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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 ganes. 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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OPTICAL IMAGING OF LIPOPOLYSACCHARIDE-INDUCED OXIDATIVE STRESS IN ACUTE LUNG INJURY FROM HYPEROXIA AND SEPSIS

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Journal of Innovative Optical Health Sciences 2013年第3期6卷 11-20页

作者： REYHANEH SEPEHR SAID H.AUDI SEPIDEH MALEKI KEVIN STANISZEWSKI ANNIE L.EIS girija G.konduri MAHSA RANJI Biophotonics Laboratory Department of Electrical Engineering and Computer ScienceUniversity of Wisconsin Milwaukee 3200 N Cramer St.MilwaukeeWI 53211USA Department of Biomedical Engineering Marquette University 1515 W Wisconsin Avenue MilwaukeeWI 53233USA Department of Pediatrics Cardiovascular Research Center Medical College of Wisconsin8701 Watertown Plank Rd MilwaukeeWI 53226USA Department of Pediatrics Cardiovascular Center and Children's Research InstituteMedical College of Wisconsin CCCSte C410999 N92 StMilwaukeeWI 53226USA

Reactive oxygen species(ROS)have been implicated in the pathogenesis of many acute and chronic pulmonary disorders such as acute lung injury(ALI)in adults and bronchopulmonary dysplasia(BPD)in premature *** infection and oxygen toxicity,which result in pulmonary vascular endot belial injury,contribute to impaired vascular growth and alveolar simplfcation soen in the lungs of premature infants with *** induces ALI,reduces cell proliferation,cauuses DNA damage and promotes cell death by causi ng mitochondrial dys *** objective of this study was to use an optical imaging technique to evaluate the variations in fluorescence intensities of the auto fuorescent mitochondrial metabolic coenzymes,NADH and FAD in four different groups of *** ratio of these fluorescence signals(NA DH/FAD),referred to as NADH redox ratio(NADH RR)has been used as an indicator of tisue metabolism in ***,we investigated whether the changes in metabolic state can be used as a marker of oxidative stress caused by hyperoxia and bacterial lipopolysaccharide(LPS)exposure in neonatal mat *** examined the tissue redox states of lungs from four groups of rat pups:nomoxic(21%O_(2))pups,hyperoxic(90%O_(2))pups,pups treated with LPS(normoxic+LPS),and pups treated with LPS and hyperoxia(hyperoxic+LPS).Our results show that hyperoxia oxidized the respiratory chain as reflected by a~31%docrease in lung tssue NADH RR as compared to that for normoxic *** treatment alone or with hyperoxia had no significant ffect on lung tssue NADH RR as compared to that for normoxic or hyperoxic lungs,***,NADH RR serves as a quantitative marker of oxidative stress level in lung injury caused by two clinically important conditions:hyperoxia and LPS exposure.

关键词： Fluorescence imaging NADH FAD LPS Hyperoxia

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