Arsenic Exposure and Methylation Efficiency in Relation to Oxidative Stress in Semiconductor Workers

Round 1

Reviewer 1 Report

 In this study, Pan et al. measured arsenic exposure and oxidative stress in semiconductor workers. The study was very well designed, with a an appropriate control group (administration workers) and two different exposure groups (maintenance and production workers). Many different forms of arsenic were measured in urine, as well as two different measures of oxidation (8-oxodG and MDA). Total arsenic levels were also measured in fingernails and hair.

Although well designed, this study has a considerable overlap with previously published reports, which diminishes its relevance.

Specifically:

1. At line 94-96, the authors mention the absence of measures of chronic exposure to arsenic and their correlation to oxidative stress, by which I believe they mean measurements in fingernails and hair as opposed to urine ("acute"). However, their own study fails to show any correlation between total arsenic measured in fingernails and hair with either 8-oxodG or MDA (Table 4). The authors should highlight that they found that fingernail and hair samples do not perform as well as urinary samples to measure oxidative stress.
2. The use of MDA as a measure of oxidative stress was innovative. Although it is increased in exposed workers, it did not correlate with either MMA or PMI. The authors should highlight the fact that does not perform as well as 8-oxodG in this context. 
3. In the paragraph at lines 91-97, the authors mention "studies" but reference only one (Byun et al.). Please list all studies referred to here.
4. The y axis in figure 2 seems mislabeled and should probably display MDA
5. The authors do not show the correlation between 8-oxodG and SMI. Please add the SMI data.
6. In general, the positive results presented here overlap with those presented in Hu et al. 2006 in semiconductor workers. Although this study is cited, Pan et al. does not address the similarities between the two studies. This is particularly puzzling as Dr. Pan is also an authors in Hu et al 2006, which makes the current study really a follow-up work. Please fully discuss the similarities and differences between the two studies.
7. Please correct the Greek letter "mu" being displayed as @

Author Response

Reviewer 1

 

Overall statement. In this study, Pan et al. measured arsenic exposure and oxidative stress in semiconductor workers. The study was very well designed, with an appropriate control group (administration workers) and two different exposure groups (maintenance and production workers). Many different forms of arsenic were measured in urine, as well as two different measures of oxidation (8-oxodG and MDA). Total arsenic levels were also measured in fingernails and hair. Although well designed, this study has a considerable overlap with previously published reports, which diminishes its relevance.

 

Answer: We believe that this study has its scientific merits and its findings can expand our understanding of arsenic toxicity. As we stated in the text, a few studies have examined relations between As exposure using speciated As metabolites in semiconductor workers and occupational conditions. Also, these studies only measured urinary As to recent exposure in semiconductor workers without well-defined occupational classification [6, 23-24]. Furthermore, those studies did not examine associations between speciated As metabolites from chronic exposure and As exposure-mediated oxidative effects in semiconductor workers. To our knowledge, few studies conducted to assess oxidative stress effects for both short-and long term exposure to As in semiconductor workers. This study aimed to examine oxidative effects of both short-and long-term exposure to As in semiconductor workers based on job classification. Furthermore, we have added additional text to increase the relevance of our manuscript; “However, the study in Hu et al. [23] did not include multiple biological matrices, including fingernails and hair, to evaluate long-term health effects of As exposure. Also, Hu’s study didn’t evaluate the correlation between urinary As species and MDA, and didn’t assess oxidative stress effects for long term exposure to As in semiconductor workers."

 

1.  At line 94-96, the authors mention the absence of measures of chronic exposure to arsenic and their correlation to oxidative stress, by which I believe they mean measurements in fingernails and hair as opposed to urine ("acute"). However, their own study fails to show any correlation between total arsenic measured in fingernails and hair with either 8-oxodG or MDA (Table 4). The authors should highlight that they found that fingernail and hair samples do not perform as well as urinary samples to measure oxidative stress.

 

Answer: We agree with the comments. We have highlighted that fingernail and hair samples do not perform as well as urinary samples to measure oxidative stress induced by As exposure, as shown in the Abstract section at Lines 31-32 and in the Discussion section at Lines 362-363. Indeed, arsenic does accumulate in human fingernail and hair. However, inorganic arsenic is the dominant form in fingernail and hair, while only small levels of DMA and MMA are present. That suggests that some arsenic in fingernail and hair may not undergo the methylation process, which oxidation process takes place.

