Article

In vivo real-time measurement of superoxide anion radical with a novel electrochemical sensor

Advanced Medical Emergency and Critical Care Center, Yamaguchi University Hospital, Ube 755-8505, Japan; Department of Pure and Applied Chemistry, Faculty of Science and Technology, Tokyo University of Science, Noda 278-8510, Japan; First Department of Internal Medicine, Kyoto Prefectural University of Medicine, Kyoto 602-8566, Japan
Free Radical Biology and Medicine (Impact Factor: 5.27). 10/2009; DOI: 10.1016/j.freeradbiomed.2009.07.012

ABSTRACT The dynamics of superoxide anion (O2−) in vivo remain to be clarified because no appropriate method exists to directly and continuously monitor and evaluate O2− in vivo. Here, we establish an in vivo method using a novel electrochemical O2− sensor. O2− generated is measured as a current and evaluated as a quantified partial value of electricity (Qpart), which is calculated by integration of the difference between the baseline and the actual reacted current. The accuracy and efficacy of this method were confirmed by dose-dependent O2− generation in xanthine–xanthine oxidase in vitro in phosphate-buffered saline and human blood. It was then applied to endotoxemic rats in vivo. O2− current began to increase 1 h after lipopolysaccharide, and Qpart increased significantly for 6 h in endotoxemic rats, in comparison to sham-treated rats. These values were attenuated by superoxide dismutase. The generation and attenuation of O2− were indirectly confirmed by plasma lipid peroxidation with malondialdehyde, endothelial injury with soluble intercellular adhesion molecule-1, and microcirculatory dysfunction. This is a novel method for measuring O2− in vivo and could be used to monitor and treat the pathophysiology caused by excessive O2− generation in animals and humans.

1 Bookmark
 · 
86 Views
  • Sensors and Actuators B Chemical 03/2014; 176:884-892. · 3.84 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Heatstroke is generally considered as a sepsis-like syndrome induced by hyperthermia leading to multiorgan dysfunction. High-mobility group box 1 (HMGB1) has recently been identified as a mediator of systemic inflammation leading to multiorgan dysfunction in sepsis and nonsepsis. Elevation of plasma HMGB1 in heatstroke has been suggested in experimental models and clinical patients. By far, whether HMGB1 could be a potential therapeutic target in heatstroke is unknown. The objectives of this study are to use HMGB1 monoclonal antibody to specifically inhibit the activity of extracellular HMGB1 and to observe the possible protection of liver injury in a rat heatstroke model. After treatment with neutralizing antibodies to HMGB1, rats were exposed to a high-temperature and high-humidity environment. At the time of heatstroke onset, the plasma and liver cytoplasm HMGB1 levels were detected by enzyme-linked immunosorbent assay. The histopathology of liver tissue was observed under light microscopy and transmission electron microscopy. Plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities were determined using the commercially available kits. Plasma tumor necrosis factor-α, interleukin-1β (IL-1β), and IL-6 were determined using enzyme-linked immunosorbent assay kits. HMGB1 levels in plasma and liver cytoplasm were both elevated in heatstroke rats, which were both associated with increased plasma ALT and AST levels. Histopathologic results showed that HMGB1 monoclonal antibody pretreatment could obviously alleviate the pathologic impairments of heatstroke rats. HMGB1 monoclonal antibody pretreatment could also downregulate plasma AST and ALT levels in heatstroke rats. Plasma tumor necrosis factor-α, IL-1β, and IL-6 levels in heatstroke rats were elevated, which could be significantly suppressed by HMGB1 antibody pretreatment. HMGB1 could be a potentially effective treatment target in heatstroke. The pathogenic mechanism of heatstoke is complicated, which needs comprehensive prevention and treatment.
    The journal of trauma and acute care surgery. 03/2013; 74(3):801-7.
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Superoxide anion is the key radical that causes intracellular oxidative stress. The lack of a method to directly monitor superoxide concentration in vivo in real time has severely hindered our understanding on its pathophysiology. We made transgenic zebrafish to specifically express yellow fluorescent proteins, a reversible superoxide-specific indicator, in the liver and used a fiber-optic fluorescent probe to noninvasively monitor the superoxide concentration in real time. Several superoxide-inducing and scavenging reagents were administrated onto the fish to alter superoxide concentrations. The distinct biochemical pathways of the reagents can be discerned from the transient behaviors of fluorescence time courses. These results demonstrate the feasibility of this method for analyzing superoxide dynamics and its potential as an in vivo pharmaceutical screening platform.
    Biomedical Optics Express 01/2013; 4(9):1702-9. · 3.18 Impact Factor