Exploring the Impact of Training Methods on Repeated Sprints in Hypoxia Training Effects

Round 1

Reviewer 1 Report

General

The current manuscript looked to examine the impact of CO2, temperature, and relative humidity on physiological and perceived parameters using different methods of hypoxia. The researchers used repeated sprint ability was compared between those in a controlled tent versus those who wore a mask. They found that RSH caused increases in CO2, relative humidity, and RPE during tent training, while also affecting physiological parameters of performance. This manuscript was very well written but needs some revisions before it can be published.

Introduction

Page 2, Line 63-65: Needs a reference.

Methods

Page 2, Line 86: You need to define the ‘well-trained cyclist’ criteria.

Page 2, Lines 88-90: This sentence needs to be reworded for clarity. Also, was the 180s rest between sets active recovery or total rest?

Page 2, Line 91-93: You need to explain what mask was used and who manufactured it.

Discussion

Page 4, Line 173: Remove the track change.

Author Response

First of all, we would like to thank you for the review and your comments to improve the work.

We have made changes to the document based on your suggestions. Every point has been discussed. We hope our answers can satisfy your comments.

Below, to facilitate the identification of revisions in the text, the responses to reviewers are highlighted in blue letters, and the changes made in the manuscript can be observed in red letters.

General

The current manuscript looked to examine the impact of CO2, temperature, and relative humidity on physiological and perceived parameters using different methods of hypoxia. The researchers used repeated sprint ability was compared between those in a controlled tent versus those who wore a mask. They found that RSH caused increases in CO2, relative humidity, and RPE during tent training, while also affecting physiological parameters of performance. This manuscript was very well written but needs some revisions before it can be published.

Introduction

Page 2, Line 63-65: Needs a reference.

Response: Citation and reference added:

“Jacobson, T. A., Kler, J. S., Hernke, M. T., Braun, R. K., Meyer, K. C., & Funk, W. E. (2019). Direct human health risks of increased atmospheric carbon dioxide. Nature Sustainability, 2(8), 691-701.2

Methods

Page 2, Line 86: You need to define the ‘well-trained cyclist’ criteria.

Response: The criterion was agreed according by Jeukendrup et al 2000:

Where indicates that a maximal oxygen consumption >64 (ml/kg/min) and relative power >5.0 (W/kg) can be considered well-trained athletes.

the reference was added to the text: Jeukendrup, A. E., Craig, N. P., & Hawley, J. A. (2000). The bioenergetics of world class cycling. Journal of science and medicine in sport, 3(4), 414-433.

Page 2, Lines 88-90: This sentence needs to be reworded for clarity. Also, was the 180s rest between sets active recovery or total rest?

Response: Thanks, for the suggestion, the sentence has been rewritten:

“Each RSH session consisted of a 10-minute warm-up of continuous cycling (100 watts/80-90 rpm). Then, the RSH session began, which consisted of performing three sets of five repetitions of 10 s (maximum sprints), with an active recovery of 20 s be-tween repetitions and 180 s between sets”.

Page 2, Line 91-93: You need to explain what mask was used and who manufactured it.

Response: Thanks, added to the document Facial Mask(Im Sachsenhausen, Bickenbach, Germany).

Discussion

Page 4, Line 173: Remove the track change.

Response: Thanks, the track change was removed

Reviewer 2 Report

Although this manuscript tackles an interesting topic, there are significant concerns from this reviewer. The study design does not allow the conclusions as outlined in the manuscript. The two conditions varied by wearing a mask which might have an influence on several of the investigated parameters.

Some details:

-        The title and the use of ‘CO₂’ in the manuscript might be confusing. The authors tried to investigate the effect of increased fraction in inspiratory CO₂ (FiCO2)

-        Abstract, ln. 20: The abbreviation RSH must be introduced in the in the abstract

-        Abstract, ln. 23: Do the authors refer to the core temperature? Why is ‘CORE’ given in capital letters? In this context: the abbreviation for core temperature occurs differently (“CoreT°”, ln. 58; “T°Core” ln. 116).

