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Published on: February 7, 2018
Development of a cell culture model system for routine testing of substances inducing oxidative stress
D Ritter1, J W Knebel, M Aufderheide
1Fraunhofer Institute of Toxicology and Aerosol Research, Department of In Vitro Toxicology, Nikolai-Fuchs-Str. 1, D-30625 Hannover, Germany.
Abstract:
The toxicological model of oxidative stress is a mechanism which is currently thought to be involved in the formation and development of many serious human diseases. Little is known about cellular responses to oxidative damage and the related specific toxicological properties of substances such as chemicals or components of the polluted urban atmosphere. On the basis of biological pathways involved in cellular antioxidative mechanisms, we developed a biological model for studying the oxidative properties of substances. This includes human lung cells and methods for biochemical analysis of cellular endpoints. Antioxidative and glycolysis-related enzyme activities (glutathione peroxidase, glutathione reductase, superoxide dismutase, catalase, phosphofructokinase, enolase, glucose-6-phosphate dehydrogenase), ATP/ADP/AMP and glutathione (oxidized and reduced) are determined. Routine testing of substances with high reproducibility and fast screening is provided by adapting methods for biochemical analysis to determinations using cells grown on microtitre plates. First experiments with standard model substances inducing oxidative stress such as H(2)O(2) and tert-butylhydroperoxide show that enzymatic activities can be measured with good reproducibility and their changes can be followed kinetically. The results indicate the relevance of the determined parameters for such toxicological events and the usefulness of the biological indicator system for routine testing.
Insights
Researchers developed a novel biological model using human lung cells to assess oxidative stress from chemicals. This model enables reproducible, high-throughput screening of cellular responses to oxidative damage, aiding toxicological assessments.
Area of Science:
- Toxicology
- Cell Biology
- Biochemistry
Background:
- Oxidative stress is implicated in numerous human diseases.
- Cellular responses to oxidative damage and the toxicological properties of environmental substances remain poorly understood.
- Existing toxicological models lack comprehensive analysis of cellular antioxidative mechanisms.
Purpose of the Study:
- To develop a reproducible biological model for assessing the oxidative properties of substances.
- To investigate cellular responses to oxidative stress using human lung cells.
- To establish a high-throughput screening system for toxicological evaluations.
Main Methods:
- Utilized human lung cells cultured on microtitre plates.
- Developed biochemical assays to measure antioxidative and glycolysis-related enzyme activities.
- Quantified cellular energy status (ATP/ADP/AMP) and glutathione levels (oxidized and reduced).
Main Results:
- Demonstrated reproducible measurement of enzymatic activities in response to oxidative stress inducers like H(2)O(2).
- Showcased the ability to kinetically monitor changes in enzyme activities.
- Validated the model's utility with standard oxidative stress agents.
Conclusions:
- The developed biological model provides a reliable system for routine toxicological testing.
- Key biochemical parameters are relevant indicators of oxidative stress events.
- The system facilitates fast screening and reproducible assessment of substance toxicity.
