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BEHANDLING-God og treffsikker diagnostikk, behandling og rehabilitering

Using pluripotent stem cells (iPSC) to model mitochondrial disease, study tissue specific manifestations and investigate treatments

Alternative title: null

Awarded: NOK 3.9 mill.

Mitochondria are fundamental to diverse biological processes including energy production, apoptosis and ageing, and mitochondrial dysfunction plays a role in common neurodegenerative diseases such as Parkinson?s disease. Primary mitochondrial diseases are the commonest inborn errors of metabolism and we showed that those due to mutations in the mitochondrial DNA polymerase (POLG) are common in Norway. POLG mutations cause abnormalities of mitochondrial DNA (mtDNA) making them a paradigm for mitochondrial disease in general. Nevertheless, despite the presence of mitochondria in all cells, POLG related diseases particularly affect brain and liver. In order to study the mechanisms of disease, we generated induced pluripotent stem cells (iPSC) from patients with POLG mutations and differentiated these into neurones and cardiomyocytes. In collaboration with Dr Gareth Sullivan, Oslo, we have also generated liver cells from these iPSC to study the liver dysfunction seen in patients treated especially the toxic response to the anticonvulsant sodium valproate. This NFR funded project has allowed us to develop a new method for culturing cardiomyocytes in 96 well plates that will facilitate later experiments for drug testing etc. We have also found changes in mitochondrial DNA during differentiation and this important discovery has shed light on what happens during the development of cardiomyocytes. We have also generated neural stem cells (NSC) and investigated the changes in mitochondria and mitochondrial DNA that occur during differentiation both the NSC and further to fully differentiated neurones. Changes in gene expression (transcriptomics) are completed, and analysis is ongoing. Further studies including proteomics are planned. Including making both dopaminergic and spinal neurones, we have also developed methods to differentiate NSC to astrocytes and oligodendrocytes. These are being co-cultured with neurones to investigate how these cell-types modulate normal function and disease development in neurones. Co-culturing of astrocytes with control neurones or those that have POLG mutations is being studied using functional studies and transcriptomics and proteomics. We are also developing 3D cultures to investigate the interplay between astrocytes and more fully developed neurones. Our studies have provided greater understanding of not only disease mechanisms, but also what happens during normal development. The research is original, based on cutting edge technology and, given the high prevalence of POLG mutations here in Norway, highly relevant. Collaboration between Bergen and Oslo Stem Cell Centres is now well established and this will form the basis for future work.

Mitochondria are fundamental to diverse biological processes including energy production and apoptosis, and dysfunction of mitochondria is known to cause a wide range of disease. Further, mitochondria are linked to several neurodegenerative diseases notab ly Parkinson's disease and mutations in mitochondrial DNA (mtDNA) are thought to play a role in ageing. Mitochondrial diseases are the commonest inborn errors of metabolism and we have shown that one group, due to mutations in the mitochondrial DNA polyme rase (POLG), are common in Norway. POLG mutations cause abnormalities of mtDNA and are therefore, a paradigm for mitochondrial disease in general. Induced pluripotent stem cell (iPSC) technology offers a unique opportunity to model human disease in a rene wable and tissue specific manner. We will use human iPSC from patients with POLG mutations to model disease in cells such as neurones, hepatocytes and cardiomyocytes, tissues that we know do, and do not, manifest disease. We will study the mechanisms resp onsible for tissue specific involvement and how this evolves, specifically how it affects energy metabolism and mtDNA. We will also investigate the mechanisms behind the hepatotoxicity seen in these patients treated with the first line anticonvulsant sodi um valproate. Subsequently, we will explore ways of using these iPS cells to develop treatments. This will involve medium/low throughput screening for potentially therapeutic molecules and, by genetic manipulation of the primary cells, generating patient cells (neurones, hepatocytes etc.) free of the mutation to use for treatment. The planned studies will increase our understanding of disease mechanisms and basic mitochondrial function. The research is original, based on cutting edge technology and, given the high prevalence of POLG mutations here in Norway, highly appropriate. The combination of clinical questions and basic science approach means that this research is by all criteria translational.

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BEHANDLING-God og treffsikker diagnostikk, behandling og rehabilitering