-->PONE-D-25-31350-->-->Efficacy and safety of invasive laser acupuncture (650 and 830 nm) on knee osteoarthritis: A pilot randomized clinical trial-->-->PLOS ONE

Dear Dr. Kim,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

==============================

Can you please answer the following

Control design: The study lacks a non-acupoint laser control group, making it difficult to determine whether effects are acupoint-specific or due to nonspecific laser stimulation.

BMI imbalance: Significant baseline BMI differences (p = 0.0072) were not adjusted for; this could confound the outcomes.

Sample size: The small sample (n = 45) limits statistical power and generalizability; findings should be interpreted as preliminary.

Missing data handling: Use of LOCF is suboptimal; more robust methods such as multiple imputation or mixed-effects modeling are recommended.

Interpretation of results: The conclusions appear overstated given the pilot design; results should be described more cautiously as exploratory.

thank you

==============================

Please submit your revised manuscript by Nov 27 2025 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

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-->If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

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We look forward to receiving your revised manuscript.

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Ramada Rateb Khasawneh

Academic Editor

PLOS ONE

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[This work was supported by a grant (RS-2023-KH139215) from the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health and Welfare, Republic of Korea.].

Please state what role the funders took in the study.  If the funders had no role, please state: "The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript."

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Please include this amended Role of Funder statement in your cover letter; we will change the online submission form on your behalf.

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5. If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise.

6. Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

Additional Editor Comments:

Can you please answer the following

Control design: The study lacks a non-acupoint laser control group, making it difficult to determine whether effects are acupoint-specific or due to nonspecific laser stimulation.

BMI imbalance: Significant baseline BMI differences (p = 0.0072) were not adjusted for; this could confound the outcomes.

Sample size: The small sample (n = 45) limits statistical power and generalizability; findings should be interpreted as preliminary.

Missing data handling: Use of LOCF is suboptimal; more robust methods such as multiple imputation or mixed-effects modeling are recommended.

Interpretation of results: The conclusions appear overstated given the pilot design; results should be described more cautiously as exploratory.

thank you

[Note: HTML markup is below. Please do not edit.]

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

Reviewer #3: Yes

**********

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

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

-->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

Reviewer #3: Yes

**********

-->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: No

Reviewer #3: Yes

**********

-->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: This pilot randomized clinical trial investigates the efficacy and safety of invasive laser acupuncture (ILA) at wavelengths of 650 nm and 830 nm compared to a sham control in patients with knee osteoarthritis (KOA). The study employs a robust design, adhering closely to CONSORT and SPIRIT guidelines, including proper randomization and effective blinding of patients and outcome assessors. Clinically relevant outcomes such as pain (VAS), functional impairment (WOMAC), and quality of life (EQ-5D-5L) were assessed using validated tools. Findings indicate significant short-term improvements in pain relief, functional outcomes, and quality of life with ILA treatment, and no substantial adverse events were reported. However, I have the following concerns:

1. The small sample size (n=45) is adequate for a pilot study but insufficient for drawing definitive conclusions regarding efficacy and safety. I would recommend explicitly acknowledging this limitation in the manuscript, providing clear justification for the chosen sample size, and including preliminary sample-size estimations for future large-scale studies based on the variability and effect sizes observed.

2. The use of the last observation carried forward (LOCF) method for handling missing data may introduce bias and potentially affect the validity of findings. I suggest employing multiple imputation or conducting sensitivity analyses to examine the robustness of the results to different methods of missing-data handling, thereby strengthening the credibility of your conclusions.

3. Due to the intervention’s nature, investigators delivering the treatment were not blinded, potentially introducing performance bias. I recommend clearly discussing this limitation and proposing strategies, such as independent outcome assessments or objective outcome measures, to mitigate bias in future trials.

4. There was a significant baseline imbalance in body mass index (BMI) among groups (p=0.0072), which could confound the outcomes related to pain and function. I suggest explicitly addressing this issue in the manuscript and performing analyses adjusted for BMI to account for possible confounding effects.

