Interleukin-23 levels in umbilical cord blood are associated with neurodevelopmental trajectories in infancy

Machiko K. Asaka; Tomoko Nishimura; Hitoshi Kuwabara; Hiroaki Itoh; Nagahide Takahashi; Kenji J. Tsuchiya

doi:10.1371/journal.pone.0301982

Peer Review History

Original SubmissionDecember 8, 2023
29 Jan 2024 Decision Letter - Wen-Jun Tu, Editor PONE-D-23-33306IL-23 levels in umbilical cord blood associated with neurodevelopmental trajectories in infancyPLOS ONE Dear Dr. Kuwabara, 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 14 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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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 [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: Yes Reviewer #3: Yes Reviewer #4: Partly ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes Reviewer #4: No ****** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes Reviewer #4: 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 Reviewer #3: Yes Reviewer #4: 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: Comments. Title - **** All abbreviations should have been provided in full on first mention and this applies to the title, running (short) title, abstract, impact statement, main text and each table / figure independently as they will be read independently. Please use the abbrevations correctly and effectively so all the abbrevations should be checked. Abstract - Please give the number of the participants for each group. - The abstract must contain enough data (figures) to enable the reader (researcher) to judge its content and therefore relevance. It can not include undefined terms or contradictory statements. - The manuscript should have ended with a clear conclusion that sums up the value of the study within its limitations. Keywords - Please add the keywords according to alpbabetical order. * Introduction - An introduction should include information on why the subject under investigation is important, what is currently known on the topic and how the current investigation will improve our knowledge. Materials and Methods Results Tables and Figures Discussion - In the discussion the authors should summarise and qualify their findings, compare outcomes with other publications and draw conclusion. The implications of the study and the future direction of study should be discussed. Importantly the study’s limitations should be considered. - The manuscript should have ended with a clear conclusion that sums up the value of the study within its limitations. References - The number of the references should not be exceeded 30 (n<30) and also should be cited in the last five years. ** All abbreviations should have been provided in full on first mention and this applies to the title, running (short) title, abstract, impact statement, main text and each table / figure independently as they will be read independently. Please use the abbrevations correctly and effectively so all the abbrevations should be checked. * Passive sentences should be prefferred instead of active sentences starting with we. * Language and presentation require improvement. * Need several grammar/wording corrections (There are a lot of words with no space between them, there are missing spaces after commasin the manuscript and also, there are a lot of gramatical error. Language could be improved). Reviewer #2: Thank you very much to the authors for presenting this interesting study on the association between cord blood cytokine levels and neurodevelopmental outcome. The research objectives, methodology and results are clearly presented. My questions and comments to the authors are as follows: 1) Baseline demographics: Did the authors compare baseline demographics between the groups to see if they are comparable? Although all the babies are delivered full term, other factors including birth weight, antenatal problems, Apgar scores, chorioamnionitis, neonatal course, family history of developmental delay etc…. may all affect neurodevelopmental outcome. Can the authors elaborate more on this? 2) Neurodevelopmental outcomes: The authors have used the Mullen Scale of Early Learning to assess neurodevelopmental outcome in the first two years of life. For infants who are assessed to be in the “markedly delayed” or “delayed” group, would there be interventions or further investigations? And are there ongoing studies to look at neurodevelopmental outcome at a later age (eg 5 years old and later)? 3) Cytokine results: In previous studies on neurodevelopment of neonates (both preterm and term), cytokines including IL-6, IL-17, TNF- γ and TNF-a are associated with inflammation and injury to the developing brain, leading to poor neurodevelopmental outcome. In this study, only IL-23 is found to have significant association. The authors have postulated that the difference in TNF- γ may be due to difference in socioeconomic factors between subjects in various studies. Can the authors explain the difference between these results and previous findings? Reviewer #3: Dear Authors, i would like to congratulate You to describe such interesting subject I would be satisfied if You could desribe in the Introduction IL23 referring to development. In the line 113 You wrote when child was assessed: could You explain or correct why you wrote: the ages of 1, 24, 6, 10, 14, 18, or 24 months . 