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After revisions, all reviewers agreed to publish the manuscript. I also reviewed the manuscript and found no obvious risks to publication. Therefore, I also approved the publication of this manuscript.
[# PeerJ Staff Note - this decision was reviewed and approved by Gwyn Gould, a PeerJ Section Editor covering this Section #]
The authors have addressed the majority of comments.
The authors have claimed the limitations of the current reseach.
The authors have addressed the comments.
The authors have resubmitted their manuscript in response to the review comments. I appreciate their efforts to address the concerns raised. While the authors did not conduct additional experiments as initially suggested, they have provided a more detailed discussion addressing the points raised in the previous review. The manuscript’s overall scientific merit remains intact, and the expanded discussion provides sufficient context to address the original concerns.
There was some confusion with the line numbers mentioned in the rebuttal letter, as they do not correspond accurately to the revised manuscript. This discrepancy made it challenging to cross-reference the authors’ changes with the original comments. For future submissions, I recommend ensuring that such details are accurately aligned.
The authors have adequately addressed the key concerns, even though further experiments were not conducted. I recommend accepting the manuscript for publication.
no comment
no comment
The authors are requested to carefully revise the manuscript and answer the questions raised by the reviewers.
Overall, the writing is clear and understandable for a scientific paper. It presents complex information in a logical manner. However, there are a few issues that could be improved:
Incomplete Sentences: A few sentences are incomplete or grammatically incorrect. For example, the sentence on line 687 starts with "Transferrin (TF), an iron-binding transport protein reported to be |involved with..." and the sentence on line 698 states "In a previous study demonstrated..."
Sentence Structure: A few sentences are quite long and complex, which can make them harder to follow. Breaking these into shorter sentences could improve readability.
Repetitive Phrasing: Some phrases are used repeatedly (e.g., "However, further investigation is required..." or "These findings suggest..."). Varying the language would make the writing more engaging.
Technical Terms: The paper assumes a high level of familiarity with immunology and cell biology. While this is appropriate for a scientific audience, a few additional explanations might be helpful for readers outside the field (e.g., a brief definition of "inflammasome" and "macrophage polarization").
Figure References: In Figure 1, 6-day incubation was indicated to induce TAM. The time is not consistent with the "7 days" described in Line 339. This should be clarified. The cartoons used for the last two conditions in Figure 1 are the same. It could be improved to distinguish those two conditions.
In Figure 2, assessing macrophage differentiation could be more effectively achieved by quantifying M0/M1/M2 marker expression using flow cytometry, rather than relying solely on morphological changes, which were not uniformly observed. Flow cytometry would allow for precise quantification of differentiated cell percentages, facilitating a more accurate comparison of induction efficiency under different conditions. This approach is also recommended for the experiments presented in Figure 3.
The statistical methods in this manuscript are partially described and the majority of the conclusions are supported by the presented data.
This is an interesting study that investigates the effects of oleamide on the repolarization of tumor-associated macrophages (TAMs) and their impact on breast cancer cell viability. The authors used a co-culture system to generate TAMs and treated them with oleamide. They then performed proteomic analysis to identify differentially expressed proteins and evaluated the effects of oleamide-treated TAM-conditioned medium on breast cancer cell viability and apoptosis. The study provides valuable insights into the potential role of oleamide in modulating TAM polarization and its implications for cancer treatment.
1. The authors should provide more information about the rationale behind using oleamide and its potential mechanism of action in the context of TAM polarization and breast cancer. Why was the breast cancer cell line MDA-MB-231 specifically selected for this study? Is the observed effect specific to breast cancer, or can it be extrapolated to other types of cancer cell line coculture models? The authors should also discuss the limitations of this study, including the use of a single breast cancer cell line and the potential influence of various factors present in the tumor microenvironment.
2. The authors should double-check the formatting and consistency of the references throughout the manuscript. For instance, line 1039-1043 are the same reference but repeated.
1. Regarding donors:
• Line 102 mentions that the experiment was carried out based on “a donor”. Was this donor one of the donors AB, O1, and O2? If it was one donor, how could the authors perform “three independent experiments”? Did they mean three technical replicates? Please clarify.
• For the LC-MS/MS experiment, MDM from three donors AB, O1, and O2 were used. Why do the results only show AB and O? Please clarify.
2. Regarding the macrophge markers:
• The marker evaluation is biased towards M2 markers. As mentioned in the introduction (lines 62-63), M1 markers include CD80/86, CXCL10, among others (such as doi: 10.1016/j.immuni.2014.06.008). For ELISA, markers like IL12 can be used, among others. More M1 marker evaluation following oleamide treatment will help understand if the polarization effect is mainly through suppressing the M2 phenotype or also increasing the M1-like phenotype.
• Since there were proteome data from three donors AB, O1, and O2, why not summarize all well-accepted markers (such as those cited in the manuscrip, as well as doi: 10.1016/j.immuni.2014.06.008) that can be detected in the current study to see the marker changes in untreated TAM and oleamide-treated TAMs? This will offer more comprehensive insights into macrophage marker alteration instead of focusing on a few selected markers.
1. The authors should elaborate on the biological significance of the differentially expressed proteins identified in the proteomic analysis and their potential roles in TAM polarization and breast cancer progression. For instance, Lines 511-523: What do these pathways indicate? How are they relevant to their potential roles in TAM polarization and breast cancer progression? Draw a conclusion for the results.
2. The authors should discuss the potential clinical implications of their findings and suggest future research directions to further elucidate the therapeutic potential of oleamide in cancer treatment.
1. The authors should ensure that all abbreviations are defined upon first use in the manuscript. For instance, Line 234: Liquid chromatography-Mass Spectroscopy (LC-MS/MS) was not defined when it appeared in the text the first time.
2. The authors should carefully proofread the manuscript to correct any minor grammatical or typographical errors, among others. For instance:
• Typo in legend of figure 3: "anti-xinflammatory" should be "anti-inflammatory".
• Line 105: "1x105" should be "1×10^5".
• Line 245: Mention that "AGC" stands for "Automatic Gain Control".
• The figure legends were inserted in both the results section and the figure section, causing repetition.
• Lines 481-483: These lines belong to the methods section. Please specify the accession number.
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