Plasma midkine concentrations in healthy children, children with increased and decreased adiposity, and children with short stature

Youn Hee Jee; Kun Song Lee; Shanna Yue; Ellen W. Leschek; Matthew G. Boden; Aysha Jadra; Anne Klibanski; Priya Vaidyanathan; Madhusmita Misra; Young Pyo Chang; Jack A. Yanovski; Jeffrey Baron

doi:10.1371/journal.pone.0224103

Peer Review History

Original SubmissionJune 2, 2019
15 Jul 2019 Decision Letter - Kathleen E. Bethin, Editor PONE-D-19-15659 Plasma midkine concentrations in healthy children, children with increased and decreased adiposity, and children with short stature PLOS ONE Dear Dr Baron, 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. ============================== You need to address comments # 1,2 and 4 by the reviewer and comments #1-4 from me. Comment #3 from the reviewer and #5-6 from me are suggestions. ============================== We would appreciate receiving your revised manuscript by Aug 29 2019 11:59PM. When you are 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. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. To enhance the reproducibility of your results, we recommend that if applicable you deposit your laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols 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). This letter should be uploaded as separate file and labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. This file should be uploaded as separate file and labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. This file should be uploaded as separate file and labeled 'Manuscript'. Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out. We look forward to receiving your revised manuscript. Kind regards, Kathleen E. Bethin, MD, PhD Academic Editor PLOS ONE Journal Requirements: 1. When submitting your revision, we need you to address these additional requirements. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at http://www.journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and http://www.journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf 2. You indicated that you had ethical approval for your study. In your Methods section, please ensure you have also stated whether you obtained consent from parents or guardians of the minors included in the study. 3. We note that you have included the phrase “data not shown” in your manuscript. Unfortunately, this does not meet our data sharing requirements. PLOS does not permit references to inaccessible data. We require that authors provide all relevant data within the paper, Supporting Information files, or in an acceptable, public repository. Please add a citation to support this phrase or upload the data that corresponds with these findings to a stable repository (such as Figshare or Dryad) and provide and URLs, DOIs, or accession numbers that may be used to access these data. Or, if the data are not a core part of the research being presented in your study, we ask that you remove the phrase that refers to these data. Additional Editor Comments: 1. The authors state,“All relevant data are within the manuscript and its Supporting Information files. “ However, the raw data are not included in the manuscript and there are no supporting files. 2. Figure 1 legend does not belong on the figure itself. 3. Figure 2 legend- must be reworked. A figure legend is not a description of the results. Please refer to reviewer comments. 4. For Figure 2 B please explain why/how the age brackets shown were chosen. 5. I agree with reviewer #1 assessment- The results section does not flow with the numbered sections. I would remove the numbers and the titles and add some transitions from one section to the other. 6.On page 11, lines 240-242 the MDK in children with ISS is compared to “healthy children.” This is implying that children with ISS are not healthy. I would suggest rewording the sentence. . [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: The submitted manuscript by Lee et al describes plasma midkine levels in different cohorts of children with normal weight, obesity and underweight, as well as short stature. This study was done because of prior observation that midkine levels were higher in obesity and short stature. The authors set out to clarify the midkine concentrations in these populations. The cohorts are well characterized and described. The data is clear and the manuscript is well-written. There a few items which could be addressed by the authors. 1) Page 7, line 133. The authors describe the Tanner staging for the pediatric cohort. However, they used different Tanner staging procedures based on sex -- females were staged for gonadarche, while males were staged by pubarche. The authors then evaluated midkine levels based on Tanner stage. However, the authors are combining 2 different pubertal assessments into one category and gonadarche is biochemically different than pubarche. The authors should discuss the rationale for this categorization. Alternatively, if they have data that would be consistent across both sexes, then midkine levels could be presented based on that Tanner staging. 