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Find out more on how to host your own Frontiers Research Topic or contribute to one as an author by contacting the Frontiers Editorial Office: researchtopics@frontiersin.org 2 January 2018 | High-Intensity Exer cise in Hypoxia Frontiers in Physiology HIGH-INTENSITY EXERCISE IN HYPOXIA - BENEFICIAL ASPECTS AND POTENTIAL DRAWBACKS Image: Maridav/Shutterstock.com Topic Editors: Olivier Girard, Qatar Orthopaedic and Sports Medicine Hospital, Qatar Donald R. McCrimmon, Northwestern University, United States Gregoire P. Millet, University of Lausanne, Switzerland In the past, ‘traditional’ moderate-intensity continuous training (60-75% peak heart rate) was the type of physical activity most frequently recommended for both athletes and clinical populations (cf. American College of Sports Medicine guidelines). However, growing evidence indicates that high-intensity interval training (80-100% peak heart rate) could actually be associated with larger cardiorespiratory fitness and metabolic function benefits and, thereby, physical performance gains for athletes. Similarly, recent data in obese and hypertensive individuals indicate that various mechanisms – further improvement in endothelial function, reductions in sympathetic neural activity, or in arterial stiffness – might be involved in the larger cardiovascular protective effects associated with training at high exercise intensities. 3 January 2018 | High-Intensity Exer cise in Hypoxia Frontiers in Physiology Concerning hypoxic training, similar trends have been observed from ‘traditional’ prolonged altitude sojourns (‘Live High Train High’ or ‘Live High Train Low’), which result in increased hemoglobin mass and blood carrying capacity. Recent innovative ‘Live Low Train High’ meth- ods (‘Resistance Training in Hypoxia’ or ‘Repeated Sprint Training in Hypoxia’) have resulted in peripheral adaptations, such as hypertrophy or delay in muscle fatigue. Other interventions inducing peripheral hypoxia, such as vascular occlusion during endurance/resistance training or remote ischemic preconditioning (i.e. succession of ischemia/reperfusion episodes), have been proposed as methods for improving subsequent exercise performance or altitude tolerance (e.g. reduced severity of acute-mountain sickness symptoms). Postulated mechanisms behind these metabolic, neuro-humoral, hemodynamics, and systemic adaptations include stimulation of nitric oxide synthase, increase in anti-oxidant enzymes, and down-regulation of pro-inflamma- tory cytokines, although the amount of evidence is not yet significant enough. Improved O 2 delivery/utilization conferred by hypoxic training interventions might also be effective in preventing and treating cardiovascular diseases, as well as contributing to improve exercise tolerance and health status of patients. For example, in obese subjects, combining exer- cise with hypoxic exposure enhances the negative energy balance, which further reduces weight and improves cardio-metabolic health. In hypertensive patients, the larger lowering of blood pressure through the endothelial nitric oxide synthase pathway and the associated compensatory vasodilation is taken to reflect the superiority of exercising in hypoxia compared to normoxia. A hypoxic stimulus, in addition to exercise at high vs. moderate intensity, has the potential to further ameliorate various aspects of the vascular function, as observed in healthy populations. This may have clinical implications for the reduction of cardiovascular risks. Key open questions are therefore of interest for patients suffering from chronic vascular or cellular hypoxia (e.g. work-rest or ischemia/reperfusion intermittent pattern; exercise intensity; hypoxic severity and exposure duration; type of hypoxia (normobaric vs. hypobaric); health risks; magnitude and maintenance of the benefits). Outside any potential beneficial effects of exercising in O 2-deprived environments, there may also be long-term adverse consequences of chronic