-        Introduction, ln. 62 ff: The physiological effect can not be used to explain the motivation of this study! Environmental fraction of CO₂ is increasing in the range of 0.04% whereas here the effect of a 10-fold higher FCO2 is subject of the experiments.

-        Materials and Methods, ln. 92: the mask is an additional respiratory resistant and may effect HR as well as FiCO2 and thermal sensation.

-        Materials and Methods, ln. 130ff: since RPE and thermal sensation are rating scales this should not be analysed by a MANOVA developed for metric parameters. A power analysis should be calculated.

-        Results, table 1: Since ambient temperature did not vary how can the temperature effect be investigated?

-        Discussion: Since no effects were found to power variables, the effect of this training is useless?

Author Response

First of all, we would like to thank you for the review and your comments to improve the work.

We have made changes to the document based on your suggestions. Every point has been discussed. We hope our answers can satisfy your comments.

Below, to facilitate the identification of revisions in the text, the responses to reviewers are highlighted in blue letters, and the changes made in the manuscript can be observed in red letters.

Although this manuscript tackles an interesting topic, there are significant concerns from this reviewer. The study design does not allow the conclusions as outlined in the manuscript. The two conditions varied by wearing a mask which might have an influence on several of the investigated parameters.

Some details:

-        The title and the use of ‘CO₂’ in the manuscript might be confusing. The authors tried to investigate the effect of increased fraction in inspiratory CO₂ (FiCO2)

Response: Thanks for your attention. We have modified the text to clarify that the measurement we conducted pertained to the hypoxic condition with increase the inspiratory fraction of oxygen (FiO₂), rather than CO₂. Inside the tent, we created an environmental with increased levels of CO₂, which cannot be achieved with the use of a facial mask. For a more comprehensive explanation, please refer to the study conducted by Vasquez-Bonilla et al., 2021.

Reference: Vasquez-Bonilla, A. A., Rojas-Valverde, D., González-Custodio, A., Timón, R., & Olcina, G. (2021). Tent versus mask-on acute effects during repeated-sprint training in normobaric hypoxia and normoxia. Journal of Clinical Medicine, 10(21), 4879.

-        Abstract, ln. 20: The abbreviation RSH must be introduced in the in the abstract

Response: Thanks, added abbreviation to abstract.

-        Abstract, ln. 23: Do the authors refer to the core temperature? Why is ‘CORE’ given in capital letters? In this context: the abbreviation for core temperature occurs differently (“CoreT°”, ln. 58; “T°Core” ln. 116).

Response: Thank you, and it was modified to CoreTº, which is the correct.

-        Introduction, ln. 62 ff: The physiological effect can not be used to explain the motivation of this study! Environmental fraction of CO₂ is increasing in the range of 0.04% whereas here the effect of a 10-fold higher FCO2 is subject of the experiments.

Response: Thanks, we have attempted to modify the motivation of the study, which is to observe if increases in CO₂, temperature, and relative humidity influence the performance of repeated sprint training in hypoxia.

-        Materials and Methods, ln. 92: the mask is an additional respiratory resistant and may effect HR as well as FiCO2 and thermal sensation.

Response: Thanks for your observation, certainly, the mask could modify internal CO₂ levels, causing hypercapnia. However, the objective of this study was to observe the changes in CO₂, temperature and relative humidity of the environment.

Furthermore, the semi-permeable mask used in this study is not a closed-circuit air mask; instead, it allows for direct inspiration of hypoxic air from the generator.

To get a clearer idea, you can refer to Figure 2 of this study:

“Vasquez-Bonilla, A. A., Rojas-Valverde, D., González-Custodio, A., Timón, R., & Olcina, G. (2021). Tent versus mask-on acute effects during repeated-sprint training in normobaric hypoxia and normoxia. Journal of Clinical Medicine, 10(21), 4879”.

