Abstract
38 - Quantitative Changes in Dihydroethidium (DHE) Oxidation Products from Isolated Mitochondria While Respiring on Select Substrates and the Effects Mitochondrial Inhibitors Commonly Used in Bioenergetic Profiling
Free radical biology & medicine, Vol.100, pp.S31-S31
11/2016
DOI: 10.1016/j.freeradbiomed.2016.10.079
Abstract
Dihydroethdium (DHE) oxidation is commonly used as a method for monitoring cellular production of “reactive oxygen species (ROS)”. Usually changes in DHE florescence due to oxidation in cells and tissues are measured by microscopy, flow cytometry and occassionaly by HPLC analysis. Rarely are functional intact mitochondria while respiring on known mitochondrial substrates used, nor changes in “ROS” and DHE oxidation products analyzed while using bioenergetics profiling strategies like those used in Seahorse type experiments. We have studied quantitative changes in DHE oxidation products, as measured by HPLC with electrochemical detection, from intact, functional mitochondria that were simultaneously analyzed by Seahorse bioenergetic profile analysis. Combining these 2 types analysis allows for a more thorough and quantitative approach to understanding the relationship of mitochondrial “ROS” and mitochondrial bioenergetics. Forinstance in functional isolated mouse brain mitochondria we measured the rates of molecular oxygen consumption and used the same mitochondria for measuring the quantitative production of the superoxide specific DHE oxidation product, 2-hydroxyethidium, the non-specific DHE oxidation product ethidium and select DHE mixed dimers. The quantitative yields of products do not appear to be limited by DHE substrate availablity but represent the actual oxidation product yield under these experimental and kinetic conditions: the product yield are quite different than those observed from intact cells and tissues.
Supported NIH grants R01 CA169046, R01 CA184051, R21 CA137230, P30 CA086862, R01 GM073929, P42 ES013661 and VA Merit Award 1101BX001318.
Details
- Title: Subtitle
- 38 - Quantitative Changes in Dihydroethidium (DHE) Oxidation Products from Isolated Mitochondria While Respiring on Select Substrates and the Effects Mitochondrial Inhibitors Commonly Used in Bioenergetic Profiling
- Creators
- Brett A Wagner - University of IowaGarry R Buettner - University of Iowa
- Resource Type
- Abstract
- Publication Details
- Free radical biology & medicine, Vol.100, pp.S31-S31
- DOI
- 10.1016/j.freeradbiomed.2016.10.079
- ISSN
- 0891-5849
- eISSN
- 1873-4596
- Publisher
- Elsevier Inc
- Language
- English
- Date published
- 11/2016
- Academic Unit
- Radiation Oncology
- Record Identifier
- 9984314287702771
Metrics
16 Record Views