Abstract
Oxidative stress in a biological system is often defined as a redox imbalance within cells or groups of cells within an organism. Reductive-oxidative (redox) imbalances in cellular systems have been implicated in several diseases, such as cancer. To better understand the redox environment within cellular systems, it is important to be able to characterize the relationship between the intensity of the oxidative environment, characterized by redox potential, and the biomolecular consequences of oxidative damage. In this study, we show that an in situ electrochemical potential gradient can serve as a tool to simulate exogenous oxidative stress in surface-attached mammalian cells. A culture plate design, which permits direct imaging and analysis of the cell viability, following exposure to a range of solution redox potentials, was developed. The in vitro oxidative stress test vessel consists of a cell growth flask fitted with two platinum electrodes that support a direct current along the flask bottom. The applied potential span and gradient slope can be controlled by adjusting the constant current magnitude across the vessel with spatially localized media potentials measured with a sliding reference electrode. For example, the viability of Chinese Hamster Ovary cells under a gradient of redox potentials indicated that cell death was initiated at approximately 0.4 V vs. standard hydrogen electrode (SHE) media potential and this potential could be modified with antioxidants. This experimental platform may facilitate studies of oxidative stress characteristics on different types of cells by enabling imaging live cell cultures that have been exposed to a gradient of exogenous redox potentials.
Highlights
Oxidative stress (OS) in biological systems is defined as an imbalance between reactive oxygen species (ROS) and the organism’s antioxidant defenses
We investigated the effect of the extracellular reduced glutathione on CHO cell viability when subjected to an electrochemical potential gradient
In this study we show that an electrochemical potential gradient can serve as a tool to simulate exogenous oxidative stressthat on surface attached mammalian cells
Summary
Oxidative stress (OS) in biological systems is defined as an imbalance between reactive oxygen species (ROS) and the organism’s antioxidant defenses. By measuring the cytosolic concentrations of reduced and oxidized glutathione [9,10] or other electrochemical couples [9] the intracellular redox potential can be calculated Such data allows the establishment of the relationship between the biochemical effects and the thermodynamic and kinetic models, even if, strictly speaking, the biological systems do not operate in equilibrium. Electrochemical methods in biological systems are often used for the detection of various ROS species during the cellular redox processes [19], yet are rarely used for inducing redox potentials on cells [20]. With our experimental approach we can impose a stable, well-controlled linear potential profile over a cell monolayer and as a result observe a cellular response to a gradient-generated exogenous redox environment. Cells subjected to increasingly higher oxidative potentials undergo cell death, as confirmed by live/dead fluorescent probe emission profiles
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