-        Materials and Methods, ln. 130ff: since RPE and thermal sensation are rating scales this should not be analysed by a MANOVA developed for metric parameters. A power analysis should be calculated.

Response: Thanks, the statistical power calculation was added using the G*Power software. The information was introduced in the section of statistical analysis and the tables.

-        Results, table 1: Since ambient temperature did not vary how can the temperature effect be investigated?

-        Discussion: Since no effects were found to power variables, the effect of this training is useless?

Response: Thanks, you for your feedback, while the isolated and controlled conditions of this study did not allow ambient temperatures to reach dangerous levels, previous studies has shown that exercising in normobaric hypoxic tents can lead to increased temperature and humidity (Vasquez-Bonilla et al., 2021). These factors may hinder neural thermoregulation and affect comfort, particularly when individuals experience heightened thermal sensations, as indicated by the results of this study.

In response to your question, we acknowledge the challenge of relying solely on p-values as an indicator of positive results. Instead, we examined the effect size and found a moderate effect size (0.087). This suggests a moderate impact on the interaction of conditions and the practical implications of our findings.

While you may already be familiar with statistical inference, we would like to provide additional information in response to this review:

"Calculating effect size provides a valuable alternative to assess the magnitude or practical significance of the observed difference or association between the variables under study. The p-value focuses on statistical significance, which is the probability of obtaining similar or more extreme results under the null hypothesis. However, the practical relevance of the results, such as the importance and impact of the differences or associations found, is often overlooked when solely relying on statistical significance (Held, 2019; Wasserstein and Lazar, 2016)”.

“Publication and reporting biases exist, favoring the publication and selective reporting of results with low p-values. This bias can lead to distortions in the scientific literature and an overestimation of effects (Metze et al., 2018)".

Therefore, it is important that you take into consideration that the p value with the complement of effect size and statistical power is not so low in the temperature and relative power (w/kg) of our participants.

References:

Metze, K., da Silva, F. A. B., & Lorand-Metze, I. (2018). Lowering the P-Value from 0.05 to 0.005 Conflicts with the 3R Rules–an Advocacy for Alternatives to Hypothesis Testing with the P-Value Approach. ALTEX-Alternatives to animal experimentation, 35(4), 516-517.

Open Science Collaboration. (2015). Estimating the reproducibility of psychological science. Science, 349(6251), aac4716.

Held, L. (2019). The assessment of intrinsic credibility and a new argument for p< 0.005. Royal Society open science, 6(3), 181534.

Wasserstein, R. L., & Lazar, N. A. (2016). The ASA statement on p-values: context, process, and purpose. The American Statistician, 70(2), 129-133.

Reviewer 3 Report

Dear Authors

As one of the reviewers, I express my personal scientific opinion on your work. I would like to reassure you that I was trying to be positive and constructive but particularly as fair and honest as possible to your work. The clear explanation provided in Method’s section is appreciated. I should also note that the originality of the study, the statistical approach used, the calculation of the Effect Size and the work done on tables are all positive points. However, the lack of test-retest reliability and the CI calculation as well as the absent of reporting any potential limitation of the study are somewhat negative points.

Please accept my judgment with a positive and constructive way.

General comments:

1.      Why you did not measure the cyclists’ performance, let’s say using a time-trial in normal environment, prior to and following the intervention period, in an attempt to see potential training adaptations during the RSH with either in the tent or using mask?  

2.      Although the article is in general understandable and well-presented, it is written in slightly an informal and not such an academic way. I would like to suggest you to revise the whole paper accordingly in order to meet the scientific criteria of written academic language.

