**4. Autophagy, exercise and altitude exposure**

Attractive data concerning the role of autophagy during exercise are emerging. Autophagic vacuole formation during physical exercise was observed for the first time by Salminen and coworkers in 1984 with electron microscopy [156]. Nevertheless, there were no further studies on the topic until recently. In the last decade, data supporting the importance of autophagy in muscle homeostasis in response to exercise have been numerous, starting with a study from Bonaldo's team that showed that mice presenting impaired autophagy develop severe muscle weakness (i.e., accumulation of defective mitochondria, exacerbated apoptosis, muscle degeneration, and atrophy) [157]. Thereafter, other studies highlight that chronic inactivation of autophagy leads to a loss of metabolic effects related to exercise and drastic decreases in endurance performance [158].

Concerning autophagy modulation in response to acute exercise in humans, studies by Jamart and colleagues [159] were the first to demonstrate a raise of autophagy-regulatory genes and autophagic flux markers after ultraendurance exercise. By showing that AMPK and FoxO3a regulate in a coordinated way autophagy and ubiquitin-proteasome pathways during ultraendurance exercise, the authors gave an important picture of the cross-regulation of both degradation pathways in response to long-lasting endurance exercise [160]. Regarding more common endurance exercises, the modulation of muscle protein turnover, autophagy, and mitochondrial dynamics markers has been investigated thereafter in mice in response to different exercises conducted or not until exhaustion. Endurance exercise quickly initiates the autophagy pathway through Ulk1 activation resulting in an increase of autophagic flux, especially near exhaustion [161]. A rise in the phosphorylation of DRP1, a GTPase essential for mitochondrial fission, quickly occurred during exercise without any change in the expression of fusion markers (OPA1 and Mfn2). These data are consistent with an increase in mitophagy (i.e., the degradation of mitochondria by autophagy) since exacerbated fission can lead to mitochondrial fragmentation. Noteworthy, exercise decreases the activity of the main protein synthesis pathway (i.e., Akt/MTOR signaling pathway), from 90 min of moderate exercise (40– 50 % of VO2max), concomitantly to an increase in the phosphorylation of a marker of endoplas‐ mic reticulum stress (eiF2α Ser-51) [161]. Others studies reported an increase of endoplasmic reticulum stress by the evaluation of the content of the double-stranded RNA-activated protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK), the ER stress-induced transcription factor C/EBP homologous protein (CHOP), and the X-box binding protein 1 (XBP1s), in response to both ultraendurance [162] and moderate-intensity exercise [163]. However, reticulophagy has not been studied yet in skeletal muscle, especially in response to exercise. Further works are needed to clarify the possible clearance of important organelles, such as ribosomes or endoplasmic reticulum, during physical exercise. Exercise promotes better "cell health"; it would be not surprising to discover that exercise increases the turnover of such organelles like it is strongly suggested for mitochondria [21].

with AMPK; upon AICAR treatment, the complex remains stable for 3 h and then dissociates. As suggested by another group, Ulk1 dissociation from AMPK could permit to Ulk1 complex to be more active [154]. Conversely, this dissociation can constitute a negative regulatory feedback as proposed by Loffler et al. [155]. The authors showed that Ulk1 could mediate phosphorylation of AMPK on the regulatory subunits, constituting an inhibitory feedback control. Further works have to define the molecular mechanisms for these events, especially

Attractive data concerning the role of autophagy during exercise are emerging. Autophagic vacuole formation during physical exercise was observed for the first time by Salminen and coworkers in 1984 with electron microscopy [156]. Nevertheless, there were no further studies on the topic until recently. In the last decade, data supporting the importance of autophagy in muscle homeostasis in response to exercise have been numerous, starting with a study from Bonaldo's team that showed that mice presenting impaired autophagy develop severe muscle weakness (i.e., accumulation of defective mitochondria, exacerbated apoptosis, muscle degeneration, and atrophy) [157]. Thereafter, other studies highlight that chronic inactivation of autophagy leads to a loss of metabolic effects related to exercise and drastic decreases in

Concerning autophagy modulation in response to acute exercise in humans, studies by Jamart and colleagues [159] were the first to demonstrate a raise of autophagy-regulatory genes and autophagic flux markers after ultraendurance exercise. By showing that AMPK and FoxO3a regulate in a coordinated way autophagy and ubiquitin-proteasome pathways during ultraendurance exercise, the authors gave an important picture of the cross-regulation of both degradation pathways in response to long-lasting endurance exercise [160]. Regarding more common endurance exercises, the modulation of muscle protein turnover, autophagy, and mitochondrial dynamics markers has been investigated thereafter in mice in response to different exercises conducted or not until exhaustion. Endurance exercise quickly initiates the autophagy pathway through Ulk1 activation resulting in an increase of autophagic flux, especially near exhaustion [161]. A rise in the phosphorylation of DRP1, a GTPase essential for mitochondrial fission, quickly occurred during exercise without any change in the expression of fusion markers (OPA1 and Mfn2). These data are consistent with an increase in mitophagy (i.e., the degradation of mitochondria by autophagy) since exacerbated fission can lead to mitochondrial fragmentation. Noteworthy, exercise decreases the activity of the main protein synthesis pathway (i.e., Akt/MTOR signaling pathway), from 90 min of moderate exercise (40– 50 % of VO2max), concomitantly to an increase in the phosphorylation of a marker of endoplas‐ mic reticulum stress (eiF2α Ser-51) [161]. Others studies reported an increase of endoplasmic reticulum stress by the evaluation of the content of the double-stranded RNA-activated protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK), the ER stress-induced transcription factor C/EBP homologous protein (CHOP), and the X-box binding protein 1 (XBP1s), in response to both ultraendurance [162] and moderate-intensity exercise [163]. However,

in skeletal muscle.

