Thermal Induction of Oxidative Stress
What happens to the cell when it is exposed to elevated ambient temperatures? In this project, I aimed to examine how heat stress affected oxidative stress levels in fibroblast cell lines. In this first undergraduate research project, I refined my skills in cell culture and experimental design while cultivating a love for the process of biochemistry.
Reactive oxygen species (ROS) are highly reactive oxygen-containing molecules that rapidly interact with other cellular molecules, causing oxidative stress. ROS are produced during mitochondrial respiration, in which one-electron transfer produces free radical oxygen species. Low levels of ROS are required for cell signaling pathways; however, when ROS concentrations exceed the cell’s antioxidant capacity, chain reactions occur, damaging DNA, RNA, and proteins.
When a cell is exposed to elevated temperatures, the HSF1 transcription factor initiates the transient expression of a family of Heat Shock Proteins (HSPs) responsible for refolding or degrading damaged proteins. Here is a simplified diagram of the heat shock response.
2,7-Dichlorofluorescin diacetate (DCFH-DA) is commonly used as an oxidative stress quantification tool. Here is a flow diagram showing how DCFH-DA is used. When DCFH is oxidized to DCF by an ROS, the molecule becomes fluorescent, providing a quantitative signal of ROS that we can measure and visualize.
DCF fluoresces in the green region of the visible spectrum, and we can visualize its distribution within the cell. To the right is a fluorescence microscopy image of the cells loaded with DCFH-DA. I am grateful for the introductory knowledge I gained and for what I learned from working on this project. It provided a foundation for cultivating curiosity and diligence in biochemistry in the future.
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Check out my poster for this research project, presented in Spring 2025!