The Modulation of Phospho-Extracellular Signal-Regulated Kinase and Phospho-Protein Kinase B Signaling Pathways plus Activity of Macrophage-Stimulating Protein Contribute to the Protective Effect of Stachydrine on Acetaminophen-Induced Liver Injury
, Huang-Ping Yu 1,2,†, Hung-Chen Lee 1,2, Chun-Yu Chen 1,2 and Chia-Chih Liao 1,2,*Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThe study by Liu et al. investigated the hepatoprotective effects of stachydrine, an alkaloid from Leonurus heterophyllus, in an acetaminophen-induced liver injury mouse model. Stachydrine treatment showed promise in reducing elevated serum alanine transferase levels, pro-inflammatory cytokines, malondialdehyde activity, and inhibiting the ERK and AKT pathways, suggesting its potential as a preventive measure against APAP-induced liver damage. Some revisions are suggested to improve the quality of the manuscript.
1. The mechanistic study on the ERK and AKT pathways is superficial, the downstream markers should be measured to provide convincing data that this pathway is involved in the effect of stachydrine.
2. The study on macrophage is confusing and does not add anything. Monocyte/Macrophage is commonly infiltrated and activated in variety of liver injury. And it is not a surprise that anti-inflammatory and anti-oxidant subject could inhibit the process, more studies should be conducted on the underlying mechanism and interaction between stachydrine and macrophages.
3. In spite of the protective effect on liver injury, does stachydrine improve the survival rate of animals chanllenged with APAP?
4. In figure 3B, higher magnification as well as arrows should be used to indicate the necrotic area and the inflammatory cell infiltration.
5. Some discussions on the approval status of stachydrine, or its present clinical application are encouraged.
Comments on the Quality of English Languageminor editing
Author Response
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Author Response File: 
Author Response.pdf
Reviewer 2 Report
Comments and Suggestions for AuthorsLiu et al. showed the hepatoprotective effect of stachydrine after toxic dose administration of APAP in mice. The presented results indicated that stachydrine may be a promising beneficial target in the prevention 22 of APAP-induced liver damage through attenuation of the inflammatory response, inhibition of the 23 ERK and AKT pathways, and expression of macrophage-stimulating proteins. Please, find my comments below.
1. In the present study, the author used stachydrine (ST) concentration in the range of 2.5-10 mM concentration. Please, explain whether the selected concentration of ST is clinically relevant.
2. How was the 30-minute post-dose APAP time decided for stachydrine administration in mice? Is the 30-minute time sufficient to cause hepatotoxicity by APAP?
3. In Figure 1, I don’t see a dose-dependent increase in cell viability with an increase in ST concentration from 2 to 10 mM. Please, highlight what could be the reason. On the other hand, in Figure 2 we do see a dose-dependent decrease in p-ERK and p-AKT protein expression.
4. In Figure 2, why only 10 mM APAP and ST treatment arm compared with the APAP group? Why not 2 mM compared equally? Two different controls being used for 2- and 10-mM ST treatment analysis.
5. In Figures 3 and 5, I don’t see a dose-dependent decrease in ALT, IL-1β, and IL-6 levels with an increase in concentration of ST. Please, highlight the plausible reason.
6. In the introduction, the author mentioned ST is a major alkaloid in the Leonurus heterophyllus plant. Please, mention the % of ST in the plant.
7. Why only male mice were used in the study? Rodent usually shows sex-dependent differences in toxicity. I feel 50-50% male and female should be included in the study.
8. Please, make the referencing style consistent.
Author Response
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Author Response File: Author Response.pdf
Round 2
Reviewer 1 Report
Comments and Suggestions for AuthorsMy questions have been addressed.