24 month is after 1st month, I do not understand? Could You write more about classification of The Mullen Scale of Early Learning (MSEL) due to next authors or readers have clear information when a child is :"low normal" delayed? Could You write more about exclussion criteria? Were there children with any inflamantory process because of sepsa or sth? Were there any USG of Transparietal ultrasound? Could You write about birth weight, week of gestation or Apgar score in inclussion criteria (VLBW, ELBW, Apgar score under 7 points)? Could You write in disccusion why You suspect ( in which process) children could have abnormal IL23 level? Your sincerely, Reviewer #4: This study, using data from the Mullen Scale of Early Learning on over 1000 Japanese infants followed from birth to 2 years, identified five distinct classes of early neurodevelopmental trajectories: high normal, normal, low normal, delayed, and markedly delayed. This classification was proved to be able to associate the cord blood cytokine concentrations with neurodevelopmental outcomes. However, some methodological issues required a revised analysis for robust and conclusive findings. 1) The paper did not mention how the neurodevelopmental trajectories were classified. as the classification of the 5 classes is the key in this paper, detailed explanation of the criteria and methods used to classify infants into the five neurodevelopmental trajectories is necessary. Also please ensure the robustness of the classification techniques. 2) There are about 30 cytokine concentrations considered in the association analyses. However, in the regression analyses p-values were not adjusted for multiple comparison in Table 3. Application of p-value adjustment methods in the regression analysis is necessary, specifically in Table 3. 3) is there a specific reason why the infants' age was not controlled as a confounding factor? 4) It is better to show some figures such as boxplots or histograms for the identified cytokine concentrations among the 5 classes. Generation of boxplots or histograms to visually compare cytokine concentration distributions across the five identified classes will give us more intuitive comparison of the distributions and outliers. ****** 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: Yes: Hasan Ali İnal Reviewer #2: Yes: Genevieve Po Gee Fung Reviewer #3: Yes: Roksana Malak Dep of Rheumatology Rehabilitation and Internal Medicine Poland Reviewer #4: 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.0301982.r001
Revision 1
8 Mar 2024 Author Response We would like to thank the Reviewers for their insightful comments and suggestions, which we believe have greatly helped us to improve our manuscript and provide a more balanced account of our research. We have carefully reviewed our manuscript in accordance with these comments, and made all necessary changes, which have shown using the track changes function in the revised manuscript to facilitate the review process. Please find below our point-by-point responses to all of the reviewer comments. Reviewer #1: Title All abbreviations should have been provided in full on first mention and this applies to the title, running (short) title, abstract, impact statement, main text and each table / figure independently as they will be read independently. Please use the abbreviations correctly and effectively so all the abbreviations should be checked. Response: We would like to thank you for this valuable suggestion, we have revised the manuscript accordingly. Abstract Please give the number of the participants for each group. The abstract must contain enough data (figures) to enable the reader (researcher) to judge its content and therefore relevance. It cannot include undefined terms or contradictory statements. Response: We would like to thank you for this valuable suggestion, we have revised the manuscript accordingly. (Lines 1-16) Keywords Please add the keywords according to alphabetical order. Response: Thank you for pointing this out, we have now rearranged the order of keywords. An introduction should include information on why the subject under investigation is important, what is currently known on the topic and how the current investigation will improve our knowledge. In the discussion the authors should summarize and qualify their findings, compare outcomes with other publications and draw conclusion. The implications of the study and the future direction of study should be discussed. Importantly the study’s limitations should be considered. The manuscript should have ended with a clear conclusion that sums up the value of the study within its limitations. Response: We would like to thank you for your