Particulate Matter 2.5 Level Modulates Brachial Artery Flow-Mediated Dilation Response to Aerobic Exercise in Healthy Young Men
Round 1
Reviewer 1 Report
Thank you for inviting me to review this interesting manuscript exploring the effect of air pollution on endothelial function before and after exercise. The work is novel and relevant and the discussion is comprehensive. There are some minor edits to make for clarity of expression (lines 48, 61/62, 107, 277/278, 281/282).
Author Response
We would like to thank the reviewers and the editorial office for their time and effort to provide the thoughtful comments and suggestions. We believe that the revised manuscript is much improved following the suggested revisions.
Thank you for inviting me to review this interesting manuscript exploring the effect of air pollution on endothelial function before and after exercise. The work is novel and relevant and the discussion is comprehensive. There are some minor edits to make for clarity of expression (lines 48, 61/62, 107, 277/278, 281/282).
Response:
Consistent with your suggestion, we revised all the above sentences to improve the clarity of expression. Thank you again.
Author Response File: 
Author Response.docx
Reviewer 2 Report
This is an interesting study, and my suggestions are as follows,
1.In the statistical analyses section, why both used Wilcoxon signed rank test and paired t-test? For the two methods, they have similar functioning, but one for nonparameter statistics, and the other for parameter statistics. The authors need to clearify. Besides, why both used two-way ANOVA and GEE? For smaller sample, one-way ANCOVA would be suitable, OF course, the authors can still discuss the differences between “pre-exercise in HPM2.5 and pre-exercise in LPM2.5” additionally using t-test or the like (Wilcoxon signed rank test).
2.Line 198-200, “Additionally, the post-exercise scaled FMD in the LPM2.5 condition was significantly higher than pre- and post-exercise scaled FMD in the HPM2.5 condition (P < 0.03; Figure 1)” why only one p values? What is the meaning for comparison between post-exercise LPM2.5 vs. pre-exercise HPM2.5?
3.Line 202-203, The description “The baseline diameter significantly increased after the aerobic exercise in the HPM2.5 condition (P = 0.001; Table 3)” was not consistent with the denotation below the Table 3 “* P = 0.001 vs. pre is from pairwise multiple comparisons adjusted by Bonferroni correction.” Did the authors meant “* P = 0.001 vs. pre-exercise HPM2.5 is from pairwise multiple comparisons adjusted by Bonferroni correction”?
4. For a smaller sample like the study, did the authors consider the discussion and presentation of effect size, not only hypothesis testing outcomes?
Author Response
We would like to thank the reviewers and the editorial office for their time and effort to provide the thoughtful comments and suggestions. We believe that the revised manuscript is much improved following the suggested revisions.
- In the statistical analyses section, why both used Wilcoxon signed rank test and paired t-test? For the two methods, they have similar functioning, but one for nonparameter statistics, and the other for parameter statistics. The authors need to clearify. Besides, why both used two-way ANOVA and GEE? For smaller sample, one-way ANCOVA would be suitable, OF course, the authors can still discuss the differences between “pre-exercise in HPM2.5 and pre-exercise in LPM2.5” additionally using t-test or the like (Wilcoxon signed rank test).
Response:
Thank you for your comments on statistical analyses. We used both Wilcoxon signed rank test and paired t-test to assess the baseline differences between HPM2.5 and LPM2.5 condition visits because the two dependent variables (i.e., DBP and FMD/SR) did not meet the normality requirement for performing a parametric test.
To consider the influence of baseline diameter on FMD, the allometric scaling is recommended in the current FMD analysis guidelines [1,2]. As mentioned in the guidelines, treating the logarithmically transformed baseline diameter as a covariate to analyze FMD is not applicable for two-way ANOVA with repeated measures. Therefore, this study used GEE with lnDbase and lnSRAUC as covariates to follow the current FMD analysis guideline. This statistical analysis method was introduced and used in the previous studies [3,4].
- Line 198-200, “Additionally, the post-exercise scaled FMD in the LPM2.5 condition was significantly higher than pre- and post-exercise scaled FMD in the HPM2.5 condition (P < 0.03; Figure 1)” why only one p values? What is the meaning for comparison between post-exercise LPM2.5 vs. pre-exercise HPM2.5?
