Oxidative Effects in Early Stages of Embryo Development Due to Alcohol Consumption

Round 1

Reviewer 1 Report

Authors should revise the manuscript.

Comments for author File: Comments.pdf

Language is fine.

Author Response

Firstly, thanks for your comments and recommendations. We have thought about your revision and we would like to answer every point:

MAJOR REVISION

1. We agree with you. The topic was not well defined in the title. The new title, "Oxidative effects in early stages of embryo development due to alcohol consumption", is clearer than the previous one.
2. We have included new data with references that confirm the previous statistics. 
3. We have discussed the possibility of including a new section about animal studies. Even though we had prepared a sketch, we did not eventually include it, since it appeared to us that a great part of the information could be redundant. We think that the text is clear about the data obtained from animal studies. 
4. You are right, the figures could be improved. We have re-made them, improving both the quality and the orthography. 
5. We have included some remarks indicating the need for new studies over the text.
6. We have added new recent references. We understand that some references are quite old, but we want to remark that they are useful in terms of narrative. We would have liked to find more recent studies, but there is some lack of specific novel research. Precisely for this reason it is interesting for us to highlight this issue and encourage new approaches.

MINOR REVISION

Thanks for your help, we have corrected all these mistakes. 

REMARK

We have highlighted in some parts of the text that the main goal of the manuscript is the study of alcohol consumption, and consequently, the effects of ethanol contained in spirits. We think there is no ambiguity in the new version of the manuscript. Please, notice that we have tried to simplify some parts of the text for better understanding. We have also strengthened the parts of genes and treatments.

Thanks again for your help and we hope you find interesting all the changes we have made. 

Reviewer 2 Report

·         Main Research Question: The main question addressed by this research is the impact of ethanol (alcohol) on the development of embryos, specifically regarding oxidative stress and its role in FASD. The study seeks to understand how ethanol-induced oxidative stress contributes to developmental anomalies and disorders in embryos.

·         Originality and Relevance: This research is highly relevant to the field of developmental biology and public health, given the widespread consumption of alcohol and the significant effects of ethanol exposure on fetal development. It contributes original insights by focusing on oxidative stress mechanisms, offering a detailed examination of how ethanol metabolism disrupts cellular and molecular processes, leading to developmental disorders. The paper addresses a critical gap in understanding the specific biochemical pathways through which alcohol exerts its teratogenic effects.

·         Contribution to the Field: Compared with other published material, this paper adds value by providing a comprehensive review of ethanol's oxidative effects on early embryo development, consolidating knowledge about the role of Reactive Oxygen Species (ROS) in mediating ethanol's harmful effects. It synthesizes evidence from various studies to propose potential therapeutic interventions, such as the administration of metformin during critical developmental periods, highlighting a novel approach to mitigating FASD's adverse outcomes.

·         Methodology Improvements: The paper does an excellent job of detailing ethanol's metabolic pathways and the resultant oxidative stress. However, to further strengthen the research, the authors could consider incorporating more in vivo studies to validate the proposed mechanisms and therapeutic interventions. Additionally, a broader range of antioxidants and their specific protective mechanisms against ethanol-induced damage could be explored to provide a more comprehensive overview of potential treatment strategies.

·         Consistency of Conclusions with Evidence: The conclusions drawn in this study are consistent with the evidence presented, clearly linking ethanol-induced oxidative stress to developmental disorders associated with FASD. The paper effectively addresses the research questions posed, supported by a robust discussion of ROS's role and the potential for therapeutic interventions. It acknowledges the limitations and complexities of translating these findings into clinical practice, calling for further research in this area.

·         Appropriateness of References: The references provided are appropriate and cover a wide range of studies, from molecular mechanisms of ethanol metabolism to clinical implications and potential interventions. They support the paper's arguments well and demonstrate a thorough review of the current literature on the subject.

·         Additional Comments on Tables, Figures, and Data Quality: The tables and figures effectively illustrate the metabolic pathways of ethanol and the oxidative effects on embryo development. They provide a clear visual representation of complex biochemical processes, enhancing the reader's understanding. The data presented, particularly regarding oxidative stress markers and the impact of therapeutic interventions, are compelling and underscore the paper's key messages.

Wording

• Deficiencies: The paper generally maintains a clear and precise use of language, suitable for an academic publication in the field of developmental biology and toxicology. However, there are instances where wording could be streamlined for clarity and brevity. For example, some sentences are overly complex, potentially obscuring the intended message.
• Modification Suggestions: Consider simplifying complex sentences and ensuring that terminology is consistently defined at its first occurrence. This approach will enhance readability for a broader audience, including those whose primary research focus may not align exactly with the subject matter.
• Modification Examples: Change "Ethanol interferes in periods of greater embryonic susceptibility and organogenesis, producing congenital defects" to "Ethanol exposure during critical periods of embryonic development, such as organogenesis, can lead to congenital defects."

Tone

• Deficiencies: The tone of the paper is appropriately academic and matches the conventions of the field. It maintains an objective stance and uses passive constructions where suitable to focus on the research findings rather than the researchers. However, the introduction could benefit from a slightly more engaging tone to better capture the reader's interest and emphasize the significance of the research.
• Modification Suggestions: While keeping the formal academic tone, incorporate a brief overview of the real-world implications of the findings in the introduction to immediately engage the reader with the relevance of the study.
• Modification Examples: Change "Alcohol has an important toxic effect, especially on embryonic development" to "The toxic effects of alcohol, particularly on embryonic development, pose significant public health challenges."

