Cyclophilin D knockout significantly prevents HCC development in a streptozotocin-induced mouse model of diabetes-linked NASH

Winston T. Stauffer; Michael Bobardt; Daren R. Ure; Robert T. Foster; Philippe Gallay

doi:10.1371/journal.pone.0301711

Peer Review History

Original SubmissionDecember 25, 2023
6 Feb 2024 Decision Letter - Vadim Ten, Editor PONE-D-23-41197Cyclophilin D knockout significantly prevents HCC development in a streptozotocin-induced mouse model of diabetes-linked NASH PLOS ONE Dear Dr. Stauffer, 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 by the reviewer #1 during the review process. Your manuscript was found to be interesting for the field. However, several points raised by the reviewer #1 need to be addressed. Please, discuss the role of CypD in your paradigm, considering excessive cellular death in CypD KO tissue (Reviewer #2). Please submit your revised manuscript by Mar 22 2024 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 rebuttal 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. We look forward to receiving your revised manuscript. Kind regards, Vadim Ten Academic Editor PLOS ONE Journal Requirements: When submitting your revision, we need you to address these additional requirements. 1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf 2. Please expand the acronym “NIH/NIAID” (as indicated in your financial disclosure) so that it states the name of your funders in full. This information should be included in your cover letter; we will change the online submission form on your behalf. 3. Thank you for stating the following in the Competing Interests section: "Daren R. Ure and Robert T. Foster are employees of Hepion Pharmaceuticals, the developers of Reconfilstat/CRV431." Please confirm that this does not alter your adherence to all PLOS ONE policies on sharing data and materials, by including the following statement: "This does not alter our adherence to PLOS ONE policies on sharing data and materials.” (as detailed online in our guide for authors http://journals.plos.org/plosone/s/competing-interests). If there are restrictions on sharing of data and/or materials, please state these. Please note that we cannot proceed with consideration of your article until this information has been declared. Please include your updated Competing Interests statement in your cover letter; we will change the online submission form on your behalf. 4. We note that you have included the phrase “data not shown” in your manuscript. Unfortunately, this does not meet our data sharing requirements. PLOS does not permit references to inaccessible data. We require that authors provide all relevant data within the paper, Supporting Information files, or in an acceptable, public repository. Please add a citation to support this phrase or upload the data that corresponds with these findings to a stable repository (such as Figshare or Dryad) and provide and URLs, DOIs, or accession numbers that may be used to access these data. Or, if the data are not a core part of the research being presented in your study, we ask that you remove the phrase that refers to these data. 5. Please include your full ethics statement in the ‘Methods’ section of your manuscript file. In your statement, please include the full name of the IRB or ethics committee who approved or waived your study, as well as whether or not you obtained informed written or verbal consent. If consent was waived for your study, please include this information in your statement as well. 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: Partly Reviewer #2: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: I Don't Know Reviewer #2: 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 ****** 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: No Reviewer #2: 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: The format of the presented manuscript is confusing. This reviewer failed to find individual figure legends, since it seems that they are randomly inserted to the main body of the manuscript. The labelling is confusing, since all the Sets (1,2,3) are different in different figures, but not properly explained in the figure legends (assuming these were figure legends). I will be willing to read the ms after reformatting and clarification of the experimental groups assignment. Maybe it would be better to explain the Sets directly in the figure panels. Reviewer #2: Overall, this is very interesting manuscript. The experimental plan is sounds and data presented are clear. My only concern is the author's conclusion regarding participation of the mPTP in the observed effects. According to the data CypD KO tissues (which presumable have inhibited mPTP) have even more death then WT tissues, which actually argues against the mPTP which is known to be a strong activator of cell death. This is in mind I believe that authors should discuss the possibility of physiological roles of CypD, which are independent of mPTP, which would explain the effect that they observe. ****** 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. 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 ******** [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. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0301711.r001
Revision 1