intermittent severe hypoxia. Sleep apnea syndrome, for instance, leads to oxidative stress and the production of reactive oxygen species, and ultimately systemic inflammation. Postulated pathophysiological changes associated with intermittent hypoxic exposure include alteration in baroreflex activity, increase in pulmonary arterial pressure and hematocrit, changes in heart structure and function, and an alteration in endothelial-dependent vasodilation in cerebral and muscular arteries. There is a need to explore the combination of exercising in hypoxia and association of hypertension, developmental defects, neuro-pathological and neuro-cognitive deficits, enhanced susceptibility to oxidative injury, and possibly increased myocardial and cerebral infarction in individuals sensitive to hypoxic stress. The aim of this Research Topic is to shed more light on the transcriptional, vascular, hemody- namics, neuro-humoral, and systemic consequences of training at high intensities under various hypoxic conditions. Citation: Girard, O., McCrimmon, D. R., Millet, G. P., eds. (2018). High-Intensity Exercise in Hypoxia - Beneficial Aspects and Potential Drawbacks. Lausanne: Frontiers Media. doi: 10.3389/978-2-88945-406-8 4 January 2018 | High-Intensity Exer cise in Hypoxia Frontiers in Physiology Table of Contents 06 Editorial: High-Intensity Exercise in Hypoxia: Beneficial Aspects and Potential Drawbacks Grégoire P . Millet and Olivier Girard High-Intensity, Continuous Exercise in Hypoxia 10 Twin Resemblance in Muscle HIF-1 a Responses to Hypoxia and Exercise Ruud Van Thienen, Evi Masschelein, Gommaar D’Hulst, Martine Thomis and Peter Hespel 21 Prediction of Critical Power and W ¢ in Hypoxia: Application to Work-Balance Modelling Nathan E. Townsend, David S. Nichols, Philip F . Skiba, Sebastien Racinais and Julien D. Périard 30 Maximal Oxygen Uptake Is Achieved in Hypoxia but Not Normoxia during an Exhaustive Severe Intensity Run Matthew I. Black, Christopher R. Potter, Jo Corbett, Cain C. T. Clark and Stephen B. Draper 37 Task Failure during Exercise to Exhaustion in Normoxia and Hypoxia Is Due to Reduced Muscle Activation Caused by Central Mechanisms While Muscle Metaboreflex Does Not Limit Performance Rafael Torres-Peralta, David Morales-Alamo, Miriam González-Izal, José Losa-Reyna, Ismael Pérez-Suárez, Mikel Izquierdo and José A. L. Calbet 52 Increased P I O2 at Exhaustion in Hypoxia Enhances Muscle Activation and Swiftly Relieves Fatigue: A Placebo or a P I O 2 Dependent Effect? Rafael Torres-Peralta, José Losa-Reyna, David Morales-Alamo, Miriam González-Izal, Ismael Pérez-Suárez, Jesús G. Ponce-González, Mikel Izquierdo and José A. L. Calbet Repeated Sprint Training in Hypoxia 67 High-Intensity Exercise in Hypoxia: Is Increased Reliance on Anaerobic Metabolism Important? Brendan R. Scott, Paul S. R. Goods and Katie M. Slattery 71 Changes in Muscle and Cerebral Deoxygenation and Perfusion during Repeated Sprints in Hypoxia to Exhaustion Sarah J. Willis, Laurent Alvarez, Grégoire P . Millet and Fabio Borrani 83 Variations in Hypoxia Impairs Muscle Oxygenation and Performance during Simulated Team-Sport Running Alice J. Sweeting, François Billaut, Matthew C. Varley, Ramón F . Rodriguez, William G. Hopkins and Robert J. Aughey 5 January 2018 | High-Intensity Exer cise in Hypoxia Frontiers in Physiology 94 High Altitude Increases Alteration in Maximal Torque but Not in Rapid Torque Development in Knee Extensors after Repeated Treadmill Sprinting Olivier Girard, Franck Brocherie and Grégoire P . Millet 107 Nitrate Intake Promotes Shift in Muscle Fiber Type Composition during Sprint Interval Training in Hypoxia Stefan De Smet, Ruud Van Thienen, Louise Deldicque, Ruth James, Craig Sale, David J. Bishop and Peter Hespel 118 Similar Inflammatory Responses following Sprint Interval Training Performed in Hypoxia and Normoxia Alan J. Richardson, Rebecca L. Relf, Arron Saunders and Oliver R. Gibson 128 Hypoxic Repeat Sprint Training Improves Rugby Player’s Repeated Sprint but Not Endurance Performance