Review Reports

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Reviewer's Comments on the Manuscript by Kyung-Hyun Cho et al.

 

The manuscript presents a cross-sectional analysis of lipoprotein characteristics and blood pressure in a middle-aged Korean cohort. While the authors have conducted extensive laboratory measurements, the study suffers from fundamental limitations that severely undermine its potential contribution to the field. The core premise-that HDL quality is crucial-is not novel, and the presented data, in its current form, largely confirms existing paradigms without providing new mechanistic insight or robust, independent associations.

 

Major Concerns:

1.Lack of Conceptual Novelty: The investigation is framed around the well-established hypothesis that HDL functionality is more important than HDL-cholesterol quantity in cardiovascular risk. This is a current consensus in the field, as thoroughly referenced by the authors themselves. The study applies established techniques (measurement of PON, FRA, glycation, size) to a new sample but fails to ask or answer a novel scientific question. The findings do not challenge, refine, or significantly advance the existing model; they primarily provide a new dataset consistent with it.

 

2.Limited Mechanistic Insight and Superficial Discussion: The discussion does not move beyond a summary of the results and a restatement of known literature. It lacks depth in exploring how the observed alterations in lipoprotein glycation or antioxidant activity might mechanistically lead to hypertension. The study remains descriptive at the phenotype level, offering no novel insights into molecular pathways, signaling events, or physiological mechanisms linking dysfunctional HDL/LDL to blood pressure regulation. This severely limits its value to the research community.

 

3.Study Design Constraints That Inherently Limit Impact: The combination of a small sample size (n=50) and a cross-sectional design confines the study to generating hypotheses, not testing them. The authors cannot establish temporality or causality. While such studies have value in early-phase research, their conclusions must be exceedingly cautious.

Author Response

Thank you for your insightful comments. Following the reviewer’s suggestion, we made point-to-point response and reflected on revision.

Please find attached file as our response.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors,

The present manuscript analyzes how HDL quality (namely composition, particle morphology/size, glycation extent, and antioxidant functions such as PON and ferric ion reducing ability) relates to blood pressure and LDL oxidation or glycation in 50 middle-aged Korean volunteers (in this study 25 male, 25 female individuals) recruited via nationwide advertisement. Interestingly, the authors report sex differences in BP and adiposity (e.g., higher SBP/DBP and BMI/visceral fat in males) and use correlation/regression analyses to connect LDL particle size and glycation/oxidation to SBP/DBP, and HDL2/HDL3 functional measures (PON, FRA) to LDL oxidation/glycation readouts. In reviewer’s opinion, this is an interesting study that moves the field significantly forward and can be considered for publication provided the authors address the major concerns raised:

Major Comments:

1. State the study design explicitly as cross sectional and observational in both the Abstract and Discussion, so readers could understand that the analyses are based on a single time point and do not test directionality. Accordingly, revise the wording of key conclusions to describe the findings as associations or correlations rather than implying cause and effect or definitive mechanisms, particularly when discussing HDL2 and HDL3 roles and “hypertension risk” reduction.
2. Clearly define all blood pressure terminology and categories used in the study, including the exact cutoff values and guideline source for terms such as “prehypertension,” “hypertension,” and “normotension”. In addition, use the same blood pressure language consistently throughout the manuscript so the reader can easily follow how participants were classified and how BP outcomes were interpreted.
3. Standardize terminology for key measurements across the entire manuscript to avoid ambiguity. For example, the fluorescence-based glycation readout should be referred to using a single, consistent term (such as “glycation extent” or “AGE-like fluorescence”), and it should be defined clearly in the Method section (including the assay principle, excitation/emission settings, and what the signal is intended to represent). Use the same term in the Results, Figure legends, and Discussion, and avoid switching between different labels that could misinterpreted as different assays.
4. The authors are suggested to soften the mechanistic language in both the schematic figure and discussion sections so that the proposed HDL2 and HDL3 “roles” are presented as a working, hypothesis-generating model based on correlational associations rather than as established causal mechanisms. In particular, statements implying that HDL2 or HDL3 directly “drives,” “inhibits,” or “prevents” small-dense LDL formation or hypertension should be revised to phrasing such as “is associated with,” “is consistent with,” or “may reflect,” and the figure legend should explicitly note that the schematic summarizes an interpretive model derived from cross-sectional correlations. This will better align the narrative with the study design and reduce the risk of over-interpreting observational findings.
5. For reproducibility, add more methodological detail for the key quantitative readouts. For particle sizing, clarify exactly how size was measured (e.g., TEM image analysis workflow), how many particles were quantified per sample or per participant, how many images/fields were analyzed, whether selection and measurement were performed blinded to group or BP status, and what software and criteria were used (including any thresholding, exclusion rules, or binning strategy). For the antioxidant activity assays, provide complete reaction conditions and reporting units, including reagent identities and final concentrations, buffer composition and pH, temperature, incubation time, sample volume or protein amount loaded, and how activity was calculated and normalized; also double-check that all concentrations and units are correct and consistently reported across Methods, figure legends, and Results.
6. Add a short transparency statement noting that the study performs multiple correlation analyses and should be interpreted as exploratory and hypothesis generating, and that formal multivariable adjustment was not performed. This can be framed neutrally, for example by stating that the analyses provide an initial map of associations that motivates future studies with larger cohorts and prespecified models, without presenting it as a major barrier to publication.
7. The TEM images are visually compelling and informative, but they are currently presented in a largely semi-quantitative manner. Include clearly visible scale bars consistently on all TEM panels and specify how particle size was quantified, including the number of particles analyzed per sample and how those particles were selected for the analysis.

