Pine pollen reverses the function of hepatocellular carcinoma by inhibiting α-Enolase mediated PI3K/AKT signaling pathway

Yanhong Luo; Chun Guo; Caixia Ling; Wenjun Yu; Yuanhong Chen; Lihe Jiang; Qiuxiang Luo; Chunfang Wang; Weixin Xu

doi:10.1371/journal.pone.0312434

Peer Review History

Original SubmissionMay 20, 2024
4 Jul 2024 Decision Letter - Turki Talal Turki, Editor PONE-D-24-17156Pine pollen reverses the function of hepatocellular carcinoma by inhibiting α-Enolase mediated PI3K / Akt signaling pathwayPLOS ONE Dear Dr. Xu, 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 Aug 18 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, Turki Talal Turki, Ph.D. 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. We note that the grant information you provided in the ‘Funding Information’ and ‘Financial Disclosure’ sections do not match. When you resubmit, please ensure that you provide the correct grant numbers for the awards you received for your study in the ‘Funding Information’ section. 3. Thank you for stating the following financial disclosure: "This study was supported by the National Natural Science Foundation of China (No. 81960303); Project of Guangxi Key Laboratory of Molecular Pathology of Hepatobiliary Diseases (No. [2021]61); the Foundation of Modern Industrial College of Biomedicine and Great Health, Youjiang Medical University for Nationalities, Baise, Guangxi, China; School level project of Youjiang Medical University for Nationalities (No. yy2021sk012). " 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." If this statement is not correct you must amend it as needed. Please include this amended Role of Funder statement in your cover letter; we will change the online submission form on your behalf. 4. 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Please ensure that your ethics statement is included in your manuscript, as the ethics statement entered into the online submission form will not be published alongside your manuscript. 8. Please remove your figures from within your manuscript file, leaving only the individual TIFF/EPS image files, uploaded separately. These will be automatically included in the reviewers’ PDF." 9. Please include captions for your Supporting Information files at the end of your manuscript, and update any in-text citations to match accordingly. Please see our Supporting Information guidelines for more information: http://journals.plos.org/plosone/s/supporting-information. [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 ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: No ****** 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 manuscript entitled pollen reverses the function of hepatocellular carcinoma by inhibiting α-Enolase mediated PI3K PI3K/Akt pathway” pathway to investigate the role of ENO1 in hepatocellular carcinoma cells and its relationship with the PI3K/Akt signalling signaling pathway. The focus was ENO1 ENO1, which highly expressed in tumour tumor and correlated with tumour tumor and metastasis. The Inhibiting ENO1 could reduce the of the PI3K/Akt signalling signaling pathway, inhibiting the proliferation and migration of hepatocellular carcinoma cells. The manuscript is well-organized and clearly stated. I would suggest accept accepting it the following major concerns are addressed: 1.The article discusses the role of a metalloenzyme, ENO1, in glycolysis and the regulation of its expression in tumor cells. The significance of ENO1 and the related research background could have been presented in more detail with more clarity in the introductory section. 2.The use of the CPTAC database to study the phosphorylation sites and levels of ENO1 in tumour tissues is mentioned in the article, and the methods and results of data analysis need to be explained in more detail to ensure the reliability and accuracy of the data. 3.Figure 6 is not clear and the need to avoid duplicates in Figure 6C. 4.Suggested supplementary tumour volume images in nude mice. 5.References need to be renewed. Some important recent studies regarding the hepatocellular carcinoma should be cited and discussed. For example, PMID: 38827325, 33987373, 38739668, etc. 6.It is suggested to add a supplementary figure with a graphic abstract which could better clarify the significance of this study. Reviewer #2: Luo et al. presented a study applying various computational and experimental analysis to investigate the biological functions of Pine pollen in hepatocellular carcinoma. They found that Pine pollen inhibits HCC via ENO1, MBP-1 and PI3K/Akt pathway. The analysis and results appear intriguing and promising; however, there are several major issues in the current manuscript that undermine the study's clarity and conclusion. 1. The manuscript's English writing is quite poor, which significantly affects its readability. For example, on page 2 line 39, it is confusing to read “We analyzed the bioinformatics of ENO1”, which should be revised to “We applied various bioinformatic analysis of open-access data to study the expression of ENO1”. It is recommended that the authors seek assistance from a professional editing service or a native English speaker to enhance the quality of the writing. 