Michael J. Hamlin, Peter D. Olsen, Helen C. Marshall, Catherine A. Lizamore and Catherine A. Elliot Resistance Training in Hypoxia 138 Heavy Resistance Training in Hypoxia Enhances 1RM Squat Performance Mathew W. H. Inness, François Billaut, Emily J. Walker, Aaron C. Petersen, Alice J. Sweeting and Robert J. Aughey 146 Sex-Specific Impact of Ischemic Preconditioning on Tissue Oxygenation and Maximal Concentric Force Pénélope Paradis-Deschênes, Denis R. Joanisse and François Billaut Therapeutic Use of Hypoxia 155 Walking in Hypoxia: An Efficient Treatment to Lessen Mechanical Constraints and Improve Health in Obese Individuals? Olivier Girard, Davide Malatesta and Grégoire P . Millet 161 Endurance Training in Normobaric Hypoxia Imposes Less Physical Stress for Geriatric Rehabilitation Stephan Pramsohler, Martin Burtscher, Martin Faulhaber, Hannes Gatterer, Linda Rausch, Arn Eliasson and Nikolaus C. Netzer EDITORIAL published: 04 December 2017 doi: 10.3389/fphys.2017.01017 Frontiers in Physiology | www.frontiersin.org December 2017 | Volume 8 | Article 1017 | Edited and reviewed by: François Billaut, Laval University, Canada *Correspondence: Grégoire P. Millet gregoire.millet@unil.ch Specialty section: This article was submitted to Exercise Physiology, a section of the journal Frontiers in Physiology Received: 31 October 2017 Accepted: 23 November 2017 Published: 04 December 2017 Citation: Millet GP and Girard O (2017) Editorial: High-Intensity Exercise in Hypoxia: Beneficial Aspects and Potential Drawbacks. Front. Physiol. 8:1017. doi: 10.3389/fphys.2017.01017 Editorial: High-Intensity Exercise in Hypoxia: Beneficial Aspects and Potential Drawbacks Grégoire P. Millet 1 * and Olivier Girard 2 1 Faculty of Biology and Medicine, Institute of Sport Sciences, University of Lausanne, Lausanne, Switzerland, 2 Athlete Health and Performance Research Centre, Qatar Orthopaedic and Sports Medicine Hospital, Doha, Qatar Keywords: altitude training, deoxygenation, repeated sprint training in hypoxia, resistance training in hypoxia, muscle activation, HIF-1 α , ischemic preconditioning, hypoxia Editorial on the Research Topic High-Intensity Exercise in Hypoxia: Beneficial Aspects and Potential Drawbacks RECENT DEVELOPMENTS IN HYPOXIC TRAINING With the recent development of new altitude training methods (Millet et al., 2013; Girard et al., 2017), the question of the specific central and peripheral adaptations to high-intensity exercise in hypoxia is now crucial. This research topic investigated the beneficial aspects and potential drawbacks of these methods and would undoubtedly be of interest for many exercise physiologists. A total of 16 papers have been accepted, arising from 18 different research groups from 12 countries. Four different main areas have been investigated: 1. High-intensity, continuous exercise in hypoxia. 2. Repeated sprint training in hypoxia. 3. Resistance training in hypoxia. 4. Therapeutic use of hypoxia. HIGH-INTENSITY, CONTINUOUS EXERCISE IN HYPOXIA Van Thienen et al. investigated the HIF-1 pathway (from vastus lateralis biopsies) in 11 monozygotic twin pairs who performed an experimental trial in both normoxia and hypoxia. They tested the hypothesis that this pathway and its downstream targets in energy metabolism are regulated in a genotype-dependent manner. A key observation was that hypoxic exercise-induced increment of muscle HIF-1a mRNA content was about 10-fold more similar within monozygotic twins than between the twins. Authors concluded that genetic factors play an important role in the muscular responses to acute hypoxic stress at rest and during exercise and that the regulation of HIF-1a stabilization in acute hypoxia is genotype-dependent. Townsend et al. computed the critical power (CP) and the work above CP (W’) in male cyclists performing time trials in normoxia and at five different altitudes from 250 to 4,250 m. They predicted performance during a high-intensity intermittent test performed in normoxia and in normobaric hypoxia (simulated altitude: 2,250 m). They reported a curvilinear decrease in CP with increase in altitude severity as well as a significant decrease in W ′ occurring only at 4,250 m. Practically, this study enables the prescription of equivalent relative intensity interval training workouts in hypoxic conditions compared with normoxia. 