Identification of PPARG as key gene to link coronary atherosclerosis disease and rheumatoid arthritis via microarray data analysis

Zhenzhen Zhang; Yupeng Chen; Xiaodan Fu; Linying Chen; Junlan Wang; Qingqiang Zheng; Sheng Zhang; Xia Zhu

doi:10.1371/journal.pone.0300022

Peer Review History

Original SubmissionSeptember 7, 2023
28 Sep 2023 Decision Letter - Abozar Ghorbani, Editor PONE-D-23-28394Identification of PPARG as key gene to gap CAD and RA via microarray data analysisPLOS ONE Dear Dr. zhang, 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 12 2023 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. 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If applicable, please specify in the figure caption text when a figure is similar but not identical to the original image and is therefore for illustrative purposes only. Additional Editor Comments: 1. Fig 1. Need more explanation in legend. 2. The text need to be improved as English Grammar and scientific frame. 3. Space and “,” should be checked in the whole text. 4. Some DE genes should be validated using qRT-PCR or other techniques. 5. Quality of figures should be improved and the text and details should be clear. [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: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: N/A 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: Yes 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: This study identifies 737 DEGs associated with diseased groups in CAD datasets and 2008 DEGs associated with diseased groups in RA datasets. The author observed that TUBB2A, FKBP5, PPARG were significantly associated with RA and CAD. Major revisions are requested for this paper to be accepted. Major comments: 1. Although the authors made some efforts attempting to integrate several existing gene expression and datasets via GO and GSEA enrichment that they chose for over-represented functional gene sets, validation experiments that could support the authors' conclusion are missing, sufficient new/validated information is lacking. 2. This study need more work in order to identify more datasets or use data from authors' laboratory to show replication of the results. 3. The authors need to use a PCA and a regression SVM (R-SVM) to obtain a global view and evaluate the performance of DEGs in classifying control conditions versus CAD and RA conditions. 4. Lines 75 to 85 are not related to the introduction. 5. Most of the Figures are not clear eg, Fig 2 (A,B), Fig 4 (A,D,E), Fig 5 (A,B,C,D,F) , Fig 6 (E) and Fig 7 (A,B,C,D,E). The authors provide clearer Figures. Reviewer #2: It was a valuable study. However, some punctuation and grammatical errors are included within the manuscript. Therefore, with a minor revision and correction of the mistakes, this manuscript is absolutely suitable and recommended for publication. ****** 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: comment.docx https://doi.org/10.1371/journal.pone.0300022.r001
Revision 1
29 Dec 2023 Author Response Major comments: 1.Although the authors made some efforts attempting to integrate several existing gene expression and datasets via GO and GSEA enrichment that they chose for over-represented functional gene sets, validation experiments that could support the authors' conclusion are missing, sufficient new/validated information is lacking. Dear Reviewer, Thank you for your valuable feedback. In addition to bioinformatics analysis, our immunohistochemistry data validate the differential expression and localization of PPARG on synovial tissue and arterial wall samples from RA and CAD patients. The lack of validation in vivo and in vitro experiments in indeed the limitation of our study. Unfortunately, due to time constraints, reagent availability, and current financial limitations, we were unable to perform the validation experiments as part of this study.However, we want to assure you that the validation experiments are an integral part of our future research plan. Moreover, we speculated that the identified hub genes PPARG can gap RA and CAD through macrophages mediated Fc-gamma receptor-mediated endocytosis based on the following reference 1 Chawla A, Barak Y, Nagy L, Liao D, Tontonoz P, Evans RM. PPAR-gamma dependent and independent effects on macrophage-gene expression in lipid metabolism and inflammation. Nat Med. 2001 Nov;7(1):48-52. doi: 10.1038/83336. PMID: 11135614. Which support PPARG function in lipid metabolism induced inflammation, including cornary artery disease. 2 Toobian D, Ghosh P, Katkar GD. Parsing the Role of PPARs in Macrophage Processes. Front Immunol. 2021 Dec 22;12:783780. doi: 10.3389/fimmu.2021.783780. PMID: 35003101; PMCID: PMC8727354. Which provide evidence to support that PPARG has been implicated in regulating macrophage polarization, phagocytosis, inflammatory cytokine production, and lipid metabolism . 