Modelling retinal degenerative mechanisms and exploring novel neuroprotective therapies using 3D organoids

• Autor(a) principal: Carreira, Olga
• Data de Publicação: 2023
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10362/160940
• topic: 3D-organoids; Parkinson's Disease; Polyphenols; Neurodegeneration; Retina; α-Synuclein; Domínio/Área Científica::Engenharia e Tecnologia::Outras Engenharias e Tecnologias

Visual implications in Parkinson's Disease (PD) are supported by various evidence. For instance, α-Synuclein (αSyn) modifications and accumulation in the patients’ retina have been reported. However, the consequences of these deposits are not fully understood. To study the retina, the development of cultures where the cell behavior is closer to their natural counterparts is extremely important. Thus, 3D models, as retinal organoids (ROs), are used to mimic the composition, structure, complexity, and function of the human retina. This thesis aimed to further characterize the effects of αSyn overexpression in retinal degenerative mechanisms, by generating PD-patient hiPSCs-derived ROs, according to well-known Capowski et al. protocol. Cellular characterization and molecular alterations of the 3D cultures were evaluated by Western-Blot and Immunofluorescence assays. The results indicate that ROs were efficiently generated, achieving complex structures with phenotypic differences between control and PD model. αSyn levels were increased in PD patient-derived ROs, however it was not possible to identify which retinal cell population is overexpressing this toxic protein. Neurodegenerative hallmarks were evaluated in the PD model, revealing a tendency for increased oxidative stress, neuroinflammation and neurodegeneration, whereas autophagic and apoptotic pathways decreased along differentiation of the organoids. Controversially, mitochondrial number increased in PD patient-derived ROs, and the characterization of organoids in later developmental stages showed increased and more dispersed photoreceptor progenitor cells, but less rod photoreceptors in the PD model. As proof of concept, PD patient-derived ROs were treated with tauroursudeoxycholic acid, Catechol and Pyrogallol, known to modulate pathologic mechanisms in neurodegeneration, revealing that all treatments had an impact in neurodegenerative hallmarks. Overall, this work supports 3D ROs as a model to study molecular mechanisms in retinal degenerative diseases, mimicking the αSyn neurotoxic effects in this context. It also validates the use of ROs as tools for drug screening against the here mentioned neurodegenerative hallmarks.