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I confirm that you have addressed the points reasonably and I am happy to recommend the acceptance of this manuscript to the section editor. Congratulations!
[# PeerJ Staff Note - this decision was reviewed and approved by Paula Soares, a PeerJ Section Editor covering this Section #]
Three reviewers now recommend its acceptance, while one (reviewer 5) still suggests changing the title. I myself feel that the current title may sound too general compared with its content. Although this is not a strong request, could reconsider this issue? I appreciate your patience.
The introduction is overall well written, with Literature references, and sufficient field background/context.
Well-established protocols were used for the execution of the research study.
Research findings are novel
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The authors have made substantial improvements to their manuscript, addressing the reviewers’ comments thoughtfully and effectively.
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The title is still misleading. It should have a clear reference to the model used
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While the mouse model effectively mimics TTC features, its translatability to human disease is limited due to differences in hormonal regulation, stress response, and myocardial physiology. Including a discussion on these limitations and suggesting future studies involving human samples would strengthen the clinical relevance. The study lacks validation of findings in human TTC patients. While the murine model offers insights, incorporating human data, such as serum biomarkers or cardiac biopsies, would enhance the study’s applicability and credibility. Expanding this aspect could be a significant improvement for translational research.
The study investigates the molecular mechanisms underlying Takotsubo cardiomyopathy (TTC) using a murine model induced by isoproterenol (ISO). Through proteomic analysis, the authors identified 81 differentially expressed proteins (DEPs), highlighting pathways associated with oxidative stress, inflammation, and metabolic dysregulation. Upregulation of pathways such as ferroptosis, oxidative phosphorylation, and collagen fibril organization. Identification of hub proteins (e.g., Ntrk2, Fdft1, Serpine1) that may contribute to myocardial remodeling and stress responses. Evidence of myocardial recovery over time, coupled with persistent structural damage. The study provides insights into potential therapeutic targets to mitigate stress-induced myocardial injury.
Several reviewers have reviewed your manuscript. Most of them recommend you revise the manuscript more or less. Please read their comments and revise the manuscript accordingly. Despite the number of comments, I believe there is no essential difficulty in addressing them.
[# PeerJ Staff Note: The review process has identified that the English language must be improved. PeerJ can provide language editing services if you wish - please contact us at [email protected] for pricing (be sure to provide your manuscript number and title). Your revision deadline is always extended while you undergo language editing. #]
Overall Manuscript was meticulously designed and executed.
The protocols used for this study are well established, experimental design is suitable for research study execution.
In this research, authors investigated the pathophysiological mechanisms associated with ISO-induced TTS-like cardiomyopathy in female mice. Proteomic analysis offers new perspectives on the molecular processes that contribute to stress-induced myocardial damage. The discovery of critical proteins related to oxidative stress, inflammation, and energy metabolism underscores the complex responses of the myocardium to acute stress. By combining functional, structural, and proteomic data, understanding of TTC, which sets the stage for future research focused on creating targeted therapies to alleviate the prolonged impacts of acute stress on cardiac health.
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I acknowledge that you submitted a review on "Proteomics of stress-induced cardiomyopathy: insights from differential expression, protein interaction networks, and functional pathway enrichment."
As the reviewer for this article, I would like to commend the authors for selecting a relevant and intriguing topic for their manuscript. Their expertise in this area is evident, and I appreciate their hard work and dedication to producing a paper that is both comprehensive and concise. The article is not only well-written but also highly relevant, successfully capturing the reader's attention.
The text is written in clear English. It is also well-referenced with relevant literature. The structure conforms to PeerJ standards. The figures are relevant.The research question is clearly defined and significant. It explains how the study addresses a specific knowledge gap. The investigation was conducted meticulously, adhering to high technical and ethical standards. The methods are described in enough detail to allow for replication. The conclusions are clearly stated, connected back to the original research question, and supported by the results.
I reviewed the manuscript to my satisfaction and found it engaging; I read it in one sitting without any complaints. My current research focus is on the ISO model of TTC in rodents, and this manuscript has significantly enhanced my understanding of this scientific field. I sincerely thank the authors for their contributions and the Editorial board of the Peer J for allowing me the opportunity to review this work.
With warm regards,
Assistant Professor Tanja Šobot,
Department of Physiology, Faculty of Medicine University of Banjaluka,
Banjaluka, Bosnia and Herzegovina
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This manuscript explores the proteomic landscape of stress-induced cardiomyopathy using a murine model. The study employs label-free proteomics, GO/KEGG pathway analysis, and protein-protein interaction networks to identify key pathways such as ferroptosis and oxidative stress, and hub proteins like Ntrk2 and Serpine1. While the research is timely and employs advanced techniques, the manuscript is hampered by several issues. These include inconsistent language and figure formatting, insufficient experimental validation of key findings, limited discussion of recovery mechanisms, and a lack of contextualization within the broader field.
1. "Proteomic" in the manuscript title and text should be corrected to "Proteomics".
2. While the manuscript mentions the use of bioinformatics tools such as MaxQuant, STRING, and Cytoscape, it does not specify the software versions or configurations used. Providing these details is essential for reproducibility.
3. The study mentions 30 mice divided into two groups but does not specify how many biological replicates were used for proteomic analyses and validation. Clarifying this information is important for assessing the robustness of the findings.
4. The authors used the STRING database for constructing the PPI network, but the confidence score threshold for interactions is not provided. Including this information would enhance transparency and reliability.
5. While structural changes in the myocardium are described, no quantitative analysis (e.g., fibrosis area percentage) is provided. Incorporating quantitative metrics would strengthen the evidence for remodeling.
