Thioredoxin Reductase-2 Is Essential for Keeping Low Levels of H2O2 Emission from Isolated Heart Mitochondria

Stanley, Brian A.; Sivakumaran, Vidhya; Shi, Sa; McDonald, Iain; Lloyd, David; Watson, Walter H.; Aon, Miguel A.; Paolocci, Nazareno

doi:10.1074/jbc.m111.284612

Cited by 160 publications

(161 citation statements)

References 56 publications

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“…The total GSH or GSSG was determined with the commercially available kit from Bioxytech (GSH/GSSG-412), with slight modifications for 96-well microplate reader using a Flex Station 3 (Molecular Devices). The absolute concentrations of GSH or GSSG present in the samples were estimated using a mitochondrial volume of 2 l/mg mitochondrial protein (4,49) and the amount of protein in the sample determined according to the bicinchoninic acid method (BCA protein assay kit; Thermo Scientific).…”

Section: Methodsmentioning

confidence: 99%

Restoring redox balance enhances contractility in heart trabeculae from type 2 diabetic rats exposed to high glucose

Bhatt

Aon

Tocchetti

et al. 2015

American Journal of Physiology-Heart and Circulatory Physiology

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Bhatt NM, Aon MA, Tocchetti CG, Shen X, Dey S, Ramirez-Correa G, O=Rourke B, Gao WD, Cortassa S. Restoring redox balance enhances contractility in heart trabeculae from type 2 diabetic rats exposed to high glucose. Am J Physiol Heart Circ Physiol 308: H291-H302, 2015. First published December 3, 2014; doi:10.1152/ajpheart.00378.2014.-Hearts from type 2 diabetic (T2DM) subjects are chronically subjected to hyperglycemia and hyperlipidemia, both thought to contribute to oxidizing conditions and contractile dysfunction. How redox alterations and contractility interrelate, ultimately diminishing T2DM heart function, remains poorly understood. Herein we tested whether the fatty acid palmitate (Palm), in addition to its energetic contribution, rescues function by improving redox [glutathione (GSH), NAD(P)H, less oxidative stress] in T2DM rat heart trabeculae subjected to high glucose. Using cardiac trabeculae from Zucker Diabetic Fatty (ZDF) rats, we assessed the impact of low glucose (EG) and high glucose (HG), in absence or presence of Palm or insulin, on force development, energetics, and redox responses. We found that in EG ZDF and lean trabeculae displayed similar contractile work, yield of contractile work (Ycw), representing the ratio of force time integral over rate of O2 consumption. Conversely, HG had a negative impact on Ycw, whereas Palm, but not insulin, completely prevented contractile loss. This effect was associated with higher GSH, less oxidative stress, and augmented matrix GSH/thioredoxin (Trx) in ZDF mitochondria. Restoration of myocardial redox with GSH ethyl ester also rescued ZDF contractile function in HG, independently from Palm. These results support the idea that maintained redox balance, via increased GSH and Trx antioxidant activities to resist oxidative stress, is an essential protective response of the diabetic heart to keep contractile function. contractile work; rate of respiration; redox environment; antioxidant systems; oxidative phosphorylation; mitochondrial ros emission; Zucker diabetic fatty rat A COMMON COMPLICATION OF TYPE 2 diabetes mellitus (T2DM) is cardiomyopathy, characterized by diastolic and systolic dysfunction, which is likely impacted by alterations in metabolic substrate availability. Although the healthy heart is flexible regarding fuel selection, the high levels of glucose and fat in T2DM lead to questions about which factors contribute to dysfunction and which are beneficial as energy source or redox donors (35).Fatty acids (FA) and glucose are the two major fuels driving heart contraction, and in T2DM and obesity existing evidence indicates increased FA oxidation (12, 15). The idea that FAs excess negatively regulates glucose oxidation in diabetes according to the Randle mechanism (33) remains a central postulate explaining some negative consequences of substrate shift in this metabolic disorder. However, it is now increasingly appreciated that hyperglycemia per se can trigger cellular damage, independently from FA utilization (11). The multiple mechanisms throug...