13 Feb 2024 Author Response Dear PLOS One Editors and Reviewers: We thank the editors and reviewers for taking the time to carefully read and comment on our manuscript. Below are each of the journal requirements and reviewer comments with our responses following. For clarity, our responses begin and end with a * symbol. Journal Requirements: When submitting your revision, we need you to address these additional requirements. 1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf Thank you, we have made corrections as per the style requirements. 2. Please expand the acronym “NIH/NIAID” (as indicated in your financial disclosure) so that it states the name of your funders in full. This information should be included in your cover letter; we will change the online submission form on your behalf. NIH/NIAID stands for National Institutes of Health/National Institute of Allergy and Infectious Diseases. We will wait for PLOS One to change the online submission form on our behalf. Thanks! 3. Thank you for stating the following in the Competing Interests section: "Daren R. Ure and Robert T. Foster are employees of Hepion Pharmaceuticals, the developers of Reconfilstat/CRV431." Please confirm that this does not alter your adherence to all PLOS ONE policies on sharing data and materials, by including the following statement: "This does not alter our adherence to PLOS ONE policies on sharing data and materials.” (as detailed online in our guide for authors http://journals.plos.org/plosone/s/competing-interests). If there are restrictions on sharing of data and/or materials, please state these. Please note that we cannot proceed with consideration of your article until this information has been declared. Daren Ure and Robert Foster were employees of Hepion Pharmaceuticals at the time of writing of this manuscript. This does not alter our adherence to PLOS ONE policies on sharing data and materials. Additionally, the manuscript does not contain any data produced or funded by Hepion Pharmaceuticals, or any other commercial entity. and the authors thus are not reporting any competing interests, see the Competing Interests statement below. Please include your updated Competing Interests statement in your cover letter; we will change the online submission form on your behalf. Competing Interests Statement: The authors have no conflicts to declare. 4. We note that you have included the phrase “data not shown” in your manuscript. Unfortunately, this does not meet our data sharing requirements. PLOS does not permit references to inaccessible data. We require that authors provide all relevant data within the paper, Supporting Information files, or in an acceptable, public repository. Please add a citation to support this phrase or upload the data that corresponds with these findings to a stable repository (such as Figshare or Dryad) and provide and URLs, DOIs, or accession numbers that may be used to access these data. Or, if the data are not a core part of the research being presented in your study, we ask that you remove the phrase that refers to these data. Because the data is not essential to the research presented here, we have elected to remove the wording mentioning the irrelevant data. 5. Please include your full ethics statement in the ‘Methods’ section of your manuscript file. In your statement, please include the full name of the IRB or ethics committee who approved or waived your study, as well as whether or not you obtained informed written or verbal consent. If consent was waived for your study, please include this information in your statement as well. Thank you, we have updated our ethics statement to include our Animal Use Protocol in the Methods section. 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: Partly Reviewer #2: Yes Thank you for your comments. See below for our responses to “5. Review Comments to the Author”. 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: I Don't Know Reviewer #2: Yes Thank you, see below for our responses to “5. Review Comments to the Author”. 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 Thank you for your responses. 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: No Reviewer #2: Yes Thank you, please see below for our responses to “5. Review Comments to the Author”. 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: The format of the presented manuscript is confusing. This reviewer failed to find individual figure legends, since it seems that they are randomly inserted to the main body of the manuscript. The labelling is confusing, since all the Sets (1,2,3) are different in different figures, but not properly explained in the figure legends (assuming these were figure legends). I will be willing to read the ms after reformatting and clarification of the experimental groups assignment. Maybe it would be better to explain the Sets directly in the figure panels. Thank you for your valuable comments. Rather than being randomly inserted in the manuscript, the figure legends are inserted after the paragraph in which the corresponding figure is first cited. While we regret any confusion, we are unable to change this formatting because it is a requirement of the PLOS One editors (as is all formatting in the manuscript). Figure citations in the text are in parentheses and read (Fig. #). Figure legends are a separate paragraph from the main text and are identifiable by bolded text that begins “Figure #.” followed by a bolded sentence corresponding to the figure’s title. The full legend then follows in regular unbolded text. I am copying the Figure 1 legend directly from the manuscript as an example: Figure 1. CypD KO mice have similar liver morphology to WT mice except for the appearance of HCC tumors in the STZ-WD model. WT and CypD KO mice were separated into three sets. Set 1 mice were naïve, Set 2 received twice weekly CCl4 via intraperitoneal (IP) injection for thirty weeks, and Set 3 mice received a single STZ IP injection and were maintained on western diet (WD) for thirty weeks. Mice were sacrificed and then weighed, and the livers removed, weighed, and imaged. Set 1 livers were typically dark red with smooth surfaces. Set 2 livers were similar in size and color but had a slightly scaly surface. Many of the livers in Set 3 developed HCC tumors. Set 3 livers without tumors were larger and tan in color. Set 3 livers with tumors tended to be much smaller. Likewise, Set 3 mice tended to be larger overall, but mice with extensive HCC weighed less than mice without HCC. Representative livers for each group are shown here. *p≤0.001 significance between a condition and WT control by unpaired students t-test. The actual figures are submitted separately as TIF files, again, as required by the journal. We have double checked and confirmed that Sets 1, 2, and 3 refer to the same sets of mice in all figures. We added extra labeling to two of the figures to make this abundantly clear. To reiterate, Set 1 mice were nourished on ad libitum normal water and chow and served as a non-diseased baseline. Set 2 mice also received normal water and chow but was