Talk:WikiJournal of Medicine/Extract of Laurus nobilis attenuates inflammation and epithelial ulcerations in an experimental model of inflammatory bowel disease
WikiJournal of Medicine
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DOI: 10.15347/WJM/2023.002
QID: Q102344224
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Suggested citation format:
Natalie S. Correa; Robert A. Orlando (20 January 2023). "Extract of Laurus nobilis attenuates inflammation and epithelial ulcerations in an experimental model of inflammatory bowel disease". WikiJournal of Medicine 10 (1): 2. doi:10.15347/WJM/2023.002. Wikidata Q102344224. ISSN 2002-4436. https://upload.wikimedia.org/wikiversity/en/c/c1/Extract_of_Laurus_nobilis_attenuates_inflammation_and_epithelial_ulcerations_in_an_experimental_model_of_inflammatory_bowel_disease.pdf.
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This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction, provided the original author and source are credited.
Alaa Najjar 
(handling editor) contact
Athikhun Suwannakhan (handling editor) contact
Article information
Plagiarism check
Pass. Report from WMF copyvios tool: Flagged terms, like “Inflammatory bowel diseases (IBD), including Crohn's disease and ulcerative colitis”, “both acute and chronic colitis” and “DSS, 40,000 Da, 3.5% (wt/vol), ICN Biochemicals” were appropriate. --Alaa :)..! 19:24, 3 December 2020 (UTC)
Editorial note
Contacted handling editor to see if they can make any progress with reviewers Rwatson1955 (discuss • contribs) 10:17, 21 June 2022 (UTC)
First peer review
Review by anonymous peer reviewer , Published papers on inhibition of breast and colorectal cancer cells
These assessment comments were submitted on , and refer to this previous version of the article
- In the last paragraph of the introduction, hypothesis and main objective of this study should be stated. It is not clear at the moment.
We have included a new sentence in the last paragraph of the introduction to directly address our hypothesis and main objective of this study.
- In materials and methods, the subsection “Standardized preparation of herbal tea extracts” should be changed to “Preparation of Laural Nobilis extracts”
We have made this change to the subsection.
- The authors mentioned about Day 0 in the methods section, but in the results section did not show ana results on Day 0, particularly Figure 1.
The results obtained on day 0 were used as a measure of starting values to calculate percent change from the beginning of treatment (either DSS or DSS+Laurel extract), this being Day 0. A typo was corrected in the figure legend to make this point clear--originally the figure legend read "day 1", this was an error and now correctly reads "day 0".
- In the results section, in my opinion its first paragraph is more likely a discussion. Please consider moving it to the discussion. Otherwise remove it.
This comment was also raised by Reviewer 2. We agree with both Reviewers and have moved the first paragraph in the Results section describing the DSS murine model to the first section of the Discussion section.
- For Figure 2 which represents the length of colon, why the bar for “no tx” group does not have error bars. Please add or clarify.
Three animals were included in the "no tx" group and did not receive DSS or LN extract. The standard deviation for colon length for these three animals measured less than 1mm. This small standard deviation resulted in error bars too small to visualize on the graph.
- Arrowheads should be added to Figure 3 to make readers understand the area that you would like to point out more easily. In this figure, author would like to show the attenuation of leukocyte infiltration and morphological damage.
Arrows have been added to fig 3 as suggested by the Reviewer, as well as scale bars and magnifications as requested by Reviewer 2.
- In Figure 4, I suggest that author change it into colors with arrows. Also, I suggest quantify the finding. Software such as ImageJ can measure the relative intensity which can be presented quantitatively. Please also add the methods section after doing so.
We have changed the image to conventional green corresponding to the FITC-secondary antibody conjugate. We considered using ImageJ software to quantify the immunofluorescence data, however, after changing the color to conventional green the data shows more clearly the infiltration by CD4+ cells. Based on this finding, we do not believe that ImageJ quantification would add additional information to the result beyond what is presented visually. We have also added arrows to fig 4 to highlight our positive immunofluorescence results.
General questions:
- Why did the authors choose 200µl concentration of laurel extract for their experiment in mice? What is the rationale behind it? Please clarify.
- What is the solution used to dissolve/dilute the extract (water, PBS or etc.)? This should be added to the materials and methods section?
- What is the sex of the mice used in this experiment?
We chose 200uL as the gavage volume due to prior experience with the most active amount of LN extract in our in vitro assays. We have included the following sentence to the Methods section to clarify our rationale: "A volume of 200 μL was chosen because this amount of LN extract demonstrated maximal activity in our previous in vitro assays[21] and allows for an acceptable volume that does not cause unnecessary distress to mice when orally administered." The extract was prepared using ultrapure water. This information is included in Methods. The mice used in this study were all male. This information is included in Methods.