6. The study uses a single high dose of isoproterenol (ISO) without testing variable dosages, raising questions about dose-response effects. The authors should briefly justify the selected dose and clarify whether it represents a clinical or EC50 level or others.
7. In Figure 1C, there are no any annotations in the figure and scale bar.
8. The H&E results in Figure 2 show changes across different time points (0, 1, 3, and 7 days), but these are not annotated. Adding annotations to highlight specific changes over time would enhance clarity.
9. The histological micrographs in some subfigures of Figure 2 are missing scale bars, which are critical for accurate interpretation of structural changes in the myocardium. The authors should add these annotations.
10. The two panels in Figure 3 have different sizes, which disrupts visual balance. Ensuring both subfigures share the same width would improve presentation consistency.
11. The gene names in Figure 4C are displayed unclearly and appear flattened. The authors should adjust the formatting to make the text more legible.
12. Many pathways in Figure 6 have p-values greater than 0.05, indicating non-significance. Including non-significant pathways in the figure could confuse readers. It is recommended to display only statistically significant pathways (p < 0.05) to maintain clarity.
13. The protein-protein interaction network in Figure 7 is poorly labeled. Node labels are unclear, making it difficult for readers to identify specific hub proteins. Improved labeling and clarity are recommended.
14. Figure 8 lacks proper annotations, making it difficult to interpret the red and blue regions in each subfigure. The authors should clearly label these elements to indicate their significance (e.g., red and blue regions representing upregulated and downregulated pathways). Additionally, the number of subfigures in each column should be consistent.
The study utilized LC-MS/MS to identify differentially expressed proteins and gene ontology pathwyas in a muring model of takostubo syndrome. Authors induced takostubo syndrome with a single dose of isoproterenol. The study identified that key proteins involved in energy metabolism, oxidative stress and cell death pathways were differentially expressed in this model.
Authors have provided sufficient background for the study.
All the raw data and figures shared
Major concerns:
1. Authors used a single dose of ISO , intraperitoneal injection of ISO (Sigma-Aldrich, Germany) at a dose of 200 mg/kg . The survival curve for the ISO toxicity should be included during the study.
2. Echocardiography measurements were conducted using 9MHz transduced. Authors should include the details of the machine used for the study.
3. Which tracing parameter used to trace the PSLAX view? It seems like Fig 2C systole-control is traced in diastole mode. Verify the image.
4. Ther results for ejection fraction, cardiac output and diastolic function needs to incorporated to explain the cardiac function in control vs ISO.
5. All the figure legends needs adequate explanation with number of animals used, statistical significance details
6. IN H7E staining, high light the area with arrow marks to show the changes in control vs ISO
7. Give the details for fold change cut off ratio, p value cut off used to create the heat map and volcano plot.
8. Give details of the illustration packages used to create heat map and volcano plots
9. Why authors performed GO enrichment and kegg enrichment. Both pathways gives different directions for the analysis. To conclude the findings either one pathway and highlighting the protein interaction is sufficient.
10. How the GSEA correlate with the to and down regulated pathways. Discuss.
Novelty of the findings needs to be highlighted
Validity of the findings needs to mentioned with number of animals in each experiments and statistical explanation.
The manuscript is generally clear. However, editing is required to correct some typographical errors and some repeated sentences.
The experiments are well designed and the results are clearly presented. However, the title is misleading, since this is a model of isoproterenol-induced cardiomyopathy not stress-induced cardiomyopathy. The authors need to change this title to something like: Proteomics of catecholamine stress-induced cardiomyopathy or Proteomics of isoproterenol-induced Takotsubo-like cardiomyopathy
The findings of the proteomics are valid. However, confirmation should be done by western blotting (not by PCR). The authors need to measure the protein expressions of the most important DEPs by western blotting.
The figure legends should include more details: a detailed descriptive title, and a summary of statistics
The KEGG and GSEA findings are presented but lack a strong biological narrative. Explain the relevance of ferroptosis and collagen reorganization in the context of myocardial injury.
The methods for protein identification and threshold for DEPs could be explained more clearly. For example, explicitly state how missing data in the proteomics workflow was handled.
Hub proteins like Ntrk2, Fdft1, Serpine1, and Cyp1a1 are identified but not sufficiently discussed. Explain their specific roles in stress response or cardiac remodeling.
The study investigates stress-induced cardiomyopathy (Takotsubo cardiomyopathy) in a murine model using a high dose of isoproterenol (ISO). The authors employ label-free quantitative proteomics to analyze myocardial tissues, revealing 81 differentially expressed proteins (DEPs). Pathway analyses, including GO, KEGG, and GSEA, highlight disruptions in energy metabolism, oxidative stress, and ferroptosis. The protein-protein interaction (PPI) network identifies key proteins like Ntrk2, Fdft1, Serpine1, and Cyp1a1. The study provides insights into the molecular mechanisms of stress-induced myocardial damage, paving the way for identifying therapeutic targets.
1. Since female mice were used, discuss how gender influences stress-induced cardiomyopathy and why females are more susceptible.
2. The study highlights transient myocardial dysfunction with eventual recovery. Explore the potential mechanisms underlying recovery and whether they could be targeted therapeutically.
3. Some figures (e.g., Figure 5 and Figure 6) are crowded and could be simplified. Provide clearer legends explaining the key findings and their biological importance.
4. Highlight the limitations of the murine model, especially its relevance to human TTC. Include how differences in stress responses and myocardial metabolism may affect generalizability.
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