2) Page 8, line 169. What does the term "parallelism" mean ? Do they mean to say correlation or consistency? 3) Results (starting on page 9) Do the individual results categories need to be numbered?? 4) Figure legend for Figure 2 should be reworked. Would recommend as follows: This figure legend could be reworked Fig 2A) Scatter-plot of log-transformed plasma MDK values (ng/ml) vs. age in healthy children (N= 222). B) MDK values (mean +/- sem) stratified by age (?? any stats for this, ie difference between <1 year and other age cohorts??) C &D) Scatterplots of plasma MDK concentrations vs. height (C) and BMI (D) (Show the r-value on the curves) E) MDK values (mean +/- sem) stratified by Tanner Stage. ****** 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 to be viewed.] 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 us at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0224103.r001
Revision 1
3 Sep 2019 Author Response August 5, 2016 Kathleen E. Bethin, MD, PhD Academic Editor PLOS ONE Re: PONE-D-19-15659 Title: Plasma midkine concentrations in healthy children, children with increased and decreased adiposity, and children with short stature Dear Dr. Bethin, We would like to thank the reviewer and editor for their careful evaluation of our manuscript. As requested, we are providing the point-by-point response to the reviewer and editor and the revised version of our manuscript with track changes. The reviewer and editor made a number of important comments, which we carefully addressed, as detailed below. With these revisions, we believe that the paper is strengthened and therefore hope that the reviewer and editor will find that it is now suitable for publication in PLOS ONE. Sincerely, Jeffrey Baron, MD Chief, Section on Growth and Development Reviewer comments: Journal requirements #1: Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. Response: We believe that our manuscript meets PLOS One’s style including the file names. Please let us know if any adjustments are needed. Journal requirements #2: In your Methods section, please ensure you have also stated whether you obtained consent from parents or guardians of the minors included in the study. Response: The language of consent process was revised as below. Page 4: Written informed consent was obtained from all subjects and from parents or legal guardians of minors. Assent was also obtained from minors as appropriate. Journal requirements #3: “Data not shown” in your manuscript does not meet our data sharing requirements. PLOS does not permit references to inaccessible data. Response: The “data not shown” is not critical to the manuscript and therefore was simply deleted. Page 8: Plasma was obtained from subjects regardless of fasting or time of day because these factors did not affect plasma MDK concentrations based on our evaluation (data not shown). Measurement of MDK in serum did not show good parallelism (data not shown) and the concentrations of MDK were far lower than in plasma, therefore only plasma MDK concentrations are reported. Editor comments #1: The authors state, “All relevant data are within the manuscript and its Supporting Information files.” However, the raw data are not included in the manuscript and there are no supporting files. Response: The raw data in excel is now submitted as a supporting information file. Editor comments #2: Fig 1 legend does not belong on the figure itself. Response: The fig 1 legend is revised as below. Page 4: Fig 1. Subject groups. Healthy children without obesity, children and young adults with abnormal adiposity (obesity and anorexia nervosa), children with short stature, and healthy adults were studied. Numbers of subjects in each group are shown in parentheses. SS, short stature; AN, anorexia nervosa; ISS, idiopathic short stature; SGA, small for gestational age; GH, growth hormone; NIH, National Institutes of Health; MGH, Massachusetts General Hospital; U.S., United States (studied at NIH) Editor comment #3: Fig 2 legend- must be reworked. A figure legend is not a description of the results. Please refer to reviewer comments. Response: Thank you for the editor’s and reviewer’s comment. Fig 2 legend is revised as below. Page 10: Fig 2. Plasma midkine (MDK) concentrations in healthy children. Plasma was obtained from healthy, non-obese children younger than 18 years (n=222) from the United States or South Korea. Plasma MDK concentration was measured by sandwich enzyme-linked immunosorbent assay (ELISA). A) Scatterplot of plasma midkine (MDK) concentrations (ng/mL) vs age. B) MDK concentrations (mean ± SEM) stratified by age. C) Scatterplot of plasma MDK concentrations vs. height standard deviation score (SDS) and D) Scatterplot of plasma MDK concentrations vs. body mass index (BMI) SDS. E) Plasma MDK concentrations (mean ± sem) stratified by Tanner stage in those subjects who had pubertal staging performed at the time of blood collection (n = 132). Editor comment #4: For Fig 2 B please explain why/how the age brackets shown were chosen. Response: The