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1 Medicine, Medical University of Ohio, Toledo, Ohio, United States
2 Medicine, Friendship Hospital, Beijing, China
3 Chinese Academy of Medical Sciences, The Institute of Nutrition and Food Hygiene, Beijing, China
4 Proteomics Core Laboratory, Medical University of Ohio, Toledo, Ohio, United States
* To whom correspondence should be addressed. E-mail: jshapiro{at}meduohio.edu.
The effect of cardiac glycosides to increase cardiac inotropy by altering Ca2+ cycling is well known but still poorly understood. The studies described in this report focus on defining the effects of ouabain signaling on sarcoplasmic reticulum Ca2+ ATPase function. Rat cardiac myocytes treated with 50 µM ouabain demonstrated substantial increases in systolic and diastolic Ca2+ concentrations. The recovery time constant for the Ca2+ transient, TauCa2+, was significantly prolonged by ouabain. Exposure to 10 µM H2O2, which causes a similar increase in intracellular reactive oxygen species as 50 µM ouabain, caused a similar increase in TauCa2+. Concurrent exposure to 10 mM N-Acetyl Cysteine or an aqueous extract from green tea (50 mg/ml) both prevented the increases in TauCa2+ as well as the changes in systolic or diastolic Ca2+ concentrations. We also observed that 50 µM ouabain induced increases in developed pressure in addition to diastolic dysfunction in the isolated perfused rat heart. Co-administration of ouabain with N-Acetyl Cysteine prevented these increases. Analysis of sarcoplasmic reticulum Ca2+ ATPase protein revealed increases in both the oxidation and nitrotyrosine content in the ouabain treated hearts. Liquid chromatography/mass spectrometric analysis confirmed that the sarcoplasmic reticulum Ca2+ ATPase protein from ouabain treated hearts had modifications consistent with oxidative and nitrosative stress. These data suggest that ouabain induces oxidative changes of the sarcoplasmic reticulumCa2+ ATPase structure and function which may, in turn, produce some of the associated changes in Ca2+ cycling and physiological function.
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