Differential alternative splicing in skeletal muscle after 14 days immobilization-induced atrophy in young men

Abstract

Skeletal muscle disuse during injury decreases muscle mass and function. Previous studies have determined differential gene expression associated with atrophy, but changes in alternative splicing (AS) in response to unloading is poorly defined. This study aimed to identify global changes in differential alternative splicing (DAS) in skeletal muscle following unloading. Twenty-one healthy males (26.0 ± 5.7 years; 81.2 ± 11.6 kg) underwent 14 days unilateral knee-brace immobilization. Participants underwent strength and muscle mass assessment before and after the immobilization and vastus lateralis muscle biopsies were taken pre-immobilization, and on days 3 and 14. RNA-sequencing was performed, and DAS events were identified using the rMATS package. Leg strength decreased − 21.5 kg (P < 0.0001) while mid-thigh circumference was reduced ~ 2% (P = 0.016). A total of 157,464 and 159,395 splicing events affecting 15,219 and 15,374 genes identified on day 3 and 14, respectively. Most DAS events occurred in coding sequence regions and were primarily due to exon skipping. A higher number of DAS events were observed by day 14 than day 3, indicating time-dependent regulation. Gene Ontology analysis of DAS at day 14 revealed 37 enriched GO biological processes related to MAPK signalling, mitochondrial function, and organelle organization. Unloading-induced atrophy was associated with widespread changes in AS, particularly in genes linked to muscle maintenance and protein synthesis. AS appears to promote down-regulation of target genes and subsequent differentially enriched biological pathways associated with the functional outcomes of decreased muscle mass and strength. These findings indicate changes in alternative splicing is a regulatory mechanism in muscle wasting during immobilization.