Motor Neuron Progenitor Cells Prevent Early Atrophy of Denervated Muscle in a Syngeneic Model

A major obstacle to functional recovery following peripheral nerve injury is the time-dependent loss of the ability of muscle fibers to successfully receive reinnervation and regain function. The authors have previously shown that the implantation of motor neuron progenitor (MNP) cells, derived from murine embryonic (ES) stem cells, into denervated muscle, can prevent muscle atrophy and preserve structure in an in vivo adult rat model for 7 days. This study investigated the potential of MNP cell transplants to preserve muscle mass and architecture for up to 21 days using a syngeneic transplant model of peripheral nerve injury.

SVJ-129 ES cells were differentiated using retinoic acid and sonic hedgehog peptide to induce cholinergic MNP cells. Fluorescent-labeled MNP cells were injected into the gastrocnemius muscle of strain SVJ-129 mice following denervation by ipsilateral sciatic nerve transection. Control mice were subjected to identical unilateral sciatic nerve injuries, but received injection of phosphate buffered saline (PBD) solution only. Gastrocnemius muscles were weighed and analyzed at 7 and 21 days. Frozen sections of the muscle were stained for alpha-bungarotoxin, HB-9 and NF-68. Muscles were also imaged in vivo at 7, 14, and 21 days post transplant using confocal microscopy.

Seven days after ES-MNP transplant, little or no atrophy of the denervated muscle had occurred, as evidenced by 90% conservation of muscle mass. By 21 days, the muscle atrophy was noted to be 52%. In contrast, control muscles had atrophied significantly after 7 and 21 days, with muscle mass reduced to 70% and 46%, respectively. Confocal microscopy showed that the MNP cells had dispersed throughout the surface of the muscle. By days 14 and 21, the implanted cells had aligned themselves along muscle fibers and had penetrated within the tissue. The presence of the cells was confirmed by immunofluorescence.

This study demonstrated that neuronal precursors derived from embryonic stem cells can prevent muscle atrophy after denervation. The cells can survive up to day 21 post transplant. MNP transplants may improve clinical outcome by protecting denervated muscle from irreversible atrophy and by maintaining muscle responsiveness to eventual reinnervation by regenerating host motor neurons.