Response:
First, we apologize for the error on the P value in Line 198-200. After finding the error, we rechecked and corrected all the P values in the manuscript to ensure they were all correct.
Even though the post-exercise scaled FMD in the LPM2.5 condition was significantly increased than pre- and post-exercise scaled FMD in the HPM2.5 condition (P=0.001; P=0.009), we presented one P value range reflecting the two different P values to make the sentence as concise as possible.
According to your suggestion, we deleted the expression on the comparison between post-exercise LPM2.5 vs. pre-exercise HPM2.5. We deeply appreciate your keen comments.
- Line 202-203, The description “The baseline diameter significantly increased after the aerobic exercise in the HPM2.5 condition (P = 0.001; Table 3)” was not consistent with the denotation below the Table 3 “* P = 0.001 vs. pre is from pairwise multiple comparisons adjusted by Bonferroni correction.” Did the authors meant “* P = 0.001 vs. pre-exercise HPM2.5 is from pairwise multiple comparisons adjusted by Bonferroni correction”?
Response:
Thank you for your comment. Yes, the P = 0.001 vs. pre-exercise HPM2.5 is from pairwise multiple comparisons adjusted by Bonferroni correction. We have corrected the sentence to improve clarity (Line 211).
- For a smaller sample like the study, did the authors consider the discussion and presentation of effect size, not only hypothesis testing outcomes?
Response:
Thanks much for your comment. Since we found a significant condition x time interaction in our primary outcome variable, we decided not to report the Partial Eta Squared values (effect size). Moreover, we thought that adding the Partial Eta Squared values to the tables makes them too busy. For your information, we have attached the Partial Eta Squared table here.
Additionally, the concerns of small sample size were mentioned in the discussion section as a study limitation. Due to the small sample size, we cautiously presented our conclusions based on the results of this study.
|
|
HPM2.5 |
LPM2.5 |
η2 value |
||||
|
|
pre |
post |
pre |
post |
C |
T |
C × T |
|
SBP, mmHg |
114.2 ± 2.0 |
120.3 ± 2.3 |
116.0 ± 2.3 |
119.7 ± 2.4 |
0.07 |
0.89 |
0.18 |
|
DBP, mmHg |
70.3 ± 2.6 |
73.6 ± 2.6 |
69.8 ± 3.0 |
73.0 ± 2.7 |
0.03 |
0.70 |
>0.01 |
|
PP, mmHg |
43.9 ± 1.5 |
46.8 ± 1.5 |
46.2 ± 1.3 |
46.7 ± 2.0 |
0.06 |
0.36 |
0.15 |
|
HR, bpm |
59.3 ± 2.4 |
76.9 ± 1.9 |
60.3 ± 2.8 |
76.1 ± 2.8 |
>0.01 |
0.96 |
0.10 |
|
Baseline |
3.68 ± 0.11#† |
3.78 ± 0.10* |
3.76 ± 0.11 |
3.74 ± 0.11 |
0.06 |
0.73 |
0.61 |
|
Peak diameter, mm |
3.97 ± 0.11† |
4.07 ± 0.10 |
4.04 ± 0.10 |
4.06 ± 0.12 |
0.09 |
0.47 |
0.25 |
|
Absolute FMD, mm |
0.29 ± 0.04 |
0.29 ±0.04 |
0.28 ± 0.03 |
0.32 ± 0.03 |
>0.01 |
0.09 |
0.23 |
|
Relative FMD, % |
8.10 ± 1.08 |
7.86 ± 1.20 |
7.54 ± 1.02 |
8.60 ± 0.91 |
>0.01 |
0.06 |
0.36 |
|
FMD/SRAUC, %/ s-1 × 10-3 |
0.14 ± 0.02 |
0.11 ± 0.01 |
0.12 ± 0.01 |
0.15 ± 0.02 |
0.12 |
0.20 |
0.13 |
|
SRAUC, s-1 × 10-3 |
54.13 ± 5.14 |
63.88 ± 3.83 |
64.75 ± 7.11 |
65.74 ± 7.95 |
0.14 |
0.17 |
0.21 |
|
TTP, s |
58.8 ± 1.0† |
57.4 ± 3.2 |
52.1 ± 2.8 |
55.3 ± 3.5 |
0.21 |
0.01 |
0.13 |
References
- Atkinson, G.; Batterham, A.M. Allometric scaling of diameter change in the original flow-mediated dilation protocol. Atherosclerosis 2013, 226, 425-427, doi:10.1016/j.atherosclerosis.2012.11.027.