5. Although significant differences were observed in primary outcomes (VAS and WOMAC), the responder rate defined by OMERACT-OARSI criteria was not significantly different among groups, raising questions regarding clinical significance. I recommend clearly interpreting these results in the context of minimal clinically important differences (MCID) to enhance clinical relevance and interpretability.

6. The short follow-up period (6 weeks post-intervention) limits assessment of the sustained effects of ILA treatment. I suggest clearly addressing this limitation and recommending longer follow-up durations (3–6 months or longer) in future studies to determine long-term treatment effects.

7. The evaluation of safety data is limited due to the study’s small sample size and short duration. I recommend explicitly noting this limitation in the manuscript and advising larger-scale trials with extended follow-up periods to robustly assess treatment safety and potential adverse events.

8. This was a single-center study, limiting the generalizability of the results. I suggest clearly discussing this limitation and recommending that future research involve multicenter studies with more diverse patient populations to improve external validity and applicability of the findings.

Reviewer #2: To meet STRICTA Guidelines:

1. Identify depth of insertion

2. Name as well as number acupuncture points

3. Description of participating acupuncturists (qualification or professional affiliation, years in acupuncture practice, other relevant experience)

Language/clarity issues:

Line 67: Define invasive laser acupuncture – in intro – needle size, diode used, etc.

Line 127 "daily mean pain visual analogue scale [VAS] score during activity and at rest over the past week up to screening is between 35 and 74." VAS is usually a score of 0-10. If you are using a 0-100 VAS, this needs to be explained here.

Line 158 assigned to three groups (control, 650, or 830) – need to add nm not just the number. And "control" should also be expressed as "0"

Line 165: During the period of this trial, the assessor did not approach any participant except the

assessment time – this is unclear. What is meant by approach? Interact? Speak? See?

258: 45 participants survived and... SURVIVED is the wrong word - perhaps replace with "were accepted as trial participants"

Line 329: asthenia – are you using this to indicate feeling faintness? Was this during treatment?

In your discussion, you suggest significant outcomes. Then in your Sample Size Calculation, you correctly note that sample size was too limited to evaluate effectiveness. be clearer about the issue with sample size in the discussion.

Reviewer #3: 1. The text indicates that rescue medications were administered during Visit 1 and Visit 13. Were these provided to all participants, or exclusively to those exhibiting severe symptoms? If limited to a subset of participants, could this have influenced the observed outcomes?

2. This study employed a relatively small sample size. Is the comparison involving 15 cases statistically significant? Please provide supporting references or statistical justification for this finding.

3. Differences in body mass index exist across the three groups. Given that clinical practice widely acknowledges an association between body weight and KOA, were potential confounding effects related to BMI adjusted for in the analysis? Additionally, can the robustness of the statistical methods employed be confirmed?

**********

-->6. 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.

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Reviewer #1: No

Reviewer #2: Yes: Jennifer Brett

Reviewer #3: No

**********

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-->PONE-D-25-31350R1-->-->Efficacy and safety of invasive laser acupuncture (650 and 830 nm) on knee osteoarthritis: A pilot randomized clinical trial-->-->PLOS One

Dear Dr. Kim,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

==============================

My principal concern remains the absence of an appropriate sham control, specifically both a true sham acupuncture group and a sham laser group. In my view, such controls should involve the application of acupuncture (with or without laser) at non-acupoint locations. Without these comparator groups, the current study design does not adequately exclude the possibility that the observed effects are nonspecific and could similarly occur with stimulation at non-acupoint sites.

