Review for 10.1186/s12876-024-03176-0 (2024)

Title: The Effects of Low-Dose IL-2 on Th17/Treg Cell Imbalance in Primary Biliary
Cholangitis Mouse Models
Version: 3 Date: 14 Dec 2023
Reviewer's report:
The manuscript titled "Therapeutic Potential of Low-Dose IL-2 Through Targeting the Th17/
Treg Axis in Primary Biliary Cholangitis Mice," authored by Wang et al., focuses on addressing
the observed imbalance in the Th17/Treg cells ratio reported in patients with primary biliary
cholangitis (PBC). This imbalance is believed to play a crucial role in the disease's pathogenesis.
The authors aim to test the hypothesis that a low dose of IL-2 can alleviate the severity of the
disease.
The study primarily employs a PBC model involving female C57BL/6 mice, subjected to 2OA-
BSA injections twice at two-week intervals and poly I:C every three days. The impact of low-
dose IL-2 therapy on the Th17/Treg ratio is assessed using flow cytometry, H&E staining of
tissue paraffin sections, and ELISA tests. Additionally, the results are analyzed in the context of
an existing RNA-seq database, which includes Ursodeoxycholic acid (UDCA) responders and
non-responders, compared to non-liver disease controls.
In my personal opinion, prioritizing the results obtained from animal subjects is crucial, given
the sacrifices made by living creatures. Furthermore, the findings hold significant importance
for the scientific community, as many potential therapeutics are often tested in vitro but may fail
to demonstrate significant effects in an animal model, thereby limiting their clinical translation.
However, I have identified several concerns in the manuscript that warrant thorough revision.
The current version is challenging to understand, and addressing these issues will enhance its
clarity and scientific rigor. I believe, the manuscript needs serious re-writing and it would benefit
from language editing service.
Major issues
- The introduction occasionally appears too superficial, containing some errors and mental
shortcuts, for instance “The Th17 and Treg cells were a pair of cells with similar developmental
pathways and opposite functions, and their imbalance is often associated with autoimmune
diseases”
- The final paragraph of the introduction should provide a roadmap for the article, which is
currently missing. Additionally, the sentence "Therefore, the present study is to investigate the
potential role of Th17/Treg cells in the pathogenesis of PBC and the regulatory role of IL-2
in the PBC mouse model" is overly broad, considering that this information is already covered
elsewhere in the manuscript. Does it effectively highlight the manuscript's key strengths?
- The mouse experiment requires a more comprehensive description. It may be beneficial for the
authors to consider relocating some figures from the supplementary information, particularly
Figure S3, which contains crucial details about the experiment, including the IL-2 dose.
Additionally, please review the spelling in the figures.
- The experiment's description lacks concise information about the substances injected at
three different time points. Specifically, when did the administration of low-dose IL-2 to the
mice commence? Please include references to prior studies where these methods have been
successfully employed.
- The statement "The control group was injected with PBS instead of 2OA-BSA and poly I:
C." raises questions about whether the control group received PBS only. This aspect should be
clarified in the Material and Methods section. Maybe the authors should consider adding a table
containing the details about each group?
- Since the figures are not embedded in the text and do not contain appropriate descriptions
underneath, it is difficult to assess how it would look like in the final text and whether it would
be correctly described. This is against the instruction for authors that states “Figure titles (max
15 words) and legends (max 300 words) should be provided in the main manuscript”
- Since “The imbalance of Th17/Treg cells was progressively aggravated with the extension of
modeling time.”, it should be shown in supplementary information.
- Figures should be numbered in the order they appear in the text; for example, Figure 3A
should precede Figure 3C. Additionally, the figures should be labeled sequentially as A, B, C,
etc. Currently, they appear chaotic, and I recommend reviewing all figures to ensure proper
numbering and labeling.
- In Figure 3A, have the authors assessed the statistical significance of the increase in Treg cell
population and the decrease in Th17 following low-dose IL-2 treatment? A similar question
applies to the bar graphs in 3D. If the answer is "yes," please indicate the significance on the
graphs.
- Figure 4 contains groups marked as “E-therapy”, “M-therapy”, “L-therapy” and “pbc”. It is
not clear what the different therapies mean. The whole paragraph should be rewritten so that the
reader understands and follows the authors analysis. Again, the description of a figure is a must.
- Discussion appears too general. For instance, the authors mention the imbalance of Th17/Treg
cells as a potential therapeutic target for diseases but omit specific disease names, rendering the
argument invalid. In my opinion, the discussion should be more concise and focused than its
current form.
Minor issues
- Several details are missing from the Materials and Methods section, including the source of
reagents used in the mouse experiments, the microscope's name, photo magnification, antibody
names, ELISA source, company, and catalogue numbers, as well as the R version and the
package source.
- When discussing the balance/imbalance of Th17/Treg, please be precise, as these are cell
populations, not individual cells. Statements like "the imbalance recovered" (lines 179-180) are
ambiguous and should be rephrased for clarity.
- Information such as "Poly I: C is an inducer of type-1 interferon (IFN). In clinical practice,
it has been found that type-1 interferon induces autoimmune characteristics in patients with
viral hepatitis [29], while 2OA-BSA, as a high-affinity reactant of AMA [30]," belongs in the
Materials and Methods or Introduction sections. Provide more details about the mouse model
used, as it is not the only model of PBC. Additionally, the discussion should acknowledge any
limitations associated with the chosen model.