Myofibroblasts: A key promoter of tumorigenesis following radiofrequency tumor ablation

Marwan Moussa; David Mwin; Haixing Liao; M. Fatih Atac; Aurelia Markezana; Eithan Galun; S. Nahum Goldberg; Muneeb Ahmed

doi:10.1371/journal.pone.0266522

Peer Review History

Original SubmissionJune 27, 2021
7 Jan 2022 Decision Letter - Yanlin Yu, Editor PONE-D-21-21027 Myofibroblasts: A key promoter of tumorigenesis following radiofrequency tumor ablation. PLOS ONE Dear Dr. Ahmed, 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 Feb 20 2022 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, Hanna Landenmark Senior Editor, PLOS ONE on behalf of Yanlin Yu Academic Editor, PLOS ONE Journal Requirements: 1) Please include a complete ethics statement in the Methods section. Please ensure that you state for each experiment conducted the name of the ethics committee that approved the study, the method of euthanasia, the method of anesthesia and any efforts to alleviate suffering. Please also provide for each experiment the following information: 1. maximum sizes tumors grew, 2. how tumor size was calculated, 3. the post-operative and post tumor implantation care received by the animals, including the frequency of monitoring and the criteria used to assess animal health and well-being. Please also state whether the IACUC provided approval for any large tumor sizes. 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. 3) Please note that concerns were raised about image quality, however, in the event of publication, all images will be uploaded in the original resolution rather than that seen in the PDF, so there is no need to enhance your figures if the original resolution is sufficiently clear. Additional Editor Comments (if provided): The authors report the role of activated myofibroblasts in RFA-induced tumorigenesis using murine models. The study is well designed, the results are interesting. The manuscript is well written, but the images of the figures are not clear. So, I suggest publishing the vision of the manuscript with clear figures images. [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 ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: 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 ****** 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 ****** 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: RF ablation was reported to increase growth of distant untreated tumor foci following local tumor ablation. Thus the researchers were looking to demonstrate that undesirable pro-tumorigenic effects induced by RF ablation, is carried out by myofibroblasts and that their suppression might affect tumor growth and progression. They report on the key role of activated myofibroblasts in RFA-induced distant metastases growth and successful pharmacologic blockade in mouse and rat models of colon, renal and breast experimental models. Activated myofibroblasts are a key cell population in post RF ablation distant tumor growth, primarily through induction of the HGF/c-MET/STAT3 axis. Post RFA distant tumor growth was observed in different phenotypes of ablated primary organ such as liver or kidney or phenotype of tumors such as colorectal cancer or breast cancer. They also observed that RF induced distant tumor growth can be achieved by inhibition of activated myofibroblasts using atorvastatin, an established lipid lowering medication, and anti-fibroblast agent. This is a well executed research with interesting and important results. I suggest that the authors will discuss the possibility that macrophage inhibition will achieve similar effects? ****** 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: Yes:** Yona Keisari [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.0266522.r001
Revision 1
20 Jan 2022 Author Response Dear Drs. Landemark and Yu: Thank you very much for your favorable review of our submitted original manuscript, PONE-D-21-21027, entitled “Myofibroblasts: A key promoter of tumorigenesis following radiofrequency tumor ablation”. Enclosed you will find our revised manuscript in which we have made the necessary requested changes. We will now detail the changes made in response to all comments on a point-by-point basis. Journal Requirements: 1. Please include a complete ethics statement in the Methods section. Please ensure that you state for each experiment conducted the name of the ethics committee that approved the study, the method of euthanasia, the method of anesthesia and any efforts to alleviate suffering. Please also provide for each experiment the following information: 1. maximum sizes tumors grew, 2. how tumor size was calculated, 3. the post-operative and post tumor implantation care received by the animals, including the frequency of monitoring and the criteria used to assess animal health and well-being. Please also state whether the IACUC provided approval for any large tumor sizes. Response: We now include a complete ethics statement in the Methods section. All of the methods and studies reported were approved by our Institutional Animal Care and Use Committee. We now include the additional information requested, including maximum tumor sizes (all of which were within the approved tumor size limit in our study protocol), how these were calculated, and the post-operative and post-tumor implantation care received. “All of the methods and studies reported were approved by our Institutional Animal Care and Use Committee. Specifically, rat studies were performed at Beth Israel Deaconess Medical Center, while mouse experiments were performed at Hadassah Hebrew University Medical Center in accordance with protocols approved by the respective Institutional Animal Care Committees. For rat studies, the maximum permissible diameters for R3230 breast adenocarcninomas, as per approved