182 Muscle Cell and Tissue

endurance performance [158].

**4. Autophagy, exercise and altitude exposure**

The rise of autophagy is essential to prevent mitochondrial damage during endurance exercise. Although acute inhibition of autophagy prior to exercise seems to not significantly affect performance, it leads to accumulation of dysfunctional mitochondria and augmentation of oxidative stress especially during eccentric contraction [164]. Thus, autophagy has a critical role in mitochondria quality control during acute exercise. In addition, autophagy is highly involved in exercise training-induced adaptations. Autophagy-deficient mice present attenu‐ ated improvement of endurance capacity in response to endurance training. In parallel to lower increases of basal autophagy flux, these mice show attenuated raises of mitochondrial content and angiogenesis [165], explaining the poor response to the training program.

The dietary factors have to be considered in autophagy response to acute exercise since essential amino acids (EAA) or carbohydrate (CHO) intake modulate protein turnover. Jamart et al. (2014) found that exercise performed in the fasted state permits a higher raise in auto‐ phagic flux indexes compared with the fed state. Concerning resistance exercise, few works showed a depression of autophagy markers after such an exercise like the study by Fry et al. conducted in humans [166]. In addition, autophagic flux markers can be depressed following EAA and carbohydrate (CHO) ingestion after resistance exercise [167]. To date, autophagy seems non-critical for muscle adaptations to resistance training [161]. However, an exception occurs during aging in which both endurance and resistance training are able to reverse the drop of autophagy regulatory proteins that occurs [168,169] (Fig.4).

Little is known regarding protein turnover pathways, especially autophagy, in response to exercise performed during altitude exposure in humans. Such an environment can induce a state of hypoxia that is exacerbated according to the level of altitude considered. Hypoxia results in decreased oxygen availability and leads to several hormonal, cardiorespiratory, and muscular adjustments in order to preserve cell homeostasis. Long-lasting hypoxia can cause a diminution of skeletal muscle mass and a reduction of muscle oxidative capacity. In agree‐ ment with an alteration of protein synthesis flux, hypoxia impairs the overload-induced increase of the PI3K/Akt/MTOR signaling pathway in rats [170]. Regarding degradation pathways in humans, while ubiquitin-proteasome system seems not positively modulated by environmental hypoxia, an upregulation of skeletal muscle autophagic flux markers has been found during acute normobaric hypoxia (10.7 % O2) and after exercise conducted in such an environment [171,172]. Another study investigated the effects of acute high-altitude exposure (at 5,300 m altitude) in the course of the Caudwell Research Expedition to Mt. Everest [173]. The authors notably found an upregulation of heat shock cognate 71 kDa protein involved in chaperone-mediated autophagy and a reduction of protein translation markers. Taken together, these studies seem to highlight a preventive role of autophagy for energy expenditure and an activation of chaperone-mediated autophagy during acute high-altitude exposure. However, the effects of chronic altitude exposure on autophagy and its combination with training remain to be characterized to date. Chronic hypoxia leads to a change in oxidative

**Figure 4.** Exercise and autophagy in skeletal muscle. Adapted from Sanchez AMJ et al. 2014. Autophagy is essential to support skeletal muscle plasticity in response to endurance exercise. American Journal of Physiology – Regulatory, In‐ tegrative and Comparative Physiology 307(8): R956-R969 [21]. Essential amino acids (EAA); carbohydrate (CHO)

metabolism [174,175]; it is likely that autophagy, especially mitochondrial autophagy, be amended during such an exposure. Consistent with this, experiments performed in cells showed that mitochondrial autophagy is induced by chronic hypoxia through HIF-1 (hypoxiadependent factor-1) and BNIP3, and this regulation constitutes a preventive response that is necessary to avoid accumulation of reactive oxygen species and cell death [176]. The effects of exposition to moderate altitude (i.e., 1,500–3,000 m) have also to be definite since it concerns a larger population (athletes, general tourist population, and highlanders) compared to high altitude.

Research of this type is leading to a better understanding of the autophagy-mediated turnover of organelles like mitochondria in response to exercise and altitude exposure. Finding optimal training strategies represents an important objective to enhance exercise adaptations in both athletes and patients with metabolic or muscle diseases, including COPD (chronic obstructive pulmonary disease) that can cause systemic hypoxia and loss in muscle capability.