I find this particularly important given that, in the Discussion section, the authors attribute the therapeutic efficacy of both conventional and laser acupuncture to stimulation of defined acupoints. Moreover, the authors’ response to the previous reviewer comment on this issue appears inconsistent with their own interpretation presented in the Discussion. please answer this

==============================

Please submit your revised manuscript by May 13 2026 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:-->

• A letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

As the corresponding author, your ORCID iD is verified in the submission system and will appear in the published article. PLOS supports the use of ORCID, and we encourage all coauthors to register for an ORCID iD and use it as well. Please encourage your coauthors to verify their ORCID iD within the submission system before final acceptance, as unverified ORCID iDs will not appear in the published article. Only  the individual author can complete the verification step; PLOS staff cannot  verify ORCID iDs on behalf of authors.

We look forward to receiving your revised manuscript.

Kind regards,

Ramada Rateb Khasawneh

Academic Editor

PLOS One

Journal Requirements:

If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise.

Additional Editor Comments :

My principal concern remains the absence of an appropriate sham control, specifically both a true sham acupuncture group and a sham laser group. In my view, such controls should involve the application of acupuncture (with or without laser) at non-acupoint locations. Without these comparator groups, the current study design does not adequately exclude the possibility that the observed effects are nonspecific and could similarly occur with stimulation at non-acupoint sites.

I find this particularly important given that, in the Discussion section, the authors attribute the therapeutic efficacy of both conventional and laser acupuncture to stimulation of defined acupoints. Moreover, the authors’ response to the previous reviewer comment on this issue appears inconsistent with their own interpretation presented in the Discussion. .. please answer this

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

-->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 #4: All comments have been addressed

**********

-->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 #4: Yes

**********

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

Reviewer #1: Yes

Reviewer #4: 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 #4: Yes

**********

-->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 #4: 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 carefully addressing all the major concerns from my previous review. It is optional that if you want to further strengthen the manuscript, I suggest explicitly reinforcing the necessity of sensitivity analyses for missing data handling in future research, as well as clearly emphasizing the importance of assessing sustained long-term outcomes.

Reviewer #4: After reviewing the revised manuscript and the authors’ responses to the reviewers’ comments, I believe that the authors have satisfactorily addressed the concerns raised in the previous round of review. The revisions have improved the clarity of the manuscript, particularly with regard to methodological explanations, reporting according to relevant guidelines, and discussion of the study limitations.

Although the study has inherent limitations associated with its pilot design and relatively small sample size, these issues are now appropriately acknowledged and discussed by the authors. The statistical analyses and presentation of results are generally adequate for an exploratory pilot randomized clinical trial, and the conclusions have been revised to reflect the preliminary nature of the findings.

Overall, the manuscript provides useful preliminary evidence regarding the potential efficacy and safety of invasive laser acupuncture for knee osteoarthritis and may serve as a basis for future larger-scale trials.

Therefore, I recommend acceptance of the manuscript for publication.

**********

-->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 #4: Yes: Dr.Fawwaz Al-Smadi

**********

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To ensure your figures meet our technical requirements, please review our figure guidelines: https://journals.plos.org/plosone/s/figures

You may also use PLOS’s free figure tool, NAAS, to help you prepare publication quality figures: https://journals.plos.org/plosone/s/figures#loc-tools-for-figure-preparation.

NAAS will assess whether your figures meet our technical requirements by comparing each figure against our figure specifications.

-->PONE-D-25-31350R2-->-->Efficacy and safety of invasive laser acupuncture (650 and 830 nm) on knee osteoarthritis: A pilot randomized clinical trial-->-->PLOS One

Dear Dr. Kim,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please submit your revised manuscript by Jun 26 2026 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:-->

• A letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

-->

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

As the corresponding author, your ORCID iD is verified in the submission system and will appear in the published article. PLOS supports the use of ORCID, and we encourage all coauthors to register for an ORCID iD and use it as well. Please encourage your coauthors to verify their ORCID iD within the submission system before final acceptance, as unverified ORCID iDs will not appear in the published article. Only  the individual author can complete the verification step; PLOS staff cannot  verify ORCID iDs on behalf of authors.