3 Brosig L, Hong J, Wallis BB, Giacomini JC, Assimes TL, Goronzy JJ, Weyand CM. Hypermetabolic macrophages in rheumatoid arthritis and coronary artery disease due to glycogen synthase kinase 3b inactivation. Ann Rheum Dis. 2018 Jul;77(7):1053-1062. doi: 10.1136/annrheumdis-2017-212647. Epub 2018 Feb 3. PMID: 29431119. Which support that the macrophages implicated in the pathogenesis of both RA and CAD 2. This study need more work in order to identify more datasets or use data from authors' laboratory to show replication of the results. Dear Reviewer, Thank you for your valuable feedback. In this study, we have carried out several verification steps. Firstly, 25 DEGs obtained from the intersection of DEGs in RA (GSE5523+GSE55457) and CAD (GSE42148) datasets and WGCNA were validated in validation datasets(RA GSE12021 and CAD GSE20680), referenced to (Fig.2)Additionally, we performed PCA and SVM analysis ( Fig.3 ) and ROC analysis to identify diagnostic discrimination ability in disease group and normal group of the 25 core genes and the three hub genes on both training and validation datasets(Fig.5) . Then qRT-PCR was conducted to validate the mRNA expression level of three hub genes in RA and CAD disease compared with their matched normal tissues. Finally, we conducted immunohistochemistry experiments to confirm the expression and localization of the core gene PPARG in synovial tissue and arterial wall macrophages from patients with chronic synovitis and coronary artery disease to support our bioinformatic study from clinical aspects (Fig.9) We acknowledge the importance of replicating our findings to enhance the robustness of our study. However, we carefully considered various factors in our analysis, including sample size matching, sample types, result integration of the article logic, which led us to prioritize the mentioned datasets. Furthermore, the experimental validations will be supplemented in future studies and grant applications. 3.The authors need to use a PCA and a regression SVM (R-SVM) to obtain a global view and evaluate the performance of DEGs in classifying control conditions versus CAD and RA conditions. Dear Reviewer, Thank you for your valuable feedback and suggestions. We obtained the 25 core genes from the intersection of DEGs in RA (GSE5523+GSE55457) and CAD (GSE42148) datasets and WGCNA. To validate the discriminatory ability of 25 core genes in distinguishing disease group from normal group in both RA and CAD, the expression matrix was extracted from the training and validation datasets of RA and CAD . PCA analysis was performed on the new datasets using R packages FactoMineR and factoextra . SVM analysis was then conducted using functions from the R package e1071. Finally, the classification performance of the 25 core genes was visualized by ROC curves using pROC package.Fig.5 showed the 25 core genes distinguished the disease group from the normal group in both the RA and CAD training and validation datasets with AUCs above 0.8. 4.Lines 75 to 85 are not related to the introduction. Dear Reviewer, Thank you for your valuable feedback. We have deleted the content of line75 to 85 in the introduction and revised as follows: Potential targets and regulatory biological pathways were identified in RA and CAD datasets respectively using microarray bioinformatics technology in recent years, however, there is a dearth of studies that investigate the macrophages related inflammatory processes between RA and CAD. This research aims to fill this gap by employing GSEA analysis to identify common signaling pathways on RA and CAD datasets.WGCNA and ssGSEA will identify module genes linked to Fc-gamma receptor phagocytosis. Three hub genes will be extracted using STRING and Cytoscape, then assessing their predictive ability in both diseases through ROC analysis. GeneMANIA and the Cibersort algorithm will provide insights into immune infiltration and transcription factors associated with the hub genes. Notably, the involvement of PPARG and macrophages in RA and CAD will be highlighted. Understanding these processes could offer new insights into the pathogenesis shared by both diseases and potentially aid in the development of targeted therapies. 5.Most of the Figures are not clear eg, Fig 2 (A,B), Fig 4 (A,D,E), Fig 5 (A,B,C,D,F) , Fig 6 (E) and Fig 7 (A,B,C,D,E). The authors provide clearer Figures. Thank you for your feedback on the clarity of our figures. We appreciate your attention to detail and would like to clarify the issue you mentioned. We have compared the raw figures in TTF format and the figures of PDF versions, the latter indeed have lower resolution and pixel quality compared to the TTF versions. I think the review can try to accesse the raw figures in TTF format using the provided link in the top right corner of the PDF figures as marked in the following attached figures.We apologize for any inconvenience caused and appreciate your understanding. If you have any further questions or concerns, please do not hesitate to let us know. We appreciate your time and effort in reviewing our work. Pdf figure Editor Comments 1.Fig 1. Need more explanation in legend. Thank you for your feedback on Fig 1 legend. We have revised as follows: ＂Fig 1. Data analysis flowchart. Schematic flowchart of data aquirement, processing ,analysis and validation.＂ 2.The text need to be improved as English Grammar and scientific frame. Thank you for your suggestions. We have made revise by the editors and the proofreading certificate was uploaded in the supplementary materials. 