Alzheimer’s disease (AD) is seen as a the deposition of senile plaques (SPs) and neurofibrillary tangles (NFTs) in vulnerable brain regions. Aβ42 staining was exhibited within cultured neurons by confocal immunofluorescence microscopy and within neurons of PS1 mutant transgenic mice. A central question about the role of Aβ in disease issues whether extracellular Aβ deposition or intracellular Aβ accumulation initiates the disease process. beta-Pompilidotoxin Here we statement that human neurons in AD-vulnerable brain regions specifically accumulate γ-cleaved Aβ42 and suggest that this intraneuronal Aβ42 immunoreactivity appears to precede both NFT and Aβ plaque deposition. This study suggests that intracellular Aβ42 accumulation is an early event beta-Pompilidotoxin in neuronal dysfunction and that preventing intraneuronal Aβ42 aggregation may be an important therapeutic direction for the treatment of AD. Alzheimer’s disease (AD) neuropathology is usually classically characterized by the accumulation of senile plaques (SPs) and neurofibrillary tangles (NFTs) in vulnerable brain regions. SPs are composed of parenchymal and cerebrovascular aggregates of β-amyloid (Aβ) 40/42(43) peptides. Increasing evidence indicates that Aβ plays a central role in the pathophysiology of AD. Individuals with Down’s syndrome (DS) have an extra copy of chromosome 21 where the gene encoding the β-amyloid precursor protein (βAPP) is usually localized and invariably develop AD pathology at an early age. Mutations in βAPP segregate with some forms of autosomal dominant familial AD (FAD). Transgenic mice bearing FAD βAPP mutations develop striking AD-like senile plaque pathology. 1 FAD mutations in βAPP and presenilin 1 (PS1) lead to elevated secretion of Aβ especially the more amyloidogenic Aβ42. In addition immunohistochemical studies have underscored the importance of Aβ42 as the initiator of plaque pathology in AD and DS. 2 3 Over the past few years cell biological studies support the view that Aβ is usually generated intracellularly 1 4 from your endoplasmic reticulum (ER) 1 7 8 to the trans-Golgi network (TGN) 4 and the endosomal-lysosomal system. 10 Recently endogenous Aβ42 staining was exhibited within cultured main neurons by confocal immunofluorescence microscopy 9 and within neurons of human PS1 mutant transgenic mice by immunocytochemical light microscopy. 11 A central question on the role of Aβ in AD is whether extracellular Aβ deposition or intracellular Aβ accumulation is initiating the disease process. Several groups had postulated the presence of intraneuronal Aβ immunostaining. However the Aβ immunoreactivity observed in these studies was compromised by that of full-length βAPP because beta-Pompilidotoxin these Aβ antibodies also recognize full-length βAPP. 12-14 In addition NFTs had previously been reported to be immunoreactive to Aβ. 15-16 This association of Aβ with NFTs was Rabbit Polyclonal to Collagen V alpha1. subsequently believed to be the result of artifactual “shared” epitopes. 17 We now report that human neurons in AD-vulnerable brain regions specifically accumulate γ-cleaved Aβ42 but not the more abundantly secreted Aβ40. We also demonstrate intraneuronal Aβ42 staining in neurons in both the absence and presence of NFTs. Our observations in adjacent sections of intraneuronal Aβ42 staining and hyperphosphorylated tau staining suggest that neuronal Aβ42 staining is more abundant and therefore may precede NFTs which would exclude the possibility of cross-reactivity of shared epitopes. Furthermore we observe the earliest Aβ42 beta-Pompilidotoxin immunoreactive SPs developing along the projections and at terminals of early Aβ42 accumulating neurons suggesting a mechanism for the previously hypothesized regional specificity of AD disease progression within the brain. 18 Materials and Methods Antibodies Polyclonal rabbit Aβ40 (RU226) and Aβ42 (RU228) C-terminal specific antibodies were generated at Rockefeller University (RU). Polyclonal rabbit Aβ40 and Aβ42 C-terminal antibodies were obtained commercially (QCB) also. The results acquired with both of these models of antibodies had been similar and had been verified using well-characterized polyclonal rabbit Aβ40 (FCA3340) beta-Pompilidotoxin and Aβ42 (FCA3542) antibodies 19 (kindly supplied by F. Checler). Antibody 4G8 identifies proteins 17-24 of Aβ (Senetek). Hyperphosphorylated tau was identified by antibody AT8 (Polymedco). ApoE was visualized having a mouse monoclonal anti-ApoE antibody (Boehringer-Mannheim). Immunocytochemistry Postmortem mind tissue was analyzed from representative neurologically regular controls (age groups three months and 3 30 44 58 and 79 years); seniors nursing home occupants without dementia (Clinical.