Second peer review
Review by anonymous peer reviewer ,
These assessment comments were submitted on , and refer to this previous version of the article
The manuscript presents potential effect of Laurus nobilis (LN) that attenuates inflammation and epithelial ulcerations in Inflammatory Bowel Disease (IBD). This work contributes the part of current data carried out by in vivo experiment to prove the anti-inflammatory activities of LN in the colonic tissues, though its wide metabolic benefits have been studied both in vitro and in animal models.
The manuscript is well-descripted to the methodology and related results. Here are some minor points for further improvement: The active ingredients in these extracts are stable. Consistent values were obtained in our bioactivity assays after freeze-drying and reconstituting the extract. Did the authors have previous or current data referencing the use of freezing-thawing extract in this manuscript or from elsewhere? In the current data, which form of the extract was applied to the mice?
Yes, the freeze-thaw results were examined in our previous work referenced in citation #21. We have added this citation to the sentence in Methods under subheading "Preparation of Laurus nobilis extracts". We have also added the words, "Freshly prepared" to the description of Laurel extract administered to animals in Methods, subsection "Animal treatments".
It is wise to add brief description of statistical analysis in Materials and Methods.
A description of statistical methods is included in figure legends where needed.
The first paragraph of Result is likely placed in Discussion.
We have moved the first paragraph of Results to the beginning of Discussion as suggested.
Figure 3 representing the micrographs of randomly selected parts of the colonic tissue. By reliability and standard interpretation, all micrographs in each panel (A-C) require proper scale bar and annotations. Image panels of each figure do not seem to be recorded at the same magnifications. That may be tricky to indicate the clear epithelial change from one to the others. For instance, image in Figure 3B indicates the epithelial damage, including ulcerations. Did the authors find breach of epithelial covering with some vascular damages? Figure 3A (control animals) do not show the appearances of villi. Murine colon shouldn’t have villi in colon. Figure 3B (DSS treated animals) shows tissue ulcerations, extensive neutrophil infiltration into the mucosal layer, and granuloma formation. Did this appearance have a proof of chronic inflammation? Did the authors demonstrate epithelioid cells surrounded by T-cells in the image panel? Figure 3C (DSS + LN-treated animals) shows almost exact appearance of the colonic tissue to the control group. It is such a surprising phenomenon that there are no remaining of damaged mucosa or granuloma, while in lamina propria is clear from infiltrated lymphoid. That potential effect of LN is very impressive. The last paragraph of Result has one typo, cryosections.
We have added scale bars to fig 3 and indicated magnifications to the accompanying figure legend. We did not find epithelial breach or significant vascular damage likely due to our need to terminate the animal treatments prior to seeing such significant damage. The DSS model is very aggressive and requires close monitoring of animal wellbeing and typically early experimental termination to comply with our animal welfare protocol. We definitely agree with Reviewer 2 that the substantial effect of LN extract preventing severe colonic damage is remarkable. The LN extracts were given to the animals in parallel with DSS treatment, rather than subsequent to DSS-induced colonic damage, thereby likely providing a protective effect against DSS pathologies. Testing the properties of LN extract to reverse DSS-induced damage was not possible due to the severe, short-term aggressive progression of DSS damage. Also, we have corrected our error as indicated by the Reviewer and removed the word villi and replaced it with "colonic folds".
Result of immunofluorescence analysis should be presented in color micrographs. It is worth representing the positive reactivity of CD4+ antibody in the frozen tissues. Was the immunoreactivity localized in mucosal cells? That may indicate the CD4+ T helper cell activation.
We have replaced the original figure 4 with a color image better reflecting the FITC-conjugated secondary antibody used for this experiment. The color image better shows immunoreactivity and the presence of substantial CD4+ cell infiltration into the mucosal layer. This is also better described by the addition of arrows pointing out immunoreactivity near the epithelial layer.
Discussion is rather weak point of this manuscript. The authors should carefully consider the similar finding or in related potential effect of other herbs to the inflamed tissues. The second paragraph of the manuscript seem to be fit in the Background. Deep down, we all believe that LN could provide an anti-inflammatory activity in one way or another when sufficient data with clinical trial are evident.
We have elected to limit our Discussion primarily to the results we observed in this study and also offer a brief description of the components found in Laurel extract by other studies. We believe the next step in this study is to fractionate the Laurel extract to identify the primary active compound(s) that offer the potent anti-inflammatory activity we report in this study. We believe the numerous published reviews defining anti-inflammatory activities of natural products offer a better platform to describe the wonderful healing powers of natural compounds that are so often overlooked.
Robert A Orlando, PhD (discuss • contribs) 21:53, 28 December 2022 (UTC)