2. The structure of the paper is currently disorganized, such that there are nine figures and seven tables totally, making it difficult for readers to follow the authors’ arguments and findings. The authors should reorganize the figures/tables to provide a coherent flow of information, such as putting some figures into supplementary file. Also, the index in Figure 3 is incorrect, please revise that. 3. The statistical analysis presented in the manuscript is not adequately justified and appears unreasonable in parts. For example, on page 18 line 184, the authors claimed that they applied one-way ANOVA test; however, at least in Figure 2 the test should be t-test or Wilcoxon test depends on the distribution of data. Besides, is the p-value in this study adjusted? The authors should revisit their analytical methods to ensure they are appropriate for their data. 4. The interpretation of the figures in the manuscript needs to be more accurate and detailed. Each figure and panel should be clearly explained. For example, how does each part in Figure 4 support the conclusion “Establishment of stable cell line overexpressing ENO1”? Overall, the current manuscript cannot be considered for publication. The authors should improve the quality and clarity substantially so that the manuscript can meet the publication standards. ****** 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. Attachments Attachment Submitted filename: Review-Comments-PlosOne.docx https://doi.org/10.1371/journal.pone.0312434.r001
Revision 1
14 Aug 2024 Author Response 1. The English has been polished by authoritative institutions. 2. The table of cell scratch experiment has been included as a supplementary document. The index of Figure 3 has been modified. 3. The inter group analysis with time overlap adopts multiple analysis of variance. The numerical change was misread at the time, and the P-value has not been changed. I consulted with statistical experts, and they believe that this is correct. 4. The chart has been reorganized: Figure 4 (A) White light and fluorescence photos of overexpressed ENO1 cells transfected for 72 hours, with fluorescence cells accounting for approximately 80% of the white light cell count; Fluorescence represented successfully transfected cells. (B) White light and fluorescence photos of empty vector cells transfected for 72 hours, with fluorescent cells accounting for approximately 80% of the white light cell count; Fluorescence represented successfully transfected cells. (C) White light and fluorescence photos of non transfected cells cultured for 72 hours, but the cells did not show fluorescence; (D) Photos of overexpressed ENO1 cells screened with 20 μg/μl purinomycin for 3 days showed good cell growth; photos of empty transfected cells screened with 20 μg/μl purinomycin for 3 days showed good cell growth; and photos of non transfected cells screened with 20 μg/μl purinomycin for 3 days showed that all cells were killed by puromycin; Screening with puromycin involves killing untransfected cells that are not resistant to puromycin, while screening stable transfected cells that are resistant to puromycin. (E) RT-qPCR dissolution curve of overexpression ENO1 cells showed that the amplification product was ENO1; The histogram of relative expression of ENO1 RT-qPCR in overexpression ENO1 cells and empty vector cells showed the expression level of ENO1 in overexpressing ENO1 transfected cells was 25.09 times higher than that in empty transfected cells. These results indicated that ENO1 could be overexpressed in overexpressing ENO1 transfected cells. Figure 5 (A) shows that the empty transfected cells were cultured in wells without PP solution for 12 hours as a negative control; Overexpression stable transformed cells were cultured in 5 μ L PP solution wells for 12 hours; Overexpression stable transformed cells were cultured in 75 μ L PP solution wells for 12 hours; (B) The empty transfected cells were cultured in wells without PP solution for 24 hours as a negative control; Overexpression stable transformed cells were cultured in 5 μ L PP solution wells for 24 hours; Overexpression stable transformed cells were cultured in 75 μ L PP solution wells for 24 hours. This indicates that compared to the negative control, as the concentration of PP solution increases, the degree of inhibition of overexpression stable transformed cells becomes more severe. (C) The relationship between the logarithm of drug concentration and the corresponding cell inhibition rate of PP solution after 12 hours of action was plotted. The calculated IC50 was 5.78 μ g/μ l, which was used as the PP solution concentration and corresponding action time for subsequent experiments. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0312434.r002