Abstract:

3.      Line 20: What is the RSH (repeated sprints in hypoxia)? Please consider to report it prior to abbreviating it.

4.      I would expect to see some numbers and/or percentages, and actual P values within the abstract.

Introduction:

5.      Lines 42: Please eliminate the one of the two commas.

6.      Lines 70-71: Please check grammar for clarity.

7.      Paragraph 5: I found a bit irrelevant to the study the information provided concerning air pollution and CO2 production to the atmosphere due to human activities since the study did not examine the effect of air pollution on athletes’ performance during RSA. However, this particular point could be discussed in the discussion section, if it is necessary.

Methods:

8.      Why you did not use any invasive method for evaluating core body temperature?

Results:

9.      Good work has been done concerning the content of the tables but the presentation of the tables should be improved.

Moderate editing of English language is required.

Author Response

First of all, we would like to thank you for the review and your comments to improve the work.

We have made changes to the document based on your suggestions. Every point has been discussed. We hope our answers can satisfy your comments.

Below, to facilitate the identification of revisions in the text, the responses to reviewers are highlighted in blue letters, and the changes made in the manuscript can be observed in red letters.

Dear Authors

As one of the reviewers, I express my personal scientific opinion on your work. I would like to reassure you that I was trying to be positive and constructive but particularly as fair and honest as possible to your work. The clear explanation provided in Method’s section is appreciated. I should also note that the originality of the study, the statistical approach used, the calculation of the Effect Size and the work done on tables are all positive points. However, the lack of test-retest reliability and the CI calculation as well as the absent of reporting any potential limitation of the study are somewhat negative points.

Please accept my judgment with a positive and constructive way.

Response: Thanks for your feedback, consequently a limitations section is added indicating the lack of test-retest reliability and the CI calculation.

Added to the text: The main limitation of this study is the small sample size; however, the study results are promising for future investigations of hypoxic training under different conditions. Likewise, another limitation is the lack of biochemical measures that would give us results of the physiological response at the cellular level. Future research should assess the health risk of athletes by conducting studies with pulmonary ventilation measurements when using a tent to train in hypoxia. Finally, in the future of the research they need to be more rigorous in the study design, for example a calculation of the test-retest reliability and of the confidence intervals would improve the results.

General comments:

1. Why you did not measure the cyclists’ performance, let’s say using a time-trial in normal environment, prior to and following the intervention period, in an attempt to see potential training adaptations during the RSH with either in the tent or using mask? 

Response: Thank you for your suggestion. Initially, we considered conducting a performance test in a natural environment. However, due to the utilization of normobaric hypoxia-inducing instruments such as the tent and masks, our study was confined to small, controlled spaces. Therefore, conducting the experiment in a natural environment was not feasible. Nevertheless, we believe that in future research, this approach could yield significant benefits for investigating training in hypoxia. 

2. Although the article is in general understandable and well-presented, it is written in slightly an informal and not such an academic way. I would like to suggest you to revise the whole paper accordingly in order to meet the scientific criteria of written academic language.

Response: We appreciate your suggestion, we tried to modify the text so that it was written in a more academic way possible.

Abstract:

3. Line 20: What is the RSH (repeated sprints in hypoxia)? Please consider to report it prior to abbreviating it.

Response: Thanks, added abbreviation to abstract.

3. I would expect to see some numbers and/or percentages, and actual P values within the abstract.

Response: added the p value and the effect size to the abstract

Introduction:

4. Lines 42: Please eliminate the one of the two commas.

Response: Thanks, it was removed

5. Lines 70-71: Please check grammar for clarity.

Response: Revised and improved the sentence grammar

6. Paragraph 5: I found a bit irrelevant to the study the information provided concerning air pollution and CO2 production to the atmosphere due to human activities since the study did not examine the effect of air pollution on athletes’ performance during RSA. However, this particular point could be discussed in the discussion section, if it is necessary.

Response: Thanks for the feedback, it was removed from the text.

Methods:

7. Why you did not use any invasive method for evaluating core body temperature?

Response: Thanks for the suggestion, we agree that conducting invasive measurement of core temperature would be an interesting study. However, this study also wants to look at the implementation of a new non-invasive CoreTº measurement technology (Green Eye, TechGrow, The Hague, The Netherlands). Likewise, the study design, using invasive measures, requires a more rigorous follow-up of the participants than the use of the non-invasive sensor.

