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Quantitative optical measurement of mitochondrial superoxide dynamics in pulmonary artery endothelial cells

Journal of Innovative Optical Health Sciences 2018年第1期11卷 90-105页

作者： Zahra Ghanian Girija Ganesh Konduri Said Halim Audi Amadou K.S.Camara Mahsa Ranji Department of Electrical Engineering University of Wisconsin-Milwaukee MilwaukeeWisconsinUSA Department of Pediatrics Division of Neonatology Medical College of WisconsinMilwaukeeWisconsinUSA Department of Biomedical Engineering Marquette University MilwaukeeWisconsinUSA Department of Anesthesiology and Anesthesia Research Medical College of Wisconsin MilwaukeeWisconsinUSA

Reactive oxygen species(ROS)play a vital role in cell signaling and redox regulation,but when present in excess,lead to numerous *** quantitative characterization of mitochondrial superoxide anion(O^(·-)_(2))production in fetal pulmonary artery endothelia cells(PAECs)has never been *** aim of this study is to assess mitochondrial O^(·-)_(2)pro-duction in cultured PAECs over time using a novel quantitative optical *** rate,the sources,and the dynamics of O^(·-)_(2)production were assessed using targeted metabolic modulators of the mitochondrial electron transport chain(ETC)complexes,specifically an uncoupler and inhibitors of the various ETC complexes,and inhibitors of extra-mitochondrial sources of O^(·-)_(2).After stabilization,the cells were loaded with nanomolar mitochondrial-targeted hydroethidine(Mito-HE,MitoSOX)online during the experiment without washout of the residual ***-lapse fuorescence microscopy was used to monitor the dynamic changes in O^(·-)_(2)fluorescence intensity over time in *** transient behaviors of the fuorescence time course showed exponential increases in the rate of O^(·-)_(2) production in the presence of the ETC uncoupler or *** most dramatic and the fastest increase in O^(·-)_(2)production was observed when the cells were treated with the uncoupling agent,*** also showed that only the complex IV inhibitor,KCN,attenuated the marked surge in O^(·-)_(2)production induced by *** results showed that mitochondrial respiratory complexes I,III and IV are sources of O^(·-)_(2) production in PAECs,and a new observation that ROS production during uncoupling of mitochondrial res-piration is mediated in part via complex *** novel method can be applied in other studies that examine ROS production under stress condition and during ROS mediated injuries in vritro.

关键词： Fluorescence microscopy time-lapse imaging oxidative stress superoxide pentachlorophenol sodium salt rotenone antimycin A potassium cyanide MitoSOX.

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