In Vitro Radioenhancement Using Ultrasound-Stimulated Microbubbles: A Comparison of Suspension and Adherent Cell States
Round 1
Reviewer 1 Report
Present work discuss about the use of ultrasound mediated microbubble use and radiation treatment. The work looks interesting.
1. Please provide details about the microbubbles, were they prepared in lab and characterized?
2. Pls provide graphical abstract
I think English is clear and simple to read.
Author Response
- Please provide details about the microbubbles, were they prepared in lab and characterized?
Author response: We used the commercially available Definity® perflutren lipid microspheres (mean diameter 1.3 – 3.3 µm) (Lantheus Medical Imaging) which were purchased from Global Medical Solutions (Melbourne, Australia). We have added this information to Line 129-130.
- Pls provide graphical abstract
Author response: We have attached a graphical abstract as requested, and is attached to the manuscript on Page 13, and below.
Author Response File: 
Author Response.pdf
Reviewer 2 Report
This article is a great example of original articles. I only have a couple comments.
1) Regarding the "Statistical Analysis" paragraph, the authors have not explain the normalization process (e.g. mathematical models utilized). Please give some more information.
2) It would be great to include a table that clearly presents the difference resulting in the survival of cells by the implementation of each different method. This way there will be a clear presentation of the conclusion of this article.
3) Is there any observation regarding the application of this method to healthy cells? What happen if this technique is applied to an organism? Are the healthy cells also influenced negatively?
Author Response
- Regarding the "Statistical Analysis" paragraph, the authors have not explain the normalization process (e.g. mathematical models utilized). Please give some more information
Author response: Thank you for pointing this out. We had included this information at the end of the previous section in Lines 180-187, and on Lines 190-191.
- It would be great to include a table that clearly presents the difference resulting in the survival of cells by the implementation of each different method. This way there will be a clear presentation of the conclusion of this article
Author response: We have added Table 1 which summarises the effect of USMBs, radiation exposure and culture state on the survival of the cells following treatment. This table is seen on page 9 of the manuscript.
- Is there any observation regarding the application of this method to healthy cells? What happen if this technique is applied to an organism? Are the healthy cells also influenced negatively?
Author response: As the context for this study is around methods to enhance radiation therapy using x-rays, which is used primarily as a treatment option for cancer, we did not focus on exploring the potential for this technique to be used to treat non-cancerous cells, rather as the potential for this technique to act as a radiosensitising agent alongside other agents already under investigation [1]. However, it is an important consideration as the benefit of x-ray treatment does rely on the presence of a therapeutic window and more research needs to be done to investigate this further. For future work, such microbubbles can be conjugated with cancer markers to accumulate them in the tumour prior irradiation.
References
[1] Reda M, Bagley AF, Zaidan HY, Yantasee W. Augmenting the therapeutic window of radiotherapy: A perspective on molecularly targeted therapies and nanomaterials. Radiother Oncol 2020;150:225–35. https://doi.org/10.1016/j.radonc.2020.06.041.