6 Millet and Girard High-Intensity Exercise in Hypoxia Black et al. explored whether there is a difference in the percentage of VO2max achieved (during a 2-min exhaustive run) in normoxia and hypoxia in 14 middle distance runners. Compared to normoxia, VO2max was lower during a ramp test and VO2 kinetics (greater time constant of the primary phase) were slower in hypoxia. Whereas the runners were unable to reach VO2max during the exhaustive constant work- rate run lasting ∼ 2 min in normoxia, they were able to achieve the reduced VO2max in a hypoxia despite slower VO2 kinetics. Torres-Peralta et al. investigated the contribution of central and peripheral mechanisms during exercise to exhaustion in normoxia and hypoxia. Following the exercise to exhaustion, legs circulation was occluded during 10 or 60s for impeding recovery and increasing the metaboreflex. The fact that task failure was apparently not due to muscle peripheral fatigue, but instead primarily resulted from reduction in muscle activation, highlights the importance of central mechanisms. The same research group Torres-Peralta et al. investigated the role played by different levels of inspired pressure in O2 (P I O 2 ) on muscle activation during exhaustive exercise. A unique observation was that the increase in P I O 2 at exhaustion reduced fatigue and allowed exercise continuation. This study therefore indicates that severe hypoxia induces larger central fatigue (decrease in muscle activation). REPEATED SPRINT TRAINING IN HYPOXIA Decrease in convective factors, in turn leading to a reduced training intensity (i.e., not sufficiently intense to stress O 2 delivery and maximized adaptations), is an inherent characteristic of interval-training in hypoxia (IHT) compared with normoxia. To overcome this limitation, the so-called “repeated-sprint training in hypoxia” or RSH has been developed as a new intervention in our laboratory in Lausanne (Faiss et al., 2013a): with exercise intensity being maximal during RSH we have postulated that this would allow a better recruitment of fast-twitch muscle fibers (Faiss et al., 2013a,b). An up-regulation of circulating microRNAs levels was observed only when exercise was performed at high-intensity and high altitude (i.e., and not at lower intensity) and therefore RSH training is based on the repetition of short ( < 30 s) “all-out” sprints with incomplete recoveries in hypoxia (Vogt et al., 2001; Faiss et al., 2013a). Hence, a lower rate of O 2 delivery to the muscles increases the stress on glycolytic flux, which may stimulate the up-regulation of this energy pathway. Compared with repeated-sprint training in normoxia (RSN), RSH could induce beneficial adaptations at the muscular level, along with improved blood perfusion, which may lead to greater improvements in repeated-sprint ability. Superior repeated-sprint ability in normoxic conditions has been associated with completion of RSH vs. RSN in cohorts of rugby players (Galvin et al., 2013), well-trained cyclists (Faiss et al., 2013b), cross-country skiers (Faiss et al., 2015), soccer players (Gatterer et al., 2014; Brocherie et al., 2015a), field hockey players (Brocherie et al., 2015b). The effectiveness of RSH was confirmed by a recent meta-analysis based on 9 controlled studies (all published in the past 4 years) showing larger mean performance (Brocherie et al., 2017). Blood lactate accumulation is higher at a simulated altitude of 4,000 m when compared with more moderate simulated altitudes during repeated treadmill sprints (Goods et al., 2014). In an opinion letter, Scott et al. stated that RSH led to a greater reliance on anaerobic metabolism. This increased metabolic stress is likely to promote peripheral fatigue resistance