Mendelian randomization analysis of plasma levels of CD209 and MICB proteins and the risk of varicose veins of lower extremities

Alexandra S. Shadrina; Elizaveta E. Elgaeva; Ian B. Stanaway; Gail P. Jarvik; Bahram Namjou; Wei-Qi Wei; Joe Glessner; Hakon Hakonarson; Pradeep Suri; Yakov A. Tsepilov

doi:10.1371/journal.pone.0268725

Peer Review History

Original SubmissionFebruary 4, 2022
16 Mar 2022 Decision Letter - Renato Polimanti, Editor PONE-D-22-02162Mendelian randomization analysis of plasma levels of CD209 and MICB proteins and the risk of varicose veins of lower extremitiesPLOS ONE Dear Dr. Shadrina, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by Apr 30 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. 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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. Additional Editor Comments: Reviewer #1 highlighted several major issues in the analytic design of the study. I strongly recommend to the authors to fully address Reviewer's 1 requests. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes ****** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes ****** 5. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: Shadrina et al. performed a Mendelian Randomization (MR) study to replicate their previous findings on the effects of the plasma levels of MICB and CD209 proteins on varicose veins of lower extremities (VVs). I consider this study to be sound, and the topic is highly relevant to elucidate the etiology of VVs. I have the following recommendations to improve the manuscript: Major issues This study was limited by the small number of instrumental variables used in the analysis (two SNPs). Thus, I suggest performing a sensitivity analysis with suggestive variants. Authors can use the MR–Robust Adjusted Profile Score (MR-RAPS) approach to account for the possible biases introduced by using weaker instrumental variables (IVs). Please, evaluate the presence of horizontal pleiotropy in the IVs as this could bias the MR estimates. Please consider evaluating other MR methods (MR-Egger, weighted median, simple mode, and weighted mode) to compare the concordance of the IVW estimates with those obtained with the other MR methods. Minor issues Abstract Please, indicate the meaning of MICB and CD209 Please include the confidence intervals of the effect estimates Please indicate that the study was performed only in European descents Introduction I consider the Introduction to be very long. It includes details that are not necessary to understand the present study, for example, the release of the data and the ICD-code used to define VV in their previous MR. Please, consider a more brief description of this previous study. Also, the extensive description and benefits of MR could be more suitable for the Discussion section. Methods Line 156. Please, consider changing the term White by European descents, as the latter term better describes the sample's ancestral origins. Line 162. I suggest using PCs instead of principal components to be consistent with the abbreviation of PC previously used in this manuscript. Did the authors evaluate whether the SNPs associated with the exposure were associated with the outcome? Results Please, include the confidence intervals for all the reported estimates. Please report the F-statistic and include the proportion of variance explained (R2) of the IVs. Discussion It would be interesting if the authors discuss if there is any potential confounders in the causal association between CD209 and VVs. Please include as a potential limitation that analyzed data were obtained only from European descents and results could not be generalizable to other populations. ****** 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 [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.0268725.r001
Revision 1
30 Apr 2022 Author Response RESPONSE TO COMMENTS Additional Editor Comments: Reviewer #1 highlighted several major issues in the analytic design of the study. I strongly recommend to the authors to fully address Reviewer's 1 requests. Response: Thank you. We performed additional analyses and corrected our manuscript to address Reviewer's 1 requests and improve our study. Reviewers' comments: Reviewer #1: Shadrina et al. performed a Mendelian Randomization (MR) study to replicate their previous findings on the effects of the plasma levels of MICB and CD209 proteins on varicose veins of lower extremities (VVs). I consider this study to be sound, and the topic is highly relevant to elucidate the etiology of VVs. Response: Thank you. We were very pleased to learn your positive opinion about our study. Reviewer #1: I have the following recommendations to improve the manuscript: Major issues This study was limited by the small number of instrumental variables used in the analysis (two SNPs). Thus, I suggest performing a sensitivity analysis with suggestive variants. Authors can use the MR–Robust Adjusted Profile Score (MR-RAPS) approach to account for the possible biases introduced by using weaker instrumental variables (IVs). Response: Thank you for this important suggestion. We performed a sensitivity analysis with an extended set of IVs associated with CD209 level at a suggestive level of statistical significance (5 IVs). The increased number of IVs allowed us to conduct other MR methods as well as to perform the heterogeneity tests, horizontal pleiotropy tests, and the test for the correct direction of effect. We also used the MR-RAPS approach to account for the possible biases introduced by selecting weaker IVs. We updated the manuscript text with a description of the methods used (added the subsection “Sensitivity analyses” to the Materials and Methods section), added three Supplementary tables (S3 Table, S6 Table, and S7 Table) with the