IACUC protocol, was a mean diameter of 2 cm. Tumor sizes were assessed using manual caliper measurements of two largest perpendicular dimensions and calculating mean diameters. After surgical procedures, the animals were monitored on a daily basis to determine any specific clinical signs of postoperative pain and distress. Specific signs of weakening or distress included, but were not limited to: total combined tumor burden greater than 2 cm or 10% of body weight, decreased appetite, reduced ambulation (interfering with or hampering with the animal's ability to obtain food and water, bear its own weight, or regain normal posture if placed on the back), weight loss (20% loss of body weight within 7 days, measured every 2-3d), tumor ulceration or necrosis for subcutaneous tumors. Post-surgically, if signs of distress were detected, a single dose of buprenorphine SR (72 hour duration of action) was administered at the site of surgery. In cases of tumor burden exceeding approved limits, animal were euthanized. For mouse experiments, general condition and animal well-being were used as metrics of tumor burden in lieu of a maximum permissible size due to microscopic nature of CRC liver deposits. Similar to rat experiments, post-operatively, animals were monitored on a daily basis to determine any specific clinical signs of postoperative pain and distress. For post-operative pain and distress, a single dose of buprenorphine SR was administered at the site of surgery. If signs of tumor burden overgrowth were detected, such as decreased appetite, reduced ambulation and or weight loss animals were euthanized.” 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. Response: There are no changes in the reference list. 3. Please note that concerns were raised about image quality, however, in the event of publication, all images will be uploaded in the original resolution rather than that seen in the PDF, so there is no need to enhance your figures if the original resolution is sufficiently clear. Response: We have now uploaded images of higher resolution. We are happy to work with the editorial staff on these as required. Additional Editor Comments: The authors report the role of activated myofibroblasts in RFA-induced tumorigenesis using murine models. The study is well designed, the results are interesting. The manuscript is well written, but the images of the figures are not clear. So, I suggest publishing the vision of the manuscript with clear figures images. Response: See response above. Reviewer #1 - Specific comments: “RF ablation was reported to increase growth of distant untreated tumor foci following local tumor ablation. Thus the researchers were looking to demonstrate that undesirable pro-tumorigenic effects induced by RF ablation, is carried out by myofibroblasts and that their suppression might affect tumor growth and progression. They report on the key role of activated myofibroblasts in RFA-induced distant metastases growth and successful pharmacologic blockade in mouse and rat models of colon, renal and breast experimental models. Activated myofibroblasts are a key cell population in post RF ablation distant tumor growth, primarily through induction of the HGF/c-MET/STAT3 axis. Post RFA distant tumor growth was observed in different phenotypes of ablated primary organ such as liver or kidney or phenotype of tumors such as colorectal cancer or breast cancer. They also observed that RF induced distant tumor growth can be achieved by inhibition of activated myofibroblasts using atorvastatin, an established lipid lowering medication, and anti-fibroblast agent. This is a well-executed research with interesting and important results. I suggest that the authors will discuss the possibility that macrophage inhibition will achieve similar effects?” Response: We appreciate the Reviewer’s comments and enthusiasm for our work. We now address the potential role of macrophage suppression as well in the Discussion. “Furthermore, this observation makes a strong case for cell specific targeting strategies, especially, of other abundant cell populations with known tumor promoting properties within the tumor microenvironment such as M2 macrophages.” We have uploaded clean and annotated versions of the manuscript with tracked changes and figures, as requested. Once again we thank you for a favorable reply. Sincerely, Muneeb Ahmed, M.D. on behalf of all authors Attachments Attachment Submitted filename: PONE-D-21-21027.Response to Reviewers.doc https://doi.org/10.1371/journal.pone.0266522.r002
23 Mar 2022 Decision Letter - Yanlin Yu, Editor Myofibroblasts: A key promoter of tumorigenesis following radiofrequency tumor ablation. PONE-D-21-21027R1 Dear Dr. Ahmed, 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 for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, 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, Yanlin Yu Academic Editor PLOS ONE Additional Editor Comments (optional): Dear Dr. Ahmed, Thank you for submitting your revised manuscript to PLOS ONE. I am delighted to inform you that your manuscript, " Myofibroblasts: A key promoter of tumorigenesis following radiofrequency tumor ablation", has now been accepted for publication in PLOS ONE. Kind regards, Yanlin Reviewers' comments: https://doi.org/10.1371/journal.pone.0266522.r003
Formally Accepted
24 May 2022 Acceptance Letter - Yanlin Yu, Editor PONE-D-21-21027R1 Myofibroblasts: A key promoter of tumorigenesis following radiofrequency tumor ablation. Dear Dr. Ahmed: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. 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 plosone@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 Dr. Yanlin Yu Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0266522.r004

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 .