Protective effects of Colla Corii Asini Collagen Peptides on D-galactose injection combined with UVB irradiation-induced aging in mice

Qingdi Luo; Song Zhang; Zhuo Sun; Zhihao Wang; Qiulin Yue; Xin Sun; Li Tian; Baojun Li; Kunlun Li; Chen Zhao; Lin Zhao; Le Su

doi:10.1371/journal.pone.0317302

Peer Review History

Original SubmissionSeptember 27, 2024
9 Dec 2024 Decision Letter - Arumugam Muthuvel, Editor PONE-D-24-41650Protective effects of Colla Corii Asini Collagen Peptides on D-galactose injection combined with UVB irradiation-induced aging in micePLOS ONE Dear Dr. Zhao, 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 Jan 23 2025 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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(Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: In this study, the authors have investigated the protective role of Colla Corii Asini collagen peptides on D-Galactose injection combined with UVB irradiation-induced aging in mice. The study is interesting and may appeal to wider audience. Comments 1. Authors must explain, how they determined low dose(67mg/kg) and high dose (200mg/kg) of CCACPs 2. Figure 1(C), what is the unit of measurement for Autonomic activity? or define the scale 3. In all the bar graphs, authors can move the significant stars from control group to aging model group to better represents the significant changes in model group compared to control group. 4. In 3.10. Indicate the appropriate figure number in the text, instead for just mentioning Figure 5. Mention figure number for Il-6 data 5. Typo Figure 3, (C) Is it MMP-1 or MMP-3? Figure 2 (C) axis label (U/mg prot) instead of (U/mgprot) Materials and methods, 2.3 Animal testing, line 9. Reframe the sentence “There is no expected unexpected deaths” Reviewer #2: Explain in detail the uniqueness of employing a combined D-galactose and UVB-induced aging paradigm, setting it apart from other research that used just one of these stressors. Give further background information on how the results might apply to aging-related diseases in people. Explain the rationale behind the 8-week exposure period. Would a longer time frame yield more thorough insights? Give further information about the methods used to regulate the UVB exposure so that each individual experienced the same level of intensity. To contextualize the peculiarity of CCACP, update Table 2 to include a comparison of its amino acid content with that of other collagen sources. And additional if some references are added from latest research if available it will be nice ****** 6. 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Revision 1
17 Dec 2024 Author Response Dear Editors and Reviewers, Thank you for your valuable comments and suggestions on our manuscript titled “Protective effects of Colla Corii Asini Collagen Peptides on D-galactose injection combined with UVB irradiation-induced aging in mice” (ID: PONE-D-24-41650). The comments provided have been very valuable for revising and improving our manuscript, and they have been of significant guidance to our research. We have carefully reviewed the comments and made the necessary corrections, hoping for approval. The revised sections are highlighted in the updated manuscript. The main corrections and our responses to the editor and reviewers' comments are as follows: Reviewer #1: In this study, the authors have investigated the protective role of Colla Corii Asini collagen peptides on D-Galactose injection combined with UVB irradiation-induced aging in mice. The study is interesting and may appeal to wider audience. 1. Authors must explain, how they determined low dose (67mg/kg) and high dose (200mg/kg) of CCACPs. We greatly appreciated your professional review of our manuscript. The low dose (67 mg/kg) and high dose (200 mg/kg) of CCACPs were determined based on a comprehensive literature review and our preliminary human studies. To begin with, previous studies have reported the use of collagen peptides in animal models at doses ranging from 50 to 800 mg/kg [1–3]. Moreover, we conducted preliminary human studies to identify effective and well-tolerated doses of CCACPs (data not shown). Based on the effective doses observed in these human studies, we applied standard body surface area–based conversion methods to translate the human doses into equivalent animal doses, thereby establishing the appropriate dosing for the mouse experiments. As a result, we selected 67 mg/kg as the low dose and 200 mg/kg as the high dose, ensuring that these doses were both within the range reported in the literature and represented the most effective doses of CCACPs. 