 

2.  The use of MDA as a measure of oxidative stress was innovative. Although it is increased in exposed workers, it did not correlate with either MMA or PMI. The authors should highlight the fact that does not perform as well as 8-oxodG in this context.

Answer: We have added the text that “8-OHdG could serve as a more reliable biomarker for predicting As methylation than MDA did.” and "MDA concentration is regarded as a systemic oxidative stress indicator, however MDA reacts readily with other substances in the body, and its level, unlike that of urinary 8-OHdG, may not accurately reflect oxidative stress throughout the whole body." Please see Lines 32-33and Lines 439-443.

 

3. In the paragraph at lines 91-97, the authors mention "studies" but reference only one (Byun et al.). Please list all studies referred to here.

 

Answer: We have listed all the references 6, 23-24. Please see Line 95.

 

4.  The y axis in figure 2 seems mislabeled and should probably display MDA. 

 

Answer: We agree with the comment that the y axis in Figure 2 mislabeled and should be displayed as MDA. However, the Reviewer 2 recommended to remove Figure 2, since the reported results sound not so significant, and it is not useful for the next discussion of Section 4 to show some plots reporting not correlated data. Based on the reviewer’s recommendation, we have removed Figure 2. However, we stated that urinary MDA concentrations of the As-exposed group were not significantly correlated with urinary MMA concentrations (P = 0.169, n = 427). Please see Lines 288-290.

 

5.  The authors do not show the correlation between 8-oxodG and SMI. Please add the SMI data.

Answer: We have added the text "Urinary 8-OHdG concentrations in the As-exposed group were not significantly associated with SMI (P=0.582, n = 427)." Please see Lines 294-295, Page 10.

6. In general, the positive results presented here overlap with those presented in Hu et al. 2006 in semiconductor workers. Although this study is cited, Pan et al. does not address the similarities between the two studies. This is particularly puzzling as Dr. Pan is also an authors in Hu et al 2006, which makes the current study really a follow-up work. Please fully discuss the similarities and differences between the two studies.

Answer: We agree with the comments. We have added the text " Our study was consistent with Hu’s study that MMA concentrations in semiconductor workers positively correlated with urinary 8-OHdG. However, the study in Hu et al. did not include multiple matrics, including fingernails and hair evaluate both short-term and long-term health effects from As exposure. Also, the study didn’t examine the correlation between urinary As species and MDA. Finally, they did not assess oxidative stress effects from long-term exposure to As in semiconductor workers. ", Please see Lines 423-426.

 

7.  Please correct the Greek letter "mu" being displayed as@

Answer: We have revised the unit to be μ at Lines 142 and 145.

 

Reviewer 2 Report

GENERAL COMMENTS

I believe that the submitted manuscript from Pan et al. is widely in the scope of the Atmosphere journal. Their work addresses a very important topic and, to my knowledge, very few previous works dealt with similar aspects of the As exposure. The number of workers involved in their study is very high and represents a correct amount of data to write a significant paper from the statistical point of view. Therefore, their work is surely potentially important for the scientific community, but at the current stage it needs some major revisions to be published in the Atmosphere journal. The Authors should better explain the novelty of their work and better present their data and figures.

 

SPECIFIC COMMENTS

Line 23. Delete “were”.

Lines 98-104. The Authors could better specify the novelty of their work in this Section.

Line 107. I believe that the total number of workers involved in your study is 609, not 518 as reported in this sentence.

Lines 141 and 144. Please correct the unit of measure of the As concentration.

Lines 189-190. This assumption should be justified.

Line 191. Replace “data was” with “data were”.

Line 208. In my opinion, Section 3.1 is too descriptive. I am aware that most of the comments are in Section 4, but I believe that some comments on the results from Tables 1 and 2 could be inserted also in Section 3.1 in order to make this section clearer.

Lines 290-294. I believe it could be better to insert the correlation coefficients in Tables 3 and 4, and then discuss them in Sections 3.2 and 3.3.

Line 301. Please do not repeat in each sentence that r is the Pearson correlation coefficient. Mention it only the first time you use it.