induced by RSH. Using near-infrared spectroscopy it was previously reported that prefrontal cortex, but not muscle, oxygenation is impaired when ten, 10-s sprints (with 10 s of rest) are completed at 13 vs. 21% oxygen (Smith and Billaut, 2010). For the first time, Willis et al. compared muscle and cerebral oxygenation trends during a one-off RSH trial performed to exhaustion in normoxia (400 m) and at two different simulated altitudes (2,000 and 3,800 m). There was a continual decrease in convective factors of oxygen delivery (e.g., decreases in pulse oxygen saturation and peak oxygen uptake) with increased hypoxia severity, which was linked with impairment in performance (number of sprints and total work) across conditions. Cerebral deoxygenation demonstrated greater changes at 3,800 m compared with 400 and 2,000 m, as well As well as larger deoxygenation/reoxygenation levels during sprints/recoveries near exhaustion. These results show that central autoregulation (i.e., increase in cerebral perfusion near exhaustion) occurs in order to continue exercise despite limited peripheral and cerebral oxygen delivery, until a certain point of limited diffusion at which protective mechanisms cause exercise cessation. Sweeting et al. reported that repeated-sprint and single-sprint efforts are compromised at 3,000 m simulated altitude, possibly due to limited muscle O 2 availability during recovery periods. Whilst repeated-sprint and single-sprint efforts were maintained at 2,000 m, the elevated physiological demands at 3,000 m may have been overwhelming. Girard et al. investigated the neuromuscular adjustments following repeated treadmill sprints at simulated altitudes of 1,800 and 3,600 m or in normoxia. Post-exercise decrease in voluntary strength of knee extensors was greater at 3,800 m than at 1,800 m and normoxia. However, the exercise-induced alterations in rapid torque development were similar between the three conditions. De Smet et al. investigated if oral nitrate intake influenced buffering capacity and fiber type distribution (vastus lateralis biopsies) after 5 weeks of sprint interval training in hypoxia or in normoxia. Altogether, sprint interval training in hypoxia did not lead to enhanced aerobic or anaerobic endurance exercise performance but oral nitrate supplementation increased the proportion of type IIa muscle fibers. Richardson et al. reported that the improvement in VO 2peak and the inflammatory responses (IL-6 and TNFa) were similar after 2 weeks of repeated sprint performed in normoxia or in hypoxia. However, improvement in anaerobic threshold was observed only after RSH. Frontiers in Physiology | www.frontiersin.org December 2017 | Volume 8 | Article 1017 | 7 Millet and Girard High-Intensity Exercise in Hypoxia Hamlin et al. reported the performance changes after six sessions of RSH vs RSN in 19 well-trained male rugby players. These authors confirmed the effectiveness of RSH since repeated sprint performance was enhanced to a larger extent whereas the aerobic performance did not change. RESISTANCE TRAINING IN HYPOXIA There is growing research interest focusing on the so-called “resistance training in hypoxia” (RTH) (Scott et al., 2014). A classical reasoning is that, in an O 2 -deprived environment, the low partial pressure of O 2 would increase metabolite (e.g., blood [La] and anabolic hormones [e.g., growth hormone]) accumulation, leading to an accelerated recruitment of higher threshold motor units (Manimmanakorn et al., 2013) and a subsequent higher hypertrophy with eventually greater improvements in muscle strength (Scott et al., 2015). Here, Inness et al. examined the effects of 20 sessions of heavy resistance training performed either in hypoxia (RTH) or in normoxia. RTH induced a larger enhancement in absolute and relative strength as well as 1RM. Paradis-Deschênes et al. investigated the effects of ischemic preconditioning on knee extensions in strength-trained male vs. female athletes. Males reported a greater peripheral oxygen extraction and greater