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

Study designExperimental Animal Study.ObjectiveTo 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.SettingUniversity of Florida laboratory in Gainesville, USA.MethodsSprague Dawley rats received either a unilateral 150kdyne C4 contusion (n = 4 females, n = 5 males) or a laminectomy control surgery (n = 2 females, n = 3 males). 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 NF-L (NF-L-Ct), to label healthy axons, or an antibody against amyloid precursor protein (APP), considered the current "gold standard" for identifying axonal injury. The pattern of ongoing axonal degeneration was assessed.ResultsSpinal cord and brainstem cross-sections from injured rats had punctate MCA-6H63 positive fibers with a pathological appearance, loss of anti-NF-L-Ct colabeling, 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.ConclusionsThe MCA-6H63 antibody labels degenerating axons following SCI and offers a tool to quantify axonal degeneration.