3.Space and “,” should be checked in the whole text. Thank you for your remind. We have checked the whole text to correct this mistake. 4.Some DE genes should be validated using qRT-PCR or other techniques. Thank you for your suggestions. We have performed qRT-PCR to validate TUBB2A, PPARG and FKBP5 mRNA expression in RA and CAD tissues and their mathced adjacent normal tissues.The results was demonstrated and in the results “GO and KEGG pathway analysis of 25 susceptible genes in RA and CAD”and showed in Fig.6G-H 5.Quality of figures should be improved and the text and details should be clear. Thank you for your suggestions. We have assembled our figures with adobe illustration software and upload the figure again. If there is something unsatisfactory, please don’t hesitate to contact us. Best wishes! Yours sincerely! Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0300022.r002
11 Jan 2024 Decision Letter - Abozar Ghorbani, Editor PONE-D-23-28394R1Identification of PPARG as key gene to link coronary atherosclerosis disease and rheumatoid arthritis via microarray data analysisPLOS ONE Dear Dr. zhang, 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 25 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, Abozar Ghorbani, Ph.D 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 #3: 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 #3: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #3: 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 #3: 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 #3: 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: 1. Briefly refer to the used data in the abstract of the method section 2. The reference genes should be specified in the real-time PCR section 3. What method has been used to analyze real-time PCR data? Reviewer #3: The study is very interesting, as well as, the manuscript is improved very well, in my opinion, you can accept it. However, it is recommended to edit the following comments. ****** 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 #3: Yes: Fatemeh Yaghoobizadeh ******** [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: Comment 2.docx Attachment Submitted filename: comments.docx https://doi.org/10.1371/journal.pone.0300022.r003
Revision 2
14 Jan 2024 Author Response Reviewer #1: The study is very interesting, as well as, the manuscript is improved very well, in my opinion, you can accept it. However, it is recommended to edit the following comments. 1.Briefly refer to the used data in the abstract of the method section Response:Thank you for your comment. I have added the names of the datasets in the revised abstract of the method section as follows“The study utilized five raw datasets: GSE55235 , GSE55457 , GSE12021 for RA patients, and GSE42148 and GSE20680 for CAD patients in line 24-25” 2.The reference genes should be specified in the real-time PCR section Response: Thank you for your comment. I have specified the reference genes as follows”GAPDH was used as reference gene” in line 165 3.What method has been used to analyze real-time PCR data? Response: Thank you for your comment. I have mentioned the method usde to analyze real-time PCR data as follows”Gene expression was calculated according to the 2-ΔΔCT method .” in line 165-166. Reviewer #3 The study is very interesting, as well as, the manuscript is improved very well, in my opinion, you can accept it. However, it is recommended to edit the following comments. 1.Generally, the pronouns (such as I, we, us) are avoided to be written in a technical research article. In the regard, try to reframe the text, e.g., line 132, 173, 216, 277, 281, 303, etc. Response: Thank you for your comments. We have made revisions to the text accordingly. And the “revised manuscript with track changes” has been submitted for your review. 2.There is a blank page between the 154 and 156 lines. Please edit this error. Response: Thank you for pointing out the error. We apologize for the oversight. We have now removed the blank page between lines 154 and 156 as shown in the “revised manuscript with track changes”. 3.The diagrams quality is quite low. It is required to include high-resolution pictures for further comments. Response: Thank you for your comments. We have already recombined the PDF images using Adobe Illustrator to generate vector graphics, exported them in TTF format with a PPI of 600dp, and re-uploaded them in the submitting system, The original images remain clear even when enlarged by 150%. I hope it will be OK. 4.Please review the numbers of figures. I think you mean “Figure 6” in line: 278, 282, 287, etc. and after the above-mentioned lines. Response:Thank you for your comment. We have now reviewed the number of figures in the manuscript and made the necessary changes in lines 278, 282, 287, and other relevant lines, . The revised manuscript with track changes has been submitted for your review. 5.Please double check for spelling and English grammar errors (e.g., Space and “,”, and integral and correct time tense for whole document) in the manuscript. Response: Thank you for your comment. We have now thoroughly reviewed the manuscript for spelling and English grammar errors. This includes checking for spaces, commas and the correct tense .The revised manuscript with track changes has been submitted for your review. 