Mmu-let-7a-5p inhibits macrophage apoptosis by targeting CASP3 to increase bacterial load and facilities mycobacterium survival

Xuehua Zhan; Wenqi Yuan; Rong Ma; Yueyong Zhou; Guangxian Xu; Zhaohui Ge

doi:10.1371/journal.pone.0308095

Peer Review History

Original SubmissionSeptember 27, 2023
9 Jan 2024 Decision Letter - Francesco Sessa, Editor PONE-D-23-31553mmu-let-7a-5p inhibits macrophage apoptosis by targeting CASP3 to increase bacterial load and promote facilities mycobacteria survivalPLOS ONE Dear Dr. Ge, 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 23 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. 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When you submit your revised manuscript, please ensure that your figures adhere fully to these guidelines and provide the original underlying images for all blot or gel data reported in your submission. See the following link for instructions on providing the original image data: https://journals.plos.org/plosone/s/figures#loc-original-images-for-blots-and-gels. In your cover letter, please note whether your blot/gel image data are in Supporting Information or posted at a public data repository, provide the repository URL if relevant, and provide specific details as to which raw blot/gel images, if any, are not available. Email us at plosone@plos.org if you have any questions. Additional Editor Comments: This study aims to explore the impact of mmu-let-7a-5p on Raw264.7 mononuclear macrophage apoptosis and its influence on intracellular Bacillus Calmette-Guérin (BCG) load in vitro. While the study holds potential interest for readers, there are several significant changes required. The abstract section needs enhancement, specifically in introducing the theme, outlining the aims, and summarizing methods and results. Additionally, the conclusion section should outline future research directions, as the current presentation is unclear. The introduction section should also be improved with background information relating to the study's aims. Furthermore, the material and methods section would benefit from the inclusion of a schematic diagram illustrating the study's protocol and a justification for using GAPDH as a reference gene for qPCR detection of mRNA. Additionally, the authors should provide the original figures of the western blot as supplementary files for the results section and submit the main raw data as supplementary files. In the discussion section, the authors should refrain from comparing the present study's data to their previous findings and expand the analysis to include international data. Moreover, it should be important to insert the section limitations. Lastly, the conclusion section should suggest future research directions. Minor points: - English should be reviewed by a native speaker. - Please, check the use of reference manager software (Error! Reference source not found..-> line 121). - The short form should be introduced by the extension form the first time; subsequently, it should be used through the text. Please, check all abbreviations. [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: No Reviewer #2: Partly ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: I Don't Know Reviewer #2: N/A ****** 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: I am sorry that I have little understanding of the content of this article and cannot make scientific and correct judgments about it. Please find another reviewer. I'm sorry that I clicked on 'accept review' earlier (before seeing the manuscript) because it caused me to review the manuscript without being familiar with it, which is unfair to the author. Reviewer #2: This study found that mmu-let-7a-5p regulates the mycobacterial load in macrophages through apoptosis, which is of certain value and innovation, because in addition, reports about it focus on tumors and cardiovascular diseases, and few reports are related to infectious diseases. But there are some issues in the article that need to be revised 1. The figure legends were not clear, and there was no explanation of the production method or number of replicates for each group of samples. 2. There are A-H graphs in Figure 8, but only A-B is explained and does not match 3. There are some columnar statistical charts, although the p-value is less than 0.05, the actual difference is not significant, which usually means that there may be some deviation between the statistical significance and the actual biological significance. As shown in Figure 8F, Figure 7B, etc 4. Figure 8G and Figure 8H, qRT PCR to detect mRNA of Caspase-3 Pro and Caspase-3 cleared. In fact, qRT PCR cannot distinguish between Caspase-3 Pro and Caspase-3 cleared. You need to explain. ****** 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 comment1.docx https://doi.org/10.1371/journal.pone.0308095.r001
Revision 1
19 Mar 2024 Author Response Dear Editors and Reviewers: Thank you for your letter and for the reviewer’s comments concerning our manuscript entitled “mmu-let-7a-5p inhibits macrophage apoptosis by targeting CASP3 to increase bacterial load and promote facilities mycobacteria survival” [manuscript number: PONE-D-23-31553]. First of all, please let me express my respect and sincere thanks for your conscientious work attitude. Those comments are all valuable and very helpful for revising and improving our manuscript, as well as the important guiding significance to our researchers. Review comments This study found that mmu-let-7a-5p regulates the mycobacterial load in macrophages through apoptosis, which is of certain value and innovation, because in addition, reports about it focus on tumors and cardiovascular diseases, and few reports are related to infectious diseases. But there are some issues in the article that need to be revised 1. The figure legends were not clear, and there was no explanation of the production method or number of replicates for each group of samples. 