Reinnervation of denervated muscle by implantation of nerve‐muscle‐endplate band graft to the native motor zone of the target muscle

Abstract

IntroductionMotor endplate reinnervation is critical for restoring motor function of the denervated muscle. We developed a novel surgical technique called nerve‐muscle‐endplate band grafting (NMEG) for muscle reinnervation.MethodsExperimentally denervated sternomastoid muscle in the rat was reinnervated by transferring a NMEG from the ipsilateral sternohyoid muscle to the native motor zone (NMZ) of the target muscle. A NMEG pedicle contained a block of muscle (~ 6 × 6 × 3 mm), a nerve branch with axon terminals, and a motor endplate band with numerous neuromuscular junctions. At 3 months after surgery, maximal tetanic muscle force measurement, muscle mass and myofiber morphology, motoneurons, regenerated axons, and axon‐endplate connections of the muscles were analyzed.ResultsThe mean force of the reinnervated muscles was 82% of the contralateral controls. The average weight of the treated muscles was 89% of the controls. The reinnervated muscles exhibited extensive axonal regeneration. Specifically, the mean count of the regenerated axons in the reinnervated muscles reached up to 76.8% of the controls. The majority (80%) of the denervated endplates in the target muscle regained motor innervation.ConclusionsThe NMZ of the denervated muscle is an ideal site for NMEG implantation and for the development of new microsurgical and therapeutic strategies to achieve sufficient axonal regeneration, rapid endplate reinnervation, and optimal functional recovery. NMEG‐NMZ technique may become a useful tool in the treatment of muscle paralysis caused by peripheral nerve injuries in certain clinical situations.

Nerve growth factor and basic fibroblast growth factor promote reinnervation by nerve−muscle−endplate grafting

“…(C) Average maximal force of the SM muscles treated with NMEG−NMZ and factors was 91% of the controls (left bar). The right dotted bar illustrates average percent rate of maximal force of the SM muscles treated with NMEG−NMZ alone (82% of the control), as demonstrated in our previous study …”

“…Immediately after muscle denervation, the rats in the reinnervation group underwent NMEG−NMZ surgery and neurotrophic factor therapy. The details regarding NMEG−NMZ transplantation have been described previously . First, the NMZs in the right SM and SH muscles were outlined according to the motor point and location of intramuscular nerve terminals and MEP band (Fig.…”

“…Some sagittal sections were immunostained with monoclonal antibody SMI‐31 as a marker for all axons (Covance Research Products, Berkeley, California) as described elsewhere . Briefly, the sections were treated in phosphate‐buffered saline (PBS) containing 0.3% Triton and 2% bovine serum albumin (BSA) for 30 minutes, incubated with SMI‐31 primary antibody (1:800) in PBS containing 0.03% Triton at 4 °C overnight, treated for 2 hours with anti‐mouse biotinylated secondary antibody (1:1,000; Vector Labs, Burlingame, California), processed with the avidin−biotin complex method using a VectaStain ABC kit (1:1,000; ABC Elite; Vector), and treated with diaminobenzidine−nickel as chromogen to visualize peroxidase labeling.…”

“…The stained sections were examined under a fluorescence microscope (Axioplan‐1; Carl Zeiss, Gottingen, Germany) and photographed with a USB 3.0 digital microscope camera (Infinity 3‐3URC; Lumenera Corp., Ottawa, Ontario, Canada). The extent of axonal regeneration and muscle reinnervation was evaluated by quantifying the neurofilament‐immunoreactive (NF‐ir) axons within the treated muscles as described in our previous publications . The intramuscular axonal density was assessed by computing the number of the NF‐ir axons and the area fraction of the axons within a section area (1.0 mm 2 ).…”

“…MEPs and their innervating axons were labeled with double‐fluorescence staining, as described previously, with some modifications. Briefly, some sagittal sections were fixed for 20 minutes in Zamboni fixative with 5% sucrose at 4 °C.…”