We look forward to receiving your revised manuscript.

Kind regards,

Ramada Rateb Khasawneh

Academic Editor

PLOS One

Journal Requirements:

1. If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise.

2. Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

Additional Editor Comments:

Could you please address the issues raised by the reviewer?

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

-->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 #5: (No Response)

**********

-->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 #5: Partly

**********

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

Reviewer #1: Yes

Reviewer #5: No

**********

-->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 #5: Yes

**********

-->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 #5: 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: The authors have sufficiently addressed all of my questions. I do not have any additional concerns or questions.

Reviewer #5: General Assessment

This manuscript presents a pilot randomized, placebo-controlled clinical trial evaluating the efficacy and safety of invasive laser acupuncture (ILA) at two wavelengths (650 and 830 nm) in patients with knee osteoarthritis (KOA). The topic is clinically relevant, and the randomized design with a sham control is a strength. However, several methodological and reporting limitations reduce confidence in the robustness and interpretation of the findings. In its current form, the manuscript would benefit from substantial revision.

Major Comments

Sample Size and Statistical Power

The study includes a total of 45 participants (approximately 15 per group). The manuscript does not clearly report an a priori sample size calculation. Given the small sample size, the study is likely underpowered, increasing the risk of both type I and type II errors.

Multiple Comparisons and Risk of False Positives

Multiple outcomes are assessed across several time points without clear indication of correction for multiple comparisons. This raises concern about inflated type I error rates.

Interpretation of Findings

As a pilot study, conclusions should be more cautious and emphasize the exploratory nature of the findings rather than suggesting efficacy.

Randomization and Allocation Concealment

The manuscript lacks sufficient detail regarding the randomization process and allocation concealment.

Blinding

Further clarification is needed on how blinding was implemented and whether its effectiveness was assessed.

Clinical Relevance

The manuscript should address whether observed differences meet minimal clinically important differences (MCID).

Biological Plausibility

A stronger discussion of the biological mechanism underlying the intervention is recommended.

Minor Comments

Clarify treatment duration wording

Define acupuncture point abbreviations

Clarify rescue medication use

Include CONSORT flow diagram

Minor language edits recommended

**********

-->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 #5: No

**********

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

To ensure your figures meet our technical requirements, please review our figure guidelines: https://journals.plos.org/plosone/s/figures

You may also use PLOS’s free figure tool, NAAS, to help you prepare publication quality figures: https://journals.plos.org/plosone/s/figures#loc-tools-for-figure-preparation.

NAAS will assess whether your figures meet our technical requirements by comparing each figure against our figure specifications.

-->

-->PONE-D-25-31350R3-->-->Efficacy and safety of invasive laser acupuncture (650 and 830 nm) on knee osteoarthritis: A pilot randomized clinical trial-->-->PLOS One

Dear Dr. Kim,

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Ramada Rateb Khasawneh

Academic Editor

PLOS One

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Additional Editor Comments:

Could you please address the issue raised by the reviewer?

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Reviewers' comments:

Reviewer's Responses to Questions

-->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 #6: All comments have been addressed

Reviewer #7: (No Response)

Reviewer #8: 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 #6: Yes

Reviewer #7: (No Response)

Reviewer #8: Yes

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

Reviewer #1: Yes

Reviewer #6: Yes

Reviewer #7: (No Response)

Reviewer #8: Yes

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Reviewer #1: Yes

Reviewer #6: Yes

Reviewer #7: (No Response)

Reviewer #8: Yes

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Reviewer #1: Yes

Reviewer #6: Yes

Reviewer #7: (No Response)

Reviewer #8: Yes

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-->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: The authors have fully addressed each of my comments and concerns, I do not have any other questions.

Reviewer #6: Thank you for the detailed revision and thoughtful point-by-point responses. The manuscript has improved substantially, addressing all comments of reviewers.