- Thijssen, D.H.J.; Bruno, R.M.; van Mil, A.; Holder, S.M.; Faita, F.; Greyling, A.; Zock, P.L.; Taddei, S.; Deanfield, J.E.; Luscher, T.; et al. Expert consensus and evidence-based recommendations for the assessment of flow-mediated dilation in humans. Eur Heart J 2019, 40, 2534-2547, doi:10.1093/eurheartj/ehz350.
- Birk, G.K.; Dawson, E.A.; Batterham, A.M.; Atkinson, G.; Cable, T.; Thijssen, D.H.J.; Green, D.J. Effects of Exercise Intensity on Flow Mediated Dilation in Healthy Humans. Int J Sports Med 2013, 34, 409-414, doi:10.1055/s-0032-1323829.
- Cheng, J.L.; Williams, J.S.; Hoekstra, S.P.; MacDonald, M.J. Improvements in vascular function in response to acute lower limb heating in young healthy males and females. J Appl Physiol 2021, 131, 277-289, doi:10.1152/japplphysiol.00630.2020.
Author Response File: Author Response.docx
Reviewer 3 Report
Introduction
L33 PM2.5 was associated with the risks of CVD. low/moderate/high, Be spacific
A strong rationale should be added for this study
Materials and Methods
Subjects: There is a small number of participants in this study. Why it so?
Why sample size was not calculated for this study?
There is more likely type 1 or type 2 errors.
Mention the reliability and validity of equipments used in this study.
All the testing protocol followed in this study was based upon what?
All the data were collected by whom ? researcher or lab technicians.
Result:
Discussion:
The hypothesis of the study was not confirmed.
Author Response
We would like to thank the reviewers and the editorial office for their time and effort to provide the thoughtful comments and suggestions. We believe that the revised manuscript is much improved following the suggested revisions.
Introduction
L33 PM2.5 was associated with the risks of CVD. low/moderate/high, Be spacific
A strong rationale should be added for this study
Response:
Consistent with your suggestion, we revised Line 32-34. Additionally, we added the sentence to provide a stronger rationale for testing the study hypothesis (Line 59-61). Thanks for your thoughtful comments.
Materials and Methods
Subjects: There is a small number of participants in this study. Why it so? Why sample size was not calculated for this study? There is more likely type 1 or type 2 errors.
Response:
We understand your concern about the sample size. Frankly, the small sample size is one of our study limitations, and we already mentioned it in the discussion section (Line 303-307). We could not calculate a proper sample size with enough power because the previous study that has the most similar study design with our study did not offer all values needed to calculate the sample size by G-Power [1]. We aimed to recruit 12 participants when planning this study, considering the sample size in the most similar previous study and the expected number of dropouts. However, the planned 12 participants could not be recruited due to difficulties in deciding whether or not to perform the planned experimental procedures while monitoring outdoor PM2.5 levels on a daily basis under the COVID-19 pandemic.
Due to the small sample size, this study might not draw firm (strong) conclusions. Thus, we made very cautious conclusions based on the results of this study. In addition, one of our study purposes was to investigate the feasibility on whether we could conduct this type of research study in a laboratory setting with ambient air without artificial air quality manipulation in an environmental chamber. Moreover, even with a small sample size, we met all assumption criteria for statistical analyses that are used in our study. Therefore, we don’t think that there would be any type 1 or type 2 errors.
Mention the reliability and validity of equipments used in this study.