strength enhancement than females. That said, a recent meta-analysis (Ramos-Campo et al., 2017) concluded that RTH did not provide significant benefit for muscle size and strength over resistance training in normoxia. This highlights the need for additional research on this burgeoning area. THERAPEUTIC USE OF HYPOXIA Potential benefits of using hypoxia exposure therapeutically have been recently suggested for elderly, obese or hypertensive patients (Millet et al., 2016). Here, Girard et al. postulated that hypoxic walking would be beneficial in obese patients since it might lead to a decreased walking speed and subsequently a lower biomechanical load. Along the same lines, Pramsohler et al. assessed the physical effort in geriatric patients (age > 65 years) for the same HR response in normoxia or in hypoxia (simulated altitude: 3,000 m). The main benefits of the hypoxic sessions were a lower stress on the locomotor systems for a similar physiological strain than in normoxia. CONCLUSION As confirmed by this research topic, high-intensity exercise in hypoxia is a growing area of interest. Ergogenic effect? Stamped! Adaptive mechanisms? More investigation needed! Therapeutic usefulness? The future! AUTHOR CONTRIBUTIONS All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication. REFERENCES Brocherie, F., Girard, O., Faiss, R., and Millet, G. P. (2015a). High-intensity intermittent training in hypoxia: a double-blinded, placebo-controlled field study in youth football players. J. Strength Cond. Res. 29, 226–237. doi: 10.1519/JSC.0000000000000590 Brocherie, F., Girard, O., Faiss, R., and Millet, G. P. (2017). Effects of repeated-sprint training in hypoxia on sea-level performance: a meta-analysis. Sports Med. 47, 1651–1660. doi: 10.1007/s40279-017- 0685-3 Brocherie, F., Millet, G. P., Hauser, A., Steiner, T., Rysman, J., Wehrlin, J. P., et al. (2015b). “Live high-train low and high” hypoxic training improves team-sport performance. Med. Sci. Sports Exerc. 47, 2140–2149. doi: 10.1249/MSS.0000000000000630 Faiss, R., Girard, O., and Millet, G. P. (2013a). Advancing hypoxic training in team sports: from intermittent hypoxic training to repeated sprint training in hypoxia. Br. J. Sports Med. 47(Suppl. 1), i45–i50. doi: 10.1136/bjsports-2013-092741 Faiss, R., Léger, B., Vesin, J. M., Fournier, P. E., Eggel, Y., Dériaz, O., et al. (2013b). Significant molecular and systemic adaptations after repeated sprint training in hypoxia. PLoS ONE 8:e 56522. doi: 10.1371/journal.pone.0056522 Faiss, R., Willis, S., Born, D. P., Sperlich, B., Vesin, J. M., Holmberg, H. C., et al. (2015). Repeated double-poling sprint training in hypoxia by competitive cross-country skiers. Med. Sci. Sports Exerc. 47, 809–817. doi: 10.1249/MSS.0000000000000464 Galvin, H. M., Cooke, K., Sumners, D. P., Mileva, K. N., and Bowtell, J. L. (2013). 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Conflict of Interest Statement: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. Copyright © 2017 Millet and Girard. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Frontiers in Physiology | www.frontiersin.org December 2017 | Volume 8 | Article 1017 | 9 ORIGINAL RESEARCH published: 18 January 2017 doi: 10.3389/fphys.2016.00676 Frontiers in Physiology | www.frontiersin.org January 2017 | Volume 7 | Article 676 | Edited by: Olivier Girard, Aspetar Orthopedic and Sports Medicine Hospital, Qatar Reviewed by: Tyler John Kirby, Cornell University, USA Gregoire P. Millet, University of Lausanne, Switzerland *Correspondence: Peter Hespel peter.hespel@faber.kuleuven.be † These authors have contributed equally to this work. Specialty section: This article was submitted to Exercise Physiology, a section of the journal Frontiers in Physiology Received: 15 September 2016 Accepted: 20 December 2016 Published: 18 January 2017 Citation: Van Thienen R, Masschelein E, D’Hulst G, Thomis M and Hespel P (2017) Twin Resemblance in Muscle HIF-1 α Responses to Hypoxia and Exercise. Front. Physiol. 