Review for 10.1186/s12876-025-03940-w (2025)

Title: An elevated percentage of CD4#CD25#CD127low regulatory T cells in peripheral blood
indicates a poorer prognosis in hepatocellular carcinoma after curative hepatectomy
Version: 2 Date: 22 Mar 2025
Reviewer's report:
This manuscript explores the role of peripheral CD4+CD25+CD127low Tregs in predicting
prognosis after curative hepatectomy in HCC patients. This study is innovative, employing flow
cytometry to assess Treg levels and utilizing X-tile software to determine the optimal cutoff
value. The findings provide clinical insights into postoperative monitoring and individualized
treatment strategies for HCC patients. However, several aspects require revision to improve
the clarity, depth, and rigor of the study:1.The manuscript would benefit from professional
language editing to enhance clarity and readability. For instance, the sentence in the abstract,
“which may be a significant predictor of HCC clinical outcome”, lacks precision. A more
refined version could be “which has been suggested as a potential prognostic indicator in HCC.”
Additionally, ensure uniform formatting for P-values throughout the manuscript (e.g., “P =
0.008” instead of “P=0.0079” or “P<0.05”). 2. While the introduction provides a comprehensive
overview of HCC epidemiology and immune microenvironment, further discussion on the
controversial role of Tregs in HCC prognosis would strengthen the manuscript. Some studies
indicate that Tregs may have different prognostic implications across various cancers (e.g.,
colorectal and head and neck squamous cell carcinomas). Expanding this discussion with
relevant citations would clarify the knowledge gap this study aims to address. 3. The
methodology for flow cytometry should include more specific details regarding the reagents
and instruments used. Specify the manufacturer of the antibodies (e.g., BD Biosciences)
and the flow cytometer model. Additionally, mention whether single-staining controls were
used to confirm antibody specificity. Furthermore, provide details on the gating strategy
and data analysis software (e.g., FlowJo or Kaluza) to enhance reproducibility. 4. The
Cox regression model is appropriately applied to assess the prognostic significance of Treg
levels; however, it is unclear whether multicollinearity among covariates was evaluated. It
is recommended to report the variance inflation factor (VIF) values to confirm the absence
of multicollinearity. Additionally, clarify the variable selection method used in the Cox
model (e.g., stepwise regression). 5. Some cited studies are outdated (e.g., references [7]
and [9] from 2005 and 2020). To ensure the study is well-grounded in the latest research,
incorporate recent publications (2022–2024) from Hepatology or J Hepatology on HCC
immune microenvironment and Tregs, particularly regarding their prognostic significance and
therapeutic implications. 6.The discussion effectively highlights the role of Tregs in HCC
recurrence; however, the prognostic differences across subgroups (e.g., AFP < 400 ng/mL,
MVI-negative patients) should be explored in greater depth. Consider discussing potential
mechanisms underlying these differences, such as whether Tregs mediate recurrence through
suppressing cytotoxic T lymphocyte or NK cell activity. Moreover, while the manuscript briefly
mentions the impact of postoperative treatment on prognosis, it would be valuable to explore
whether Treg levels could serve as predictive biomarkers for immunotherapy response (e.g.,
PD-1/PD-L1 inhibitors).