Biochemical and Biophysical Research Communications
Febuxostat suppressed renal ischemia–reperfusion injury via reduced oxidative stress
Highlights
► Xanthine oxidase inhibitor, febuxostat, blocked reperfusion-induced oxidative stress. ► Febuxostat reduced nitrotyrosine, TBARS and urine 8-isoprostane. ► Febuxostat inhibited ER stress, assessed by GRP-78, ATF4, and CHOP. ► Febuxostat suppressed increased serum creatinine under ischemia–reperfusion injury. ► Febuxostat inhibited macrophage infiltration and apoptosis.
Introduction
Renal ischemia–reperfusion (I/R) injury, frequently associated with shock or surgery, is a major cause of acute renal failure [1]. Reactive oxygen species (ROS) have been implicated as a major pathophysiological component in I/R injury in several tissues including kidney [2]. Several lines of insights have focused on xanthine oxidoreductase (XOR) inhibitor as a therapeutic tool for I/R injury. XOR inhibits the generation of uric acid (UA) as the final product of purine catabolism, as well as the resulting generation of superoxide. Under ischemic condition, adenosine triphosphate (ATP) is degraded to xanthine, and hypoxanthine, which are substrates for XOR. XOR functions as either xanthine dehydrogenase (XDH) form, which transfers electron to NAD+, and generates NADH or xanthine oxidase (XO) form, which transfers electron to O2 and generates oxidative stress. Because ischemia-induced cellular calcium overload convert XDH to XO [3], under reperfusion phase, enhanced XO can produce more ROS, such as superoxide, hydrogen peroxide, and hydroxyl radicals. These ROS can exaggerate cellular damages.
Recently, apoptosis is triggered by ROS-mediated activation of endoplasmic reticulum (ER) stress requiring involvement of CHOP pathway [4]. Disturbances such as hypoxia and oxidative stress may lead to ER dysfunction, which can induce ER stress in kidney [5]. Oxidative stress can cause aberrant unfolded and malfolded proteins, which in turn induces ER stress. Some unfolded protein responses enhance the protein-folding capacity by activating the transcription of target genes, such as glucose-regulated protein-78 (GRP-78) [6]. ER stress-induced apoptosis is mainly mediated C/EBP homologous protein-10 (CHOP). CHOP is a transcription factor, which induces several proapoptotic factors, and is downstream of activating transcription factor-4 (ATF4). Severe ER stress preferentially induces proapototic CHOP expression as compared to mild ER stress [5].
On the basis that XO produces ROS, XOR inhibitor might have a protective effect under renal I/R injury. Allopurinol, one of XOR inhibitor, is a classic “suicide inhibitor,” as its binding to and reduction of the molybdenum (Mo) cofactor induces self-oxidation to form oxypurinol, an active inhibitory metabolite. Reduction of the Mo cofactor by allopurinol ultimately leads to electron transfer to the FAD, resulting in the ROS production [7]. In addition, oxypurinol binding and resultant inhibition also requires the Mo cofactor to be reduced [8]. In this point of view, both allopurinol and oxypurinol require enzyme turnover resulting in ROS formation before inhibition is attained. On the contrary, a new XOR inhibitor, febuxostat, is not affected by the above enzyme redox state and interaction with XO, and thereby produces less ROS. In this study, we examined whether treatment with febuxostat could protect the kidney from tubular ROS production under renal ischemia–reperfusion injury and, thereby, inhibit subsequent tubulointerstitial injury.
Section snippets
Experimental design
Male Sprague–Dawley rats weighing 200 g were purchased from Japan SLC Inc. (Shizuoka, Japan) and were maintained under standard conditions until the experiments were done. All studies were performed in accordance with the principles of the Guideline on Animal Experimentation of Osaka University. The rats were randomly allocated into three groups: (1) vehicle-treatment group (Veh group); (2) febuxostat-treatment group (Feb group) and (3) sham-operated group (sham group). Vehicle and febuxostat
Febuxostat suppressed renal XO and XDH activity
Renal tissue XO activity was not changed in vehicle-treated ischemia–reperfusion (I/R)-injured kidneys (1124 ± 187 μU/mg protein) compared with sham-operated kidney (1390 ± 397 μU/mg protein) 4 h after I/R injury. In contrast, XO activity was not detected in febuxostat-treated kidneys. Concomitant with the reduction of XO activity, febuxostat significantly reduced UA levels (0.3 ± 0.0 mg/dL) compared with vehicle treatment (1.4 ± 0.1 mg/dL) 4 h after I/R injury, but no difference was observed at 24 h between
Discussion
We demonstrated that febuxostat suppressed XO activity, reduced oxidative stress, and thereby ameliorated tubulointerstitial injury in a rat model of I/R injury. Untreated I/R-injured kidneys exhibited increased plasma creatinine, tubular apoptosis, interstitial macrophage infiltration and interstitial SMαA expression, while administration of febuxostat ameliorated these manifestations. Importantly, febuxostat reduced oxidative stress, assessed by nitrotyrosine, TBARS and urine 8-isoprostane,
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