Ultrasound stimulation of hindlimb skeletal muscle is associated with selective differences in splenic metabolites during hindlimb unloading in mice

Physical inactivity and unloading induce skeletal muscle atrophy and systemic metabolic and immune dysregulation. We examined associations of skeletal muscle ultrasound (US) stimulation with splenic metabolite profiles and explored plasma extracellular particle (EP) characteristics in a hindlimb unloading (HU) model. Male C57BL/6J mice were assigned to control, HU, or HU with US stimulation (HUUS). Blood, gastrocnemius, and spleen were collected for particle analysis, transcriptomics, histology, and metabolomics. HU and HUUS had lower body weight, gastrocnemius weight, spleen weight, and muscle fiber cross-sectional area than control. Exploratory analysis showed numerically higher particle concentrations in polymer-precipitated plasma EP-containing fractions in HUUS than in HU, although the difference was not statistically significant ( p = 0.050). In gastrocnemius muscle, gene sets for endoplasmic reticulum protein processing and proteostasis were positively enriched in HUUS versus HU. In the spleen, HU showed nominally higher upstream glycolytic intermediates and nominally lower flavin adenine dinucleotide (FAD), whereas erythrose 4-phosphate (E4P) and GDP were lower and cis-aconitate was higher after FDR correction. Compared with HU, HUUS showed lower glucose 1-phosphate (G1P) after FDR correction and nominal differences in F6P, fumarate, and FAD. Overall, US stimulation under HU conditions was associated with pathway-level transcriptional remodeling and selective splenic metabolite differences.