Measurement of Reactive Oxygen Species in Cardiovascular Studies

Abstract

Several mammalian enzymes are capable of transferring electrons to molecular oxygen, sequentially forming the 1 electron-reduction product superoxide (O2·−) and the 2 electron-reduction product hydrogen peroxide (H2O2). These serve as progenitors for other reactive oxygen species (ROS), including peroxynitrite (ONOO−), hypochlorous acid, the hydroxyl radical, lipid peroxides, lipid peroxy- radicals, and lipid alkoxyl radicals. Another relevant group of molecules is the reactive nitrogen species, including NO, the nitrogen dioxide radical, and the nitrosonium cation. In the cardiovascular system, the most important enzymes that produce ROS are the Nox-based reduced nicotinamide-adenine dinucleotide phosphate (NADPH) oxidases, xanthine oxidase, the mitochondrial electron transport system and, under certain circumstances, NO synthase. Conditions such as hypertension, atherosclerosis, hypercholesterolemia, diabetes, and insulin resistance increase either the activity or the expression of these enzymes, leading to elevated ROS production. ROS, in turn, contribute to these disorders. As examples, virtually every aspect of atherosclerotic lesion formation is augmented by oxidative events. Via several mechanisms involving vessels, the kidney, and the central nervous system, ROS augment hypertension. ROS have been implicated in causing insulin resistance and pancreatic damage leading to diabetes. Moreover, ROS, when produced in specific subcellular compartments in controlled amounts, can act as signaling molecules to regulate normal cellular functions. These reactions have been reviewed in depth elsewhere recently.1 Given the importance of ROS and reactive nitrogen species in physiology and pathophysiology, it has become essential that methods be adapted and standardized to quantify the levels of these molecules in cells and tissues. Numerous methods have been used for ROS detection, each with potential pitfalls and advantages. All of these techniques can yield errors, and it is advisable to use ≥2 methods to avoid a mistaken conclusion. In this review, we will discuss various methods used to detect and quantify …