Expressing Constitutively Active Rheb in Adult Neurons after a Complete Spinal Cord Injury Enhances Axonal Regeneration beyond a Chondroitinase-Treated Glial Scar.

Abstract

After a spinal cord injury (SCI), CNS axons fail to regenerate, resulting in permanent deficits. This is due to: (1) the presence of inhibitory molecules, e.g., chondroitin sulfate proteoglycans (CSPG), in the glial scar at the lesion; and (2) the diminished growth capacity of adult neurons. We sought to determine whether expressing a constitutively active form of the GTPase Rheb (caRheb) in adult neurons after a complete SCI in rats improves intrinsic growth potential to result in axon regeneration out of a growth-supportive peripheral nerve grafted (PNG) into the SCI cavity. We also hypothesized that treating the glial scar with chondroitinase ABC (ChABC), which digests CSPG, would further allow caRheb-transduced neurons to extend axons across the distal graft interface. We found that targeting this pathway at a clinically relevant post-SCI time point improves both sprouting and regeneration of axons. CaRheb increased the number of axons, but not the number of neurons, that projected into the PNG, indicative of augmented sprouting. We also saw that caRheb enhanced sprouting far rostral to the injury. CaRheb not only increased growth rostral and into the graft, it also resulted in significantly more regrowth of axons across a ChABC-treated scar into caudal spinal cord. CaRheb(+) neurons had higher levels of growth-associated-43, suggestive of a newly identified mechanism for mTOR-mediated enhancement of regeneration. Thus, we demonstrate for the first time that simultaneously addressing intrinsic and scar-associated, extrinsic impediments to regeneration results in significant regrowth beyond an extremely challenging, complete SCI site.

SIGNIFICANCE STATEMENT:

After spinal cord injury (SCI), CNS axons fail to regenerate, resulting in permanent deficits. This is due to the diminished growth capacity of adult neurons and the presence of inhibitory molecules in the scar at the lesion. We sought to simultaneously counter both of these obstacles to achieve more robust regeneration after complete SCI. We transduced neurons postinjury to express a constitutively active Rheb to enhance their intrinsic growth potential, transplanted a growth supporting peripheral nerve graft into the lesion cavity, and enzymatically modulated the inhibitory glial scar distal to the graft. We demonstrate, for the first time, that simultaneously addressing neuron-related, intrinsic deficits in axon regrowth and extrinsic, scar-associated impediments to regeneration results in significant regeneration after SCI.

• Authors: Di Wu, Michelle C Klaw, Theresa Connors, Nikolai Kholodilov, Robert E Burke, Veronica J Tom
• Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience (J Neurosci) Vol. 35 Issue 31 Pg. 11068-80 (Aug 05 2015) ISSN: 1529-2401 [Electronic] United States
• PMID: 26245968 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2015 the authors 0270-6474/15/3511068-13$15.00/0.

Chemical References

• Neuropeptides
• Ras Homolog Enriched in Brain Protein
• Rheb protein, rat
• Monomeric GTP-Binding Proteins
• Chondroitin ABC Lyase

Topics

• Animals
• Axons (drug effects, physiology)
• Chondroitin ABC Lyase (pharmacology, therapeutic use)
• Cicatrix (drug therapy, genetics, therapy)
• Disease Models, Animal
• Female
• Monomeric GTP-Binding Proteins (genetics)
• Nerve Regeneration (drug effects, genetics)
• Neuroglia (drug effects, physiology)
• Neurons (drug effects, physiology)
• Neuropeptides (genetics)
• Ras Homolog Enriched in Brain Protein
• Rats
• Rats, Wistar
• Spinal Cord Injuries (drug therapy, genetics, therapy)
• Treatment Outcome