following sentence was added to the methods section: Page9: In healthy children, the relationship between plasma MDK concentration and age was analyzed both by treating age as a continuous variable and also by categorizing for age: infancy (< 1 year), prepubertal age range (1-7 years), peripubertal to pubertal age range (8-14 years), and late pubertal to postpubertal age range (15-18 years). Editor comment #5: I agree with reviewer #1 assessment- The results section does not flow with the numbered sections. I would remove the numbers and the titles and add some transitions from one section to the other. Response: As suggested, the numbers and the titles in result section were removed and the transitions were revised. Editor comment #6: On page 11, lines 240-242 the MDK in children with ISS is compared to “healthy children.” This is implying that children with ISS are not healthy. I would suggest rewording the sentence. Response: Thank you for the editor’s comment. We revised the sentence as below. Page 11: To explore the relationship between MDK and linear growth, plasma midkine concentrations were measured in 68 children with idiopathic short stature (ISS, median height SDS -2.2) and compared to healthy children with stature within the normal range. Lastly, we would like to inform the editor and reviewer that we revised Fig 1 for Y-axis to contain raw data in a log scale (not log-transformed data) to help readers. Reviewer #1: The submitted manuscript by Lee et al describes plasma midkine levels in different cohorts of children with normal weight, obesity and underweight, as well as short stature. This study was done because of prior observation that midkine levels were higher in obesity and short stature. The authors set out to clarify the midkine concentrations in these populations. The cohorts are well characterized and described. The data is clear and the manuscript is well-written. There a few items which could be addressed by the authors. 1) Page 7, line 133. The authors describe the Tanner staging for the pediatric cohort. However, they used different Tanner staging procedures based on sex -- females were staged for gonadarche, while males were staged by pubarche. The authors then evaluated midkine levels based on Tanner stage. However, the authors are combining 2 different pubertal assessments into one category and gonadarche is biochemically different than pubarche. The authors should discuss the rationale for this categorization. Alternatively, if they have data that would be consistent across both sexes, then midkine levels could be presented based on that Tanner staging. Response: We agree with the reviewer’s point. The data were obtained from subject’s medical record at the time of blood collection and breast development for girls and pubic hair for boys were primarily available. To address the reviewer’s point, we revised our manuscript as follows. Page 8: For 131 healthy children without obesity, Tanner staging was assessed by physical exam conducted by an experienced pediatric endocrine research nurse or pediatric endocrinologist. The Tanner stages were later obtained from the subjects’ records. Breast development for girls and pubic hair for boys were used for analysis because these were the observations most widely recorded. 2) Page 8, line 169. What does the term "parallelism" mean ? Do they mean to say correlation or consistency? Response: We used the term “parallelism” to refer to a desirable property of an immunoassay. To assess parallelism, a subject’s plasma with high endogenous MDK is serially diluted and then assayed. If the measured concentrations of MDK decrease proportionally to the dilution factor, then the assay is said to be parallel, indicating that the binding characteristic of the antibodies to endogenous MDK is the same as antibody binding to the calibrator MDK and that the blood plasma does not create a matrix effect which alters the binding. However, in the revised manuscript, we decided not to show the parallelism data because it is not critical to the study, and we therefore deleted the sentence referring to parallelism. 3) Results (starting on page 9) Do the individual results categories need to be numbered?? Response: We agree that the numbering is not necessary, and therefore we removed the numbers and subtitles and instead used the first sentence of each paragraph to aid the reader with the transitions. 4) Figure legend for Fig 2 should be reworked. Would recommend as follows: This figure legend could be reworked Response: We appreciate the reviewer’s comment and revised the legend as suggested. Fig 2. Plasma midkine concentrations in healthy children. Plasma was obtained from healthy, non-obese children younger than 18 years (n=222) from the United States or South Korea. Plasma midkine concentration was measured by sandwich enzyme-linked immunosorbent assay (ELISA) A) Scatterplot of plasma midkine (MDK) concentrations (ng/mL) vs age. B) MDK concentrations (mean ± SEM) stratified by age. C) Scatterplot of plasma MDK concentrations vs. height standard deviation score (SDS) and D) Scatterplot of plasma MDK concentrations vs. body mass index (BMI) SDS. E) Plasma MDK concentrations (mean ± sem) stratified by Tanner stage in those subjects who had pubertal staging performed at the time of blood collection (n = 132). Attachments Attachment Submitted filename: Response to reviewers .docx https://doi.org/10.1371/journal.pone.0224103.r002
