Anti-Photoaging Effect of Jawoongo via Regulating Nrf2/ARE and TGF-β/Smad Signaling in In Vitro Photoaging Model
, Shengdao Zheng 1,2, Myeongju Kim 1 and Tae-Hoo Yi 1,*Round 1
Reviewer 1 Report
The authors show the effect of Jawoongo and how it ameliorates the photoaging effects upon UVB treatment on HaCaT cells. I have some questions:
1. Have the authors tried these same experiments on other cell lines? Why HaCaT cells were only used to study the effect of photoaging?
2. Cell viability assay should be added as a panel in all the experiments to take into effect the fact that cell number is constant in all the sets.
There are some editing problem:
The version of the manuscript available for downloading had some corrections incorporated marked in red, please change those and make necessary corrections.
Author Response
1. Thank you for your point-out. In earlier dermatological studies, HaCaT cells were frequently used as a model system, especially when examining the effects of photoaging. Since photoaging of the skin served as the main symptoms of this manuscript, HaCaT cells were chosen since trials using them were believed to be sufficient to show Jawoongo's efficacy.
In order to better understand how Jawoongo impacts the dermis portion that has been activated by UV radiation, we are also setting up tests with normal human dermal fibroblasts (NHDF). Please see the publications listed below for more details regarding the study's setup. We promise to subsequently publish the results of using NHDF in a high-caliber publication.
- Kim, M.; Ha, L.-K.; Oh, S.; Fang, M.; Zheng, S.; Bellere, A. D.; Jeong, J.; Yi, T.-H. J. P., Antiphotoaging effects of damiana (turnera diffusa) leaves extract via regulation AP-1 and Nrf2/ARE signaling pathways. 2022, 11, (11), 1486. https://doi.org/10.3390/plants11111486
- Nguyen, Q. T.; Fang, M.; Do, N. Q.; Jeong, J.; Oh, S.; Zheng, S.; Kim, M.; Choi, J.; Lim, S.; Yi, T. H. J. A., Anemopsis californica Attenuates Photoaging by Regulating MAPK, NRF2, and NFATc1 Signaling Pathways. 2021, 10, (12), 1882. https://doi.org/10.3390/antiox10121882
2. I appreciate your suggestion. Additionally, we wholeheartedly concur with the reviewer that the experiment's cell survival rate should remain consistent. It was consistently validated using microscopy that the cell condition and survival rate were stable during the three repetitions of the tests. Given that the identical cells, UVB radiation, sample concentration, and culture conditions were used in each trial, we guarantee that no two studies will yield results that differ from the cell viability test results presented in the text. Line 123 describes the seeding of the HaCaT cell in order to provide more precise information.
We deeply appreciate all your help in revising this manuscript. Thanks to the reviewer, we were able to make a much better manuscript. It was such an honor to receive a review from you.
Reviewer 2 Report
The paper presents data that a preparation of Jawoongo (JAUN) inhibits UVB induced damage and sequelae to HaCat cells. The results are convincing that this particular preparation has activity in cell culture. However, the method of preparation is not described sufficiently so that others can reproduce the work. There are 13 botanicals listed, but no description of their source, leaf or root or manufacturer (Dongkwang is a distributor not a manufactuer). Were 10 g of each included in the 130 g? What does 19.23% refer to (w/w)?
The authors state no conflict of interest but 2 authors are affiliated with a cosmetic ingredient manufacturer (Snow White Factory, Korea) and support was provided by J1 Cosbio Co. Ltd - no listing is found in a Google search. Who are these companies and why is their support not a conflict of interest? A more transparent disclosure is needed.
Author Response
Please see the attachment.
Author Response File: 
Author Response.docx
Reviewer 3 Report
I had an opportunity to review a manuscript regarding Jawoongo, a herbal extract of antioxidative properties.
This article analyses the efficacy of herbal extract on cells in vitro. The results are interesting and encourage to perform further research in vivo. The authors consequently described the process and provided the original data. The results are clearly presented.
Please find my comments below. I suggest discussing the following points:
1. Incubation time: how was the length 1h justified? could prolonging/shortening this period significantly influence the results?comparisonn to literature?
