Investigating Friedreich ataxia ophthalmopathy and cardiomyopathy using induced pluripotent stem cells

© 2015 Dr. Duncan Edward CrombieFriedreich ataxia (FRDA) is the most common of the inherited ataxias. It is an autosomal recessive disease characterised by degeneration of central nervous system regions, peripheral sensory neurons and cardiomyopathy. The predominant neuronal manifestations occur through degeneration of dorsal root ganglia and cerebellar neurons, leading to development of ataxia, dysarthia and areflexia of the lower limbs (1). Other sensory neurons are also affected with all patients demonstrating optic nerve thinning, however, only 20% will present with significant visual system impairment. FRDA occurs in most cases due to homozygosity for trinucleotide GAA repeat expansions found with in first intron of the FXN gene, which results in reduced levels of the normal mitochondrial protein, FRATAXIN (FXN). Reduced FXN protein causes mitochondrial dysfunction and iron accumulation leading to increased oxidative stress and cell death in the nervous system and heart. Yet the precise functions of FXN and the precise mechanisms behind how reduced FXN levels cause disease are still largely unknown. Regardless of the mechanisms, it is generally regarded that any increase in FXN levels in key tissues will be of therapeutic benefit. The overall aim of this PhD thesis was to investigate FRDA ophthalmopathy and cardiomyopathy, predominantly using an in vitro approach with human induced pluripotent stem cells (iPSCs). The first part the thesis, described the use of three approaches - mouse models, examination of histopathology of patient eyes post mortem, and patient specific iPSCs - to gain a better picture on the health of the retina and in particular of the retinal pigmented epithelium in FRDA and to determine if any obvious defects could be modelled in vitro. The second part of this thesis, described a novel and economical way to characterise the chromosomal stability of iPSCs, a necessary step in the generation of iPSCs for disease modelling. In the third part the thesis, FRDA iPSCs were derived from three FRDA individuals, with different GAA repeat sizes and cardiac phenotypes. Utilising an efficient protocol to generate high numbers of iPSC-derived cardiomyocytes, disease-specific phenotypes were assessed with a view to developing a drug-screening platform for rescue of identified pathogenic signatures and/or increasing FXN levels. Novel disease phenotypes were observed in the patient lines, which were indicative of intracellular calcium handling abnormalities. The data generated during this PhD demonstrates that iPSCs can be used to study FRDA and gives new insight into the ophthalmopathy and cardiomyopathy of FRDA