7:676. doi: 10.3389/fphys.2016.00676 Twin Resemblance in Muscle HIF-1 α Responses to Hypoxia and Exercise Ruud Van Thienen 1 † , Evi Masschelein 1 † , Gommaar D’Hulst 1 , Martine Thomis 2 and Peter Hespel 1 * 1 Exercise Physiology Research Group, Department of Kinesiology, KU Leuven, Leuven, Belgium, 2 Physical Activity, Sports and Health Research Group, Department of Kinesiology, KU Leuven, Leuven, Belgium Hypoxia-inducible factor-1 (HIF-1) is a master regulator of myocellular adaptation to exercise and hypoxia. However, the role of genetic factors in regulation of HIF-1 responses to exercise and hypoxia is unknown. We hypothesized that hypoxia at rest and during exercise stimulates the HIF-1 pathway and its downstream targets in energy metabolism regulation in a genotype-dependent manner. Eleven monozygotic twin (MZ) pairs performed an experimental trial in both normoxia and hypoxia (FiO 2 10.7%). Biopsies were taken from m. vastus lateralis before and after a 20-min submaximal cycling bout @ ∼ 30% of sea-level VO 2 max. Key-markers of the HIF-1 pathway and glycolytic and oxidative metabolism were analyzed using real-time PCR and Western Blot. Hypoxia increased HIF-1 α protein expression by ∼ 120% at rest vs. + 150% during exercise ( p < 0.05). Furthermore, hypoxia but not exercise increased muscle mRNA content of HIF-1 α ( + 50%), PHD2 ( + 45%), pVHL ( + 45%; p < 0.05), PDK4 ( + 1200%), as well as PFK-M ( + 20%) and PPAR- γ 1 ( + 60%; p < 0.05). Neither hypoxia nor exercise altered PHD1, LDH-A, PDH-A1, COX-4, and CS mRNA expressions. The hypoxic, but not normoxic exercise-induced increment of muscle HIF-1 α mRNA content was about 10-fold more similar within MZ twins than between the twins ( p < 0.05). Furthermore, in resting muscle the hypoxia-induced increments of muscle HIF-1 α protein content, and HIF-1 α and PDK4 mRNA content were about 3–4-fold more homogeneous within than between the twins pairs ( p < 0.05). The present observations in monozygotic twins for the first time clearly indicate that the HIF-1 α protein as well as mRNA responses to submaximal exercise in acute hypoxia are at least partly regulated by genetic factors. Keywords: monozygotic twin design, HIF-1, exercise, hypoxia, muscle biopsies INTRODUCTION Whenever the human body is exposed to oxygen deficiency, numerous physiological responses are initiated. Adaptations at both the cardiovascular, respiratory, neurological and skeletal muscle level (Petousi and Robbins, 2014) aim to maintain adequate oxygen uptake and delivery so as to preserve cellular energy homeostasis and tissue integrity. At the level of skeletal muscles, differential mechanisms are involved in the response to either acute or chronic hypoxia. For instance, in acute hypoxic stress fuel selection is shifted from fatty acids to carbohydrates, which increases the ATP yield per molecule of oxygen consumed. Such mechanism not only facilitates energy homeostasis (Hoppeler and Vogt, 2001; Hoppeler et al., 2008; Murray, 2009) but also protects against excessive 10 Van Thienen et al. Genetics in HIF-1 α and Hypoxia mitochondrial production of reactive oxygen species (Zhang et al., 2008; Murray, 2009; Edwards et al., 2010). Furthermore, acute hypoxia also regulates gene expression of several rate- limiting enzymes in the primary energy pathways, i.e., glycolysis, the Krebs cycle, as well as oxidative phosphorylation (Zoll et al., 2006; Murray and Horscroft, 2016). These alterations eventually result in a downregulation of oxidative energy production, vs. upregulation of anaerobic ATP production via glycolysis, aiming to assure adequate rates of sustained ATP-production whenever abundant oxygen supply is lacking, most prominently during exercise (Murray, 2009). The effects of chronic hypoxia on skeletal muscle on the other hand serve to facilitate oxygen diffusion in muscle tissue by stimulation of neovascularization vs. decrease of muscle fiber cross-sectional area, which reduces oxygen diffusion distance (Hoppeler and Vogt, 2001; Deldicque and Francaux, 2013). Furthermore, loss of mitochondrial