Biomedical Engineering and Bioengineering Commons^™

Detection Of Hydrogen Peroxide Production In The Isolated Rat Lung Using Amplex Red, Said H. Audi, Nina Friedly, Ranjan K. Dash, Andreas M. Beyer, Anne V. Clough, Elizabeth R. Jacobs

Biomedical Engineering Faculty Research and Publications

The objectives of this study were to develop a robust protocol to measure the rate of hydrogen peroxide (H₂O₂) production in isolated perfused rat lungs, as an index of oxidative stress, and to determine the cellular sources of the measured H₂O₂ using the extracellular probe Amplex red (AR). AR was added to the recirculating perfusate in an isolated perfused rat lung. AR’s highly fluorescent oxidation product resorufin was measured in the perfusate. Experiments were carried out without and with rotenone (complex I inhibitor), thenoyltrifluoroacetone (complex II inhibitor), antimycin A (complex III inhibitor), potassium …

Hyperoxia Causes Mitochondrial Fragmentation In Pulmonary Endothelial Cells By Increasing Expression Of Pro-Fission Proteins, Cui Ma, Andreas M. Beyer, Matthew J. Durand, Anne V. Clough, Daling Zhu, Laura Norwood Toro, Maia Terashvili, Johnathan D. Ebben, R. Blake Hill, Said H. Audi, Meetha Medhora, Elizabeth R. Jacobs

Biomedical Engineering Faculty Research and Publications

Objective—We explored mechanisms that alter mitochondrial structure and function in pulmonary endothelial cells (PEC) function after hyperoxia.

Approach and Results—Mitochondrial structures of PECs exposed to hyperoxia or normoxia were visualized and mitochondrial fragmentation quantified. Expression of pro-fission or fusion proteins or autophagy-related proteins were assessed by Western blot. Mitochondrial oxidative state was determined using mito-roGFP. Tetramethylrhodamine methyl ester estimated mitochondrial polarization in treatment groups. The role of mitochondrially derived reactive oxygen species in mt-fragmentation was investigated with mito-TEMPOL and mitochondrial DNA (mtDNA) damage studied by using ENDO III (mt-tat-endonuclease III), a protein that repairs mDNA damage. Drp-1 (dynamin-related …