2. There are A-H graphs in Figure 8, but only A-B is explained and does not match 3. There are some columnar statistical charts, although the p-value is less than 0.05, the actual difference is not significant, which usually means that there may be some deviation between the statistical significance and the actual biological significance. As shown in Figure 8F, Figure 7B, etc 4. Figure 8G and Figure 8H, qRT PCR to detect mRNA of Caspase-3 Pro and Caspase-3 cleared. In fact, qRT PCR cannot distinguish between Caspase-3 Pro and Caspase-3 cleared. You need to explain. Reply We have studied comments carefully and have made correction which we hope meet with approval. Revised portion are marked in red in the paper. The main corrections in the paper and the responds to the reviewer’s comments are as flowing: 1. The figure legends were not clear, and there was no explanation of the production method or number of replicates for each group of samples. Thank you for your comments. We have revised our figure legends and annotated description of replicates for each group of samples. 2. There are A-H graphs in Figure 8, but only A-B is explained and does not match We are very sorry for our negligence of these mistakes in the manuscript writing and we have carefully revised these mistakes. 3. There are some columnar statistical charts, although the p-value is less than 0.05, the actual difference is not significant, which usually means that there may be some deviation between the statistical significance and the actual biological significance. As shown in Figure 8F, Figure 7B, etc. We are extremely grateful to Reviewer for pointing out this problem. We have double checked the raw data and re-calculated the statistics by using Graphpad Prism 9.0. There are indeed significant differences in the results of statistical charts in our manuscripts. 4. Figure 8G and Figure 8H, qRT PCR to detect mRNA of Caspase-3 Pro and Caspase-3 cleared. In fact, qRT PCR cannot distinguish between Caspase-3 Pro and Caspase-3 cleared. You need to explain. Due to our negligence, we did not explain exactly about the figure legends, especially Figure 8. In fact, Figure 8.F-H were the quantitative analysis of TNFR1, Caspase-3 Pro and Caspase-3 cleaved protein expression in Western Blotting, they were not the expression level of mRNA. Additional Editor Comments This study aims to explore the impact of mmu-let-7a-5p on Raw264.7 mononuclear macrophage apoptosis and its influence on intracellular Bacillus Calmette-Guérin (BCG) load in vitro. While the study holds potential interest for readers, there are several significant changes required. 1. The abstract section needs enhancement, specifically in introducing the theme, outlining the aims, and summarizing methods and results. Additionally, the conclusion section should outline future research directions, as the current presentation is unclear. 2. The introduction section should also be improved with background information relating to the study's aims. Furthermore, the material and methods section would benefit from the inclusion of a schematic diagram illustrating the study's protocol and a justification for using GAPDH as a reference gene for qPCR detection of mRNA. Additionally, the authors should provide the original figures of the western blot as supplementary files for the results section and submit the main raw data as supplementary files. 3. In the discussion section, the authors should refrain from comparing the present study's data to their previous findings and expand the analysis to include international data. Moreover, it should be important to insert the section limitations. 4. Lastly, the conclusion section should suggest future research directions. Reply 1. The abstract section needs enhancement, specifically in introducing the theme, outlining the aims, and summarizing methods and results. Additionally, the conclusion section should outline future research directions, as the current presentation is unclear. Thank you for your comments. Considering the Editor’s suggestion, we have re-written the abstract section part. Abstract: We have been trying to find a miRNA that can specifically regulate the function of mycobacterial host cells to achieve the purpose of eliminating Mycobacterium tuberculosis. The purpose of this study is to investigate the regulation of mmu-let-7a-5p on macrophages apoptosis and its effect on intracellular BCG clearance. After a series of in vitro experiments, we found that mmu-let-7a-5p could negatively regulate the apoptosis of macrophages. by targeting Caspase-3. The extrinsic apoptosis signal axis TNFR1/FADD/Caspase-8/Caspase-3 was inhibited after BCG infection. Up-regulated the expression level of mmu-let-7a-5p increase the cell proliferation viability and inhibit apoptosis rate of macrophages, but down-regulated its level could apparently reduce the bacterial load of intracellular Mycobacteria and accelerate the clearance of residual Mycobacteria effectively. Mmu-let-7a-5p has great potential to be utilized as an optimal candidate exosomal loaded miRNA for anti-tuberculosis immunotherapy in our subsequent research. 