subjected to a carbon tetrachloride (CCl4) model of liver fibrosis. Set 3 mice were tested in a diabetes-linked model of NAFLD/NASH. Hopefully identifying the figure legends will aid in understanding the manuscript, as Reviewer 2 was able to do. If the PLOS One editors can provide any clarifying comments regarding how required formatting affected Reviewer 1’s understanding of the manuscript, we invite them to do so. Reviewer #2: Overall, this is very interesting manuscript. The experimental plan is sounds and data presented are clear. My only concern is the author's conclusion regarding participation of the mPTP in the observed effects. According to the data CypD KO tissues (which presumable have inhibited mPTP) have even more death then WT tissues, which actually argues against the mPTP which is known to be a strong activator of cell death. This is in mind I believe that authors should discuss the possibility of physiological roles of CypD, which are independent of mPTP, which would explain the effect that they observe. Thank you, we appreciate your kind comments. While we do make mention of CypD as a member of the mPTP complex, we do not make any claims regarding increased cell death in CypD KO tissues. Indeed, the fact that CypD global KO mice are viable at all would argue against this. To our knowledge, the literature on the role of CypD in promoting or suppressing cancer cell death is ambiguous at best. We would refer the reviewer to the last paragraph of the discussion section where we cite literature referencing CypD as both a promoter of cancer cell survival (for example by binding and sequestering BCL-2) and a promoter of cell death via the mPTP. We have added additional citations to emphasize this paradoxical relationship. While future investigation into whether there is increased HCC cell death in CypD KO mice may be warranted, it is beyond the scope of this particular manuscript. Thank you again to all the PLOS One Editors and the reviewers for suggesting improvements to the overall manuscript. The article has been enhanced by your contributions. Please contact the corresponding author(s) if you have any further questions or comments. Sincerely, Winston Stauffer Ph.D. Gallay Lab Scripps Research La Jolla, CA https://doi.org/10.1371/journal.pone.0301711.r002
6 Mar 2024 Decision Letter - Vadim Ten, Editor PONE-D-23-41197R1Cyclophilin D knockout significantly prevents HCC development in a streptozotocin-induced mouse model of diabetes-linked NASHPLOS ONE Dear Dr. Gallay, 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 Apr 20 2024 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 rebuttal 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. We look forward to receiving your revised manuscript. Kind regards, Vadim Ten MD, PhD Academic Editor PLOS ONE Journal Requirements: 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. [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 #2: (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 #2: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: I Don't Know ****** 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 ****** 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: This reviewer is not a specialist in liver physiology or tumor formation. The presented results are sound and data are displayed in a proper manner. CypD is a mitochondrial component - it would be good to present any mechanistical explanations of the observed effects on CypD in tumorigenesis It would be good to supplement these nice observational studies with some experiments aiming on elucidation of the molecular mechanism(s). Or at least separate the mitochondrial effects on MPTP CypD from other members of Pro-isomerases. Reviewer #2: This referee's comments have been largely addressed with notable exception. In the abstract authors state: "Both CRV431 and NV556 inhibit several cyclophilin isoforms, among which cyclophilin D (CypD), an essential part of the mitochondrial permeability transition pore (mPTP) complex, has not been previously investigated in this context." I believe this statement needs to be clarified. As authors would probably agree this study doesn't really investigate mPTP but rather reports changes associated with CypD KO with no direct experiments focused on mPTP. With this respect mention of mPTP in the abstract as "has not been previously investigated in this context" is misleading, since similar to the previous studies, this study doesn't address it either. As such, mPTP "possibility" belongs to the discussion rather than to the abstract section. ****** 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: 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.] 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. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0301711.r003
Revision 2
8 Mar 2024 Author Response Dear PLOS One Editors and Reviewers: We thank the editors and reviewers for taking the time to again read and comment on our manuscript. Below are each of the journal requirements and reviewer comments with our responses following in blue: Journal Requirements: 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. Thank you, we have checked that the reference list is correct and does not include any retracted papers. [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 #2: (No Response) Thank you. See below for our responses to “6. Review Comments to the Author”. 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 Thank you. 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: I Don't Know Thank you for your comments. See below for our responses to “6. Review Comments to the Author”. 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 Thank you. 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 Thank you. 