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Section: Methodsmentioning

confidence: 99%

Restoring redox balance enhances contractility in heart trabeculae from type 2 diabetic rats exposed to high glucose

Bhatt

Aon

Tocchetti

et al. 2015

American Journal of Physiology-Heart and Circulatory Physiology

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“…Trpm2 may provide the necessary Ca 2ϩ for mitochondrial bioenergetics in the heart. In this context, it is important to note that low levels of H 2 O 2 emission, which occurs in normal cardiac mitochondria (40), may tonically activate Trpm2 channels, thereby realizing a virtuous cycle of bioenergetics maintenance in WT but not KO myocytes (Fig. 10A).…”

Section: Thloredoxin 2 Expressionmentioning

confidence: 99%

Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

Hoffman

Miller

Wang

et al. 2015

American Journal of Physiology-Heart and Circulatory Physiology

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Five days post-injection, gKO-GFP heart slices had higher reactive oxygen species (ROS) levels but lower oxygen consumption rate (OCR) than WT-GFP heart slices. Trpm2 but not E960D decreased ROS and restored OCR in gKO hearts back to normal levels. In gKO myocytes expressing Trpm2 or its mutants, Trpm2 but not E960D reduced the elevated mitochondrial superoxide (O 2 .Ϫ ) levels in gKO myocytes. After hypoxia-reoxygenation (H/R), Trpm2 but not E906D or P1018L (inactivates Trpm2 current) lowered O 2 .Ϫ levels in gKO myocytes and only in the presence of extracellular Ca 2ϩ , indicating sustained Ca 2ϩ entry is necessary for Trpm2-mediated preservation of mitochondrial function. After ischemic-reperfusion (I/R), cardiac-specific Trpm2 KO hearts exhibited lower maximal first time derivative of LV pressure rise (ϩdP/dt) than WT hearts in vivo. After doxorubicin treatment, Trpm2 KO mice had worse survival and lower ϩdP/dt. We conclude 1) cardiac Trpm2-mediated Ca 2ϩ influx is necessary to maintain mitochondrial function and protect against H/R injury; 2) Ca 2ϩ influx via cardiac Trpm2 confers protection against H/R and I/R injury by reducing mitochondrial oxidants; and 3) Trpm2 confers protection in doxorubicin cardiomyopathy. ischemic cardiomyopathy; doxorubicin cardiomyopathy; voltage-independent Ca 2ϩ channels; cardiac Trpm2 currents; mitochondrial superoxide; hypoxia-reoxygenation IN MAMMALIAN CELLS, THE TRANSIENT receptor potential (Trp) protein superfamily is a diverse group of voltage-independent, cation-permeable channels organized into six subfamilies based on amino acid sequence homology (9, 30). Monomeric Trp proteins have six putative transmembrane (TM) domains and intracellular NH 2 and COOH termini. To form a functional channel, Trp proteins assemble into either homo-or heterotetramers, with the putative pore formed by loops between the fifth and sixth TM domains (21, 23). The Trp-melastatin (Trpm) subfamily consists of eight mammalian members (Trpm1-Trpm8)(30), of which Trpm2 (27), -m4, -m5, and -m6 (49) are expressed in the heart. Although Trpm4 is associated with conduction abnormalities and cardiac arrhythmias (1, 36), there is little information on the physiological and pathophysiological function of Trpm2 in the heart.

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“…The supernatant was collected and prepared in a master mix (5 mM DTNB, 8% ethanol, and 300 M NADPH) and PE buffer (100 mM potassium phosphate and 2 mM EDTA, pH 7.0). TrxR2 activity was measured by the reduction of 5,5-dithio-bis (2-nitrobenzoic acid) (DTNB) by TrxR2 and NADPH at an absorbance of 412 nm (60).…”

Section: Mitochondria Were Isolated From Wt and Erbb2mentioning

confidence: 99%

ErbB2 overexpression upregulates antioxidant enzymes, reduces basal levels of reactive oxygen species, and protects against doxorubicin cardiotoxicity