2. Figure 1(C), what is the unit of measurement for Autonomic activity? Or define the scale. The "Autonomic activities" in Figure 1(C) are represented on a normalized scale without explicit units, as defined by the proprietary algorithm of the Animal Behavior Video Analysis System, Ver 20.0703. This value is calculated based on a combination of metrics, including total travel distance (mm), time spent active (s), rest time (s), and average speed (mm/s). The scale reflects the relative activity level of the animal, with higher values indicating more frequent or intense movements. Similar normalization methods have been applied in other studies to combine multiple behavioral metrics to assess animal activity levels [4]. Thus, the values presented in Figure 1(C) are intended to provide a comparative measure of autonomic activity rather than an absolute unit of measurement. 3. In all the bar graphs, authors can move the significant stars from control group to aging model group to better represent the significant changes in the model group compared to control group. Thanks for your kind suggestion. Following the reviewer’s suggestion, we have revised the bar graphs and their captions accordingly. The significant stars have been moved from the control group to the aging model group in the revised figures to more accurately represent the significant differences in the aging model compared to the control group (Figures 1-5). 4. In 3.10. Indicate the appropriate figure number in the text, instead of just mentioning Figure 5. Mention figure number for IL-6 data. In response to the reviewer’s suggestion, we have revised the text in Section 3.10 to include specific figure numbers where the relevant data are presented. For the IL-6 expression data, we have explicitly referenced Figure 5(A). Similarly, for the p16, p19, and p21 gene expression data, we have clarified the references to Figure 5(B-C) (Page 22, lines 21-22 and Page 23, lines 3 in the revised manuscript). 5. Typo: Figure 3, (C) Is it MMP-1 or MMP-3? Figure 2 (C) axis label (U/mg prot) instead of (U/mgprot). Materials and methods, 2.3 Animal testing, line 9. Reframe the sentence “There is no expected unexpected deaths”. a) MMP-1 or MMP-3: We have corrected this typo in Figure 3(C) and confirmed that the correct marker is MMP-3. Figure 3(C) in the manuscript has been revised (Page 20, line 5). b) Axis label: We have updated the axis label in Figure 2(C) to "U/mg prot" to ensure proper spacing (Page 17, Figure 2(C)). c) Sentence rephrasing: The sentence in the Materials and Methods section has been rephrased for clarity. The revised sentence is now: "There were no unexpected deaths" (Section 2.3, Page 7, line 6). Reviewer #2: 1. Explain in detail the uniqueness of employing a combined D-galactose and UVB-induced aging paradigm, setting it apart from other research that used just one of these stressors. We greatly appreciated your professional and thoughtful review of our manuscript. The combined use of D-galactose and UVB irradiation integrates both intrinsic and extrinsic aging mechanisms. D-galactose induces oxidative stress and inflammation, mimicking systemic aging, while UVB accelerates skin aging through DNA damage and degradation of the extracellular matrix [5,6]. This dual stressor model more accurately reflects the complexity of human aging and provides a more comprehensive approach compared to studies that use only one of these stressors, such as D-galactose [7] or UVB [8]). By integrating both systemic and local aging pathways, this approach provides a more comprehensive assessment of interventions like CCACPs. These points have been added to the Discussion section. (Page 24, lines 9-16). 2. Give further background information on how the results might apply to aging-related diseases in people. Major risk factors for a range of diseases are strongly linked to aging, including neurodegenerative diseases [9,10], cardiovascular disease [11,12], type 2 diabetes [13,14], and chronic inflammation [3,15]. These conditions are often driven by mechanisms such as oxidative stress, chronic inflammation, and cellular senescence, all of which are key features of the aging process itself. As such, interventions targeting these underlying mechanisms to slow aging hold great potential for mitigating the onset and progression of aging-related diseases. We have incorporated these points into the Introduction (Page 3, lines 20-22 and Page 4, lines 1-4 ). 3. Explain the rationale behind the 8-week exposure period. Would a longer time frame yield more thorough insights? Thank you for your comments. The 8-week exposure period is consistent with prior studies in this field, which commonly use a similar duration for comparable experiments [1,8,16,17]. Research has shown that this timeframe strikes a balance between capturing meaningful effects and minimizing potential confounding factors. Therefore, we adopted an 8-week exposure period for the mouse model in this study. In future experiments, we plan to extend the exposure period to further explore the long-term effects. 