Botella Ruiz, Javier ORCID: https://orcid.org/0000-0001-9722-8519 (2020) The effects of endurance exercise and training on mitochondrial dynamics and remodelling in human skeletal muscle. PhD thesis, Victoria University.

Abstract

Exercise is one of the most effective lifestyle interventions for the prevention of non-communicable diseases. Although exercise is beneficial at multiple levels, skeletal muscle represents the primary adaptive tissue. Despite the increasing understanding of the beneficial effects of exercise, it remains unknown how different types of exercise affect the molecular pathways mediating improvements in health and performance. The aims of this thesis were: i) to explore the effects of different exercise prescriptions on the regulation of early markers of autophagy following exercise and training; ii) the cellular and mitochondrial responses to two very different exercise prescriptions emphasising high-volume (moderate-intensity continuous exercise, MICE) and high-intensity (sprint-interval exercise, SIE); iii) and to study the mitochondrial remodelling following 8 weeks of two types of exercise training (moderate-intensity continuous training, MICT; and sprint-interval training, SIT). In Chapter 1, the role of mitochondrial characteristics in the context of endurance performance was reviewed. There was convincing evidence that multiple mitochondrial adaptations represent a commonly observed adaptation necessary for training-induced improvements in endurance performance. Despite the large body of evidence suggesting that mitochondrial characteristics do not limit maximal oxygen consumption, the importance of mitochondrial characteristics for endurance performance is often supported. A new framework for the determining physiological and biological factors of endurance performance, in which mitochondrial characteristics are incorporated and play a central role, was proposed. In Chapter 2, the effects of exercise on the regulation of the multiple layers of mitochondrial dynamics and mitochondrial stress were explored. Given the limited data in humans, a summary of each pathway was established and related to exercise and training studies. Furthermore, and in line with my experimental chapters, a role for the mitochondrial-specific integrated stress response was discussed in the context of skeletal muscle and exercise. Despite limited evidence on the effects of exercise, there was emerging literature highlighting the importance of mitochondrial-specific stress response in the context of skeletal muscle diseases, where chronic mitochondrial dysfunction is a distinctive feature. In the first experimental chapter (Chapter 3), I discussed the effects of exercise on the regulation of markers of autophagy. Given that a key step in mitochondrial dynamics is mitochondrial-specific autophagy (mitophagy) it was important to answer a standing question in the field of exercise in humans: is autophagy downregulated following exercise? These conclusions were based on the decreased content of lipidated LC3 (LC3-II) observed in human skeletal muscle but not in rodents. It is shown that the exercise-induced decrease in LC3B-II protein content is a consistent finding in human skeletal muscle, independently of exercise intensity. Furthermore, the divergent response observed in LC3B between rodents and humans were reproduced. Conversely, by using a novel and adapted ex vivo autophagy flux assay, this study was able to show that autophagy does not decrease after exercise, and follows a similar pattern as previously shown in rodents. This chapter suggests that making autophagy conclusions based on basal LC3-II protein content can lead to erroneous interpretation. It calls for the use of a more valid approach, with the use of the ex vivo autophagy flux assay, as well as including multiple members of the ATG8 family for a more comprehensive analysis. In the second experimental chapter (Chapter 4), the effects of different exercise prescriptions, focused on high-volume (MICE) or high-intensity (SIE), on the cellular and mitochondrial-related signalling was discussed. Transcriptomic analysis showed that the most enriched pathway divergently activated was the unfolded protein response. This pathway was enriched following SIE and included the main genes linked to the mammalian mitochondrial unfolded protein response. This was corroborated by an enrichment analysis of the ‘Mitostress’ geneset, which was significantly enriched following SIE, but not MICE. In agreement with this, the TEM analyses showed that mitochondrial morphology and structure were robustly altered following SIE, while it remained unchanged following MICE. Interestingly, this occurred independently changes in the gene expression level of the ‘master regulator’ of mitochondrial biogenesis PGC-1α. This is the first time such mitochondrial stress and transcriptional upregulation has been reported following exercise in human studies. Given the results, it was important to elucidate the mitochondrial remodelling following exercise training. In the last experimental chapter (Chapter 5), the physiological adaptations and mitochondrial remodelling following 8 weeks of exercise training with MICT or SIT were discussed. Following MICT there was predominantly an increase in variables associated with mitochondrial content, including mitochondrial volume density (mitoVD), citrate synthase (CS) activity and total OXPHOS complexes. On the other hand, SIT resulted in a significant improvement in mitochondrial respiratory function and mitochondrial size without concomitant increases in markers of mitochondrial content. Correlation analyses showed that training-induced changes in complex I was the only variable to be correlated with training-induced changes in markers of endurance performance. These results highlight that the exercise-induced mitochondrial stress observed following sprint-interval exercise is adaptive and leads to positive mitochondrial adaptations. Furthermore, it highlights the complexity of mitochondrial adaptations to multiple exercise training prescriptions, and demonstrates the relevance of these findings for optimised exercise prescription. In conclusion, these findings provide new and novel evidence about signalling pathways divergently activated following distinct exercise sessions, and how this, when repeated over time can lead to divergent mitochondrial remodelling. Improving the understanding of the molecular regulation of skeletal muscle following exercise and training not only provides new and valuable knowledge for the discovery of new pathways involved in the adaptive responses to exercise, but also aids at improving the individualised exercise prescription in health and performance.

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