Belmonte

Das

Sysa‐Shah

et al. 2015

American Journal of Physiology-Heart and Circulatory Physiology

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bergen C, Gabrielson K. ErbB2 overexpression upregulates antioxidant enzymes, reduces basal levels of reactive oxygen species, and protects against doxorubicin cardiotoxicity. Am J Physiol Heart Circ Physiol 309: H1271-H1280, 2015. First published August 14, 2015; doi:10.1152/ajpheart.00517.2014.-Levels of the HER2/ErbB2 protein in the heart are upregulated in some women during breast cancer therapy, and these women are at high risk for developing heart dysfunction after sequential treatment with anti-ErbB2/trastuzumab or doxorubicin. Doxorubicin is known to increase oxidative stress in the heart, and thus we considered the possibility that ErbB2 protein influences the status of cardiac antioxidant defenses in cardiomyocytes. In this study, we measured reactive oxygen species (ROS) in cardiac mitochondria and whole hearts from mice with cardiacspecific overexpression of ErbB2 (ErbB2 tg ) and found that, compared with control mice, high levels of ErbB2 in myocardium result in lower levels of ROS in mitochondria (P ϭ 0.0075) and whole hearts (P ϭ 0.0381). Neonatal cardiomyocytes isolated from ErbB2 tg hearts have lower ROS levels and less cellular death (P Ͻ 0.0001) following doxorubicin treatment. Analyzing antioxidant enzyme levels and activities, we found that ErbB2 tg hearts have increased levels of glutathione peroxidase 1 (GPx1) protein (P Ͻ 0.0001) and GPx activity (P ϭ 0.0031) in addition to increased levels of two known GPx activators, c-Abl (P ϭ 0.0284) and Arg (P Ͻ 0.0001). Interestingly, although mitochondrial ROS emission is reduced in the ErbB2 tg hearts, oxygen consumption rates and complex I activity are similar to control littermates. Compared with these in vivo studies, H9c2 cells transfected with ErbB2 showed less cellular toxicity and produced less ROS (P Ͻ 0.0001) after doxorubicin treatment but upregulated GR activity (P ϭ 0.0237) instead of GPx. Our study shows that ErbB2-dependent signaling contributes to antioxidant defenses and suggests a novel mechanism by which anticancer therapies involving ErbB2 antagonists can harm myocardial structure and function.ErbB2; glutathione peroxidase; reactive oxygen species; nRTK; mitochondria NEW & NOTEWORTHY This is the first study on ErbB2 overexpression in the heart that identifies cardioprotection after doxorubicin treatment. Our findings suggest that an adverse effect on redox signaling pathways necessary for maintaining mitochondrial antioxidant defenses may be an important mechanism of cardiac toxicity induced by drugs that target ErbB2 and Abl kinases.THE ERBB2 RECEPTOR TYROSINE kinase is a member of the epidermal growth factor receptor (EGFR) family and has an essential role in cardiac function and development (9,39,47,50). ErbB2 is also important in cancer biology, and dysregulated ErbB2 signaling is implicated in various types of cancer, most notably in breast and ovarian cancer (30, 49). The clinical link between ErbB2 and heart function was discovered when 27% of ErbB2-overexpressing breast cancer patients treated with doxorubicin and anti-...

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Thioredoxin Reductase-2 Is Essential for Keeping Low Levels of H2O2 Emission from Isolated Heart Mitochondria

Cited by 160 publications

References 56 publications

Restoring redox balance enhances contractility in heart trabeculae from type 2 diabetic rats exposed to high glucose

Restoring redox balance enhances contractility in heart trabeculae from type 2 diabetic rats exposed to high glucose

Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

ErbB2 overexpression upregulates antioxidant enzymes, reduces basal levels of reactive oxygen species, and protects against doxorubicin cardiotoxicity

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Thioredoxin Reductase-2 Is Essential for Keeping Low Levels of H2O2 Emission from Isolated Heart Mitochondria

Cited by 160 publications

References 56 publications

Restoring redox balance enhances contractility in heart trabeculae from type 2 diabetic rats exposed to high glucose

Restoring redox balance enhances contractility in heart trabeculae from type 2 diabetic rats exposed to high glucose

Ca2+entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance

ErbB2 overexpression upregulates antioxidant enzymes, reduces basal levels of reactive oxygen species, and protects against doxorubicin cardiotoxicity

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Ca²⁺entry via Trpm2 is essential for cardiac myocyte bioenergetics maintenance