Low-dose simulated galactic cosmic radiation exposure after hindlimb unloading has long-term impact on skeletal muscle metabolic proteins in female mice.

Spaceflight stressors, including microgravity-induced unloading and galactic cosmic radiation (GCR), acutely disrupt mitochondrial function and contribute to skeletal muscle atrophy. The long-term remodeling of skeletal muscle following combined unloading and radiation exposure remains poorly understood. We investigated protein abundance changes 9-mo postexposure to combined unloading and radiation exposure. Female, 6-mo-old, C57Bl/6J mice underwent 5 days of hindlimb unloading (HU) or weight-bearing (WB) conditions, followed by 0, 0.5, or 1.5 Gy of simulated GCR exposure using the simplified 5-ion beam exposure (simGCRsim) (n = 5/group). The gastrocnemius muscle was collected after 9 mo of WB and analyzed by data-independent acquisition mass spectrometry. Differentially abundant proteins were identified and evaluated using pathway enrichment analyses. WB mice exposed to 0.5 Gy exhibited increased abundance of electron transport system proteins and mitochondrial transport proteins, suggesting increased mitochondrial activity relative to control mice. HU mice exposed to 0.5 Gy displayed decreased glycolytic proteins, increased reliance on oxidative pathways, and reduced antioxidant proteins (glutaredoxins and peroxiredoxin) compared with WB0.5. In HU mice, a higher radiation dose (HU1.5 vs. HU0.5) led to the downregulation of 26S proteasome subunits and the upregulation of peroxisomal antioxidant, tricarboxylic acid cycle, and β-oxidation proteins, indicating dose-dependent mitochondrial adaptations. Long-term muscular remodeling after simGCRsim exposure is influenced by both muscle-loading status and radiation dose, with prolonged shifts toward oxidative metabolism and altered protein quality control persisting months after exposure. These findings provide new insights into skeletal muscle adaptation to spaceflight stressors and have important implications for astronaut health during and after long-duration missions.NEW & NOTEWORTHY Acute simGCRsim exposure causes changes in metabolic and mitochondrial protein abundance that are observable after 9 mo. Weight-bearing and unloaded muscle show unique metabolic protein profiles, with unloaded muscle experiencing reduced expression of glycolytic proteins relative to weight-bearing muscle after 9 mo. Significant dose-dependent response to acute simGCRsim exposure, with higher doses having more profound effects on metabolic and antioxidant protein abundance.