Rest assured, we will give due consideration to your suggestion for future research endeavors.

Results:

9. Good work has been done concerning the content of the tables but the presentation of the tables should be improved.

Response: Thanks, the presentation of the tables was improved.

Round 2

Reviewer 2 Report

The manuscript was changed following some remarks of this reviewer. However, some remarks and recommendations were probably misunderstood.

The authors investigated two conditions in two separate groups as stated: “The experimental condition (n=8) included sessions in a normobaric environment using a tent (CAT-12, Louisville, Colorado) and a control group (n=8) that used a facial mask (Im Sachsenhausen, Bickenbach, Germany) to avoid high temperatures, relative humidity, and elevated CO2 in the environment [5]”.

Critical points:

-         The title suggests that three conditions, i.e. FiCO₂, temperature, and humidity, were systematically changed. This was not the case. Changes in temperature and humidity are side effects of the experimental design. The temperature changes were documented as changes in core temperature. Moreover: It is unclear where changes in humidity result from.

-         “CO₂” is still used in the manuscript as synonym for FiCO₂. This should be changed consequently throughout the manuscript.

-         Introduction: The authors now documented the relevance to investigate FiCO₂ effects due to environmental changes. Even these studies cited are discussing critical fractions of CO₂ beyond 1000 ppm. As Jacobson, T.A. et al stated: this is a problem of indoor activities. This phenomenon is well known and well-studied in habitats like submarines and Space crafts. Therefore, the aim of the study is relevant but should not be discussed with changes in atmospheric CO₂ fractions.

-         Methods: Although the authors responded to the remark of an additional respiratory resistant from the mask, there were no values or documentations given for potential inspiratory resistance.

-         There was no change performed to analyse RPE and thermal sensation as non-metric rating scales. As stated in the first review:  this should not be analysed by a MANOVA developed for metric parameters.

-         Although the authors responded in detail to the remark that no effects were found to power variables this should become a point in the discussion.

Author Response

Dear Reviewer and editorial team,

We greatly appreciate your careful review of our manuscript and your insightful comments. We have carefully considered your feedback and made the necessary revisions to address the concerns you raised. We would like to address each of your points individually:

1. Experimental Conditions: We apologize for any confusion caused by the title. We have now clarified in the manuscript that changes in temperature and humidity were side effects of our experimental design, and the primary focus was on FiCO2 effects.
2. Use of "CO2": Thank you for pointing out the inconsistency in our use of terminology. We have gone through the manuscript and consistently replaced "CO2" with "FiCO2" to accurately reflect the parameter being studied.
3. Introduction: We appreciate your feedback on the relevance of investigating FiCO2 effects in relation to environmental changes. We have reworked the introduction to better emphasize the study's focus on FiCO2 and its potential effects, while acknowledging the existing knowledge about indoor CO2 levels. We have also clarified the relevance of our study in the context of physiological responses to FiCO2 variations.
4. Non-Metric Rating Scales: We understand your point about using non-metric rating scales for RPE and thermal sensation. Based on your feedback from the first review, we have revised the analysis methods for these parameters to ensure they are appropriately treated using methods suitable for non-metric data.
5. Power Variables in Discussion: We have taken your suggestion and have included a discussion of the lack of significant effects on power variables in our manuscript. This adds a valuable dimension to the interpretation of our results.

We would like to express our gratitude for your thorough and insightful review, as it has greatly contributed to the improvement of our manuscript. Your feedback has been invaluable in ensuring the accuracy, clarity, and overall quality of our research. If you have any further suggestions or concerns, please do not hesitate to let us know.

Round 3

Reviewer 2 Report

For the final version this reviewer suggests to introduce the abbreviation "ES" in the statistical methods.