-catalyzed reactive species generation in the endothelium

Abstract

A plethora of vascular pathology is associated with inflammation, hypoxia and elevated rates of reactive species generation. A critical source of these reactive species is the purine catabolizing enzyme xanthine oxidoreductase (XOR) as numerous reports over the past 30 years have demonstrated XOR inhibition to be salutary. Despite this long standing association between increased vascular XOR activity and negative clinical outcomes, recent reports reveal a new paradigm whereby the enzymatic activity of XOR mediates beneficial outcomes by catalyzing the one electron reduction of nitrite (NO2(-)) to nitric oxide (NO) when NO2(-) and/or nitrate (NO3(-)) levels are enhanced either via dietary or pharmacologic means. These observations seemingly countervail numerous reports of improved outcomes in similar models upon XOR inhibition in the absence of NO2(-) treatment affirming the need for a more clear understanding of the mechanisms underpinning the product identity of XOR. To establish the micro-environmental conditions requisite for in vivo XOR-catalyzed oxidant and NO production, this review assesses the impact of pH, O2 tension, enzyme-endothelial interactions, substrate concentrations and catalytic differences between xanthine oxidase (XO) and xanthine dehydrogenase (XDH). As such, it reveals critical information necessary to distinguish if pursuit of NO2(-) supplementation will afford greater benefit than inhibition strategies and thus enhance the efficacy of current approaches to treat vascular pathology.

Keywords: Hypoxia; Inflammation; Nitric oxide; Nitrite; Xanthine oxidoreductase.

Figure 1. Xanthine Oxidoreductase Reactions

For XDH, xanthine is oxidized to uric acid at the Mo-co and electrons transferred via 2 Fe/S centers to the FAD where NAD⁺ is reduced to NADH (left). For XO, xanthine is oxidized to uric acid at the Mo-co and electrons are transferred to the FAD where O₂ is reduced to O₂^•− and H₂O₂ (right). Conversion from XDH to XO is mediated by post-translational modification.

Figure 2. Nitrite Reductase Activity of XOR

(A) Under low O₂ tensions, NO₂⁻ undergoes a 1 electron reduction to ^•NO at the Mo-cofactor (electrons are donated directly to Mo by xanthine). The thickness of the arrows represents the degree of electron flux while the size of the font represents level of substrate or product formation. (B) Again, under low O₂ tensions, NO₂⁻ is reduced to ^•NO at the Mo-cofactor while electrons are supplied by NADH and transferred retrograde to reduce the Mo-co.

Figure 3. XOR-Catalyzed NO Production in the Vasculature

(top panel) Under oxygenated conditions, inflammation results in enhanced XDH expression and export to the circulation where XDH is rapidly converted to XO with subsequent sequestration by endothelial GAGs. With abundant O₂ and xanthine, XO-dependent ROS formation is dominant while eNOS is the primary source of ^•NO. XO-derived O₂^•− effectively reduces ^•NO bioavailability by its diffusion limited reaction with eNOS-derived ^•NO to form peroxynitrite (O=NOO⁻). Under these conditions it is possible that, in the presence of elevated NO₂⁻, intracellular XDH could serve as a NO₂⁻ reductase. (bottom panel) Under hypoxic conditions (O₂ tensions approaching and falling below the K_m for O₂ at the FAD (~27 μM or ~1.5% O₂), XO as well as XDH can assume NO₂⁻ reductase activity while eNOS-mediated ^•NO formation is diminished due to the absence of the substrate, O₂ (noted by a diminished arrow thickness). Under these conditions, eNOS may become uncoupled resulting in O₂^•− generation. It is at this point that XO and/or XDH may become a significant source of ^•NO.