Intermediate filament (IF) proteins show specific spatial and temporal expression during development of skeletal muscle. Nestin, the least known muscle IF, has an important role in neuronal regeneration. Therefore, we analyzed the expression pattern of nestin as related to that of vimentin and desmin during skeletal muscle regeneration. Nestin and vimentin appear at 6 h post-injury in myoblasts, with maximum expression around day 3-5 post-injury. Thereafter, vimentin expression ceases completely, whereas that of nestin is downregulated to remain only in the sarcoplasm next to neuromuscular and myotendinous junctions. Desmin appears at 6-12 h post-injury and becomes the predominant IF in myofibers simultaneously with the appearance of cross-striations. The expression pattern and colocalization of nestin and vimentin, known to form heteropolymers, suggests that they are essential during the early dynamic phase of the myofiber regeneration when migration, fusion, and structural modeling of myogenic cells occurs, whereas desmin is responsible for keeping myofibrils in register in mature myofibers. In conclusion, the expression of nestin is dynamically orchestrated with that of vimentin and desmin during skeletal muscle regeneration and recapitulates that seen during myogenesis, i.e. these IFs have key functional roles in the construction and restoration of skeletal myofibers.

The intermediate filament proteins nestin, vimentin, and desmin show a specific temporal expression pattern during the development of myofibers from myogenic precursor cells. Nestin and vimentin are actively expressed during early developmental stages to be later down-regulated, vimentin completely and nestin to minimal levels, whereas desmin expression begins later and is maintained in mature myofibers, in which desmin participates in maintaining structural integrity. In this study we have analyzed the expression levels and distribution pattern of nestin in intact and denervated muscle in rat and in human. Nestin immunoreactivity was specifically and focally localized in the sarcoplasm underneath neuromuscular junctions (NMJs) and in the vicinity of the myotendinous junctions (MTJs), ie, in regions associated with acetylcholine receptors (AChRs). This association prompted us to analyze nestin in neurogenically and myogenically denervated muscle. Immunoblot analysis disclosed a marked overall increase of accumulated nestin protein. Similar to the extrajunctional redistribution of AChRs in denervated myofibers, nestin immunoreactivity extended widely beyond the NMJ region. Re-innervation caused complete reversion of these changes. Our study demonstrates that the expression levels and distribution pattern of nestin are regulated by innervation, ie, signal transduction into myofibers.

Experimental myonecrosis--induced by injection of notexin into rat tibialis anterior muscle--and subsequent regeneration were studied from 1 h to 20 days postinjury with magnetic resonance imaging using conventional and magnetization transfer sequences, and these findings were correlated with histopathology. MR images revealed necrosis within 1 h postinjection. Histopathologically, necrotized fibers enlarged and intercellular spaces widened, indicating intracellular and extracellular edema, which began to decrease after 48 h, whereafter the formation of new myofibers predominated. T2 increased progressively until 7.5 h, while T1 increased until 24 h. Magnetization transfer contrast (MTC) and magnetization transfer rate (Rwm) decreased rapidly postinjection; the decrease in Rwm lasted longer than in MTC (96 h versus 48 h, respectively). Spin echo, inversion recovery and magnetization transfer sequences revealed the lesions equally effectively. MR images and relaxation parameters reflect well the extent of histopathological injury and edema in the acute phase, whereas specific tissue changes in the regenerative phase were not detectable by MRI. MT imaging and especially magnetization transfer rate are as sensitive as conventional T2 contrast to alterations in water imbalance.