Nerve Fiber Ingrowth Into Scar Tissue Formed Following Nucleus Pulposus Extrusion in the Rabbit Anular-Puncture Disc Degeneration Model: Effects of Depth of Puncture

Abstract

In vivo histologic study of nerve ingrowth in the rabbit anular-puncture disc degeneration model. To examine innervation of intervertebral discs and surrounding tissues in the rabbit anular-puncture model with various magnitudes of injury. A rabbit anular-puncture model of disc degeneration was recently established. However, to our knowledge, the neuropathologic changes in the degenerated disc and surrounding tissues in this model have not been examined. Anular punctures with an 18-gauge needle at depths of 1 and 5 mm were randomly performed at the L2-L3 or L4-L5 discs in New Zealand white rabbits (n = 16; 3.0-3.5 kg). The degree of disc degeneration was followed radiographically, and was further evaluated by magnetic resonance imaging and histology at sacrifice 4 or 12 weeks after the puncture. To identify nerve ingrowth into the discs, the L2-L3 to L4-L5 discs were immunostained with an antibody against protein gene product 9.5, a general marker for nerve fibers. Significant decreases in disc height and signal intensity in magnetic resonance imaging were observed only in the 5 mm punctured discs (P < 0.05). In all the discs, including nonpunctured control discs, protein gene product 9.5-immunoreactive fibers were only occasionally observed in the outermost part of the anulus fibrosus. In the 5 mm punctured discs, nucleus pulposus tissues were extruded, and scar tissues formed outside the discs. No nerve ingrowth into the needle track lesion at the anular puncture site was observed. However, protein gene product 9.5-immunoreactive fibers were observed in scar tissues on the surface of the puncture site. This finding was more evident in the 5 mm than in the 1 mm punctured discs (P < 0.05). This study indicates that in the rabbit anular-puncture disc degeneration model, disc degeneration associated with a higher nerve growth into the scar tissue was more evident when induced by a 5 mm than a 1 mm puncture. Although nerve ingrowth was observed in the extruded disc tissue, nerve ingrowth into the outer anulus, which has been reported in patients with discogenic pain, was not observed during the short observation period in this disc degeneration model. The limitation in assessing pain by behavior analysis or histologic evaluation of nerve ingrowth should be considered. Further studies to identify a surrogate marker of pain should be encouraged.