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Maja Munk Dethlefsen:
PGC-1α and the regulation of autophagy and apoptosis

Date: 24-04-2018    Supervisor: Henriette Pilegaard

The overall aim of this PhD thesis was to investigate autophagy, mitophagy and apoptosis and the possible connection between them, using both long-term and acute interventions. The hypotheses to be tested were 1) Training status affects the fasting-induced autophagy response in human skeletal muscle. 2) Aging affects autophagy, mitophagy and apoptosis in mice and human skeletal muscle and lifelong exercise training can prevent these changes. 3) Autophagy, mitophagy and apoptosis are regulated in human and mouse skeletal muscle during aging and lifelong exercise training through a PGC-1α-p53 axis. 4) A high fat-high fructose diet changes hepatic autophagy and mitophagy markers in mouse liver and exercise training can restore this through a PGC-1α mediated mechanism. 5) Exercise training modifies the exercise-induced regulation of autophagy and mitophagy in the liver of high fat-high fructose fed mice.

Study I demonstrated that 36h of fasting reduced LC3I, LC3II and Beclin1 protein content in skeletal muscle from untrained subjects, while ULK1 protein, Beclin1 protein and AMPKThr172 phosphorylation increased during fasting only in trained subjects. Moreover, skeletal muscle AKTThr308, AMPKThr172, ULKSer555, ULKSer757 phosphorylation, as well as Beclin1, ULK1, p62 and DRP1 protein levels were higher in trained than untrained subjects during fasting.

Study II demonstrated that aging increased LC3II, the LC3II/LC3I protein ratio and BNIP3 dimer protein and decreased PGC-1α, p53 and p21 mRNA, while lifelong exercise training prevented these changes in mouse skeletal muscle. In addition, aging decreased BAX/Bcl-2 ratio, LC3I and BAX protein in mouse skeletal musle, without effect of lifelong exercise training. PGC-1α was required for the age-induced reduction in p53 and p21 mRNA, as well as the effect of lifelong exercise training on the LC3II/LC3I ratio and BNIP3 dimer protein. In humans aging reduced PGC-1α and BAX mRNA as well as, p62 and p21 protein in skeletal muscle, but lifelong exercise training only prevented the decrease in p62 protein.

Study III demonstrated that high fat-high fructose (HFF) increased the LC3II/LC3I ratio, BNIP3 dimer and Parkin protein and reduced LC3I protein and PGC-1α mRNA in mouse liver. Exercise training reversed the change in the LC3II/LC3I protein ratio and PGC-1α mRNA, without changes in LC3II and p62 protein, as well as increased BNIP3 monomer protein and p53 mRNA in the liver. Acute exercise increased hepatic BNIP3 monomer, Parkin protein and PGC-1α mRNA in untrained HFF fed mice, while exercise training prevented these exercise-induced responses in HFF fed mice. In addition, hepatic p21 mRNA was increased by acute exercise in all groups. Moreover, there was no difference between HFF fed Lox/Lox and PGC-1α liver-specific KO mice in any of the autophagy or mitophagy markers.

In conclusion, fasting did not induce a coherent response in several autophagy markers, indicating that autophagy was not changed with fasting in human skeletal muscle. However, several markers were higher in trained subjects during fasting, indicating that the capacity for autophagy was higher in trained than untrained human skeletal muscle during fasting. The age-associated increase in autophagy and mitophagy markers were prevented by lifelong exercise training independent of PGC-1α, indicating that lifelong exercise training can prevent age-associated regulation of autophagy and mitophagy in mice. Conversely age-associated decreases in apoptotic marker BAX/Bcl-2 were not affected, while p53 mRNA was restored by lifelong exercise training, suggesting that apoptosis is not regulated by lifelong exercise training although p53 regulation may be restored. In human subjects the changes in autophagy, mitophagy and apoptosis markers were subtle, while p21-mediated muscle regeneration may be drastically reduced. High fat-high fructose may decrease the capacity for autophagy and increase BNIP3-mediated cell death, while exercise training prevent these changes, and increase the capacity for mitophagy. Moreover, exercise training prevented acute exercise-induced regulation of autophagy and mitophagy, which may indicate that damaged mitochondria were removed and mitochondrial quality was improved by exercise training.