An Edible, Decellularized Plant Derived Cell Carrier for Lab Grown Meat

Round 1

Reviewer 1 Report

The article discusses the viability of using broccoli florets as potential scaffolds for carrying cells. This is an emerging area of research, and the article is well-written and well thought out. The following few suggestions could be incorporated to improve comprehension, 

1. To an unaware reader, it might be helpful to provide a generalized schematic in the introduction to show the different steps involved in producing the final edible meat product. In the figure, it would be useful to highlight what part of the process this paper focuses on. (Figure 5 can be incorporated into the new figure to and be presented up front)
2.  It is unclear to me how these broccoli florets will be incorporated into the final meat product. 
3. Line 110 - Please correct the capitalization in "Primary Satellite Cells"
4. Line 116 - CO2 needs to be CO₂
5. Line 120 - Colon needs to be removed
6. Line 128, 170 - H₂0 needs to be H₂O
7. Line 167 - Decellularized needs to be decellularized
8. Line 173 - Scaffold Size Distribution (Check Capitalization)
9. Line 228 - Check paragraph spacing under "Statistics"
10. Results Section - Needs to be justified (formatting)
11. Line 326 - The use of the word "patent" is unclear
12. Figure 6 - How does the density of decellularized florets compare to other alternatives investigated in the meat industry?
13. Conclusions - Needs to be justified (formatting)
14. Conclusion - Please summarize findings 

Author Response

1. To an unaware reader, it might be helpful to provide a generalized schematic in the introduction to show the different steps involved in producing the final edible meat product. In the figure, it would be useful to highlight what part of the process this paper focuses on. (Figure 5 can be incorporated into the new figure to and be presented up front)

We have created a graphical abstract that is now included at the end of the introduction.

1. It is unclear to me how these broccoli florets will be incorporated into the final meat product. 

We have added a description as to how the scaffold lends itself to an unstructured product and may serve as a bulking agent, stabilizer, and because of its cellulose fiber, a fortifying agent.

1. Line 110 - Please correct the capitalization in "Primary Satellite Cells"

Corrected

1. Line 116 - CO2 needs to be CO₂

Corrected

1. Line 120 - Colon needs to be removed

Corrected

1. Line 128, 170 - H₂0 needs to be H₂O

Corrected

1. Line 167 - Decellularized needs to be decellularized

Corrected

1. Line 173 - Scaffold Size Distribution (Check Capitalization)

Corrected

1. Line 228 - Check paragraph spacing under "Statistics"

Corrected

1. Results Section - Needs to be justified (formatting)

Corrected

1. Line 326 - The use of the word "patent" is unclear

Reworded

1. Figure 6 - How does the density of decellularized florets compare to other alternatives investigated in the meat industry?

We have added a greater explanation that communicates that there are yet to be any cell carriers marketed specifically for the meat industry. Because there have been materials that are suggested, such as polysaccharides, crosslinked cellulose, and polypeptides, we have featured carriers derived from those materials in figure 6. We have improved our explanation of figure 6 to communicate that the carriers outlined , cultispher 3, cytodex I & III, and cytopore are composed of those materials.

1. Conclusions - Needs to be justified (formatting)

Corrected

1. Conclusion - Please summarize findings 

We have added a statement better summarizing our findings

Reviewer 2 Report

Please see the attachment.

Comments for author File: Comments.docx

Author Response

We appreciate the reviewer’s comments and advice, and we are confident that it will ultimately improve the paper. Our responses to the comments are bolded below:

1. The aim and importance of the study should be better emphasized in the introduction section.

Additional comments on the importance of this study were added to the final paragraph of the introduction section. These comments served to drive home the fact that cultured meat will not become accessible and will not contribute to fighting climate change until their production costs are decreased. (lines 85-87)

2. In the introduction section (r.73-75) the authors statement: “Among plants, broccoli florets have many of the desired characteristics of a cell carrier for cellular agriculture; however, the interactions between native plant cells and seeded mammalian cells remain unknown” require some references, for example

https://doi.org/10.15835/nbha47311490, in the context of improving the nutritional quality of plants and biofortification of broccoli sprouts in order to boost value added output, such as improved nutrition and food functionality.

It would be very interesting to couple the growth of mammalian cells with pre-fortified plant scaffolds, to add further nutritional support. A cited statement on this potential branch of further research has been added to the discussion section. (Line 460). The statement that is quoted serves to communicate the need for decellularization, rather than to discuss the potential for further improving the nutritional quality of the broccoli. The intracellular contents of fresh non-decellularized plants may adversely effect the growth of mammalian cells. We seek to eliminate the potential for such interactions by decellularizing the florets.

3. In the discussion section the statement “Such processes will drive up the cost of manufacturing for a final product” should be accompanied by the evaluation of costs related to proposed procedure described in the manuscript. An evaluation of the costs/benefits would be interesting….

Two additional statements were added to the paper and cited respectively. The first statement refers to the loss of cell biomass and cell harvest efficiency associated with a trypsinization step. The second statement cites the bulk cost of trypsin.