Figures 1-4. Please move the captions under the related figures. I suggest removing Figures 2 and 4 because the reported results sound not so significant. I believe that it is not useful for the next discussion of Section 4 to show some plots reporting not correlated data.

Lines 352-354. Could the Authors provide some explanations for this point?

Lines 364-366. I suggest the Authors reporting in a table of the manuscript or in the supplementary material all the correlation coefficients among the analyzed parameters. For example, in this sentence the readers need to know the correlation coefficients to understand your comment.

Lines 377-380. Some references are missing here.

Line 398. Insert the number of the reported reference.

Lines 412-414. See my comment to Lines 364-366.

Lines 461-468. I believe that the Conclusion section is very reductive. Please revise it providing a short description of the main findings discussed in the manuscript.

Author Response

Reviewer 2

 

General Comments

1. I believe that the submitted manuscript from Pan et al. is widely in the scope of the Atmosphere journal. Their work addresses a very important topic and, to my knowledge, very few previous works dealt with similar aspects of the As exposure. The number of workers involved in their study is very high and represents a correct amount of data to write a significant paper from the statistical point of view. Therefore, their work is surely potentially important for the scientific community, but at the current stage it needs some major revisions to be published in the Atmosphere journal. The Authors should better explain the novelty of their work and better present their data and figures.

Answer: We agree with the comment.

 

Specific Comments

1. Line 23. Delete “were”.

Answer: We have deleted “were”, as shown on Line 23.

 

2. Lines 98-104. The Authors could better specify the novelty of their work in this Section. Line 107. I believe that the total number of workers involved in your study is 609, not 518 as reported in this sentence..

Answer: We have added the text; “To our knowledge, few studies conducted to assess oxidative stress effects for both short-and long-term exposure to As in semiconductor workers.”, as shown on Lines 98-99.

We have stated a correct total number of workers which was 518. Please see the text; As-exposed group (n =427) consisted of 149 maintenance staff and 278 production staff…..”, as shown on Lines 111-112.

 

3. Lines 141 and 144. Please correct the unit of measure of the As concentration. Lines 189-190. This assumption should be justified. Line 191. Replace “data was” with “data were”.

Answer: We have revised the unit to be μ at Lines 142 and 145. The“data was” has been replaced with “data were” on Line 192.

 

4. Line 208. In my opinion, Section 3.1 is too descriptive. I am aware that most of the comments are in Section 4, but I believe that some comments on the results from Tables 1 and 2 could be inserted also in Section 3.1 in order to make this section clearer.

Answer: We agree with the comment. We have deleted descriptive comments on the data in Section 3.1. Please see Lines 210-229.

 

5. Lines 290-294. I believe it could be better to insert the correlation coefficients in Tables 3 and 4, and then discuss them in Sections 3.2 and 3.3.

Answer: We agree with the comment. Correlation coefficients have been added in Tables 3 and 4. Also, we have stated the findings in Sections 3.2 and 3.3 at Lines 254-263 and Lines 265-273. Finally, we have discussed them in the Discussion section at Lines 399-411 and Lines 433-443.

 

6. Line 301. Please do not repeat in each sentence that r is the Pearson correlation coefficient. Mention it only the first time you use it.

Answer: We have deleted Pearson correlation coefficients at Lines 287 and 292.

 

7. Figures 1-4. Please move the captions under the related figures. I suggest removing Figures 2 and 4 because the reported results sound not so significant. I believe that it is not useful for the next discussion of Section 4 to show some plots reporting not correlated data.

Answer: We agree with the comment. We have deleted Figure 2 and Figure 4.

 

8. Lines 352-354. Could the Authors provide some explanations for this point?

Answer: We agree with your comment. The statement described half-life of As, which may attribute to accumulate As species in different biological metrices.  

 

9. Lines 364-366. I suggest the Authors reporting in a table of the manuscript or in the supplementary material all the correlation coefficients among the analyzed parameters. For example, in this sentence the readers need to know the correlation coefficients to understand your comment.

Answer: Both of arsenic in hair and fingernail were significantly correlated with work years, as shown in Tables 3 and 4. Correlation coefficients have been added to Tables 3 and 4. Based on the correlation coefficient, we stated that “The stronger correlation between the number of years worked and total As concentrations in fingernails added additional evidence that fingernails may accumulate As for a longer period than urine and hair do.” Please see Lines 353-356.

