Neural stem cells differentiated from human induced pluripotent stem cells (iPSCs) as a novel in vitro model to study developmental pathways in Huntington Disease [Tesi di dottorato]

Huntington Disease (HD) is an autosomal dominant disorder characterized by motor, cognitive and behavioral features caused by a CAG expansion in the HTT (huntingtin) gene beyond 35 repeats. Since the discovery of the HTT mutation 24 years ago, more than 15,000 papers have been published on HD. However, both the role of the huntingtin (wtHTT) in healthy individuals and the molecular mechanisms by which the mutated huntingtin causes the disease remain unclear. The discovery of induced pluripotent stem cell (iPSC) technology offer the possibility to generate patient-specific iPS cells and to enable the development of in vitro HD models that more accurately reflect the human disease. The disease-specific iPSCs can be differentiated into relevant cell-types affected in HD, holding a great potential for disease modeling and drug screening. In the present study, we have obtained dermal fibroblasts from 16 HD patients and 6 healthy controls. Fibroblasts from a young adult healthy control, a young subject with 43 CAG repeats, in an early stage of the disease and two JHD (Juvenile Huntington disease) patients were reprogrammed into induced pluripotent stem cells. All iPS clones that show an uniform flat morphology are characterized for their stemness and pluripotency, both in vitro through embryoid bodies formation and in vivo through teratoma formation assay. iPS cells from a JHD (85CAG) and healthy control, were differentiated into neurospheres of neural precursors (NPCs) by a new protocol optimized for differentiation of iPSCs derived embryoid bodies expressing all the three germ layers (ectoderm, mesoderm and endoderm) in neurospheres of Neural Stem Cells (NSCs). Differentiation assay confirmed that they possessed the potentiality, to differentiate into subtypes of neuronal and glial cells. Gene expression profiling was performed on iPSC HD-derived neurospheres using GeneChip Human Transcriptome Array 2.0. iPSC HD-derived neurospheres exhibit an impaired brain development processes when compared with iPSC-derived neurospheres from healthy individuals. This study aimed to produce a valid model of the Huntington disease. This novel in vitro model will permit us to have a closer view to neuronal development networks by morphologically and physiological studies on the cell types obtained from the neuropheres of NPC.

diritti: info:eu-repo/semantics/openAccess
In relazione con info:eu-repo/semantics/altIdentifier/hdl/11573/1058534
tutor esterno: F. Squitieri Valutatori
esterni: F. Angelucci
G. Bolasco
Settore BIO/10 - - Biochimica

Tesi di dottorato. | Lingua: Inglese. | Paese: | BID: TD18044227