Dear Dr. Gibbs,

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.

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We look forward to receiving your revised manuscript.

Kind regards,

Xinjun Lu

Academic Editor

PLOS ONE

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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?

Reviewer #1: Yes

Reviewer #2: Yes

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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

Reviewer #1: Yes

Reviewer #2: Yes

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

Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Referring to line 82, mentioning the 3Rs could further enhance the scientific soundness of the statement: https://nc3rs.org.uk/who-we-are/3rs. Additionally, authors could mention the FDA Modernization Act 2.0.

The authors should mention the terms microphysiological system and state organ-on-chip models. (line 86)

Referring to lines 89 to 115, the authors must focus on the main thesis statement and limit the text related to organoids.

Referring to lines 123 to 126, the authors are advised to compare the models they mentioned. It would help the reader better understand the current model's novelty and limitations.

The current research topic is interesting; authors should also consider providing a graphical abstract of their study.

Authors must consistently use the same abbreviation of induced pluripotent stem cells throughout the manuscript.

iPSCs are typically cultured on matrigel-coated surfaces; why do authors prefer Lamaninin over matrigel?

The authors should mention their study's limitations in the discussion section and expand the section on its potential implications.

Authors must label all the histological images with the proper cell type/layer.

Reviewer #2: The study of Brueske et al, aimed to determine whether incorporation of the human iPSCs into the hydrogel of the human TERT-immortalized keratinocytes and fibroblasts would improve the phenotype of the reconstructed gingival epithelium. Interestingly that such reconstructed human gingiva models demonstrated augmented keratinocyte proliferation within the epithelium, enhanced the thickness of the resultant epithelium, improved differentiation and interdigitation of the epithelium into the collagen hydrogel compared to the standard model developed from TERT cells only. Based on these data the authors conclude that the new iPSC-supplemented gingival model phenotypically resembles the native gingiva much accurately than the earlier models. The study characterises the models with examining morphological changes, i.e.: H&E histology and immunohistochemistry chromosome in situ hybridization and the TUNEL assay. Overall, the data look convincing, however, it is highly recommended that in the future studies the authors will employ combined assessment of structural and physiological changes, for the bilateral characterisation of the models and matching of the form and function.

There are few aspects after considering of which the manuscript could be accepted for publication:

• The images with the histology are of very low quality. Dear authors, please provide the images of a publication quality.

• The figure legends are provided inside the chapters of the Results section, which is a bit unusual and inconvenient. Dear authors, please place the figure legends separately, after the References, or after the Figures.

• In the Figures the P-values are represented as < 0.05, whereas in the text the p-values are given as a certain number, e.g. = 0.0424. Dear authors, please make it uniform for both.

• In the Figure 6 the images A-D contain the value of the scale bars (i.e. 10um, 100um), whereas the other Figures (5 B, C), 4 (A), 3 (A, B), 2 (A, B), and 1 (A, C) do not have these values. Dear authors, please make things uniform.

• In the experiments with the lamina propria (Figure 6) the number of the experiments is not stated. Dear authors, please provide the n-numbers.

• The Latin used throughout the text must be Italic, i.e.: in situ – line 43; e.g. – lines 76, 102, 105, 128; in vitro – line 357, etc.

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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: No

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[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.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/ . PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org

<p>Incorporation of iPSCs together with TERT-immortalized keratinocytes and fibroblasts into reconstructed human gingiva enhances phenotype of gingival epithelium.

PONE-D-25-15760R1

Dear Dr. Gibbs,

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.

An invoice will be generated when your article is formally accepted. Please note, if your institution has a publishing partnership with PLOS and your article meets the relevant criteria, all or part of your publication costs will be covered. Please make sure your user information is up-to-date by logging into Editorial Manager at Editorial Manager®  and clicking the ‘Update My Information' link at the top of the page. If you have any questions relating to publication charges, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Xinjun Lu

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

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??>

Reviewer #1: Yes

Reviewer #2: Yes

**********

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

Reviewer #1: Yes

Reviewer #2: Yes

**********

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

Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: The authors revised the manuscript according to the recommendations and provided a satisfactory response to the queries raised. Considering this, the manuscript appears suitable for publication in PLOS One.

Reviewer #2: I am satisfied with the revisions made to the manuscript by Brueske et al. and have no objections to its acceptance for publication in PLOS ONE.

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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:  Dr Vsevolod Telezhkin

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