Mechanisms Underlying Hypoxic, Heat and Cross-tolerance Adaptation in Women

Most research on how the human body responds to different environmental conditions has primarily focused on men, leaving a gap in our understanding of how women adapt to these conditions. Heat exposure in particular is known to affect the well-being and performance of humans, as well as induce chronic adaptations through an acclimation/acclimatization process, which helps the body to better regulate core temperature. Moreover, contemporary research is beginning to explore the 'cross-tolerance' phenomenon; the notion that exposure (and acclimation/acclimatization) to one environmental stressor may affect the responses to another. In particular, both heat and hypoxia are known to activate common acclimatization pathways, with pulmonary, cardiovascular, hematological and muscular adaptations occurring to facilitate both oxygen transport and core body temperature regulation. In line with this background, the primary aim of this study is to investigate the effects of a heat acclimation and exercise intervention, relative to a thermo-neutral exercise control intervention, on exercise tolerance under various environmental conditions (heat, hypoxia, neutral) in healthy, active women. The secondary aim is to establish mechanisms of adaptation, by exploring the intervention-induced changes in pulmonary, cardiovascular, hematological and muscular factors, through various tests conducted at rest and during exercise.

To address these aims, 30 healthy active female participants, aged between 18 and 35 years, will be randomized to either an experimental (heat acclimation and exercise training) or control (thermo-neutral exercise training) group. The experimental group will complete a 10-day heat acclimation training intervention, exercising for 60 minutes per day in a climactic chamber set to 35°C and 50% relative humidity. The control group will complete a similar exercise intervention, but under thermo-neutral conditions (23°C and 50% relative humidity). Before and after the intervention period, both groups will complete a wide variety of tests, including exercise capacity measurements under each environmental condition (heat \[35°C\], hypoxia \[Fraction of inspired oxygen = 0.135\], neutral \[23°C, FiO2 = 0.209\]), body composition assessments, lung function testing, hemoglobin mass and blood volume quantification, skeletal muscle mitochondrial function and vascular responsiveness assessment, and venous blood sampling for a variety of sex hormone-, redox balance-, hematopoietic-, heat shock protein-, hypoxia-inducible factor- and genetic-related markers.

• Heat acclimation and exercise training (Other): 10-day intervention consisting of daily 60-min exercise sessions aimed to induce increases in core body temperature, conducted in a climactic chamber set to 35°C and 50% relative humidity.
• Thermo-neutral exercise training group (Other): 10-day intervention consisting of daily 60-min exercise sessions conducted in a climactic chamber set to 23°C and 50% relative humidity.

Primary outcomes

• Exercise capacity under thermo-neutral normoxic conditions [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Results of an incremental exercise test to exhaustion conducted at 23°C and an FiO2 of 0.209, quantified using maximal oxygen uptake
• Exercise capacity under hot normoxic conditions [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Results of an incremental exercise test to exhaustion conducted at 35°C and an FiO2 of 0.209, quantified using maximal oxygen uptake
• Exercise capacity under thermo-neutral hypoxic conditions [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Results of an incremental exercise test to exhaustion conducted at 23°C and an FiO2 of 0.135, quantified using maximal oxygen uptake

Secondary outcomes

• Core body temperature regulation during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Core body temperature measured throughout the incremental exercise tests using a rectal temperature probe.
• Skin temperature regulation during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Skin temperature measured throughout the incremental exercise tests using temperature probes attached to the calf, thigh, chest and arm, from which weighted averages are calculated.
• Sweat rates during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Sweat rate measured throughout the incremental exercise tests using a sensor attached to the forehead.
• Hemo-dynamic activity during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Cardiac output measured throughout the incremental exercise tests by bio-electrical impedance cardiography.
• Pulmonary ventilation during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Pulmonary ventilation measured throughout the incremental exercise tests using flow measurements as recorded by metabolic cart.
• Gas exchange during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Balance of oxygen uptake and carbon dioxide production measured throughout the incremental exercise tests using gas composition sensors recorded by metabolic cart.
• Muscle oxygenation during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Vastus lateralis oxygenation measured throughout the incremental exercise tests using near-infrared spectroscopy.
• Brain oxygenation during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Cerebral oxygenation measured at the left pre-frontal cortex throughout the incremental exercise tests using near-infrared spectroscopy.
• Pulse oxygen saturation during exercise [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Capillary oxygen saturation measured on the right earlobe throughout the incremental exercise tests using pulse oximetry.
• Body composition [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Body mass measured using standard weighing scales, with body fat percentage estimated using a standardized eight-site skinfold measurement protocol with associated equation.
• Lung function [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Standard spirometry procedure to assess lung function using forced expiratory volume
• Hemoglobin mass [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Hemoglobin mass estimation using the carbon monoxide re-breathing method
• Blood/plasma volume [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Blood/plasma volume estimation using the carbon monoxide re-breathing method
• Skeletal muscle mitochondrial function [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Skeletal muscle mitochondrial function quantified using a peripheral vascular occlusion test, with the rate of decrease in muscle oxygenation used to estimate mitochondrial function.
• Skeletal muscle vascular responsiveness [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Skeletal muscle vascular responsiveness quantified using a peripheral vascular occlusion test, with the rate of increase in muscle oxygenation during re-perfusion, and the oxygenation overshoot relative to pre-occlusion baseline, used to estimate vascular responsiveness.
• Blood viscosity [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Blood viscosity will be measured by obtaining venous blood samples from the participants, extracting plasma, and using a cone/plate viscometer to quantify viscosity and varying shear rates.
• Oxidative stress marker [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Venous blood samples will be obtained to quantify redox balance from the extracted plasma.
• Hematopoiesis [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Venous blood samples will be obtained to measure various markers of hematopoiesis from the extracted plasma, serum and/or whole blood.
• Heat shock proteins [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Venous blood samples will be obtained to measure heat shock proteins in the extracted plasma and serum.
• Hypoxia-inducible factor 1α [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Venous blood samples will be obtained to measure hypoxia-inducible factor 1α in the extracted plasma and serum.

Other outcomes

• Oestrogen [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Oestrogen will be quantified in the plasma extracted from venous blood samples, to identify the menstrual cycle phases of the participants.
• Progesterone [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Progesterone will be quantified in the plasma extracted from venous blood samples, to identify the menstrual cycle phases of the participants.
• Single nucleotide polymorphisms [Measured once before (pre) and once up to 1 week after (post) the intervention period]: Venous blood samples will be processed and analysed to identify single nucleotide polymorphisms

Sex: Female, ages: 18 Years to 35 Years, accepts healthy volunteers

Inclusion Criteria:

* Age: 18 - 35 years
* Body mass index: 18.5 - 25.0 kg/m\^2
* Regular physical activity (at least 30 minutes of moderate-intensity activity, three times per week)

Exclusion Criteria:

* Habitual smoker within the past 5 years
* History of metabolic disorders or any medications deemed to pose an undue risk or introduce bias in any outcome measures
* Exposure to altitude > 2000 m above sea level within four weeks of scheduled participation
* Permanent residence at altitude > 1000 m above sea level

• Tadej Debevec, PhD, Principal investigator, University of Ljubljana