Response:
SIDEPAKTM AM520, a light-scattering laser photometer device, is a validated, reliable device [2]. It produced reliable results (R2 = 0.82) when compared to a standard device (BAM 1020, Beta Ray Attenuation Monitor 1020, and MetOne Instruments). Other software and devices such as Aloka Prosound alpha 7 ultrasound machine, SphygmoCor Xcel system and Quipu Cardiovascular Suite software have been well validated and acquired FDA approval [3,4]. They have also shown an outstanding reliability in numerous clinical trials.
All the testing protocol followed in this study was based upon what?
Response:
Thank you for your comment. The blood pressure and brachial artery flow-mediated dilation measures followed the current guidelines [5,6]. Our exercise intervention protocol followed the American College of Sports Medicine’s physical activity guidelines, performing at least 30 minutes per day of moderate-intensity aerobic exercise recommended for most adult populations [7]. Additionally, we added the reference for the modified Bruce protocol that is used to assess the cardiorespiratory fitness level in this study.
All the data were collected by whom ? researcher or lab technicians.
Response:
All data except for brachial artery flow-mediated dilation (FMD) were collected and analyzed by a researcher. Brachial artery FMD was measured by another experienced researcher who had sufficient training in sonography (more than 10 years), and the recorded FMD data were reanalyzed by the researcher who measured FMD.
Discussion:
The hypothesis of the study was not confirmed.
Response:
Thanks much for your comments. We revised the first paragraph of the discussion section based on your feedback.
References
- Rundell, K.W.; Steigerwald, M.D.; Fisk, M.Z. Montelukast prevents vascular endothelial dysfunction from internal combustion exhaust inhalation during exercise. Inhalation Toxicology 2010, 22, 754-759, doi:10.3109/08958371003743254.
- Stauffer, D.A.; Autenrieth, D.A.; Hart, J.F.; Capoccia, S. Control of wildfire-sourced PM2. 5 in an office setting using a commercially available portable air cleaner. Journal of Occupational and Environmental Hygiene 2020, 17, 109-120.
- Faita, F.; Masi, S.; Loukogeorgakis, S.; Gemignani, V.; Okorie, M.; Bianchini, E.; Charakida, M.; Demi, M.; Ghiadoni, L.; Deanfield, J.E. Comparison of two automatic methods for the assessment of brachial artery flow-mediated dilation. J Hypertens 2011, 29, 85-90, doi:10.1097/HJH.0b013e32833fc938.
- Hwang, M.H.; Yoo, J.K.; Kim, H.K.; Hwang, C.L.; Mackay, K.; Hemstreet, O.; Nichols, W.W.; Christou, D.D. Validity and reliability of aortic pulse wave velocity and augmentation index determined by the new cuff-based SphygmoCor Xcel. J Hum Hypertens 2014, 28, 475-481, doi:10.1038/jhh.2013.144.
- Thijssen, D.H.J.; Bruno, R.M.; van Mil, A.; Holder, S.M.; Faita, F.; Greyling, A.; Zock, P.L.; Taddei, S.; Deanfield, J.E.; Luscher, T.; et al. Expert consensus and evidence-based recommendations for the assessment of flow-mediated dilation in humans. Eur Heart J 2019, 40, 2534-2547, doi:10.1093/eurheartj/ehz350.
- Muntner, P.; Shimbo, D.; Carey, R.M.; Charleston, J.B.; Gaillard, T.; Misra, S.; Myers, M.G.; Ogedegbe, G.; Schwartz, J.E.; Townsend, R.R.; et al. Measurement of Blood Pressure in Humans: A Scientific Statement From the American Heart Association. Hypertension 2019, 73, e35-e66, doi:10.1161/HYP.0000000000000087.
- Garber, C.E.; Blissmer, B.; Deschenes, M.R.; Franklin, B.A.; Lamonte, M.J.; Lee, I.-M.; Nieman, D.C.; Swain, D.P. American College of Sports Medicine position stand. Quantity and quality of exercise for developing and maintaining cardiorespiratory, musculoskeletal, and neuromotor fitness in apparently healthy adults: guidance for prescribing exercise. Medicine and science in sports and exercise 2011, 43, 1334-1359.