9 Sep 2019 Decision Letter - Kathleen E. Bethin, Editor PONE-D-19-15659R1 Plasma midkine concentrations in healthy children, children with increased and decreased adiposity, and children with short stature PLOS ONE Dear Dr Baron, 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. We would appreciate receiving your revised manuscript by Oct 24 2019 11:59PM. When you are 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. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. To enhance the reproducibility of your results, we recommend that if applicable you deposit your laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols 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). This letter should be uploaded as separate file and labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. This file should be uploaded as separate file and labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. This file should be uploaded as separate file and labeled 'Manuscript'. Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out. We look forward to receiving your revised manuscript. Kind regards, Kathleen E. Bethin, MD, PhD Academic Editor PLOS ONE Additional Editor Comments (if provided): All of the reviewers/ editors comments were addressed. However, 2 minor issues were discovered on this review: 1. The number of healthy children with Tanner staging is listed as 131 on lines 155-159 of the text and listed as 132 in figure 2. 2. The term "non-Hispanic Black" was used in Table 1 and "Africa American" was used in table 2. Please be consistent. Also, legend for Table 2 defines AA as African American but AA is not used in the Table. [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 to be viewed.] 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 us at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0224103.r003
Revision 2
25 Sep 2019 Author Response Reviewer’s comments: 1. The number of healthy children with Tanner staging is listed as 131 on lines 155-159 of the text and listed as 132 in figure 2. Response: We appreciate the reviewer’s careful review. We confirmed that 132 in Figure 2 is the correct number. Therefore, the text was revised to 132. 2. The term "non-Hispanic Black" was used in Table 1 and "Africa American" was used in table 2. Please be consistent. Also, legend for Table 2 defines AA as African American but AA is not used in the Table. Response: We revised our Tables to be consistent with non-Hispanic black and removed African American and AA. https://doi.org/10.1371/journal.pone.0224103.r004
7 Oct 2019 Decision Letter - Kathleen E. Bethin, Editor Plasma midkine concentrations in healthy children, children with increased and decreased adiposity, and children with short stature PONE-D-19-15659R2 Dear Dr. Baron, We are pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it complies with all outstanding technical requirements. Within one week, you will receive an e-mail containing information on the amendments required prior to publication. When all required modifications have been addressed, you will receive a formal acceptance letter and your manuscript will proceed to our production department and be scheduled for publication. Shortly after the formal acceptance letter is sent, an invoice for payment will follow. To ensure an efficient production and billing process, please log into Editorial Manager at https://www.editorialmanager.com/pone/, click the "Update My Information" link at the top of the page, and update your user information. 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 enable them to help maximize its impact. If they will be preparing press materials for this manuscript, you must inform our press team as soon as possible and 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. With kind regards, Kathleen E. Bethin, MD, PhD Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0224103.r005
Formally Accepted
18 Oct 2019 Acceptance Letter - Kathleen E. Bethin, Editor PONE-D-19-15659R2 Plasma midkine concentrations in healthy children, children with increased and decreased adiposity, and children with short stature Dear Dr. Baron: I am 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 notify them about your upcoming paper at this point, to enable them to help maximize its impact. If they will be preparing press materials for this manuscript, 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. For any other questions or concerns, please email plosone@plos.org. Thank you for submitting your work to PLOS ONE. With kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Kathleen E. Bethin Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0224103.r006

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