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Publication

Mapping and treating metabolic rewiring in mitochondrial disease

Book - Dissertation

The mitochondrial oxidative phosphorylation system (OXPHOS) generates most of the cellular energy in the form of ATP. Pathogenic variants in genes crucial for mitochondrial function cause primary mitochondrial diseases (PMD). PMD form a large, heterogeneous group of disorders that affect at least 1 in 5000 individuals. Patients may present at any age, with symptoms in any organ, at any severity. Due to the diversity in clinical presentation and the lack of a single clear biochemical profile, diagnosis of PMD remains a challenge. Most PMD have a poor prognosis and lack effective therapies. There is thus a need for the development of specific, sensitive and rapid diagnostic markers, and for the discovery of effective therapeutic interventions. We hypothesised that PMD causes important metabolic changes due to the key role of mitochondria in numerous biochemical pathways. This project aimed to understand the cellular metabolic re-wiring occurring in PMD by characterising the metabolic consequences of OXPHOS dysfunction in skin fibroblasts derived from patients with PMD and healthy controls. The objectives were 3-fold: first to develop a new diagnostic technique, second to unravel the cellular profile in OXPHOS dysfunction, and third, to identify and test novel treatment strategies. First, the potential of oxygraphy as a diagnostic tool for PMD was investigated and compared to the gold standard of enzymology. The results showed that oxygraphy is a powerful new tool for the biochemical detection of PMD. However, oxygraphy and enzymology are complementary techniques, and should ideally be combined. The biochemical results must also be interpreted in regard to the clinical presentation, the other investigations already performed, and genetic testing results. Second, the metabolic profile of OXPHOS dysfunction was described in skin-derived fibroblasts obtained from patients with diverse PMD and validated in pharmacological models of OXPHOS dysfunction. Growth rate was determined using the Incucyte technology and steady-state glucose and glutamine tracing studies were performed with LC-MS quantification of cellular metabolites. Our results revealed that OXPHOS deficiency causes a proliferation defect and is associated with a distinct metabolic profile. The main features of this signature are a depletion of aspartic acid, of TCA intermediates and increased levels of glycerol-3-phosphate. This profile seems to be related to an NAD+/NADH imbalance. Finally, the therapeutic potential of numerous nutritional supplements was tested by assessing their effect on proliferation and on the metabolomics profile. In fibroblasts, therapy with pyruvate and uridine rescued the metabolic signature of OXPHOS dysfunction and the subsequent proliferation defect. Additionally, in zebrafish, pyruvate and uridine treatment increased the survival after rotenone exposure. This project highlights the importance of the NAD+/NADH imbalance following OXPHOS dysfunction and opens the door to novel diagnostic techniques and therapeutic interventions for PMD.
Publication year:2023
Accessibility:Open