Structure

• Deficiencies: The overall structure of the paper is logical and follows a conventional format for scientific articles, including sections on introduction, methods, results, and discussion. However, some sections, particularly the results and discussion, could benefit from clearer subheadings to guide the reader through the complex data and analyses presented.
• Modification Suggestions: Use more descriptive subheadings within the results and discussion sections to improve navigability and help readers quickly find the information most relevant to their interests.
• Modification Examples: For a subsection discussing the role of oxidative stress in fetal alcohol spectrum disorders, change a generic heading like "Results" to "Oxidative Stress Indicators in Fetal Alcohol Spectrum Disorders."

Innovation

• Deficiencies: The paper provides a comprehensive review of the oxidative effects of ethanol on embryonic development, including potential therapeutic interventions like metformin administration. While the synthesis of existing research is thorough, the paper could further its contribution to the field by suggesting new avenues for research based on the gaps and questions raised by the current literature review.
• Modification Suggestions: Highlight specific, under-researched areas within the topic that future studies could address, thereby encouraging innovation in the field.
• Modification Examples: "Future research should explore the potential of targeted antioxidant therapies beyond metformin, especially those that can cross the placental barrier and directly mitigate oxidative stress in the developing fetus."

Validity

• Deficiencies: The paper is well-researched, with extensive citations supporting the discussed effects of ethanol on embryonic development and the mechanisms of oxidative stress. However, the discussion on genetic factors influencing susceptibility to fetal alcohol spectrum disorders could be expanded to include recent studies, thereby enhancing the validity of the review.
• Modification Suggestions: Integrate recent genetic studies that provide deeper insights into individual variability in susceptibility to fetal alcohol spectrum disorders.
• Modification Examples: Include recent findings from genome-wide association studies (GWAS) that identify genetic polymorphisms related to increased risk of fetal alcohol spectrum disorders.

Data Support

• Deficiencies: The review extensively cites existing literature to support its discussions. Nonetheless, the sections on pharmacological interventions could be strengthened by including more recent clinical trials or meta-analyses, if available, to support the effectiveness of proposed treatments like metformin.
• Modification Suggestions: Conduct a more recent literature search to find the latest clinical trials or systematic reviews on the effectiveness of metformin and other antioxidants in mitigating the effects of prenatal alcohol exposure.
• Modification Examples: "Recent meta-analyses indicate that metformin shows promise in preclinical models for reducing oxidative stress induced by prenatal alcohol exposure, yet clinical trials in humans remain limited."
• References Update: Check that all references are up-to-date and consider including recent studies that may have been published since the manuscript was drafted. In particular, consider including additional references to support the discussion and to provide context to the study’s findings. I suggest adding data related to recent bulk transcriptomics studies which could represent a strong substrate to enforce the role of described molecular mechanisms, such as the recent PMID: 36490268.

Overall Recommendations

• The paper performs well in providing a comprehensive review of the literature on the oxidative effects of ethanol on embryonic development and proposes intriguing potential interventions. However, it could benefit from clearer structuring within some sections, a more engaging introduction, and an expanded discussion on genetic factors and recent clinical trials on proposed treatments. It is recommended that the authors make modifications to address the aforementioned deficiencies in structure, wording, and the incorporation of recent studies, to enhance the paper's contribution to the field and its utility as a resource for researchers and clinicians alike.

The English should be deeply revised.

Author Response

We have found your extensive revision very interesting and formative. We have also considered every point of your comments very carefully. Your recommendations led us to make several changes that we explain below:

Wording

We have tried to simplify some parts of the text. We included your example and tried to apply that idea in some other parts. For example, lines 213-214 were rewritten: "This results in an increase in the respiratory chain's activity, which produces high levels of ROS...", or lines 51-52: "In recent decades there has been a change in the consumption pattern in women, because the age of onset has decreased, and the consumption has increased."

Tone

We included your example and we tried some other strategies, for example, we tried to highlight that the text talks about ethanol included in spirits, giving some context to the study.

Structure

Sorry, but we think we did not understand this point properly. In your example, you talk about changing "Results" to "Oxidative Stress Indicators in Fetal Alcohol Spectrum Disorders." However, we have not changed the headings because no "Result" heading is included in the text and consequently we are not sure what specific changes you prefer. In fact, in our text, there is no "methods" or "results" sections.

Innovation

We had some difficulties finding new trials and analyses about the use of metformin, but eventually we have included them. Precisely, this lack of specific recent studies encouraged us to carry out this work that would inform and at the same time motivate new approaches.

Validity

We have included some information about GWAS related to increased risk of fetal alcohol spectrum disorders.

Data Support

Although we have included new studies, we have not added the PMID reference: 36490268 because we have not seen a clear way of relating these data to the increased risk due to alcohol consumption.

Overall Recommendations

The new version of the manuscript contains clearer text in some difficult points. Furthermore, we have included new references with studies that support the previous information. Please, notice that we have also improved the quality of the figures, and updated the references about the statistical data provided in the introduction. Thanks again for your help and we hope that you agree with all the changes we have made.