title: Netrin-1 abrogates ischemia/reperfusion-induced cardiac mitochondrial dysfunction via nitric oxide-dependent attenuation of NOX4 activation and recoupling of NOS.
abstract: Despite an established role of mitochondrial dysfunction in cardiac ischemia/reperfusion (I/R) injury, the upstream activators have remained incompletely defined. We have recently identified an innovative role of exogenously applied netrin-1 in cardioprotection, which is mediated by increased nitric oxide (NO) bioavailability. Here, we tested the hypothesis that this "pharmacological" treatment of netrin-1 preserves mitochondrial function via novel mechanisms that are NO dependent. Freshly isolated C57BL6 mouse hearts were perfused using a Langendorff system, and subjected to a 20min global ischemia/60min reperfusion, in the presence or absence of netrin-1. I/R induced marked increases in infarct size, total superoxide and hydrogen peroxide production, activity and protein abundance of NADPH oxidase (NOX) isoform 4 (NOX4), as well as impaired mitochondrial integrity and function, all of which were attenuated by netrin-1. This protective effect of netrin-1 is attributed to cGMP, a downstream effector of NO. The protein levels of NOX1 and NOX2 were however unaffected, and infarct size from NOX1 and NOX2 knockouts was not different from wild type animals. Scavenging of NO with PTIO reversed inhibitory effects of netrin-1 on NOX4, while NO donor attenuated NOX4 protein abundance. In vivo NOX4 RNAi, or sepiapterin perfusion, resulted in recoupling of NOS, decreased infarct size, and blockade of dysfunctional mitochondrial swelling and mitochondrial superoxide production. These data demonstrate that netrin-1 induces cardioprotection through inhibition of NOX4 activity, which leads to recoupling of NOS, augmented NO bioavailability, reduction in oxidative stress, and ultimately preservation of mitochondrial function. The NO-dependent NOX4 inhibition connects with our previously established pathway of DCC/ERK1/2/eNOS/NO/DCC feed-forward mechanism, to maintain NOS in the coupling state to attenuate oxidative stress to preserve mitochondrial function. These findings may promote development of novel therapeutics for cardiac I/R injury. This article is part of a Special Issue entitled "Mitochondria: From Basic Mitochondrial Biology to Cardiovascular Disease".
authors: Divisions of Molecular Medicine and Cardiology, Department of Anesthesiology, Cardiovascular Research Laboratories, David Geffen School of Medicine at University of California Los Angeles, Los Angeles, CA, USA; Divisions of Molecular Medicine and Cardiology, Department of Medicine, Cardiovascular Research Laboratories, David Geffen School of Medicine at University of California Los Angeles, Los Angeles, CA, USA.
authors: Kin Lung
authors: KL
authors: Siu
authors: Department of Physiology, Cardiovascular Research Laboratories, David Geffen School of Medicine at University of California Los Angeles, Los Angeles, CA, USA.
authors: Christopher
authors: C
authors: Lotz
authors: Department of Physiology, Cardiovascular Research Laboratories, David Geffen School of Medicine at University of California Los Angeles, Los Angeles, CA, USA.
authors: Peipei
authors: P
authors: Ping
authors: Divisions of Molecular Medicine and Cardiology, Department of Anesthesiology, Cardiovascular Research Laboratories, David Geffen School of Medicine at University of California Los Angeles, Los Angeles, CA, USA; Divisions of Molecular Medicine and Cardiology, Department of Medicine, Cardiovascular Research Laboratories, David Geffen School of Medicine at University of California Los Angeles, Los Angeles, CA, USA. Electronic address: hcai@mednet.ucla.edu.
authors: Hua
authors: H
authors: Cai
mesh_terms: D000818:Animals; D002316:Cardiotonic Agents / Q000008:administration & dosage / Q000494:pharmacology; D004195:Disease Models, Animal; D006861:Hydrogen Peroxide / Q000378:metabolism; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008929:Mitochondria, Heart / Q000187:drug effects* / Q000378:metabolism*; D015428:Myocardial Reperfusion Injury / Q000235:genetics / Q000378:metabolism* / Q000473:pathology / Q000628:therapy; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D009414:Nerve Growth Factors / Q000008:administration & dosage / Q000494:pharmacology*; D000075388:Netrin-1; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D018384:Oxidative Stress; D011622:Pterins / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D013481:Superoxides / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000008:administration & dosage / Q000494:pharmacology*
chemical_list: D002316:Cardiotonic Agents; D009414:Nerve Growth Factors; C000617353:Ntn1 protein, mouse; D011622:Pterins; D034741:RNA, Small Interfering; D025521:Tumor Suppressor Proteins; D013481:Superoxides; D000075388:Netrin-1; D009569:Nitric Oxide; D006861:Hydrogen Peroxide; C016727:sepiapterin; D019001:Nitric Oxide Synthase; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C422943:Nox4 protein, mouse
keywords: Ischemia/reperfusion (I/R) injury; Mitochondrial dysfunction; NADPH oxidase isoform 4 (NOX4); NO synthase (NOS); NOS uncoupling; Netrin-1