PONE-D-21-28261

Elevated aqueous endothelin-1 concentrations in advanced diabetic retinopathy

PLOS ONE

Dear Dr. Kang,

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PLOS ONE

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- https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0252530

In your revision ensure you cite all your sources (including your own works), and quote or rephrase any duplicated text outside the methods section. Further consideration is dependent on these concerns being addressed.

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Partly

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

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3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Dear author, I congratulate you for this excellent study. Although it has been known that endothelin-1 levels are upregulated in hypoxic states with concomitant vasoconstriction, this study conclusively shows that the levels are higher with severe grades of DR indicating extreme levels of hypoxia, which reduced after the intravitreal injection of Anti-VEGF agents.

However, please check the manuscript for grammatical errors.

Also, the abstract needs rephrasing: purpose section mentions this as a retrospective study, and the methods section mentions this as prospective.

I also suggest if you could revise the keywords. Having macular edema in this does not make sense. However, I would like to add aqueous endothelin-1, bevacizumab etc.

Overall, an excellent study.

Thank you

Reviewer #2: Authors present a novel work. Following suggestions would help improving the manuscript.

Major Comments

1. Purpose section of Abstract needs to provide a statement about the rationale of the study.

2. Definition of early diabetic retinopathy and advanced diabetic retinopathy needs to be provided in Methods section in Abstract

3. Conclusion in Abstract is just the reiteration of the data in the Results. Conclusion should provide interpretable message of the study along with possible future implications if any.

4. “These studies found that ET-1 has an important role in the pathogenesis and progression of DR.” The role of endothelin 1 in pathogenesis of diabetic retinopathy as indicated in this sentence in Introduction needs to be further elaborated.

5. How was proliferative diabetic retinopathy diagnosed?

6. What is the meaning of “two consecutive intravitreal injections” and how was this number achieved at?

7. “The early DR group included the patients with no gross DR,………..” What is the meaning of “gross DR”?

8. Results is lengthy and needs to be shortened. It discusses the data and provides comparisons. It should include only data and tables. There should not be any discussion of the data; which is part of Discussion section.

9. “A few studies have investigated vitreous ET-1 levels in patients with DR; however,

the results are inconsistent.18-20” Do authors believe that measuring endothelin level in vitreous would have been more appropriate than aqueous levels?

10. “This ‘certain point’ may exist between moderate and severe nonproliferative DR and/or development of macular edema, based on our study results. Advancement to severe nonproliferative DR and concomitant development of macular edema both may be involved in the increase of ET-1 inside the eye” Reviewer believes that this assertion would require different study design where same set of patients are followed up over a period of time to assess endothelin 1 levels alongside evolution of diabetic retinopathy. Or it could be concluded with present study design too, had the sample size been larger enough. Hence the present study should possibly refrain from providing a continuum of endothelin 1 levels alongside evolution of diabetic retinopathy and leave it to future studies.

11. A lot has been discussed about plasma endothelin 1 levels. Reviewer feels that this is speculative and beyond the scope of present study.

12. “If we can detect the critical ‘point’ in patients with DR, interventions antagonizing ET-1 may prevent further disease progression. Then we may prevent further vision loss in these patients with DR.” As stated in previous comment, present study design is not suitable for this assertion.

Minor Comments

1. “For the control group, those who underwent uncomplicated cataract surgery and had no other concomitant ocular diseases were included” This information has been written twice in Methods section.

2. “After reaching ‘certain threshold point’ and progression to advanced DR, then intraocular ET-1 level may be various with different in accordance with each eye status.” This sentence in Discussion needs to be reframed.

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Reviewer #1: No

Reviewer #2: No

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Elevated aqueous endothelin-1 concentrations in advanced diabetic retinopathy

PONE-D-21-28261R1

Dear Dr. Kang,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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Kind regards,

Tomislav Bulum

Academic Editor

PLOS ONE

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

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3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Thank you for the revised manuscript. All the revisions are acceptable. I do not recommend any further revisions.

Reviewer #2: Authors have modified the manuscript as suggested. There are no more suggestions or modifications for the manuscript.

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7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: Yes: Kumar Saurabh