2. The introduction section should also be improved with background information relating to the study's aims. Furthermore, the material and methods section would benefit from the inclusion of a schematic diagram illustrating the study's protocol and a justification for using GAPDH as a reference gene for qPCR detection of mRNA. Additionally, the authors should provide the original figures of the western blot as supplementary files for the results section and submit the main raw data as supplementary files. Thank you for your comments and we have updated the background information of the introduction section. Furthermore, a schematic diagram was supplied to the material and methods. Glyceraldehyde-3-phosphate Dehydrogenase (GAPDH) is widely present in many organisms and is abundant in cells, accounting for 10% -20% of the total protein. The gene has a highly conserved sequence, which is highly expressed in almost all tissues. The protein expression in the same cell or tissue is generally constant, and it is not affected by the induction substances such as partial recognition check point and phorbol lipid. It has been commonly used as a reference gene in RT-qPCR assays for gene and protein expression analysis. Due to this, we chose GAPDH as a reference gene for qPCR detection of mRNA. Additionally, we have re-submitted the original figures of the western blot as supplementary files for the results section and submit the main raw data as supplementary files. 3. In the discussion section, the authors should refrain from comparing the present study's data to their previous findings and expand the analysis to include international data. Moreover, it should be important to insert the section limitations. Thank you for your comments. We have modified the discussion section according to your comments. 4. The conclusion section should suggest future research directions. Thank you for your comments. Based on our primary research, we will expect to construct a new multiple drug delivery system with exosomes for tuberculosis treatment in the future, which loaded miRNA and traditional anti-tuberculosis drug rifampin. This was mentioned in the conculsion. Conclusion: Mmu-let-7a-5p negatively regulates the apoptosis of Raw264.7 macrophages by targeting Caspase-3. Down-regulated the expression of mmu-let-7a-5p facilities the clearance of residual mycobacteria and attenuation the bacterial load of intracellular mycobacteria. Therefore, mmu-let-7a-5p has the great potential to be utilized as a candidate miRNA for the construction of novel exosomal drug delivery system loading with rifampin for anti-tuberculosis therapy in our subsequent research. Minor points: - English should be reviewed by a native speaker. - Please, check the use of reference manager software (Error! Reference source not found..-> line 121). - The short form should be introduced by the extension form the first time; subsequently, it should be used through the text. Please, check all abbreviations. Reply 1. English should be reviewed by a native speaker. Thank you for your advice. We not only invited an English-speaking friend from the United States to help polish our manuscript. Our manuscripts are also edited by one or more highly qualified native English editors of AJE for appropriate English language, grammar, punctuation, spelling and overall style. 2. Please, check the use of reference manager software (Error! Reference source not found.-> line 121). Thank you for your comments. We are very sorry for our negligence of the mistake in line 121 of the manuscript and we have revised the mistake. 3. The short form should be introduced by the extension form the first time; subsequently, it should be used through the text. Please, check all abbreviations. According to the Reviewer’s suggestion, we have double checked all the abbreviations in our manuscript. We tried our best to improve the quality of our manuscript and made some changes to the manuscript. These changes will not influence the content and framework of the paper. And we not only listed the changes in the email but also marked in red in the revised manuscript. We appreciate Editors / Reviewers’ warm work earnestly, and hope that the correction will meet with approval. Attachments Attachment Submitted filename: Response to Reviewers&#39.docx https://doi.org/10.1371/journal.pone.0308095.r002
22 May 2024 Decision Letter - Francesco Sessa, Editor PONE-D-23-31553R1Mmu-let-7a-5p inhibits macrophage apoptosis by targeting CASP3 to increase bacterial load and facilities mycobacterium survivalPLOS ONE Dear Dr. Ge, 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 Jul 06 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, Francesco Sessa, Ph.D., MS 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: The reviewer suggested minor revision and I agree with his comments. [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 #3: (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 #3: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? 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 #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 #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 #3: The re-submitted manuscript entitled “Mmu-let-7a-5p inhibits macrophage apoptosis by targeting CASP3 to increase bacterial load and facilities mycobacterium survival” by Zhan et al have made considerable modifications, however, there are still some minor concerns that need further addressed before accept. 1. Line 253 “3.7 Mmu-let-7a-5p enhances Raw264.7 cell proliferation and viability”, according to Fig 6 and its-related contents, the current subtitle of 3.7 needs to rethink and refine, especially only CCK-8 results whether could support the cell proliferation as authors claim? 2. Figure 7, if possible, suggests the authors add a group that is just Raw264.7 cells without anything as a negative control. The current apoptotic data from Nc-mimics and NC-siRNA almost reached 30%, whether the Nc-mimics and NC-siRNA could promote apoptosis? To exclude the possibility of higher apoptotic rates such as Nc-mimics and NC-siRNA groups, the cell picture from each should provide additional support. 3. In Fig 10, it’s better to add the dilution index on the top of panel A. 4. Considering the large amounts of data and results of this study, a graphic abstract is helpful if it is provided. ****** 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 #3: 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.0308095.r003
