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Development of Cellular and Animal Models for Mucopolysaccharidosis Type III
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A mucopolissacaridose tipo III (MPS IIIA, B, C e D), ou Síndrome de Sanfilippo, é um subgrupo de doenças lisossomais de sobrecarga (DLS). São doenças raras, monogénicas e neurodegenerativas causadas por mutações nos genes que codificam as enzimas responsáveis pela degradação intralisossomal do sulfato de heparano (HS). Os doentes apresentam um fenótipo clínico grave, caracterizado por demência infantil de início precoce e por morte prematura. Embora tenham sido alcançados progressos significativos no seu conhecimento e algumas novas terapias tenham avançado até à fase de ensaios clínicos, ainda existem muitos aspetos dos mecanismos patofisiológicos inerentes a esta síndrome por desvendar, não estando atualmente disponível um tratamento eficaz. Modelos celulares e animais que reproduzam o fenótipo da doença, constituem recursos de grande valor, possibilitando estudos mais aprofundados e incisivos e, a identificação de novos alvos terapêuticos, além de uma triagem mais eficiente de novos fármacos.
Neste trabalho procurámos responder a essa necessidade através de: (i) uma caracterização aprofundada de duas linhas de células estaminais pluripotentes induzidas (iPSCs, do inglês: induced pluripotent stem cells), obtidas por reprogramação de fibroblastos de doentes MPS IIIC e IIID, previamente desenvolvidas no nosso grupo; e (ii) do estabelecimento de duas novas linhas de células estaminais de dentes decíduos esfoliados (SHEDs, do inglês: stem cells from human exfoliated deciduous teeth) derivadas de doentes com MPS IIIB e IIIC. Paralelamente, desenvolvemos um novo modelo animal para MPS IIIB, através da geração de um mutante nulo funcional na geração F0 de peixe-zebra (Danio rerio) através da tecnologia de CRISPR-Cas9.
Mucopolysaccharidosis type III (MPS IIIA, B, C, and D), or Sanfilippo Syndrome, is a subgroup of lysosomal storage disorders (LSDs). These are rare, monogenic, neurodegenerative diseases caused by mutations in genes encoding the lysosomal enzymes responsible for the degradation of heparan sulfate (HS). MPS III is characterised by a severe clinical phenotype, with early-onset childhood dementia and premature death. Despite significant advances in our understanding of the disease and the progress of some therapeutic approaches into clinical trials, many aspects of the underlying pathophysiological mechanisms remain unresolved, and no effective treatment is currently available. Cellular and animal models that reproduce the disease phenotype are valuable resources, enabling more in-depth mechanistic studies, the identification of novel therapeutic targets, and more efficient preclinical drug screening. In this work, we sought to address this need by: (i) performing an in-depth characterization of two induced pluripotent stem cell (iPSC) lines derived from fibroblasts of MPS IIIC and IIID patients, previously generated in our group; and (ii) establishing two new stem cell lines from human exfoliated deciduous teeth (SHEDs) derived from MPS IIIB and IIIC patients. In parallel, we developed a novel zebrafish model of MPS IIIB by generating a functional null mutant in the F0 generation through CRISPR-Cas9 genome editing.
Mucopolysaccharidosis type III (MPS IIIA, B, C, and D), or Sanfilippo Syndrome, is a subgroup of lysosomal storage disorders (LSDs). These are rare, monogenic, neurodegenerative diseases caused by mutations in genes encoding the lysosomal enzymes responsible for the degradation of heparan sulfate (HS). MPS III is characterised by a severe clinical phenotype, with early-onset childhood dementia and premature death. Despite significant advances in our understanding of the disease and the progress of some therapeutic approaches into clinical trials, many aspects of the underlying pathophysiological mechanisms remain unresolved, and no effective treatment is currently available. Cellular and animal models that reproduce the disease phenotype are valuable resources, enabling more in-depth mechanistic studies, the identification of novel therapeutic targets, and more efficient preclinical drug screening. In this work, we sought to address this need by: (i) performing an in-depth characterization of two induced pluripotent stem cell (iPSC) lines derived from fibroblasts of MPS IIIC and IIID patients, previously generated in our group; and (ii) establishing two new stem cell lines from human exfoliated deciduous teeth (SHEDs) derived from MPS IIIB and IIIC patients. In parallel, we developed a novel zebrafish model of MPS IIIB by generating a functional null mutant in the F0 generation through CRISPR-Cas9 genome editing.
Descrição
Dissertação de Mestrado em Biologia Celular e Molecular, Faculdade de Ciências, Universidade do Porto, 2025. https://hdl.handle.net/10216/171552
Palavras-chave
Doenças Genéticas Doenças Lisossomais de Sobrecarga Mucopolissacaridose Tipo III Modelos In Vitro Modelos In Vivo Células Estaminais Pluripotentes Induzidas Células Estaminais de Dentes Decíduos Esfoliados CRISPR-Cas9 Peixe-Zebra Lysosomal Storage Disorders Mucopolysaccharidosis Type III In Vitro Models In Vivo Models Induced Pluripotent Stem Cells (iPSCs) Stem Cells From Human Exfoliated Deciduous Teeth (SHEDs) Zebrafish
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