Embryonic stem cell-derived motor neurons form neuromuscular junctions in vitro and enhance motor functional recovery in vivo.

Abstract

Transplantation of embryonic stem cell-derived motor neurons may support the biological integrity of denervated muscle by forming new neuromuscular junctions and up-regulating specific growth factors. The authors examined the functional properties of embryonic stem cell-derived motor neurons in vitro and the effect of these cells transplanted in vivo. Murine GFP/HB9 embryonic stem cells were differentiated into motor neurons. Co-cultures of motor neurons and myotubes were prepared to confirm the formation of neuromuscular junctions with synaptic markers. Athymic mice (n = 59) were assigned randomly to one of three experimental groups. A tibial nerve transection was performed without nerve repair, and motor neurons were transplanted into the gastrocnemius muscles immediately after transection (n = 24) or 3 weeks after denervation (n = 24). Quantitative and histologic assessments of gastrocnemius muscle were performed at days 7 and 21 after cell transplantation. Additional experimental groups (n = 11), where the tibial nerve underwent repair after transplantation, were formed. The effect of the transplants on motor recovery following nerve repair was investigated. Co-culture experiments showed the formation of neuromuscular junctions. In the experiment with nerve transection without nerve repair, the muscles transplanted with motor neurons were less atrophied than control phosphate-buffered saline-injected muscles at days 7 and 21. Those muscles receiving cells transplanted 3 weeks after denervation were not preserved. The motor recovery after nerve repair with cell transplantation was significantly enhanced compared with the control group. Transplantation of motor neurons prevented denervation atrophy but was not capable of rescuing already atrophied muscle. After nerve repair, motor neuron transplantation improved functional recovery.