Immunohistochemical labeling of ongoing axonal degeneration 10 days following cervical contusion spinal cord injury in the rat

Study Design: Experimental Animal Study Objective To continue validating an antibody which targets an epitope of neurofilament light chain (NF-L) only available during neurodegeneration and to utilize the antibody to describe the pattern of axonal degeneration 10 days post-unilateral C4 contusion in the rat. Setting: University of Florida Methods Sprague Dawley rats received either a unilateral 150kdyn C4 contusion (n = 6; n = 3 females, n = 3 males) or a laminectomy control surgery (n = 5; n = 3 males, n = 2 females). Ten days following SCI or laminectomy, spinal cords and brainstems were processed for immunohistochemistry. Serial spinal cord and brainstem cross-sections were stained with the degeneration-specific NF-L antibody (MCA-6H63) and dual labeled with either an antibody against the C-terminus portion of neurofilament light chain (NF-L-Ct), to label healthy axons, or an antibody against amyloid precursor protein (APP), considered the current “gold standard” for identifying degenerating axons. The pattern of ongoing axonal degeneration was assessed. Results Spinal cord and brainstem cross-sections from injured rats had punctate MCA-6H63 positive fibers with pathological appearance, loss of anti-NF-L-Ct co-labeling, and frequent colocalization with APP. Immunopositive fibers were abundant rostral and caudal to the lesion in white matter tracts that would be disrupted by the unilateral C4 contusion. This pattern of staining was not observed in control tissue. Conclusions The MCA-6H63 antibody labels degenerating axons following SCI and offers a promising tool to quantify axonal degeneration.