6.In line 317, please clearly indicate the corresponding figure. Response: Thank you for bringing this to our attention. We have revised the manuscript according to your comment and clearly indicated the corresponding figure in line 317. We have made these changes in the revised manuscript with track changes. 7.It seems lines 382-394 are redundant in the current position. It is recommended to insert the “Conclusion” section to state your total conclusion of the findings. Response: Thank you for your valuable suggestions. We have revised the manuscript accordingly. The redundant sections from lines 382-394 have been removed, and a "Conclusion" section has been inserted to provide a comprehensive summary of our findings. The revised manuscript with track changes has been submitted for your review. 8.It is recommended to use an integral format of abbreviations and full form of terms, e.g. rheumatoid arthritis in line 437. Response: Thank you for your comment. We have made the necessary changes in the revised manuscript with track changes, Except for the first occurrence in the abstract and the introduction, where 'rheumatoid arthritis' is used in full form, the abbreviation 'RA' is used throughout the rest of the manuscript. Please review the revised manuscript for detail. 9.Please double-check the author’s citation format in lines 435, 458, etc. Response: Thank you for your comment. I am not entirely sure about the issue with the citation format in lines 435 and 458, as I have checked and found that they correspond with the numbering of the references listed below. The citation format is consistent with the above ones as well and the published article in “plos one”. Could you please provide more specific details about the problem? Thank you for your patience. 10. Please double check the formats of all references. Response: Thank you for your comment. I have checked all the references and replaced reference 7 from Microsoft Azure platform with other reference. Some references have been supplemented with published articles from Plos One, including volumes (issues) and DOI links following the published article in Plos One.Please review the revised manuscript for detail. Attachments Attachment Submitted filename: Rebutt letter.docx https://doi.org/10.1371/journal.pone.0300022.r004
2 Feb 2024 Decision Letter - Abozar Ghorbani, Editor PONE-D-23-28394R2Identification of PPARG as key gene to link coronary atherosclerosis disease and rheumatoid arthritis via microarray data analysisPLOS ONE Dear Dr. zhang, 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 Mar 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, Abozar Ghorbani, Ph.D 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. Additional Editor Comments: Because of your request to correct your mistake, I am returning to you to submit the correct version.Authors comments: There are some mistakes in the figrue order and loss of figure 9 as showed in the previewed pdf file [Note: HTML markup is below. Please do not edit.] [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.0300022.r005
Revision 3
5 Feb 2024 Author Response Reviewer #1: The study is very interesting, as well as, the manuscript is improved very well, in my opinion, you can accept it. However, it is recommended to edit the following comments. 1.Briefly refer to the used data in the abstract of the method section Response:Thank you for your comment. I have added the names of the datasets in the revised abstract of the method section as follows“The study utilized five raw datasets: GSE55235 , GSE55457 , GSE12021 for RA patients, and GSE42148 and GSE20680 for CAD patients in line 24-25” 2.The reference genes should be specified in the real-time PCR section Response: Thank you for your comment. I have specified the reference genes as follows”GAPDH was used as reference gene” in line 165 3.What method has been used to analyze real-time PCR data? Response: Thank you for your comment. I have mentioned the method usde to analyze real-time PCR data as follows”Gene expression was calculated according to the 2-ΔΔCT method .” in line 165-166. Reviewer #3 The study is very interesting, as well as, the manuscript is improved very well, in my opinion, you can accept it. However, it is recommended to edit the following comments. 1.Generally, the pronouns (such as I, we, us) are avoided to be written in a technical research article. In the regard, try to reframe the text, e.g., line 132, 173, 216, 277, 281, 303, etc. Response: Thank you for your comments. We have made revisions to the text accordingly. And the “revised manuscript with track changes” has been submitted for your review. 2.There is a blank page between the 154 and 156 lines. Please edit this error. Response: Thank you for pointing out the error. We apologize for the oversight. We have now removed the blank page between lines 154 and 156 as shown in the “revised manuscript with track changes”. 