Advancing the Art of Mitochondrial Diagnostics and Therapeutics
Abstract
Objective: Mitochondrial diseases are panethnic genetic disorders, affecting 1:5000 individuals. Currently with no cure, challenges exist in patient diagnosis, measurement of mitochondrial function, and effective therapies. The primary emphasis of this dissertation was to explore diagnostic and therapeutic targets for mitochondrial disease. Investigated therapies included; mesenchymal stem cell (MSC) therapy, dietary interventions, and side alternating vibration training (SAVT). Novel diagnostic approaches employed were; the hybrid blue-clear native polyacrylamide gel electrophoresis (BCN-PAGE), and circulating cell-free mitochondrial DNA (cf-mtDNA) techniques.
Methods: C57BL/6 mice were intravenously treated with MSCs following the induction of
metabolic inflammation through prolonged high-fat (HF) feeding. A separate cohort of BTBR,
autism-phenotype, mice were treated with a ketogenic diet (KD) to examine the impact on mitochondrial dynamics in a disease commonly experiencing mitochondrial dysfunction. These projects involved a combination of mitochondrial bioenergetics (high resolution respirometry), gut microbiome (fecal and cecal), gene expression (qRT-PCR, gene microarray), and mitochondrial morphology (confocal microscopy) experimentation to evaluate the mitochondrial-specific response to each therapy. Thirdly, clinical assessment of SAVT as a therapeutic in mitochondrial disease patients was performed using a 12-week cross-over study design using peak jumping power (PJP) as a primary functional measurement of mitochondrial function. Diagnostically, mitochondria were isolated from control and mitochondrial disease patient skin fibroblasts to compare our BCN-PAGE technique to existing diagnoses. Secondly, blood was drawn from both control and mitochondrial disease patients for clinical comparisons of mtDNA hapolotyping using our cf-mtDNA technique.
Results: Therapeutic studies indicate that MSCs improve metabolic capacity, induce widespread gene shifting, and promote healthy mitochondrial morphology in a model of metabolic inflammation. Moreover, KD administration exerts tissue-specific effects including increased mitochondrial turnover in liver, while the brain remained tightly regulated. Clinically, SAVT was well tolerated in mitochondrial disease patients, with improvements in PJP being measured following therapy. Diagnostically, both BCN-PAGE and cf-mtDNA techniques were successfully able to corroborate clinical diagnostic findings.
Conclusions: Experiments completed in this dissertation provide insight into various therapeutic opportunities for both mitochondrial disease and diseases involving mitochondrial dysfunction. Furthermore, our less-invasive diagnostic techniques may be useful to monitor the response to future therapies for mitochondrial disease.
Description
Keywords
Genetics, Biology--Molecular
Citation
Newell, C. (2017). Advancing the Art of Mitochondrial Diagnostics and Therapeutics (Doctoral thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca. doi:10.11575/PRISM/24728