Phosphorylation-dependent TDP-43 antibody detects intraneuronal dot-like structures showing morphological characters of granulovacuolar degeneration

TAR-DNA-binding protein 43 (TDP-43) was considered to be a disease-specific component of ubiquitin-positive and tau-negative inclusions in the brains of patients with frontotemporal lobar degeneration and amyotrophic lateral sclerosis (ALS). However, this protein also accumulates abnormally in neurons in other neurodegenerative diseases, including Alzheimer's disease (AD). Although the role of TDP-43 deposition in these diseases is not clear, abnormal phosphorylation of the protein is suggested to be a critical step in disease pathogenesis. In this study, we generated a new phosphorylation-dependent TDP-43 antibody and examined AD brain sections from temporal lobes, including the hippocampus and temporal neocortex, by immunohistochemistry. The antibody, called A2, specifically recognized phosphorylated TDP-43 in western blotting using ALS and AD specimens, detecting a strong 45 kDa band and several shorter fragments at around 25 kDa with smears. Immunohistochemistry demonstrated neuronal cytoplasmic inclusions in AD brain sections without staining nuclei that were normal physiological TDP-43 localization sites. These results were consistent with previous reports. However, intraneuronal dot-like structures were also intensely labeled by immunohistochemistry. These structures were observed in all the AD brain sections examined and also occurred in sections from the brains of aged subjects without AD pathologies. The morphological and immunohistochemical characteristics of these granular structures were compatible with those of granulovacuolar degeneration (GVD). The A2 antibody clearly and intensely detected granular structures distributed over the hippocampus, subiculum, parahippocampus and temporal neocortex. Thus, immunohistochemistry using phosphorylation-dependent TDP-43 antibodies would be a new useful tool for identifying GVD.