3.The diagrams quality is quite low. It is required to include high-resolution pictures for further comments. Response: Thank you for your comments. We have already recombined the raw PDF images using Adobe Illustrator to generate vector graphics, exported them in TTF format with a PPI of 300dpi, then checked and adjusted in the PACE system before re-uploaded them in the submitting system, The original images remain clear even when enlarged by 150%, which can be obtained from link on the the right corner of the pdf files. I hope it will be OK. I think figures in the pdf files may be compressed too much. 4.Please review the numbers of figures. I think you mean “Figure 6” in line: 278, 282, 287, etc. and after the above-mentioned lines. Response:Thank you for your comment. We have now reviewed the number of figures in the manuscript and made the necessary changes in lines 278, 282, 287, and other relevant lines, . The revised manuscript with track changes has been submitted for your review. 5.Please double check for spelling and English grammar errors (e.g., Space and “,”, and integral and correct time tense for whole document) in the manuscript. Response: Thank you for your comment. We have now thoroughly reviewed the manuscript for spelling and English grammar errors. This includes checking for spaces, commas and the correct tense .The revised manuscript with track changes has been submitted for your review. 6.In line 317, please clearly indicate the corresponding figure. Response: Thank you for bringing this to our attention. We have revised the manuscript according to your comment and clearly indicated the corresponding figure in line 317. We have made these changes in the revised manuscript with track changes. 7.It seems lines 382-394 are redundant in the current position. It is recommended to insert the “Conclusion” section to state your total conclusion of the findings. Response: Thank you for your valuable suggestions. We have revised the manuscript accordingly. The redundant sections from lines 382-394 have been removed, and a "Conclusion" section has been inserted to provide a comprehensive summary of our findings. The revised manuscript with track changes has been submitted for your review. 8.It is recommended to use an integral format of abbreviations and full form of terms, e.g. rheumatoid arthritis in line 437. Response: Thank you for your comment. We have made the necessary changes in the revised manuscript with track changes, Except for the first occurrence in the abstract and the introduction, where 'rheumatoid arthritis' is used in full form, the abbreviation 'RA' is used throughout the rest of the manuscript. Please review the revised manuscript for detail. 9.Please double-check the author’s citation format in lines 435, 458, etc. Response: Thank you for your comment. I am not entirely sure about the issue with the citation format in lines 435 and 458, as I have checked and found that they correspond with the numbering of the references listed below. The citation format is consistent with the above ones as well and the published article in “plos one”. Could you please provide more specific details about the problem? Thank you for your patience. 10. Please double check the formats of all references. Response: Thank you for your comment. I have checked all the references and replaced reference 7 from Microsoft Azure platform with other reference. Some references have been supplemented with published articles from Plos One, including volumes (issues) and DOI links following the published article in Plos One.Please review the revised manuscript for detail. 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. Response: Thank you for your comment. I have checked all the references and replaced reference 7 from Microsoft Azure platform with other reference. Some references have been supplemented with published articles from Plos One, including volumes (issues) and DOI links following the published article in Plos One.Please review the revised manuscript for detail. None of the reference was restracted by now after throughly checked in the pubmed. Attachments Attachment Submitted filename: Rebutt letter.docx https://doi.org/10.1371/journal.pone.0300022.r006
21 Feb 2024 Decision Letter - Abozar Ghorbani, Editor Identification of PPARG as key gene to link coronary atherosclerosis disease and rheumatoid arthritis via microarray data analysis PONE-D-23-28394R3 Dear Dr. zhang, 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, Abozar Ghorbani, 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 #1: 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 ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: 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 ****** 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 ****** 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) ****** 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 ******** https://doi.org/10.1371/journal.pone.0300022.r007
Formally Accepted
26 Mar 2024 Acceptance Letter - Abozar Ghorbani, Editor PONE-D-23-28394R3 PLOS ONE Dear Dr. Zhang, 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. Abozar Ghorbani Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0300022.r008

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