NOX1 Inhibition Attenuates Kidney Ischemia-Reperfusion Injury via Inhibition of ROS-Mediated ERK Signaling

Abstract

The protective effects of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) 1 inhibition against kidney ischemia-reperfusion injury (IRI) remain uncertain. The bilateral kidney pedicles of C57BL/6 mice were clamped for 30 min to induce IRI. Madin–Darby Canine Kidney (MDCK) cells were incubated with H2O2 (1.4 mM) for 1 h to induce oxidative stress. ML171, a selective NOX1 inhibitor, and siRNA against NOX1 were treated to inhibit NOX1. NOX expression, oxidative stress, apoptosis assay, and mitogen-activated protein kinase (MAPK) pathway were evaluated. The kidney function deteriorated and the production of reactive oxygen species (ROS), including intracellular H2O2 production, increased due to IRI, whereas IRI-mediated kidney dysfunction and ROS generation were significantly attenuated by ML171. H2O2 evoked the changes in oxidative stress enzymes such as SOD2 and GPX in MDCK cells, which was mitigated by ML171. Treatment with ML171 and transfection with siRNA against NOX1 decreased the upregulation of NOX1 and NOX4 induced by H2O2 in MDCK cells. ML171 decreased caspase-3 activity, the Bcl-2/Bax ratio, and TUNEL-positive tubule cells in IRI mice and H2O2-treated MDCK cells. Among the MAPK pathways, ML171 affected ERK signaling by ERK phosphorylation in kidney tissues and tubular cells. NOX1-selective inhibition attenuated kidney IRI via inhibition of ROS-mediated ERK signaling.

Single-Dose Toxicity Study on ML171, a Selective NOX1 Inhibitor, in Mice

“…Among the seven NOX isoforms [16], NOX1-dependent ROS production contributes to cellular growth signaling, angiogenesis, motility, and endothelial function [17][18][19]. NOX1 is a therapeutic target for IRI of multiple organs, including the heart, lung, kidney, and retina [12,[20][21][22]. ML171, a selective NOX1 inhibitor, has been studied in various animal models [11,23,24].…”

“…Recently, we confirmed the renoprotective effect of ML171 in an ischemia-reperfusion injury (IRI) mouse model. ML171 attenuated kidney IRI via inhibition of ROS-mediated Extracellular Signal-Regulated Kinase (ERK) signaling and reduced oxidative stress-induced apoptosis in renal tubule cells [ 12 ]. These results suggest that ML171 could be used as a therapeutic agent for oxidative injury in many different organs.…”

“…In a preclinical study, we treated animals with a 60 mg/kg intraperitoneal (IP) injection of ML171. The IP injection was well tolerated in all the mice without any adverse events and histological changes [ 12 ]. However, the experiments were not performed in a good laboratory practice (GLP) setting.…”

Kidney Inflammation, Injury and Regeneration 2020

“…In this current Special Edition, thirteen original research articles are presented: twelve in vivo studies in a murine or rat model and one in vitro study [ 3 ]. Seven studies show results from AKI models [ 4 , 5 , 6 , 7 , 8 , 9 , 10 ], five from fibrosis models (or CKD models) [ 9 , 11 , 12 , 13 , 14 ] and two from a transplant rejection model [ 4 , 15 ] (two studies used two different in vivo models).…”

“…The authors confirmed this effect through in vitro studies with immortal tubular epithelial cells [ 10 ]. Others used an IR model to show that the NOX1-selective inhibition attenuates kidney IR injuries via the downregulation of oxidative stress-mediated kinase signaling [ 8 ] or to investigate the autophagy dynamics during an IR injury, a potential treatment strategy due to removing damaged cells, macromolecules and organelles [ 6 ]. The effect of growth differentiation factor 15 (GDF15) was investigated in a murine model of anti-glomerular basement membrane glomerulonephritis [ 7 ].…”