PMID- 27243905 OWN - NLM STAT- MEDLINE DCOM- 20170718 LR - 20220321 IS - 1932-6203 (Electronic) IS - 1932-6203 (Linking) VI - 11 IP - 5 DP - 2016 TI - MitoNEET Protects HL-1 Cardiomyocytes from Oxidative Stress Mediated Apoptosis in an In Vitro Model of Hypoxia and Reoxygenation. PG - e0156054 LID - 10.1371/journal.pone.0156054 [doi] LID - e0156054 AB - The iron-sulfur cluster containing protein mitoNEET is known to modulate the oxidative capacity of cardiac mitochondria but its function during myocardial reperfusion injury after transient ischemia is unknown. The purpose of this study was to analyze the impact of mitoNEET on oxidative stress induced cell death and its relation to the glutathione-redox system in cardiomyocytes in an in vitro model of hypoxia and reoxygenation (H/R). Our results show that siRNA knockdown (KD) of mitoNEET caused an 1.9-fold increase in H/R induced apoptosis compared to H/R control while overexpression of mitoNEET caused a 53% decrease in apoptosis. Necrosis was not affected. Apoptosis of both, mitoNEET-KD and control cells was diminished to comparable levels by using the antioxidants Tiron and glutathione compound glutathione reduced ethyl ester (GSH-MEE), indicating that mitoNEET-dependent apoptosis is mediated by oxidative stress. The interplay between mitoNEET and glutathione redox system was assessed by treating cardiomyocytes with 2-acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanylthio-carbonylamino) phenylthiocarbamoylsulfanyl] propionic acid (2-AAPA), known to effectively inhibit glutathione reductase (GSR) and to decrease the GSH/GSSG ratio. Surprisingly, inhibition of GSR-activity to 20% by 2-AAPA decreased apoptosis of control and mitoNEET-KD cells to 23% and 25% respectively, while at the same time mitoNEET-protein was increased 4-fold. This effect on mitoNEET-protein was not accessible by mitoNEET-KD but was reversed by GSH-MEE. In conclusion we show that mitoNEET protects cardiomyocytes from oxidative stress-induced apoptosis during H/R. Inhibition of GSH-recycling, GSR-activity by 2-AAPA increased mitoNEET-protein, accompanied by reduced apoptosis. Addition of GSH reversed these effects suggesting that mitoNEET can in part compensate for imbalances in the antioxidative glutathione-system and therefore could serve as a potential therapeutic approach for the oxidatively stressed myocardium. FAU - Habener, Anika AU - Habener A AD - Department of Anesthesiology and Intensive Care Medicine, Hannover Medical School, Hannover, Germany. AD - Department of Pediatric Pneumology, Allergology and Neonatology, Hannover Medical School, Hannover, Germany. FAU - Chowdhury, Arpita AU - Chowdhury A AD - Department of Anesthesiology and Intensive Care Medicine, Hannover Medical School, Hannover, Germany. AD - Department of Cellular Biochemistry, University Medical Center Gottingen, Gottingen, Germany. FAU - Echtermeyer, Frank AU - Echtermeyer F AD - Department of Anesthesiology and Intensive Care Medicine, Hannover Medical School, Hannover, Germany. FAU - Lichtinghagen, Ralf AU - Lichtinghagen R AD - Institute for Clinical Chemistry, Hannover Medical School, Hannover, Germany. FAU - Theilmeier, Gregor AU - Theilmeier G AD - Department of Anesthesiology and Intensive Care Medicine, Hannover Medical School, Hannover, Germany. AD - Department of Health Services Sciences, Faculty of Medicine and Health Sciences, University of Oldenburg, Oldenburg, Germany. FAU - Herzog, Christine AU - Herzog C AD - Department of Anesthesiology and Intensive Care Medicine, Hannover Medical School, Hannover, Germany. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20160531 PL - United States TA - PLoS One JT - PloS one JID - 101285081 RN - 0 (2-acetylamino-3-(4-(2-acetylamino-2-carboxyethylsulfanylthiocarbonylamino)phenylthiocarbamoylsulfanyl)propionic acid) RN - 0 (Antioxidants) RN - 0 (Iron-Binding Proteins) RN - 0 (Membrane Proteins) RN - 0 (RNA, Small Interfering) RN - 0 (Reactive Oxygen Species) RN - 0 (Thiocarbamates) RN - 0 (mitoNEET protein, mouse) RN - 4X87R5T106 (1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt) RN - 7W95D60F4J (S-ethyl glutathione) RN - EC 1.8.1.7 (Glutathione Reductase) RN - GAN16C9B8O (Glutathione) RN - WYQ7N0BPYC (Acetylcysteine) SB - IM MH - 1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt/pharmacology MH - Acetylcysteine/analogs & derivatives/pharmacology MH - Animals MH - Antioxidants/pharmacology MH - Apoptosis/drug effects/*genetics MH - Cell Hypoxia/drug effects/*genetics MH - Cell Line MH - Glutathione/analogs & derivatives/pharmacology MH - Glutathione Reductase/antagonists & inhibitors/metabolism MH - Iron-Binding Proteins/*genetics MH - Membrane Proteins/*genetics MH - Mice MH - Myocytes, Cardiac/*metabolism MH - Oxidation-Reduction MH - Oxidative Stress/drug effects/*genetics MH - RNA Interference MH - RNA, Small Interfering/genetics MH - Reactive Oxygen Species/metabolism MH - Reperfusion MH - Thiocarbamates/pharmacology PMC - PMC4887087 COIS- Competing Interests: The authors have declared that no competing interests exist. EDAT- 2016/06/01 06:00 MHDA- 2017/07/19 06:00 PMCR- 2016/05/31 CRDT- 2016/06/01 06:00 PHST- 2016/02/01 00:00 [received] PHST- 2016/05/09 00:00 [accepted] PHST- 2016/06/01 06:00 [entrez] PHST- 2016/06/01 06:00 [pubmed] PHST- 2017/07/19 06:00 [medline] PHST- 2016/05/31 00:00 [pmc-release] AID - PONE-D-16-04471 [pii] AID - 10.1371/journal.pone.0156054 [doi] PST - epublish SO - PLoS One. 2016 May 31;11(5):e0156054. doi: 10.1371/journal.pone.0156054. eCollection 2016.