Revision 2
15 Jul 2024 Author Response Response to Reviewers Dear Editors and Reviewers, Thank you for your response and for the reviewer’s comments concerning our manuscript entitled “Mmu-let-7a-5p inhibits macrophage apoptosis by targeting CASP3 to increase bacterial load and promote facilities mycobacteria survival” [manuscript number: PONE-D-23-31553R1]. Please let me express my respect and sincere thanks for your conscientious work attitude. Those comments are all valuable and very helpful for revising and improving our manuscript, as well as the important guiding significance to our researchers. 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. Reply We have reviewed our references and double-checked the references list, no article was retracted. Review comments Reviewer #3: The re-submitted manuscript entitled “Mmu-let-7a-5p inhibits macrophage apoptosis by targeting CASP3 to increase bacterial load and facilities mycobacterium survival” by Zhan et al. have made considerable modifications; however, there are still some minor concerns that need further addressed before accepted. 1. Line 253 “3.7 Mmu-let-7a-5p enhances Raw264.7 cell proliferation and viability”, according to Fig. 6 and its related contents, the current subtitle of 3.7 needs to rethink and refine, especially only CCK-8 results whether could support the cell proliferation as authors claim? 2. Figure 7, if possible, suggests the authors add a group that is just Raw264.7 cells without anything as a negative control. The current apoptotic data from Nc-mimics and NC-siRNA almost reached 30%, whether the Nc-mimics and NC-siRNA could promote apoptosis? To exclude the possibility of higher apoptotic rates such as Nc-mimics and NC-siRNA groups, the cell picture from each should provide additional support. 3. In Fig. 10, it is better to add the dilution index on the top of panel A. 4. Considering the large amounts of data and results of this study, a graphic abstract is helpful if it is provided. Reply We have studied comments carefully and have made corrections which we hope meet with approval. Revised portions are marked in red in the paper. The main corrections in the paper and the responses to the reviewer’s comments are as follows: 1. Line 253 “3.7 Mmu-let-7a-5p enhances Raw264.7 cell proliferation and viability”, according to Fig. 6 and its related contents, the current subtitle of 3.7 needs to rethink and refine, especially only CCK-8 results whether could support the cell proliferation as authors claim? Thank you for your comments and we are extremely grateful to the Reviewer for pointing out this problem. Considering the reviewer’s comment, we have revised our subtitle of 3.7 and figure legends of Fig. 6. The CCK-8 results are appropriate to demonstrate the cell viability. 2. Figure 7, if possible, suggests the authors add a group that is just Raw264.7 cells without anything as a negative control. The current apoptotic data from Nc-mimics and NC-siRNA almost reached 30%, whether the Nc-mimics and NC-siRNA could promote apoptosis? To exclude the possibility of higher apoptotic rates such as Nc-mimics and NC-siRNA groups, the cell picture from each should provide additional support. First of all, we thank you for your rigorous and realistic academic style. In our data, there is a blank cell group which is Raw264.7 cells without anything as a negative control as you referred to above. Because the processing of apoptosis data needs to be compared with blank cells, the histogram shown later is actually the value compared with blank cells. Although the apoptosis rate of the Nc-mimics group and the NC-siRNA group was nearly 30%, they were significantly different from the statistical analysis results between the let-7a-5p mimics group and the Casp-3 siRNA group. 3. In Fig. 10, it is better to add the dilution index on the top of panel A. Thank you for your comments. Considering the reviewer’s suggestion, we have added the dilution index on the top of panel A. 4. Considering the large amounts of data and results of this study, a graphic abstract is helpful if it is provided. Thank you so much for your affirmation and suggestions for our work. In fact, we have already prepared a graphic abstract before our first submission. We will upload the graphic abstract together this time. Please review and make clear whether it is suitable for publication. Finally, we sincerely thank the reviewers for their valuable comments and suggestions. We try our best to revise the manuscript and improve the quality of our manuscripts by making some modifications. These changes will not affect the content and framework of the paper. We sincerely thank the editors / reviewers for their enthusiastic work and hope that the review work will be recognized. Yours sincerely Zhaohui Ge Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0308095.r004
17 Jul 2024 Decision Letter - Francesco Sessa, Editor Mmu-let-7a-5p inhibits macrophage apoptosis by targeting CASP3 to increase bacterial load and facilities mycobacterium survival PONE-D-23-31553R2 Dear Dr. Ge, 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, Francesco Sessa, Ph.D., MS Academic Editor PLOS ONE Additional Editor Comments (optional): Following the reviewers' comments, the authors have improved their manuscript. I endorse its publication. Reviewers' comments: https://doi.org/10.1371/journal.pone.0308095.r005
Formally Accepted
24 Jul 2024 Acceptance Letter - Francesco Sessa, Editor PONE-D-23-31553R2 PLOS ONE Dear Dr. Ge, 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 Lecturer Francesco Sessa Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0308095.r006

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