A 3D- induced pluripotent stem cell-derived human neural culture model to study certain molecular and biochemical aspects of Alzheimer’s Disease.

Abstract

Alzheimer’s disease (AD) is the most prevalent form of dementia and currently has no cure. Induced pluripotent stem cell (iPSC) technology is an innovative, cutting-edge technique that is able to recapitulate some key features of neurodegenerative disease pathology in vitro. The major advantage of using this technology is that neurons can be generated in vitro from AD patients with inherited mutations, thereby allowing patient-specific insight into the biochemical and pathophysiological nature of AD. 3D culture models provide a physiologically and spatially relevant microenvironment, thereby aiding better differentiation and maturation of cells in vitro. In this study, we aim to develop a 3D neural model derived from control and AD patient’s iPSCs, in which cells can differentiate, self-organize and mature. AD-iPSCs with Presenilin 1 mutation (L286V, M146L, or A246E) and age-matched controls were differentiated in 3D Matrigel for 18 weeks in vitro. Characterization of cell morphology and protein profile was performed using immunofluorescence. Western blotting was used to determine disease-associated changes such as hyperphosphorylation of tau. AD and age-matched control iPSCs differentiate into neurons and astrocytes which self-organize into 3D structures by 3 weeks of differentiation in vitro. Cells express astrocytic (GFAP), neuronal (beta-3-tubulin, MAP2), glutamatergic (VGLUT1), GABAergic (GAD65/67) and pre-synaptic (Synapsin 1) markers after differentiation. The foetal 3R tau isoforms and 4R adult tau isoforms were detected at 6 weeks post differentiation. We have developed a standardised and validated in vitro human 3D iPSCs- derived neural model with mature neurons. In these AD-derived cells, we have shown early changes in AD-associated protein expression levels, presence of Aβ oligomers, an increase in the Aβ42/40 ratio, hyperphosphorylated Tau and presence of aggregated insoluble Tau, as observed in the disease. Our data indicates that this model may recapitulate the early biochemical and pathological disease features and can be a relevant platform for studying early cellular and biochemical changes and identification of drug targets

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Identifiers

ORCID for Sandrine Willaime-Morawek: orcid.org/0000-0002-1121-6419

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