Glial Cell Line–Derived Neurotrophic Factor and Chondroitinase Promote Axonal Regeneration in a Chronic Denervation Animal Model

Abstract

Functional recovery following nerve injury declines when target re-innervation is delayed. Currently, no intervention exists to improve outcomes after prolonged denervation. We explored the neuroregenerative effects of glial cell line-derived neurotrophic factor (GDNF) and chondroitinase (CDN) in a chronic denervation animal model. A fibrin-based sustained delivery method for growth factors was optimized in vitro and in vivo, and then tested in our animal model. GDNF, CDN, and GDNF+CDN were injected into the denervated stump at the time of nerve repair. Histomorphometry and retrograde labeling were used to assess axonal regeneration. The mechanisms promoting such regeneration were explored with immunofluorescence. Five weeks after repair, the GDNF+CDN group had the highest number and maturity of axons. GDNF was noted to preferentially promote axonal maturity, whereas CDN predominantly increased the number of axons. GDNF favored motor neuron regeneration, and upregulated Ki67 in Schwann cells. CDN did not favor motor versus sensory regeneration and was noted to cleave inhibitory endoneurial proteoglycans. Early measures of nerve regeneration after delayed repair are improved by activating Schwann cells and breaking down the inhibitory proteoglycans in the distal nerve segment, suggesting a role for GDNF+CDN to be translated for human nerve repairs.

Combining timed GDNF and ChABC gene therapy to promote long‐distance regeneration following ventral root avulsion and repair

“…The dual treatment strategy of neurotrophic support using GDNF and removal of inhibitory CSPGs using ChABC has been employed previously. When combined at the lesion site, GDNF and ChABC improved axonal regeneration following a spinal cord lesion 65,66 and peripheral nerve transection 67‐69 . In these studies, ChABC enzyme and GDNF protein were applied directly at the lesion sites using gelfoam, slow release systems or direct protein injections.…”

“…When combined at the lesion site, GDNF and ChABC improved axonal regeneration following a spinal cord lesion 65,66 and peripheral nerve transection. [67][68][69] In these studies, ChABC enzyme and GDNF protein were applied directly at the lesion sites using gelfoam, slow release systems or direct protein injections. Topical ChABC application at the site of reimplantation of avulsed ventral roots did result in increased axonal regeneration up to 2 cm distally, whereas motoneurons were not rescued.…”

Schwann Cell Role in Selectivity of Nerve Regeneration

“…After 3 months of denervation, the distal stump of the tibial nerve was sutured to a freshly cut peroneal nerve. Five weeks later, motoneurons had regenerated more in the group with GDNF delivery, whereas this factor did not affect sensory neuron regeneration [ 74 ]. A similar enhancement of motor axon regeneration was found after treating a denervated saphenous nerve with GDNF and grafting it to the muscle branch of the femoral nerve.…”

“…Other inhibitory molecules which are relevant in peripheral nerve regeneration are chondroitin sulfate proteoglycans (CSPGs), extracellular matrix components that are upregulated after nerve injury [ 95 ]. In the case of CSPGs, their removal using the enzyme chondroitinase ABC also improved regeneration [ 74 , 96 ], although it did not have a differential effect on motor versus sensory neurons [ 74 ]. Both MAG of SCs and CSPGs of the extracellular matrix induce growth cone collapse through the activation of the small GTPase RhoA and its downstream effector Rho-kinase (ROCK) in regenerating neurons [ 97 , 98 ] ( Figure 2 C).…”