4. Give further information about the methods used to regulate the UVB exposure so that each individual experienced the same level of intensity. The UVB irradiation experiment was conducted according to previous reports [8,18]. To ensure consistent UVB exposure across all subjects, we carefully controlled the mice's range of motion. Specifically, the mice were confined to areas smaller than the UVB exposure range, ensuring uniform exposure intensity for all individuals. Additionally, prior to irradiation, we calibrated the intensity using a UVB radiometer to maintain a consistent dose of 40 mJ/cm² throughout the study. The specific modifications in the article are as follows: Before UV irradiation, the dorsal fur of each mouse was shaved with an electric razor, followed by the application of depilatory cream. The distance from the light source to the back of the animals was maintained at 25 cm. During UVB exposure, the mice were housed in groups within the irradiation chamber, allowing free movement [8,18]. Prior to exposure, the irradiation intensity was calibrated to ensure a consistent dose of 40 mJ/cm². Throughout the irradiation process, the mice’s range of motion was kept within the designated exposure area (Page 8, lines 3-9). 5. To contextualize the peculiarity of CCACP, update Table 2 to include a comparison of its amino acid content with that of other collagen sources. Thank you for your comments. Since the amino acid data from other collagen sources are not based on our own results, they have not been included in Table 2. However, we have added a comparison of the amino acid compositions from other collagen sources in the Discussion section. The amino acid composition of CCACPs was compared with collagen peptides derived from chicken bone and bovine bone in previous studies [1,19]. CCACPs showed significantly higher levels of phenylalanine (13.26%) and glutamic acid (10.56%) compared to both chicken and bovine collagen peptides, which are known to support tissue repair, structural protein synthesis, and metabolic balance. While the glycine content in CCACPs (19.85%) was slightly lower than that in chicken bone collagen peptides (27.86%), it was comparable to bovine bone collagen peptides (19.84%), ensuring sufficient structural integrity. Proline levels were similar across all collagen sources. These compositional differences underscore the unique amino acid profile of CCACPs, which may contribute to their enhanced bioavailability and systemic anti-aging effects, distinguishing them from other collagen sources that primarily benefit skin health (Page 25, lines 10-21). 6. Additional references from latest research would be helpful. Following the reviewer’s suggestion, we have replaced several latest references to support our findings, particularly those related to the role of collagen peptides in aging and their potential therapeutic applications. The references that were changed are 20-27 [20–27]. Journal Requirements: 1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. Thank you for your feedback. I have made the necessary revisions to ensure that the manuscript complies with PLOS ONE's style requirements, including the correct file naming conventions. Please feel free to let me know if any further adjustments are needed. 2. Please ensure that you provide the correct grant numbers for the awards you received for your study in the ‘Funding Information’ section. Thank you for pointing out the inconsistency in the funding information. To address the inconsistency noted in the funding information, upon resubmission, we have provided the correct funding numbers for the awards supporting our research in the 'Funding Information' section. The manuscript has been appropriately revised to ensure consistency with the information provided in the submission system. (Page 31, Line 18-22 and Page 32, Line 1 in the revised manuscript). We have made all necessary revisions to the manuscript based on your constructive feedback. We believe these changes have significantly improved the manuscript and we hope that the revised version meets the expectations of the reviewers and editors. Thank you again for your time and helpful suggestions. Sincerely, Lin Zhao, Ph.D. School of bioengineering Qilu University of Technology Jinan, 250353, China Fax: + 86 531 89631776 Tel: + 86 531 86529099 E-mail: iahb205@163.com Reference 1. Song H, Zhang S, Zhang L, Li B. 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Evaluation of the anti‐aging e Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0317302.r002
26 Dec 2024 Decision Letter - Arumugam Muthuvel, Editor Protective effects of Colla Corii Asini Collagen Peptides on D-galactose injection combined with UVB irradiation-induced aging in mice PONE-D-24-41650R1 Dear Dr. Zhao, 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, Arumugam Muthuvel Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0317302.r003
Formally Accepted
14 Jan 2025 Acceptance Letter - Arumugam Muthuvel, Editor PONE-D-24-41650R1 PLOS ONE Dear Dr. Zhao, 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. Arumugam Muthuvel Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0317302.r004

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