comment, and apologize for the lack of clarity on this point we have also referred to the comments of Reviewer#2-4 together, and have made the corresponding modifications to our manuscript. References The number of the references should not be exceeded 30 (n<30) and also should be cited in the last five years. Response: We would like to thank you for your valuable suggestion; however, PLOS ONE does not limit the number of references, or provide guidelines on how recent the references cited should be. Passive sentences should be preferred instead of active sentences starting with we. Language and presentation require improvement. Need several grammar/wording corrections (There are a lot of words with no space between them, there are missing spaces after commas in the manuscript and also, there are a lot of grammatical error. Language could be improved). Response: We would like to thank you for your comment and apologize for this oversight, we have sent the manuscript for English proofreading by a native English speaker again. Reviewer#2 1) Baseline demographics: Did the authors compare baseline demographics between the groups to see if they are comparable? Although all the babies are delivered full term, other factors including birth weight, antenatal problems, Apgar scores, chorioamnionitis, neonatal course, family history of developmental delay etc…. may all affect neurodevelopmental outcome. Can the authors elaborate more on this? Response: We would like to thank you for your valuable suggestion. As you suggested, various confounding factors are likely to influence the results. We selected the baseline factors that showed an association with five neurodevelopmental trajectories, as identified in a previous study by Nishimura et al., 2016, for the adjustments. We have added the description of these associated factors in the method section of the revised manuscript. (Line: 145-147, Line: 186-189) 2) Neurodevelopmental outcomes: The authors have used the Mullen Scale of Early Learning to assess neurodevelopmental outcome in the first two years of life. For infants who are assessed to be in the “markedly delayed” or “delayed” group, would there be interventions or further investigations? And are there ongoing studies to look at neurodevelopmental outcome at a later age (eg 5 years old and later)? Response: We would like to thank you for your valuable suggestion. As we did not evaluate the intervention in formulated measure, we were not able to add this data to the current analyses. However, the cohort study (the Hamamatsu Birth Cohort Study for Mothers and Children) is ongoing, and we have published neurodevelopmental transition patterns from infancy to early childhood (Kato et al., Sci Rep. 2022; 12: 4822). 3) Cytokine results: In previous studies on neurodevelopment of neonates (both preterm and term), cytokines including IL-6, IL-17, INF- γ and TNF-a are associated with inflammation and injury to the developing brain, leading to poor neurodevelopmental outcome. In this study, only IL-23 is found to have significant association. The authors have postulated that the difference in INF- γ may be due to difference in socioeconomic factors between subjects in various studies. Can the authors explain the difference between these results and previous findings? Response: We would like to thank you for your valuable suggestion. The inconsistency regarding TNF-� might be explained by the difference in outcome measurements between our and previous studies (e.g. Barbosa et al., Brain Behav Immun Health. 2020; 8: 100141). However, as our current experiments did not measure IL-6, IL-17, or INF-γ, it is difficult to interpret the inconsistency between previous studies and our results. We have added these explanations in the revised manuscript. (Line: 316-318, Line: 326-336, Line: 338) Reviewer #3 I would be satisfied if You could describe in the Introduction IL23 referring to development. Response: We would like to thank you for your valuable suggestion. As our study was exploratory in nature, and as far as we know, there were no high-quality report that led us to prioritize IL-23 over other cytokines, we thought that it is rather unfair to insist on including IL-23 in the introduction. In the line 113 You wrote when child was assessed: could You explain or correct why you wrote: the ages of 1, 24, 6, 10, 14, 18, or 24 months. 