Regeneration-associated utrophin expression in skeletal muscle: implications for utrophin-targeted therapies

Duchenne muscular dystrophy (DMD) is a lethal, X-linked muscle-wasting disease caused by loss of dystrophin. Utrophin, a structural and functional paralogue of dystrophin, can compensate for dystrophin deficiency and represents a therapeutic target applicable to patients irrespective of their DMD mutation. However, utrophin is also naturally upregulated at the sarcolemma of regenerating myofibres, creating an important confounding factor when utrophin levels are used to evaluate therapeutic activity. Here, we defined the temporal relationship between utrophin expression and complementary markers of skeletal muscle regeneration in wild-type and dystrophic mdx muscle following cardiotoxin-induced injury. Utrophin protein and mRNA dynamics were assessed together with dystrophin, embryonic myosin (MyHC-emb), centronucleation, cross-sectional area (CSA), and minimal Ferets diameter (MinFeret). MyHC-emb and utrophin were strongly associated with the early regenerative phase, although MyHC-emb peaked before maximal utrophin protein expression. In contrast, CSA and MinFeret showed an inverse temporal relationship with utrophin, while centronucleation persisted after active regeneration and did not discriminate early from late regenerative stages. Utrophin also remained elevated at the dystrophic sarcolemma for longer than in wild-type muscle. These findings demonstrate that utrophin quantification cannot be interpreted independently of the regenerative state of muscle. Combining MyHC-emb with morphological indices such as CSA and MinFeret provides a complementary framework for contextualizing regeneration-associated utrophin and should improve interpretation of utrophin changes in preclinical evaluation of utrophin-targeted therapies.