Axon regeneration through scaffold into distal spinal cord after transection.
We employed Fast Blue (FB) axonal tracing to determine the origin of regenerating axons after thoracic spinal cord transection injury in rats. Schwann cell (SC)-loaded, biodegradable, poly(lactic-co-glycolic acid) (PLGA) scaffolds were implanted after transection. Scaffolds loaded with solubilized basement membrane preparation (without SCs) were used for negative controls, and nontransected cords were positive controls. One or 2 months after injury and scaffold implantation, FB was injected 0-15 mm caudal or about 5 mm rostral to the scaffold. One week later, tissue was harvested and the scaffold and cord sectioned longitudinally (30 microm) on a cryostat. Trans-scaffold labeling of neuron cell bodies was identified with confocal microscopy in all cell-transplanted groups. Large (30-50 microm diameter) neuron cell bodies were predominantly labeled in the ventral horn region. Most labeled neurons were seen 1-10 mm rostral to the scaffold, although some neurons were also labeled in the cervical cord. Axonal growth occurred bidirectionally after cord transection, and axons regenerated up to 14 mm beyond the PLGA scaffolds and into distal cord. The extent of FB labeling was negatively correlated with distance from the injection site to the scaffold. Electron microscopy showed myelinated axons in the transverse sections of the implanted scaffold 2 months after implantation. The pattern of myelination, with extracellular collagen and basal lamina, was characteristic of SC myelination. Our results show that FB labeling is an effective way to measure the origin of regenerating axons.
Department of Neurology, Mayo Clinic, Rochester, Minnesota 55905, USA.
Bing Kun
BK
Chen
Andrew M
AM
Knight
Godard C W
GC
de Ruiter
Robert J
RJ
Spinner
Michael J
MJ
Yaszemski
Bradford L
BL
Currier
Anthony J
AJ
Windebank
D020341:Absorbable Implants; D000578:Amidines; D000818:Animals; D001370:Axonal Transport / Q000502:physiology; D001672:Biocompatible Materials / Q000494:pharmacology / Q000627:therapeutic use; D001931:Brain Mapping / Q000379:methods; D002478:Cells, Cultured; D005260:Female; D005456:Fluorescent Dyes; D020439:Growth Cones / Q000502:physiology* / Q000648:ultrastructure; D048091:Guided Tissue Regeneration / Q000379:methods*; D019344:Lactic Acid / Q000494:pharmacology / Q000627:therapeutic use; D018613:Microscopy, Confocal; D009416:Nerve Regeneration / Q000502:physiology*; D056325:Neuronal Tract-Tracers; D011100:Polyglycolic Acid / Q000494:pharmacology / Q000627:therapeutic use; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D051381:Rats; D017207:Rats, Sprague-Dawley; D013119:Spinal Cord Injuries / Q000503:physiopathology / Q000601:surgery*; D033581:Stem Cell Transplantation / Q000379:methods; D013997:Time Factors; D054457:Tissue Scaffolds; D016896:Treatment Outcome
D000578:Amidines; D001672:Biocompatible Materials; D005456:Fluorescent Dyes; D056325:Neuronal Tract-Tracers; C031455:diamidino compound 253-50; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011100:Polyglycolic Acid; D019344:Lactic Acid