24 month is after 1st month, I do not understand? Response: We would like to thank you for your comment and apologize for the error, which we have now corrected. (Line: 122) Could You write more about classification of The Mullen Scale of Early Learning (MSEL) due to next authors or readers have clear information when a child is: "low normal" delayed? Response: We would like to thank you for your valuable suggestion. According to your suggestion, we have added more information regarding the low normal and delayed classes from our previous study by Nishimura et al., 2016. (Line: 142-144) Were there children with any inflammatory process because of sepsa or sth? Were there any USG of trans parietal ultrasound? Response: We would like to thank you for your comment. However, we did not assess sepsa or sth, nor did we measure USG of trans parietal ultrasound. Could You write more about exclusion criteria? Could You write about birth weight, week of gestation or Apgar score in inclusion criteria (VLBW, ELBW, Apgar score under 7 points)? Response: We would like to thank you for your valuable suggestion. According to your suggestion, we have added more information about the inclusion and exclusion criteria, which had been adopted from our previous study by Nishimura et al., 2016. (Line: 107-108, Line: 123-125) Could You write in discussion why You suspect (in which process) children could have abnormal IL23 level? Response: We would like to thank you for your valuable suggestion. Although our research design was insufficient to allow us to ascertain the true process of IL-23 affecting the neuro-developments, the review by Kollman et al. (Immunity. 2012; 37(5): 771-83) explained that after Toll-like receptors (TLRs) stimulation of whole blood, production of Th17 cell-promoting cytokines IL-6 and IL-23 dominates in term infant, while further research has shown that low production of IL-23 after phytohemagglutinin activation in children with ASD. As such, we suspected that the abnormality of responses to the stimulation of TLRs in the perinatal period might be involved in the mechanism of abnormal cord blood IL-23 levels. These discussions were already included in the discussion section. (Line: 286-307) Reviewer #4 1) The paper did not mention how the neurodevelopmental trajectories were classified. as the classification of the 5 classes is the key in this paper, detailed explanation of the criteria and methods used to classify infants into the five neurodevelopmental trajectories is necessary. Also please ensure the robustness of the classification techniques. Response: We would like to thank you for your valuable suggestion. According to your suggestion, we have added more information about the statistical methods of identification on five neurodevelopmental trajectories from our previous study by Nishimura et al., 2016. (Line: 117-136) 2) There are about 30 cytokine concentrations considered in the association analyses. However, in the regression analyses p-values were not adjusted for multiple comparison in Table 3. Application of p-value adjustment methods in the regression analysis is necessary, specifically in Table 3. Response: We would like to thank you for your valuable suggestion. Indeed, we performed two step analyses; the first step involved exploratory analyses, while the second (final) step involved multinominal logistic regression, adjusted for multiple comparison. We have added the clear description of adjustments for multiple comparison in revised manuscripts. (Line:189-190) 3) is there a specific reason why the infants' age was not controlled as a confounding factor? Response: We would like to thank you for your valuable suggestion. At the point of measuring of the Mullen Scale of Early Learning (MSEL) and the sampling of cord blood, all the participants were the same age (month), thus we did not control for age as a confounder. 4) It is better to show some figures such as boxplots or histograms for the identified cytokine concentrations among the 5 classes. Generation of boxplots or histograms to visually compare cytokine concentration distributions across the five identified classes will give us more intuitive comparison of the distributions and outliers. Response: We would like to thank you for your valuable suggestion. According to your suggestion, we have generated Fig 1 and S1 Fig. (Line: 232, Line: 248-250, Line: 575-582) https://doi.org/10.1371/journal.pone.0301982.r002
26 Mar 2024 Decision Letter - Wen-Jun Tu, Editor Interleukin-23 levels in umbilical cord blood associated with neurodevelopmental trajectories in infancy PONE-D-23-33306R1 Dear Dr. Kuwabara, 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, Wen-Jun Tu 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 Reviewer #4: 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 #4: (No Response) ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #4: (No Response) ****** 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 #4: (No Response) ****** 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 #4: (No Response) ****** 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: Dear Author(s), Thank you for your revisions. The manuscript was revised accordingly. It can be published. Reviewer #4: (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: Yes: Hasan Ali İnal Reviewer #4: No ******** https://doi.org/10.1371/journal.pone.0301982.r003
Formally Accepted
1 Apr 2024 Acceptance Letter - Wen-Jun Tu, Editor PONE-D-23-33306R1 PLOS ONE Dear Dr. Kuwabara, 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. Wen-Jun Tu Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0301982.r004

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