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Directing The Growth Of Regenerating Motor Axons Into a Specific Peripheral Nerve Branch by Selective Injection of a Lentiviral Vector Encoding Gdnf
Godard CW de Ruiter, MD; S. A. Hoyng; Ruben Eggers; M.R. Tannemaat, MD, PhD; J. Verhaagen; Martijn J.A. Malessy, PhD
Netherlands Institute for Neuroscience, Amsterdam, Netherlands
Background: Misdirection of regenerating axons is one of the factors that can explain the disappointing results often found after nerve injury and repair. The aim of the present study was to direct regenerating motor axons using a lentiviral vector encoding glial cell line-derived neurotrophic factor (LV-GDNF), which has been shown to stimulate motor axon regeneration. We selectively injected LV-GDNF into the peroneal nerve following rat sciatic nerve transection and repair, and studied the effect on the routing of regenerating motor axons.
Methods: The rat sciatic nerve was transected at the bifurcation into the tibial and peroneal nerve branches and repaired with 10-0 sutures. In 11 animals, LV-GDNF was subsequently injected in the peroneal nerve branch at 0.5cm from the bifurcation. In control group one, consisting of 8 animals, a lentiviral vector encoding for green fluorescent protein (GFP) was injected, and, in control group two no virus was injected after direct coaptation (DC). Four weeks after surgery and transfection, simultaneous retrograde tracing was performed, with fast blue (FB) application to the tibial nerve branch and diamidino yellow (DY) application to the peroneal nerve branch. The numbers of motoneurons labelled with FB and/or DY were counted.
Results: The number of DY labelled motoneurons, from which axons had regenerated to the peroneal branch, was found to be increased after LV-GDNF injection into this nerve (352+/-366) compared with LV-GFP injection (171+/- 152) and DC (168+/-192), although the difference was not statistically significant (P=0.15). The number of FB labelled motoneurons, from which axons had regenerated to the tibial branch, however, was significantly decreased after LV-GDNF injection (465+/-111) compared with LV-GFP injection (653+/-217) and DC (595+/-172) (P<0.05). This finding indicates that less motoneurons had regenerated towards the tibial branch after LV-GDNF injection into the peroneal nerve.
Discussion: This study provides a first indication that transfection with LV-GDNF may have an effect on the routing of regenerating motor axons. The results must be cautiously interpreted because the difference in number of peroneal motoneurons was not significantly increased. This may be explained by several factors, including variation in the site of injection of the lentiviral vector, and the occurrence of the “candy store” effect at the injection site. Axon counts are currently being performed to further understand these findings. This study shows, however, that viral vector-mediated expression of a neurotrophic factor has a potential to route regenerating axons after a peripheral nerve injury.
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