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Establishment of an inducible human pluripotent stem cell-based 3D model of Alzheimer's disease

dc.contributor.advisorBrüstle, Oliver
dc.contributor.authorHebisch, Matthias Alexander Sebastian
dc.date.accessioned2020-11-16T14:50:57Z
dc.date.issued16.11.2020
dc.identifier.urihttp://hdl.handle.net/20.500.11811/8780
dc.description.abstractAlzheimer’s disease (AD) is the most prevalent neurodegenerative disease in the aged population. The vast majority of cases are sporadic, however, various mutations may cause early-onset, familial AD (FAD) variants. AD is characterized by deposition of amyloid β (Aβ) plaques in the extracellular space and hyperphosphorylated tau (p-tau) within neurons. This project aimed to implement a human stem cell-based model in a 3D setting in order to more authentically model extracellular Aβ accumulation and its downstream pathogenic effects. To maximize production of aggregation-prone Aβ42, the safe-harbor locus AAVS1 of human induced pluripotent stem cells (iPSCs) was modified to conditionally overexpress FAD mutations in the genes encoding the amyloid precursor protein (APPSwe/Lon) and Presenilin-1 (PS-1ΔE9). Transgenic iPSCs were differentiated into long-term self-renewing neuroepithelial stem (lt-NES) cells, and embedded in a 200-300 µm thick Geltrex matrix for long-term differentiation in 3D. Induction with doxycycline caused 40-fold increased Aβ42 secretion in 2D cultures, whereas Aβ was efficiently entrapped in the gel matrix in 3D cultures. After 8 weeks, induced 3D cultures displayed Thioflavin T-positive Aβ deposits that strongly resemble amyloid plaques in size, structure and specific autofluorescence. After 16 weeks, induced 3D cultures contained TBS-insoluble Aβ. P-tau was present in induced 3D cultures from week 6, and fibril-specific tau epitopes could be detected after 16 weeks. As expected, p-tau pathology did not develop in the presence of γ-secretase inhibitors. Cellular energy metabolism was impaired in induced cultures as p-tau-positive neurons displayed hallmarks of mitochondrial fragmentation, alongside a reduction in essential respiratory chain complexes and lowered respiratory capacity. Induced cultures also showed neuronal degeneration compared to uninduced and γ-secretase-inhibited controls. It follows that 3D matrix cultures of lt-NES neurons represent a valid model to study intra- and extracellular AD pathology in an authentic human system.en
dc.language.isoeng
dc.rightsIn Copyright
dc.rights.urihttp://rightsstatements.org/vocab/InC/1.0/
dc.subjectAlzheimer
dc.subject3D-Kultur
dc.subjectAmyloid Phospho-tau
dc.subjectlt-NES
dc.subjectiPSC
dc.subjectMatrix
dc.subject3D-culture
dc.subject.ddc570 Biowissenschaften, Biologie
dc.subject.ddc610 Medizin, Gesundheit
dc.titleEstablishment of an inducible human pluripotent stem cell-based 3D model of Alzheimer's disease
dc.typeDissertation oder Habilitation
dc.publisher.nameUniversitäts- und Landesbibliothek Bonn
dc.publisher.locationBonn
dc.rights.accessRightsembargoedAccess
dc.date.embargoEndDate2021-11-16
dc.identifier.urnhttps://nbn-resolving.org/urn:nbn:de:hbz:5-60401
ulbbn.pubtypeErstveröffentlichung
ulbbnediss.affiliation.nameRheinische Friedrich-Wilhelms-Universität Bonn
ulbbnediss.affiliation.locationBonn
ulbbnediss.thesis.levelDissertation
ulbbnediss.dissID6040
ulbbnediss.date.accepted2020-11-06
ulbbnediss.instituteMedizinische Fakultät / Institute : Institut für Rekonstruktive Neurobiologie (IRN)
ulbbnediss.fakultaetMedizinische Fakultät
dc.contributor.coRefereeWagner, Wolfgang


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