Vepris macrophylla Essential Oil Produces Notable Antiproliferative Activity and Morphological Alterations in Human Breast Adenocarcinoma Cells
Round 1
Reviewer 1 Report
The present study describes the effect of Vepris macrophylla essential oil on breast adenocarcinoma cell line SKBR3; it is a valuable study that inspire to continue studying its antiproliferative effects, however there are some minor issues that could be addressed to improve the manuscript.
In the introduction section you mention that natural products in combination with synthetic drugs can improve the therapeutic index of drug and reduce many undesirable side effects, it would be interesting to mention some examples of this case and, if there is some approved treatment that exhibit this effect. In this lane, and as personal interest, did you evaluated this effect in your study?
Referring to the cell viability results, you could summarize if there is significant difference between concentrations as well as between time treatments.
In lane 325, please check if is it correct “…the cytotoxic effects of EOs in toto” or do you mean TTO. Also, in lane 329-330, it could be more clear mentioning “citral is generally recognized as safe (GRAS) by the FDA”.
Summarizing your results and according to previous studies, it would be interesting to analyze the possible mechanism of action of the antiproliferative activity.
Overall, please check the use SI units in a consistent fashion throughout the manuscript (for example, 24 h instead of 24h –a space must separate the digits from the symbol-) as well as the English grammar, some sentences can be improved, mainly in the introduction section.
Author Response
Dear Reviewer,
Thanks for your revisions, we agree with your comments and below you can read the answers point by point and in the revisioned manuscript you can find our revisions.
The present study describes the effect of Vepris macrophylla essential oil on breast adenocarcinoma cell line SKBR3; it is a valuable study that inspire to continue studying its antiproliferative effects, however there are some minor issues that could be addressed to improve the manuscript.
- In the introduction section you mention that natural products in combination with synthetic drugs can improve the therapeutic index of drug and reduce many undesirable side effects, it would be interesting to mention some examples of this case and, if there is some approved treatment that exhibit this effect. In this lane, and as personal interest, did you evaluated this effect in your study?: Done, you can read the new paragraph in the introduction from line 35 to 58.
- Referring to the cell viability results, you could summarize if there is significant difference between concentrations as well as between time treatments: we can say that there aren't a significant difference. The results are the same at 24, 48 and 72 hours. The biological effects of Vepris’EO are very quick.
- In lane 325, please check if is it correct “…the cytotoxic effects of EOs in toto” or do you mean TTO (note add): Done, we confirm that the correct form is “…the cytotoxic effects of EOs in toto” (see line 339).
- Also, in lane 329-330, it could be more clear mentioning “citral is generally recognized as safe (GRAS) by the FDA”: Done, see new line 342-343.
- Summarizing your results and according to previous studies, it would be interesting to analyze the possible mechanism of action of the antiproliferative activity: thanks for this comment. This is the objective of a new manuscript. In our Lab. we are starting to design the future experimental schedules.
- Overall, please check the use SI units in a consistent fashion throughout the manuscript (for example, 24 h instead of 24h –a space must separate the digits from the symbol-) as well as the English grammar, some sentences can be improved, mainly in the introduction section: Done
Author Response File: 
Author Response.docx
Reviewer 2 Report
The manuscript I was reviewing interested me very much. Interesting subject matter, presentation of the topic legible. However, some information needs to be corrected / completed before publishing.
Introduction
The authors write about products of natural origin used in chemotherapy, but only one literature item is given. This information should be supported by more examples in order to introduce the reader to the subject more deeply. After reading the introduction, the reader feels unsatisfied with knowledge. If there are no such data in the literature, it should be clearly emphasized.
Methods
The methodology for obtaining EO is insufficiently described. It is not enough to refer to the publication, it should be briefly described how the authors received the EO from V. macrophylla.
Moreover, the method of culturing the tested cell line is described too briefly. I understand the cell line was cultured without antibiotics? how it's possible? Were no other substrate additives necessary? this information should be completed.
Why was the least sensitive MTT test used in the proliferation test? therefore the results were not supplemented with the SRB test.
Why were the same EO concentrations not tested in SEM as in the MTT test? the highest, i.e. 5 and 10 µg / ml, were excluted?
Results
Picture 3 has very poor quality. Maybe the authors in the archive will find it more readable?
Discussion
The authors emphasize that EO exerts an anti-cancer effect on numerous cancer models. The word "numerous" is too general, it should be supported by more specific examples. Please expand the discussion.
With the above information added in, the work will bring light to anti-cancer research.
Author Response
Thanks for your revisions, we agree with your comments and below you can read the answers point by point and in the revisioned manuscript you can find our revisions.
The manuscript I was reviewing interested me very much. Interesting subject matter, presentation of the topic legible. However, some information needs to be corrected / completed before publishing.
Introduction
- The authors write about products of natural origin used in chemotherapy, but only one literature item is given. This information should be supported by more examples in order to introduce the reader to the subject more deeply. After reading the introduction, the reader feels unsatisfied with knowledge. If there are no such data in the literature, it should be clearly emphasized: Done, the reviewer can read the new paragraph in the introduction from line 35 to 44.
Methods
- The methodology for obtaining EO is insufficiently described. It is not enough to refer to the publication, it should be briefly described how the authors received the EO from V. macrophylla: Done, we have insert the briefly description (see line 86-87).
- Moreover, the method of culturing the tested cell line is described too briefly. I understand the cell line was cultured without antibiotics? how it's possible? Were no other substrate additives necessary? this information should be completed: Done, the reviewer can read added information from line 92-95.
- Why was the least sensitive MTT test used in the proliferation test? therefore the results were not supplemented with the SRB test: we thanks to the reviewer for the suggestion; in our laboratory we usually use the MTT assay.
- Why were the same EO concentrations not tested in SEM as in the MTT test? the highest, i.e. 5 and 10 µg / ml, were excluted?: in order to study the biological effects of one o plus compounds you need to use their sub-cytotoxic concentrations. The highest i. 5 and 10 mg/ml were excluded for this reason. At these concentrations cells were completely destroyed and none cells were observed by SEM.
Results
- Picture 3 has very poor quality. Maybe the authors in the archive will find it more readable?: in order to show a good micrograph we have changed the image with another better one.
Discussion
The authors emphasize that EO exerts an anti-cancer effect on numerous cancer models. The word "numerous" is too general, it should be supported by more specific examples. Please expand the discussion. With the above information added in, the work will bring light to anti-cancer research: Done: the reviewer can read the new insert from line 326-331.
Best Regards
Annarita Stringaro
Author Response File: Author Response.docx
Round 2
Reviewer 2 Report
I am satisfied with the corrections provided by the authors.