 

10. Lines 377-380. Some references are missing here.

Answer: We have added the references “12. Kitchin KT Recent advances in arsenic carcinogenesis: modes of action, animal model systems, and methylated arsenic metabolites. Toxicol Appl Pharmacol. 2001, 172, 249-61.” Please see Line 373 and Lines 498-499.

 

11. Line 398. Insert the number of the reported reference.

Answer: We have added the reference “36. Hu CW, Cooke MS, Tsai YH, Chao MR. 8-Oxo-7,8-dihydroguanine and 8-oxo-7,8-dihydro-2′-deoxyguanosine concentrations in various human body fluids: implicationsfor their measurement and interpretation. Arch Toxicol 2015; 89: 201–210.” Please see Line 390, Page 13; Lines 536-538, Page 16

 

12. Lines 412-414. See my comment to Lines 364-366.

Answer: We agree with the comment. Correlation coefficient have been added.

 

13. Lines 461-468. I believe that the Conclusion section is very reductive. Please revise it providing a short description of the main findings discussed in the manuscript.

Answer: We have added the text ”Increased concentrations of As in these humans’ biological matrices warrant a need to develop proper preventive measures, including adequate ventilation and the use of personal protection equipment (gloves and respirators) to protect the health of semiconductor workers. ” in the Conclusion section. Please see Lines 460-463.

Round 2

Reviewer 2 Report

I believe that the Authors did a good job in revising their manuscript. But I believe that the manuscript still needs some minor revisions before the final publication.

1) I have not seen a particular effort from the Authors in order to describe the novelty of their study with respect to previous works (especially in the Introduction section).

2) The Authors did not reply to my following comment: Lines 189-190. This assumption should be justified.

3) Conclusion section is still very reductive. Please insert all main results of your study in this section.

 

 

Author Response

Reviewer 2

 

Overall statement. I believe that the Authors did a good job in revising their manuscript. But I believe that the manuscript still needs some minor revisions before the final publication.

 

Answer: We agree with your comment.

 

1. I have not seen a particular effort from the Authors in order to describe the novelty of their study with respect to previous works (especially in the Introduction section).

 

Answer: We agree with your comment. We have added the text to the Introduction section to address the novelty of the study: “Strengths of the study included a well-classified population and a well-defined occupational health assessment. This health assessment aimed to assess As exposure by measuring speciated As metabolites in multiple biological matrices (urine, hair and fingernails), and to determine As methylation efficiency by quantifying both primary and secondary As methylation indexes. Also, the study applied two biomarkers (8-OHdG and MDA) to examine oxidative stress from As exposure. The study was innovative because of 1) the use of MDA as an oxidative biomarker; 2) an assessment of As-induced oxidative damage at both systemic and molecular levels; and 3) an association between the oxidative biomarkers and As methylation efficiency.” Please see lines 101-109, page 3. In addition, we have pointed out information gaps from other studies. We believe that findings from the study have filled in the gaps.  Please see lines 92-95, page 2; lines 96-99, page 3.

 

2. The Authors did not reply to my following comment: Lines 189-190. This assumption should be justified.

 

Answer: The text “Student t statistics were used to compare As species and urinary 8-OHdG, MDA, and other covariates between the As exposed group and the control group. Pearson correlation analysis was used to examine relationships of the concentrations of biomarkers for oxidative stress (urinary 8-OHdG and MDA) and the values of As methylation indexes (MMA and PMI) with controlling for other variables, e.g. age, BMI, alcohol consumption, and other potential chemical exposures.” in the Materials and Methods section merely describe the statistical analysis.  Please see line 195-200, page 5.

 

3. Conclusion section is still very reductive. Please insert all main results of your study in this section

Answer: We agree with your comment. We have added the text to the Conclusion section; “As exposure increased oxidative stress in semiconductor workers. Urinary 8-OHdG served as a more reliable biomarker for assessing oxidative stress induced by As exposure as compared to MDA, a biomarker for systemic oxidative stress. Urinary total As and 8-OHdG concentrations reflected As methylation efficiency.” The statement includes a clear and concise conclusion on the major findings of this study. Please see lines 459-462, page 14.