2. Antioxidative properties of active ingredients: how strong are "pure" baicalin, shikonin, and decurin? are these available in other herbal/artificial products? are any reports regarding their efficacy in other concentrations?
3. Plants used to obtain Jawoongo: were the plants for extraction cultivated in the lab or already bought grown up (bio/uncontrolled)? are there any special requirements for cultivation (e.g. fortified soil)?
5. Citations are appropriate yet not comprehensive. In-depth literature review is highly encouraged to underline the significance of antioxidative medications and to compare Jawoongo extract to other antioxidants.
6. Limitations of the study not listed.
Author Response
Please see the attachment.
Author Response File: Author Response.docx
Reviewer 4 Report
Comments for the Author (Required):
The study by Kim H et al, titled “Anti-Photoaging Effect of Jawoongo via Regulating Nrf2/ARE 2 and TGF-β/Smad Signaling in In Vitro Photoaging Model” aims to study the beneficial effect of Jawoongo on UVB mediated photodamage in HaCaT cells. Although this is an interesting article, I have some concerns about this article.
Specific comments
Page 2, Nrf2 is transferred from the cytosol …… “heme oxygenase-1 (HO-1) production of antioxidant enzymes” to remove harmful quinones and relieve oxidative stress. - heme oxygenase-1 (HO-1) production of antioxidant enzymes- what does it mean? – reformulate.
Page 2. It is primarily composed of Lithospermum erythrorhizon and Angelica gigas, with addition and reduction. What is addition and reduction? – reformulate.
Page 3, line 137. HCI and CaCI2- It should be HCl and CaCl2
Page 4, line 143. Tris-HCI- It should be Tris-HCl
Page 6. The results showed that the administration of JAUN led to a significant and dose-dependent decrease in ROS secretion. Author measured intracellular ROS levels, but it is reported as ROS secretion. How?
It would be nice to discuss the interconnection between ROS and MAP kinase / ROS and Smad, TGF beta expression in the discussion section (as reported by others Free Radic Biol Med. 2019 Nov 1;143:275-287. Free Radic Biol Med. 2008 Oct 1;45(7):1035-44)
Minor editing of English language required
Author Response
1. Thank you for your suggestions. Corresponding parts of the manuscript is now fully reformulated (Line 49-52).
‘Nrf2 is transferred from the cytosol to the nucleus to activate ARE-dependent genes and promote the production of antioxidant enzymes such as NAD(P)H quinone oxi-doreductase-1 (NQO1), dihydrolipoamide dehydrogenase (DLD), and heme oxygen-ase-1 (HO-1) to remove harmful quinones and relieve oxidative stress.’
2. Thank you for your advice. The additions and reductions in the article refer to the continuous improvement of Jawoongo in its application from ancient times to the present, but all improvements are inseparable from the two basic raw materials of Lithospermum erythrorhizon and Angelica gigas. The improvements to the Jawoongo prescription in this experiment are described in detail in 2.2. Sample preparation.
3. Thank you for your point. It was corrected immediately.
4. Thank you for your point. It was corrected immediately.
5. Thank you for your questions. We have modified the corresponding parts of the article.
6. Thank you for your recommendations. We immediately added a section on the association to each paragraph. Please refer to lines 322 - 323, 329 – 334, and 342 - 344.
We deeply appreciate all your help in revising this manuscript. Thanks to the reviewer, we were able to make a much better manuscript. It was such an honor to receive a review from you.
Author Response File: Author Response.docx
Round 2
Reviewer 3 Report
Thank you for revising your manuscript and replying to my suggestions.
Nevertheless, besides answering my comments, the authors have not provided any additional references nor added limitations of the study. A separate response to my inquiries in a private brief unfortunately fails to make much improvement to the manuscript.
Author Response
I appreciate your response. The references have been completely updated (the form; newly added references 28 and 29). On the reviewer's recommendation, the limitations of these studies are now thoroughly described in the conclusion section.
We apologize that the revision did not satisfy your needs, thus we have revised the paper once more. Because of the reviewers' insightful criticism, we were able to improve our manuscript.
