Functional and metabolic alterations in skeletal muscle in response to physiological and pathophysiological stressors

Abstract

Skeletal muscle performance is essential for our body’s movement as well as for the wholebody metabolism. In health and disease, skeletal muscle is exposed to various endogenous and exogenous stressors, influencing its physiological functions.

In paper I, we showed that exercise performance and muscle force is affected by the stressor breast cancer, in a mouse model of breast cancer (PyMT). Mimicking the experienced muscle weakness of human breast cancer patients, PyMT mice performed poorly in a treadmill exhaustion run and their muscles produced less force than wildtype (WT) mice, although no difference in morphology, fiber type distribution or diameter was found. The muscle weakness was associated with an increase of pro-inflammatory cytokines, such as TNF-α in the skeletal muscle, activating the p38 mitogen-activated protein kinase (MAPK) stress-response pathway and decreasing the expression of mitochondrial electron transport chain (ETC) genes as well as antioxidant genes. After the mice had access to four weeks of voluntary running ad libitum, skeletal muscle force as well as the time and distance of the treadmill exhaustion run improved drastically for PyMT mice. The exercise also reduced the intramuscular stress, improved both the expression of mitochondrial ETC genes and the activity of key mitochondrial enzymes, such as citrate synthase (CS) and especially β-hydroxyacyl-CoA dehydrogenase (β-HAD) and restored the antioxidant defense system including superoxide dismutase (SOD 1,2). Additionally, we could show that the breast cancer blunted the exercise-induced expression of PPARγ coactivator-1α (Pgc-1α) in PyMT mice. Our results showed that breast cancer-induced weakness is linked to increased intramuscular stress signaling and that voluntary, moderate exercise was able to counteract the weakness in PyMT mice.

Patients with breast cancer are treated with various systemic anti-cancer treatments, and while aiming to treat cancer, these treatments often cause side-effects. In paper II, we aimed to study the effect of a novel anti-tumorigenic compound (CX-5461) on the whole-body as well skeletal muscle-specific metabolism. Four weeks of CX-5461 treatment effectively reduced the breast cancer tumor in PyMT mice, but also resulted in increased food intake, energy expenditure and a higher respiratory exchange ratio (VCO2/VO2), indicative of a substrate shift towards carbohydrate utilization in both WT and PyMT mice. Moreover, basal blood glucose levels were increased, and we observed a slower glucose clearance from the blood stream in both WT and PyMT mice after CX-5461 treatment. Skeletal muscle is an important tissue involved in maintaining the body’s glucose homeostatic. In WT mice, CX-5461 treatment reduced the basal glucose uptake, whereas in PyMT mice the insulin-stimulated glucose uptake was affected in extensor digitorum longus (EDL) muscles. We found that CX-5461 not only exerts its mechanism of action, the inhibition of the RNA-Polymerase I (Pol I) pre-initiation complex, in breast cancer cells, but also directly in skeletal muscle and through that alters the glucose and fat metabolism in skeletal muscle. The results indicate that the novel drug CX-5461 affects the whole-body metabolism including elevated blood glucose levels and reduced glucose uptake into skeletal muscle, independently of the tumor development.

Skeletal muscle is a highly metabolic tissue which functions can also be regulated by oxidative stress, cause by an imbalance in the endogenous oxidative and antioxidative system. In paper III, we investigated the role of the intermuscular redox state on glycogen phosphorylase activity and glycogenolysis, which supply the muscle with energy from glycogen storage during exercise. Glycogen phosphorylase was strongly inhibited by incubation with the reactive nitrogen species (RNS) peroxynitrite (ONOO-), contrary to the reactive oxygen species (ROS) H2O2 in muscle extracts. In intact muscles, ONOOincubation resulted in inhibition of glycogenolysis in resting and contracting as well as a reduction of muscle force in slow-twitch oxidative soleus (SOL) and fast-twitch glycolytic EDL muscles, despite not exerting a direct effect on phosphorylase activity. Moreover, post-translational nitrate modification was observed in EDL muscle after ONOOincubation. Incubation with two antioxidants N-acetylcysteine (NAC) and dithiothreitol (DTT) did not affect phosphorylase activity or glycogenolysis, but reduced the force of EDL and SOL muscle. These results suggest that exogenous ONOOinhibits phosphorylase activity in muscle extracts and glycogenolysis in intact contracted muscles, whereas antioxidants such as DTT and NAC only play a minor role in inducing endogenous ROS and regulate the phosphorylase activity.

All results from these three studies in this thesis investigate how the performance and function of skeletal muscle can be affected by different stressors. Taken together, a better understanding of the responsible underlying molecular mechanisms, might lead to targeted therapy approaches for afflicted patients in the future.