results of sensitivity analyses and GWAS summary statistics for IVs and summarized new results in the Results and Discussion sections. Reviewer #1: Please, evaluate the presence of horizontal pleiotropy in the IVs as this could bias the MR estimates. Response: Thank you. We performed two horizontal pleiotropy tests: the test based on the MR-Egger regression intercept and the MR-PRESSO global test. Both tests did not provide evidence for the presence of directional horizontal pleiotropy. The results are provided in S6 Table. Reviewer #1: Please consider evaluating other MR methods (MR-Egger, weighted median, simple mode, and weighted mode) to compare the concordance of the IVW estimates with those obtained with the other MR methods. Response: Thank you. We conducted sensitivity analyses using MR-Egger, weighted median, simple mode, weighted mode as well as IVW methods. The results were concordant with each other and with those obtained in the main IVW MR analysis using two strong IVs. All results of sensitivity analyses are now presented in S6 Table and described in the text. Reviewer #1: Minor issues Abstract Please, indicate the meaning of MICB and CD209 Response: Thank you. We added the meaning of MICB and CD209 in the Abstract. Reviewer #1: Abstract Please include the confidence intervals of the effect estimates Response: Thank you. We included OR and 95% CI in the Abstract. Reviewer #1: Abstract Please indicate that the study was performed only in European descents Response: Thank you. We added this information in the Abstract. Reviewer #1: Introduction I consider the Introduction to be very long. It includes details that are not necessary to understand the present study, for example, the release of the data and the ICD-code used to define VV in their previous MR. Please, consider a more brief description of this previous study. Also, the extensive description and benefits of MR could be more suitable for the Discussion section. Response: Thank you. We shortened the Introduction according to these recommendations. Reviewer #1: Methods Line 156. Please, consider changing the term White by European descents, as the latter term better describes the sample's ancestral origins. Response: Thank you. We corrected the text of the Materials and Methods section according to this recommendation. Reviewer #1: Methods Line 162. I suggest using PCs instead of principal components to be consistent with the abbreviation of PC previously used in this manuscript. Response: Thank you for noticing this. We corrected the text, and now “PCs” is indicated instead of “principal components”. Reviewer #1: Methods Did the authors evaluate whether the SNPs associated with the exposure were associated with the outcome? Response: Thank you. Summary statistics for the association between IVs used in the main analysis and VVs (for Gene ATLAS, eMERGE, and FinnGen cohorts) is now provided in S5 Table (columns named “Association between IV and VVs”). Summary statistics for the association between IVs used in the sensitivity analysis and VVs is now provided in S3 Table (columns named “Varicose veins of lower extremities (OUTCOME)”). All IVs were either not associated with VVs or the associations were not genome-wide significant. Reviewer #1: Results Please, include the confidence intervals for all the reported estimates. Response: Thank you. We included ORs and 95% CIs in the text of the Results section, in Table 1 and Table 2 (for the meta-analysis of MR results for FinnGen, eMERGE, and Gene ATLAS cohorts), and added all ORs and 95% CIs in Supplementary tables with MR results: S4 Table, S5 Table, S6 Table, and S7 Table. Reviewer #1: Results Please report the F-statistic and include the proportion of variance explained (R2) of the IVs. Response: Thank you. We added this information in S1 Table for the main analysis and S3 Table for the sensitivity analysis. Reviewer #1: Discussion It would be interesting if the authors discuss if there is any potential confounders in the causal association between CD209 and VVs. Response: Thank you. We included this information to the end of the Discussion section. Hypothetically, potential confounders could be related to the inflammatory pathways which may promote the release of soluble form of CD209 and at the same time play a role in the promotion of VVs formation. However, as far as we are aware, to date, evidence from the literature does not allow to specify such common pathways Reviewer #1: Discussion Please include as a potential limitation that analyzed data were obtained only from European descents and results could not be generalizable to other populations. Response: Thank you. We added this limitation in the Discussion section. Attachments Attachment Submitted filename: Response to Reviewers_PONE-D-22-02162.docx https://doi.org/10.1371/journal.pone.0268725.r002
6 May 2022 Decision Letter - Renato Polimanti, Editor Mendelian randomization analysis of plasma levels of CD209 and MICB proteins and the risk of varicose veins of lower extremities PONE-D-22-02162R1 Dear Dr. Shadrina, 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, Renato Polimanti, 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: I have no further comments to the authors. The authors have adequately addressed my comments in the revised version. ****** 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.0268725.r003
Formally Accepted
13 May 2022 Acceptance Letter - Renato Polimanti, Editor PONE-D-22-02162R1 Mendelian randomization analysis of plasma levels of CD209 and MICB proteins and the risk of varicose veins of lower extremities Dear Dr. Shadrina: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Renato Polimanti Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0268725.r004

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