Measurement of Reactive Oxygen Species (ROS) and Mitochondrial ROS in AMPK Knockout Mice Blood Vessels

Methods in Molecular Biology
Qilong Wang, Ming-Hui Zou

Abstract

Reactive oxygen species (ROS) are a group of unstable and highly reactive molecules or free radicals typically generated as by-products of cellular processes involving molecular oxygen. In vascular cells, the excessive ROS generation results in the initiation and progression of cardiovascular diseases (CVD). Therefore, a dynamic, robust, and accurate ROS detection method in the blood vessels is essential for pathophysiological research studies of the cardiovascular system.In this chapter, we describe a fluorescence dye-based detection method for assaying superoxide and mitochondrial superoxide in mouse aorta using dihydroethidium (DHE) and MitoSOX. The protocol includes preparation of frozen aortic tissue sections, monitoring DHE oxidation-derived fluorescence by fluorescence microscopy, and high-performance liquid chromatograph-based analysis of MitoSOX and its oxidation products. For studying the role of AMP-activated protein kinase (AMPK) in the redox regulation, we employed AMPKα2 knockout mice and observed increased superoxide and mitochondrial superoxide levels in the aorta of AMPK knockout mice relative to the wild-type group. This novel ROS detection method will be valuable for investigating the roles of cellular and/or...Continue Reading

Citations

Nov 2, 2019·Basic & Clinical Pharmacology & Toxicology·Hui ChenSusan Wai Sum Leung
Jun 24, 2018·Cells·Gianluca SgarbiAlessandra Baracca
Dec 18, 2020·International Journal of Molecular Sciences·Woosuk KimDae Won Kim
Feb 8, 2021·Pharmacological Research : the Official Journal of the Italian Pharmacological Society·Siao Muk ChengChun Hei Antonio Cheung
Mar 2, 2021·Frontiers in Cell and Developmental Biology·Shannon LynchSamantha Giordano-Mooga
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Mar 7, 2021·Antioxidants·Marc Herb, Michael Schramm
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Jul 20, 2021·Frontiers in Cell and Developmental Biology·Jannatul AklimaYoshihiro Ohta
May 29, 2021·Otology & Neurotology : Official Publication of the American Otological Society, American Neurotology Society [and] European Academy of Otology and Neurotology·Michael T BroderickMichael J Brenner

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