20 Sep 2024 Decision Letter - Turki Talal Turki, Editor PONE-D-24-17156R1Pine pollen reverses the function of hepatocellular carcinoma by inhibiting α-Enolase mediated PI3K/AKT signaling pathwayPLOS ONE Dear Dr. Xu, 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 Nov 04 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, Turki Talal Turki, Ph.D. Academic Editor PLOS ONE [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: (No Response) 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: (No Response) Reviewer #2: Partly ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: (No Response) Reviewer #2: N/A ****** 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: (No Response) 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: (No Response) 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: (No Response) Reviewer #2: The authors addressed part of my previous comments and improved the manuscript to some extent. However, there are still some major issues with this paper: Major comments: 1. When preparing the “Response To Reviewer Comments”, the authors should copy and paste the original comments from last round and then provide the point-by-point response to each comment. This is to maintain an easy and straightforward tracking of all comments and responses. Also, as the revision guidelines indicated, the authors should upload a marked-up copy of manuscript that highlights changes made to the original version (upload this as a separate file labeled 'Revised Manuscript with Track Changes') 2. “The English has been polished by authoritative institutions.” The English writing has indeed improved in current submission; However, there are still some sentences appear incorrect, such as in Abstract, Conclusion, the authors wrote “PP inhibits HCC by regulating the expression of ENO1 and MBP-1 and inhibiting the 53 PI3K/AKT pathway by inhibiting C-MYC and erb-B2 receptor tyrosine kinase 2.” It is confusing to read all three “inhibiting”, please revise that to make it clearer and more concise. 3. The format of reference numbers is incorrect. If the numbers are listed as superscript, then there should not be brackets around them. Please revise that and follow the official format by the journal. ****** 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.0312434.r003
Revision 2
3 Oct 2024 Author Response 1. The manuscript's English writing is quite poor, which significantly affects its readability. For example, on page 2 line 39, it is confusing to read “We analyzed the bioinformatics of ENO1”, which should be revised to “We applied various bioinformatic analysis of open-access data to study the expression of ENO1”. It is recommended that the authors seek assistance from a professional editing service or a native English speaker to enhance the quality of the writing. Answer: The English has been polished by authoritative institutions. 2. The structure of the paper is currently disorganized, such that there are nine figures and seven tables totally, making it difficult for readers to follow the authors’ arguments and findings. The authors should reorganize the figures/tables to provide a coherent flow of information, such as putting some figures into supplementary file. Also, the index in Figure 3 is incorrect, please revise that. Answer: The table of cell scratch experiment has been included as a supplementary document. The index of Figure 3 has been modified. 3. The statistical analysis presented in the manuscript is not adequately justified and appears unreasonable in parts. For example, on page 18 line 184, the authors claimed that they applied one-way ANOVA test; however, at least in Figure 2 the test should be t-test or Wilcoxon test depends on the distribution of data. Besides, is the p-value in this study adjusted? The authors should revisit their analytical methods to ensure they are appropriate for their data. Answer: Multivariate analysis of variance is used for intergroup analysis with temporal overlap in statistics. The numerical change was misread at the time, and the P-value has not been changed. I consulted with statistical experts, and they believe that this is correct. 4. The interpretation of the figures in the manuscript needs to be more accurate and detailed. Each figure and panel should be clearly explained. For example, how does each part in Figure 4 support the conclusion “Establishment of stable cell line overexpressing ENO1”? Answer: The chart has been reorganized: Figure 4 and Figure 4 (A) show the white light and fluorescence images of cells overexpressing ENO1 after transfection for 72 hours, with fluorescent cells accounting for approximately 80% of the white light cells; Fluorescence represents successfully transfected cells. (B) White light and fluorescence photos of empty vector cells transfected for 72 hours, with fluorescent cells accounting for approximately 80% of the white light cells; Fluorescence represents successfully transfected cells. (C) White light and fluorescence photos of untransfected cells cultured for 72 hours, but the cells did not show fluorescence; (D) After screening cells