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: This reviewer is not a specialist in liver physiology or tumor formation. The presented results are sound and data are displayed in a proper manner. CypD is a mitochondrial component - it would be good to present any mechanistical explanations of the observed effects on CypD in tumorigenesis It would be good to supplement these nice observational studies with some experiments aiming on elucidation of the molecular mechanism(s). Or at least separate the mitochondrial effects on MPTP CypD from other members of Pro-isomerases. Thank you for your comments. As you note and as we go over in the discussion section of the manuscript, CypD is a part of the complex forming the mPTP and thus is known under varying conditions to regulate mitochondrial permeability. However, we mention CypD is part of the mPTP only to discuss obvious avenues for future investigation. We do not claim that CypD has an effect on HCC or NAFLD/NASH progression through its role in the mPTP or its localization to mitochondria at all. Figure 5 shows that CypD KO mice have numerous perturbed genes, mostly depressed, related to HCC but mostly not directly related to the mPTP. It is thus likely that CypD KO has wide ranging effects beyond just mitochondrial permeability, only some of which have been previously reported. While future experiments will certainly be warranted to explain the precise mechanism behind CypD and cyclophilin involvement in HCC formation and progression, we believe this study is important enough to report in this journal as is, in order to guide the future studies we suggest. We also agree that isolating the effects of CypD from other cyclophilin family members is important. This study was initially intended to include comparisons with other Cyp KO mice under the same disease model. As noted in the manuscript however, these arms suffered high mortality such that they could not be included in the final comparison. Reviewer #2: This referee's comments have been largely addressed with notable exception. In the abstract authors state: "Both CRV431 and NV556 inhibit several cyclophilin isoforms, among which cyclophilin D (CypD), an essential part of the mitochondrial permeability transition pore (mPTP) complex, has not been previously investigated in this context." I believe this statement needs to be clarified. As authors would probably agree this study doesn't really investigate mPTP but rather reports changes associated with CypD KO with no direct experiments focused on mPTP. With this respect mention of mPTP in the abstract as "has not been previously investigated in this context" is misleading, since similar to the previous studies, this study doesn't address it either. As such, mPTP "possibility" belongs to the discussion rather than to the abstract section. Thank you for your valuable input. We agree with your concern and we have edited the abstract to remove mention of the mPTP, since as you note, it is not addressed in this study. 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: No Thank you again to all the PLOS One Editors and the reviewers for your comments and improvements to the overall manuscript. The article has been further enriched by your contributions. Please contact the corresponding author(s) if you have any further questions or comments. Sincerely, Philippe Gallay Professor Department of Immunology & Microbiology The Scripps Research Institute 10550 North Torrey Pines Road La Jolla, California 92037 Tel. (858) 784 8180 Fax (858) 784 8831 http://www.scripps.edu/gallay/ Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0301711.r004
20 Mar 2024 Decision Letter - Vadim Ten, Editor Cyclophilin D knockout significantly prevents HCC development in a streptozotocin-induced mouse model of diabetes-linked NASH PONE-D-23-41197R2 Dear Dr. Philippe Gallay 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 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, Vadim Ten MD, PhD Academic Editor PLOS ONE Additional Editor Comments (optional): 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 #2: 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 #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 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 ****** 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: All issues were addressed in this revision. The paper can be accepted for publication in the presented format. Reviewer #2: all good. concerns have been addressed. I have no further comments. I thank authors for their excellent work. ****** 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: No ******** https://doi.org/10.1371/journal.pone.0301711.r005
Formally Accepted
26 Mar 2024 Acceptance Letter - Vadim Ten, Editor PONE-D-23-41197R2 PLOS ONE Dear Dr. Gallay, I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now being handed over to our production team. At this stage, our production department will prepare your paper for publication. This includes ensuring the following: * All references, tables, and figures are properly cited * All relevant supporting information is included in the manuscript submission, * There are no issues that prevent the paper from being properly typeset If revisions are needed, the production department will contact you directly to resolve them. If no revisions are needed, you will receive an email when the publication date has been set. At this time, we do not offer pre-publication proofs to authors during production of the accepted work. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few weeks to review your paper and let you know the next and final steps. Lastly, if your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. 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. If we can help with anything else, please email us at customercare@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Professor Vadim Ten Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0301711.r006

Open letter on the publication of peer review reports

PLOS recognizes the benefits of transparency in the peer review process. Therefore, we enable the publication of all of the content of peer review and author responses alongside final, published articles. Reviewers remain anonymous, unless they choose to reveal their names.

We encourage other journals to join us in this initiative. We hope that our action inspires the community, including researchers, research funders, and research institutions, to recognize the benefits of published peer review reports for all parts of the research system.

Learn more at ASAPbio .