Development of a novel cellular model of Alzheimer's disease utilizing neurosphere cultures derived from B6C3-Tg(APPswe,PSEN1dE9)85Dbo/J embryonic mouse brain.

Increased production, oligomerization and aggregation of amyloid-β (Aβ) peptides are hallmark pathologies of Alzheimer's disease (AD). Expressing familial AD mutations (amyloid precursor protein and/or presenilins mutations), the Aβ-pathologies of AD has been recapitulated in animal models of AD. Very few primary cell culture-based models of AD are available and they exhibit very weak Aβ-pathologies compared to what is seen in AD patients and animal models of AD. CNS stem/progenitor cells are present in both embryonic and adult brains. They can be isolated, grown as neurospheres and differentiated into neurons, astrocytes and oligodendrocytes. It is not yet known whether CNS stem/progenitor cells can support the production of Aβ peptides in culture. In this report, we have established Aβ-pathologies such as production, secretion, oligomerization and aggregation of Aβ peptides utilizing neurosphere cultures to create a new cellular model of AD. These cultures were developed from E15 embryonic brains of transgenic mice carrying the Swedish mutations in humanized mouse APP cDNA and the exon-9 deleted human presenilin 1 cDNA both regulated by mouse prion protein gene (Prnp) promoter. Results demonstrated the expression of transgene transcripts, APPswe protein and its processed products only in transgene positive neurosphere cultures. These cultures generate and secrete both Aβ40 and Aβ42 peptides into culture medium at levels comparable to the Aβ load in the brain of AD patients and animal models of AD, and produce pathogenic oligomers of Aβ peptides. The Aβ42/Aβ40 ratio in the medium of transgene positive neurosphere cultures is higher than any known cellular models of AD. Conformation dependent immunocytochemistry demonstrated the possible presence of intracellular and extracellular aggregation of Aβ peptides in neurosphere cultures, which are also seen in AD brain and animal models of AD. Collectively, our neurosphere cultures provide robust Aβ-pathologies of AD better than existing cellular model of Alzheimer's disease.