Redox remodeling is pivotal in murine diaphragm muscle adaptation to chronic sustained hypoxia

Mechanisms underpinning chronic sustained hypoxia (CH)-induced structural and functional adaptations in respiratory muscles are unclear despite the clinical relevance to respiratory diseases. The objectives of the present study were to thoroughly assess the putative role of CH-induced redox remodeling in murine diaphragm muscle over time and the subsequent effects on metabolic enzyme activities, catabolic signaling and catabolic processes, and diaphragm muscle contractile function. C57Bl6/J mice were exposed to normoxia or normobaric CH (fraction of inspired oxygen = 0.1) for 1, 3, or 6 weeks. A second cohort was exposed to CH for 6 weeks with and without antioxidant supplementation (tempol or N-acetyl cysteine). After CH exposure, we performed two-dimensional redox proteomics with mass spectrometry, enzyme activity assays, and cellsignaling assays on diaphragm homogenates. We also assessed diaphragm isotonic contractile and endurance properties ex vivo. Global protein redox changes in the diaphragm after CH are indicative of oxidation. Remodeling of proteins key to contractile, metabolic, and homeostatic functions was observed. Several oxidative and glycolytic enzyme activities were decreased by CH. Redoxsensitive chymotrypsin-like proteasome activity of the diaphragm was increased. CH decreased phospho-forkhead box O3a (FOXO3a) and phospho-mammalian target of rapamycin content. Hypoxiainducible factor-1a and phospho-p38 mitogen-activated protein kinase content was increased in CH diaphragm, and this was attenuated by antioxidant treatment. CH exposure decreased forceand power-generating capacity of the diaphragm, and this was prevented by antioxidant supplementation with N-acetyl cysteine but not tempol. Redox remodeling is pivotal for diaphragm adaptation to CH, affecting metabolic activity, atrophy signaling, and functional performance. Antioxidant supplementation may be useful as an adjunctive therapy in respiratory-related diseases characterized by hypoxic stress.