Reviewer #7: (No Response)

Reviewer #8: As an ad-hoc reviewer invited during the third revision, I have focused my evaluation on the remaining scientific soundness, methodological queries, and logical structure of the manuscript.

1. Strengths

KOA is a major degenerative disorder affecting a significant portion of the population.

This pilot trial is well-designed to evaluate the feasibility of two different wavelengths of laser acupuncture, utilizing active acupuncture as a sham laser control for KOA patients.

The efficient delivery of laser treatment via acupuncture needles is a notable strength of this study. It suggests a potential for reducing KOA-related pain and improving QoL, thereby contributing valuable insights to the field of KOA treatment.

I hope this study, along with further follow-up research on ILA, will offer significant clinical relevance, impact, and a novel integrative approach to KOA management.

Throughout the initial and revised manuscripts, the authors have demonstrated transparency and scientific rigor by providing detailed CONSORT and STRICTA-compliant methods, comprehensive supplementary materials (including the study protocol and individual data), and plausible biological mechanisms to ensure the robustness of the trial.

Based on my review of the manuscript, I would like to provide some recommendations to further improve the paper.

2. Weaknesses and Suggestions

1) Major Issues

- The concern regarding the inflation of the Type I error rate due to multiple comparisons of the same outcome across a continuous time series against the baseline remains unaddressed.

Although the authors previously responded to a similar point by expanding the limitations section, and this exploratory perspective of this pilot trial is understood and acceptable to some extent, the statistical validity of the findings may still be compromised by this inflated error rate.

Given that the study tracks indicators over time but relies solely on independent, point-by-point ANCOVA at each time interval, please consider utilizing a linear mixed-effects model or repeated measures ANCOVA to evaluate the time-by-group interaction effect if a formal re-analysis is feasible.

If a full re-analysis is realistically difficult at this stage, the statement of this methodological limitation must be further strengthened. Furthermore, the discussion and conclusion must be rephrased in a conservative tone, explicitly cautioning readers that the individual p-values at each visit were not adjusted for multiple temporal comparisons.

- While KOA improvement would be generally expected to influence QoL, the actual improvement in QoL achieved through ILA and conventional acupuncture was limited.

Please discuss the potential ceiling effect of the EQ-5D-5L in this patient population and the limitations of MCID,

and whether EQ-VAS should be considered as complementary outcome to capture more subtle changes for further study?

- The authors provided detailed reporting on AEs with V/S and laboratory results, clarifying that no significant changes were observed among the three groups in their Discussion section and Table S3.

However, for more conservative safety evaluation, tracking individual clinical change events (or outlier shifts) is more clinically relevant than merely reporting the intergroup statistical significance of means and p-values of vital signs and lab results.

- The purpose and impact of including bilateral KOA patients in a unilateral treatment trial: "treated with ILA on one-side (more affected side)"

What was the rationale for including patients with bilateral KOA in this trial? (Please elaborate if this was chosen to enhance external validity (applicability to whole KOA population) or to facilitate favorable enrollment of the study population.)

Could the main outcomes differ significantly between patients with bilateral KOA and those with unilateral KOA?

While treating the more severely affected side, if the severity flipped (i.e., the opposite side became more painful at Visit 7 or Visit 13), could this have influenced the patient-reported outcomes?

If so, is a subgroup analysis or sensitivity analysis excluding bilateral KOA required?

Patients with bilateral KOA are likely to exhibit different weight-bearing patterns during daily activities and may have a worse prognosis or clinical outcome compared to those with unilateral KOA. Please clarify and discuss these aspects.

- If statistical significance between visits occurred at Visit 13 but not at Visit 7, does this imply that LLLT has a delayed or cumulative effect, whereas conventional acupuncture plays a major role during the early phase?

How would this timeline-related result be explained, in the background of a dose-response effect or the multiple exposures to LLLT and acupuncture treatment?

Furthermore, it is intriguing that 12 sessions of active acupuncture resulted in only a -7 mm difference on the VAS, which falls below the MCID threshold.

