Transient Hyperoxic Reoxygenation Reduces Cytochrome c Oxidase Activity by Increasing Superoxide Dismutase and Nitric Oxide

Abstract

Oxygen therapies have been shown to be cytoprotective in a dose-dependent fashion. Previously, we have characterized the protective effects of moderate hyperoxia on cell viability of ischemic human cardiomyocytes and their mitochondrial membrane potential by transient addition of oxygenated perfluorocarbons to the cell medium. Now, we report that the activity and expression of cytochrome c oxidase (COX) after prolonged ischemia depend on the amount of oxygen delivered during reoxygenation. Transient hyperoxia during reoxygenation results in a decrease of COX activity by 62 ؎ 15% and COX expression by 67 ؎ 5%, when hyperoxic tensions of ≈300 mm Hg are reached in the cell medium. This decrease in COX expression is prevented by the inhibition of inducible nitric-oxide synthase (iNOS). Immunoblot analysis of ischemic human cardiomyocytes revealed that hyperoxic reoxygenation causes a 2-fold increase of iNOS, leading to a rise in nitric oxide production by 140 ؎ 45%. Hyperoxic reoxygenation is further responsible for a 2-fold activation of hydrogen peroxide production and an increase in cytosolic superoxide dismutase expression by 35 ؎ 10%. NADPH availability has no effect on the hyperoxia-induced decrease of superoxide. Overall, these results indicate that transient hyperoxic reoxygenation in optimal concentrations increases the level of nitric oxide by activation of iNOS and superoxide dismutase, thereby inducing respiration arrest in mitochondria of ischemic cardiomyocytes.Oxygen therapies have been available for more than 100 years (1) and recently have been shown to be neuroprotective in a dose-dependent fashion (2). One mode of hyperoxic treatment is applying oxygenated perfluorocarbons (PFC) 2 locally to tissues (3). Previously, we have characterized the effects of different hyperoxic conditions on cell viability of ischemic human cardiomyocytes and their mitochondria by the addition of oxygenated PFC to the cell medium (see Fig. 1). Mitochondria have a key function in cardiomyocyte survival after ischemia and reperfusion because mitochondrial respiration produces more than 90% of the heart's energy by oxidative phosphorylation (4).Conversely, oxidative phosphorylation is a central site of reactive oxygen species production in the heart (5-7). Oxidative phosphorylation along with NADPH oxidase form the major sources of superoxide in myocytes (8) that may induce irreversible cellular damage and cell death (9). Nitric oxide (NO) can inhibit oxidative phosphorylation reversibly by blocking cytochrome c oxidase (COX) of the electron transport chain (ETC) (10). In this way, NO protects myocardial tissue from ischemia and reperfusion injury (11) possibly by defending the mitochondrion to maintain their membrane potential (12) and reducing cytotoxicity of reactive oxygen species. Sandau et al. (13) have shown that NO production by inducible nitric-oxide synthase (iNOS) can mimic a hypoxic response under normoxia. We have shown previously that hypoxia-inducible factors 1␣ and 2␣ are stabilized after hyperoxic reoxyge...

Proteomic analysis of mitochondrial proteins in the guinea pig heart following long-term normobaric hyperoxia

“…Increased formation of reactive oxygen (ROS) and nitrogen (RNS) species leads to changes in intracellular redox state and modulation of cell signaling pathways, resulting in alterations in gene expression. Data from both experimental and clinical studies suggest that beneficial effects of NBO or HBO treatment on brain or heart could be due to increased expression of antioxidant, anti-inflammatory, and anti-apoptotic proteins and reduced production of pro-oxidant, pro-inflammatory pro-apoptotic proteins [ [13][14][15], for review see [2]. However, this is in contrast with studies, which showed that hyperoxia increases cellular oxidative damage [16], for review see [11].…”

Paradoxical decrease in isoprostane and increase in superoxide dismutase following CPAP withdrawal in OSA

“…The last point is about the pathophysiological mechanism. It has been well-demonstrated in many cell models, animal studies and human diseases that SOD is activated under hyperoxic conditions rather than hypoxia, to counteract superoxide production [8,9]. Nevertheless, the hypothesis that intermittent hypoxic preconditioning may protect from subsequent ischaemia by increasing antioxidant defence cannot be ruled out.…”

STEMI therapy: Normoxic reperfusion is not good enough

“…Buja et al, for example, pointed out in a recent review paper that although reperfusion therapy in STEMI should not be unduly delayed, adjunctive strategies aiming at eliminating or at least retarding lethal reperfusion injury are required [5]. We and others recently found that cell death after prolonged ischemia of isolated human cardiomyocytes is successfully prevented via a suppression of oxygen free radical formation by applying dissolved molecular oxygen as an “add‐on therapy” during normoxic reoxygenation [7–9]. Moderate hyperoxia during reoxygenation stabilizes hypoxia‐inducible factor 1α, up‐regulates nitric oxide synthase and increases the cellular defence efforts against lethal reperfusion injury via an augmented superoxide dismutase production [7, 8].…”

“…We and others recently found that cell death after prolonged ischemia of isolated human cardiomyocytes is successfully prevented via a suppression of oxygen free radical formation by applying dissolved molecular oxygen as an “add‐on therapy” during normoxic reoxygenation [7–9]. Moderate hyperoxia during reoxygenation stabilizes hypoxia‐inducible factor 1α, up‐regulates nitric oxide synthase and increases the cellular defence efforts against lethal reperfusion injury via an augmented superoxide dismutase production [7, 8]. The proof of concept of hyperoxic postconditioning in myocardial infarction was recently provided by the AMIHOT clinical trials [10].…”