Identification of Ferroptosis-Related Hub Genes Linked to Suppressed Sulfur Metabolism and Immune Remodeling in Schistosoma japonicum-Induced Liver Fibrosis

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript presents a comprehensive bioinformatics and experimental study aimed at identifying key ferroptosis-related genes involved in the development of liver fibrosis induced by Schistosoma japonicum. The study is highly relevant, as liver fibrosis due to schistosomiasis remains a significant issue, and the mechanisms linking sulfur metabolism dysregulation, ferroptosis, and immune remodeling are not yet fully understood. The manuscript is well written and the subject is worthy of investigation. The paper is technically sound and presents interesting results.  The manuscript presents some limitations, which are clearly outlined by the authors in the Discussion. Despite this, I would like to make several comments on the study. The Discussion section requires revision, with particular attention to softening strong claims (e.g., lines 469 and 512).  Additionally, the Discussion contains repetitive statements that should be minimized. The caption of Figure 7 requires correction to comply with the publisher’s recommendations. Figure 8a should include labeled histological images indicating inflammatory infiltration, fibrosis, parasite eggs and etc. I also recommended to add the scale bars to the images.

Author Response

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Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript by Xu, Xu, Ying and collaborators investigated by bioinformatics and transcriptomics the implication of ferroptosis genes in the onset of liver fibrosis induced by Schistosomia japonicum.

They retrieved from two open access data sets transcriptomics signatures from 12 S. japonicum infected and untreated mice and 2 controls. Then they checked among 1346 genes the values of 99 uniques genes correlated to ferroptosis (FRG). The authors carried out a very careful and thorough bioinformatic analysis of gene expression, gene ontology, network interaction analysis, by interrogating several specialized databases. At the end they focused their attention on 7 genes which displayed ROC equal or very close to 1.0 and acting as hub, both for ferroptosis and fibrosis.

The bioinformatics analysis is sound, as well as the experimental verification in 4 BABL/c male mice, infected with S. japonicum cercariae.

However, the authors have never explicitly cited the name of the protein/enzyme coded by each gene. Throughout they describe the 7 genes roles in the metabolic pathways, but never their exact protein function. This information is crucial to underline and strengthen the results showed in Figures 3 and 7 and in the final paragraph of the discussion. It should be given at around line 295 to help the reader through the results. In fact, in figure 3A and 3B the Authors treat separately the “pyridoxal phosphate binding” and “vitamin B6 binding”, while it is known that vitamin B6 is the precursor of PLP.

The authors have chosen only male mice as animal model to experimentally verify their findings, however it is known that females suffer from more severe fibrosis, both in human and zoonotic schistosomiasis. Which was the rationale for their mono-gender choice?

In the heat map of figure 2C, the first 6 points of the “infected” combined groups display a trend much similar to the control than to the infected: can the Authors explain this behaviour?

In none of the 4 panels of Figure 8A a scale bar is present, indicating the dimension. The legend says 200x → but without a scale this magnification is useless.

Minor points:

line 53, line 86 → write “S. japonicum” in italics

line 203: insert a space between the reference [28] and “was”

line 366: the legend has strange spacing, please verify.

Figure 7C: the choice of colours makes the reading very difficult: light orange and light red are very similar, the same for light purple and light blue. It is almost impossible to differentiate between the predicted interaction and the physical interaction, and between the co-expression and co-localization. Please use 4 more distinct colours.

Paragraph from line 527 to line 555 is redundant and does not add much to the already long discussion. Try to summarize it.

Paragraph 589 to 598 is redundant with the conclusion (lines 600-612). Keep only one of the two.

Author Response

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Author Response File: Author Response.docx