density and mitochondrial uncoupling decreased ROS production, which might otherwise be exaggerated especially during hypoxic exercise (Murray and Horscroft, 2016). It is the prevailing opinion that hypoxia-inducible factor 1 (HIF-1) plays a pivotal role in myocellular adaptations to hypoxia (Vogt et al., 2001). HIF-1 is a heterodimeric transcription factor, built of a HIF-1 α and a HIF-1 β subunit, and serves as an intracellular oxygen-sensor to trigger cellular responses needed to cope with any drop of intramyocellular oxygen tension (Ameln et al., 2005; Mason and Johnson, 2007). In normoxic conditions, in contrast to hypoxia, the HIF-1 α subunit is not hydroxylated and is immediately degraded. Conversely, in hypoxia, HIF- 1 α accumulates in the cytosol and migrates to the nucleus to dimerize with the HIF-1 β subunit. The heterodimer so formed can bind to the regulatory domain of different target genes which in turn initiate the concerted cellular response to the hypoxic stress. The importance of HIF-1 in regulation of metabolic genes, including all glycolytic enzymes, pyruvate dehydrogenase kinase 1 (PDK1) and subunit 4-2 of cytochrome c oxidase (COX), but also regulation of angiogenesis is well established (Ameln et al., 2005; Papandreou et al., 2006; Mason and Johnson, 2007; Murray, 2009). Activation of HIF-1 in hypoxic conditions also leads to enhanced mitochondrial autophagy and a decrease in mitochondrial biogenesis and respiration (Zhang et al., 2008). However, data on the inter-individual variability of HIF-1 responses to hypoxia and the role of heritability in HIF-1 regulation in muscle are lacking. Nonetheless, we previously found genetic variants to be important in explaining some specific muscular responses to hypoxia (regulation of maximal oxygen uptake and protein metabolism) (Masschelein et al., 2014, 2015a). Furthermore, epidemiological studies in high- altitude natives to explore the incidence of gene polymorphisms that may be beneficial for survival at high altitude, have provided evidence to indicate that the HIF transcriptional system is associated with some specific loci encoding the erythropoietin and hemoglobin proteins (Simonson et al., 2010; Yi et al., 2010; Petousi and Robbins, 2014). In contrast, the role of genetic factors in modulating the response of HIF- 1 in lowlanders ascending to altitude is unknown. In fact, qualitative studies on the contribution of genetic factors in exercise performance in hypoxia are scarce (Hennis et al., 2015). To date published literature only supports a potential role of the angiotensin-converting enzyme insertion (ACE-I) and endothelial PAS domain-containing protein 1 (EPAS1) alleles in exercise performance in hypoxia (Montgomery et al., 1998; Masschelein et al., 2015a). Against this background, the hypothesis driving the current study was that the variability in response of myocellular HIF-1 and its downstream targets implicated in regulation of glycolysis as well as oxidative metabolism, is at least partly explained by genetic factors. To test this hypothesis, we conducted a well- controlled cross-over study (Masschelein et al., 2014, 2015b) in which monozygotic (MZ) twins were exposed to normoxia vs. hypoxia equivalent to ∼ 5300 m altitude, both at rest and during submaximal exercise. The data presented in this paper for the first time demonstrate that both HIF-1 mRNA and protein expression are upregulated by acute normobaric hypoxia and/or exercise in a genotype-dependent manner. METHODS Subjects The data presented in this paper are original and are part of a larger study in which 13 monozygotic twin brothers ( n = 26) were enrolled (Masschelein et al., 2014, 2015b). Inclusion criteria on admission were: non-smoking, no history of cardiovascular or respiratory disease, similar physical activity levels within twins, and no residence at altitude > 1500 m during 6 months before the study. Mono-zygosity of the twin pairs was confirmed via 8 polymorphic markers (chromosome (chr) 13, GATA30H01 and GATA85D03; chr 18, GATA2E