Mitochondrial protein S-nitrosation protects against ischemia reperfusion-induced denervation at neuromuscular junction in skeletal muscle.

Deterioration of neuromuscular junction (NMJ) integrity and function is causal to muscle atrophy and frailty, ultimately hindering quality of life and increasing the risk of death. In particular, NMJ is vulnerable to ischemia reperfusion (IR) injury when blood flow is restricted followed by restoration. However, little is known about the underlying mechanism(s) and hence the lack of effective interventions. New evidence suggests that mitochondrial oxidative stress plays a causal role in IR injury, which can be precluded by enhancing mitochondrial protein S-nitrosation (SNO). To elucidate the role of IR and mitochondrial protein SNO in skeletal muscle, we utilized a clinically relevant model and showed that IR resulted in significant muscle and motor nerve injuries with evidence of elevated muscle creatine kinase in the serum, denervation at NMJ, myofiber degeneration and regeneration, as well as muscle atrophy. Interestingly, we observed that neuromuscular transmission improved prior to muscle recovery, suggesting the importance of the motor nerve in muscle functional recovery. Injection of a mitochondria-targeted S-nitrosation enhancing agent, MitoSNO, into ischemic muscle prior to reperfusion reduced mitochondrial oxidative stress in the motor nerve and NMJ, attenuated denervation at NMJ, and resulted in accelerated functional recovery of the muscle. These findings demonstrate that enhancing mitochondrial protein SNO protects against IR-induced denervation at NMJ in skeletal muscle and accelerates functional regeneration. This could be an efficacious intervention for protecting neuromuscular injury under the condition of IR and other related pathological conditions.

Ampk phosphorylation of Ulk1 is required for targeting of mitochondria to lysosomes in exercise-induced mitophagy.

Mitochondrial health is critical for skeletal muscle function and is improved by exercise training through both mitochondrial biogenesis and removal of damaged/dysfunctional mitochondria via mitophagy. The mechanisms underlying exercise-induced mitophagy have not been fully elucidated. Here, we show that acute treadmill running in mice causes mitochondrial oxidative stress at 3-12 h and mitophagy at 6 h post-exercise in skeletal muscle. These changes were monitored using a novel fluorescent reporter gene, pMitoTimer, that allows assessment of mitochondrial oxidative stress and mitophagy in vivo, and were preceded by increased phosphorylation of AMP activated protein kinase (Ampk) at tyrosine 172 and of unc-51 like autophagy activating kinase 1 (Ulk1) at serine 555. Using mice expressing dominant negative and constitutively active Ampk in skeletal muscle, we demonstrate that Ulk1 activation is dependent on Ampk. Furthermore, exercise-induced metabolic adaptation requires Ulk1. These findings provide direct evidence of exercise-induced mitophagy and demonstrate the importance of Ampk-Ulk1 signaling in skeletal muscle.Exercise is associated with biogenesis and removal of dysfunctional mitochondria. Here the authors use a mitochondrial reporter gene to demonstrate the occurrence of mitophagy following exercise in mice, and show this is dependent on AMPK and ULK1 signaling.