While the sham control is well-designed and rigorous, there was an unavoidable lack of practitioner blinding, and this functioned as an active control.

Please discuss the potential bias arising from the practitioners' intentions or expectations when administering the control acupuncture treatment.

- In the final section of the Results: "830 nm ILA group showed a larger mean change in the primary outcome compared with the 650 nm group."

If this pilot trial was aimed at determining superiority between the 830 nm and 650 nm wavelengths, a three-group ANCOVA would not directly compare the head-to-head difference between these two active groups.

I suggest that this statement be rephrased in a more conservative tone, as it is currently based strictly on mean differences without direct intergroup statistical significance.

2) Minor Considerations

- Please consider providing a more detailed visual representation of the major outcomes would be beneficial for readers (e.g. a raincloud plot) rather than presenting them solely in the current data matrix format.

- This pilot study introduces the intervention as a specific type of LLLT in the background section. However, since the control group was designed under the premise that active acupuncture is already effective for KOA (without an inactive/placebo control), the authors should provide the underlying evidence for acupuncture's efficacy in the Introduction (or discussion) so that readers can fully understand the rationale behind this control setting.

- Please ensure the phrase "VAS during activity" is defined clearly for the readers. Does it refer to walking under weight-bearing conditions or the active range of motion of the joint? "Activity" can vary widely in terms of tasks and intensity.

- p.14, line 254: Please change "effectiveness" to "efficacy".

- Table 1: Please review the table, as it contains two duplicated rows for the VAS. Additionally, VAS data and vital sign surveillance would be more appropriately placed in the assessment/safety section.

- Table 2: Please change the "Dependent" column header to "Dependent variable".

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Reviewer #1: No

Reviewer #6: No

Reviewer #7: No

Reviewer #8: No

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Efficacy and safety of invasive laser acupuncture (650 and 830 nm) on knee osteoarthritis: A pilot randomized clinical trial

PONE-D-25-31350R4

Dear Dr. Kim,

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,

Ramada Rateb Khasawneh

Academic Editor

PLOS One

Additional Editor Comments (optional):

Good Luck

Reviewers' comments:

Reviewer's Responses to Questions

-->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 #8: All comments have been addressed

**********

-->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 #8: Yes

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

Reviewer #8: 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 #8: 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 #8: 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 #8: #. Response to Revision

- Thank you for your hard work on the revised manuscript, particularly regarding the supplementary plot visualizations, post-hoc rmANCOVA, transparent data reporting, expanded discussions with cautious interpretations, and thorough responses to the previous comments.

The authors have put tremendous effort into providing data-supported conclusions and methodological transparency, and have proactively addressed the points previously suggested.

- I fully understand the authors' point that the eligibility criteria for KOA patients were appropriate given that this trial needed to secure external validity and feasibility as a preliminary clinical study, rather than a strictly controlled experimental setup. Furthermore, the authors have expanded their discussion by referencing related literature on the potential crosstalk from contralateral pain.

It is noted that the control group was not designed to demonstrate or compare the full effect of active acupuncture, but rather to serve as a sham ILA. It was my oversight to misunderstand the control as an active control. Thank you for your response.

While practitioner open-blinding was inevitable, it was successfully minimized by securing assessor, patient, and allocator blinding. Although the authors noted that "Formal assessment of blinding effectiveness using a validated blinding index was not performed in this pilot trial" for this trial, it would be recommended to include a blinding index test in future studies to avoid similar concerns.

- From a clinical perspective, I recommend this research as it elucidates an efficient and minimally invasive ILA approach for KOA patients suffering from poor long-term prognosis and deteriorating QoL, and thus would contribute to the field of integrative medicine.

I hope the authors' upcoming confirmatory clinical trial based on this pilot study will be successful, utilizing increased statistical power and a refined, focused protocol.

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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.

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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 #8: No

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