overexpressing ENO1 with 20 μ g/μ l puromycin for 3 days, the photos showed good cell growth; After screening empty vector transfected cells with 20 μ g/μ l puromycin for 3 days, the photos showed good cell growth; After screening untransfected cells with 20 μ g/μ l puromycin for 3 days, the photos showed that all cells were killed by puromycin; Screening with puromycin involves killing untransfected cells that are intolerant to puromycin, while selecting stable transfected cells that are tolerant to puromycin. (E) The RT qPCR dissolution curve of overexpressing ENO1 cells showed that the amplified product was ENO1; The histogram of relative expression levels of ENO1 RT qPCR in overexpressing ENO1 cells and empty vector cells showed that the expression level of ENO1 in overexpressing ENO1 transfected cells was 25.09 times higher than that in empty vector transfected cells. These results indicate that ENO1 can be overexpressed in cells transfected with ENO1. Figure 5 (A) shows that the empty transfected cells were cultured in wells without PP water extract for 12 hours as a negative control; Overexpression stable transformed cells were cultured in 5 μ L PP water extract wells for 12 hours; Overexpression stable transformed cells were cultured in 75 μ L PP water extract wells for 12 hours; (B) The empty transfected cells were cultured in wells without PP water extract for 24 hours as a negative control; Overexpression stable transformed cells were cultured in 5 μ L PP water extract wells for 24 hours; Overexpression stable transformed cells were cultured in 75 μ L PP water extract wells for 24 hours. This indicates that compared to the negative control, as the concentration of PP water extract increases, the degree of inhibition of overexpression stable transformed cells becomes more severe. (C) The relationship between the logarithm of drug concentration and the corresponding cell inhibition rate of PP solution after 12 hours of action was plotted. The calculated IC50 was 5.78 μ g/μ l, which was used as the PP solution concentration and corresponding action time for subsequent experiments. Reviewer's comment response: 1. When preparing the “Response To Reviewer Comments”, the authors should copy and paste the original comments from last round and then provide the point-by-point response to each comment. This is to maintain an easy and straightforward tracking of all comments and responses. Also, as the revision guidelines indicated, the authors should upload a marked-up copy of manuscript that highlights changes made to the original version (upload this as a separate file labeled 'Revised Manuscript with Track Changes') Answer: We have responded to the comments one by one and provided a manuscript with revision marks, named "Revised Manuscript". 2. “The English has been polished by authoritative institutions.” The Englishwriting has indeed improved in current submission; However, there are still some sentences appear incorrect, such as in Abstract, Conclusion, the authors wrote “PP inhibits HCC by regulating the expression of ENO1 and MBP-1 and inhibiting the 53 PI3K/AKT pathway by inhibiting C-MYC and erb-B2 receptor tyrosine kinase 2.” It is confusing to read all three “inhibiting”, please revise that to make it clearer and more concise. Answer: The second "inhibiting" in "PP inhibitors HCC by regulating the expression of ENO1 and MBP-1 and inhibiting the PI3K/AKT pathway by inhibiting C-MYC and erb-B2 receptor tyrosine kinase 2." has been changed to "suppressing". 3. The format of reference numbers is incorrect. If the numbers are listed as superscript, then there should not be brackets around them. Please revise that and follow the official format by the journal. Answer: The superscript of the reference number has been changed to normal. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0312434.r004
8 Oct 2024 Decision Letter - Turki Talal Turki, Editor Pine pollen reverses the function of hepatocellular carcinoma by inhibiting α-Enolase mediated PI3K/AKT signaling pathway PONE-D-24-17156R2 Dear Dr. Xu, 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, Turki Talal Turki, Ph.D. 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 #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 #2: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #2: N/A ****** 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 #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 #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 #2: All comments have been addressed in this revised manuscript, and it has been improved to a large extent. ****** 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 #2: No ******** https://doi.org/10.1371/journal.pone.0312434.r005
Formally Accepted
4 Nov 2024 Acceptance Letter - Turki Talal Turki, Editor PONE-D-24-17156R2 PLOS ONE Dear Dr. Xu, 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 Dr. Turki Talal Turki Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0312434.r006

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