Minor comments

1. The title is overly long and grammatically awkward. Suggested revision (example): HDL Particle Size and Antioxidant Function Are Associated with Lower Hypertension Risk via Reduced Small Dense LDL Formation in Middle-Aged Korean Adults
2. The Introduction is quite long and partly repetitive; the main concepts (HDL quantity vs quality, sdLDL, glycoxidation) could be more concisely framed.
3. Several sentences are grammatically imprecise (e.g., “as a bystander to neglect…”). Repeated phrases such as “remarkably,” “notably,” and “interestingly” should be reduced. Some sentences exceed 40–50 words and should be simplified. A careful editing would help readability.
4. The authors should tone down overstatements such as sdLDL being “a genuine culprit to cause almost all atherogenic diseases” and ensure claims are appropriately referenced and scoped.
5. Clarify whether blood samples are serum vs plasma for lipoprotein isolation: blood was collected “without… anti-coagulant” yet density-gradient ultracentrifugation is typically described for plasma/serum explicitly.
6. Descriptives should generally be mean ± SD rather than SEM for participant characteristics (SEM can make variability look artificially small). In this manuscript its reported as mean ± SEM.
7. Please cite the specific guideline/consultation relevant to Asian BMI cutoffs and ensure wording is accurate.
8. Several places interpret nonsignificant trends as meaningful (e.g., “As shown in Figure 7, glycation extent in HDL3 was positively associated with elevation of SBP (r=0.402, p=0.004) and DBP (r=0.428, p=0.002), while TG content in HDL3 was also positively associated with elevation of BP, although statistical significance was not detected. These results suggest that an increase in glycation extent in HDL3 is a reliable risk marker for hypertension than an increase in TG content.”). Please tighten wording to match the statistics.
9. Several figures and analyses appear to use fewer participants than the full cohort (for example, N = 47 in Figures 3a, 3b, 5a, 5b, 7c, and 7d, and in some supplementary panels), but the manuscript does not explain why data are missing. These panel’s report total cholesterol and triglyceride levels measured in individual lipoprotein fractions, including LDL, HDL2, and HDL3.
10. Ensure every supplementary figure has a complete caption, consistent numbering, and clearly labeled panels that match the main text references. Notably, the main manuscript refers to four supplementary figures, but the supplementary file currently includes only three.

Major Revision Required

This manuscript has strong mechanistic depth and aligns well with the evolving concept of HDL quality over quantity. However, statistical rigor and clearer framing of limitations are strongly suggested before publication in IJMS.

Comments on the Quality of English Language

There is no major issue with the English grammar or framing of sentences, except the title that appears too long and awkward. Suggestions for some minor improvements have been indicated in the comments to authors.

Author Response

Thank you for your insightful comments. Following the reviewer’s suggestion, we made point-to-point response and reflected on revision.

Please find attached file as our response.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The study explores the relationship between HDL quality/functionality and hypertension risk in a middle-aged Korean cohort. While the focus on HDL quality rather than just quantity is timely, the manuscript has several critical data inconsistencies and methodological flaws that must be addressed before further consideration.

1. Contradictory Inclusion Criteria: The exclusion criteria state that individuals on treatment for hypertension were excluded. However, the Results and Figures explicitly compare "normotensive" vs. "hypertensive" participants. The author must clarify if the study population included undiagnosed hypertensives or if the exclusion criteria were violated.
2. Overinterpretation of Cross-Sectional Data: This is a cross-sectional study, which can only establish associations. The author frequently uses causal language, such as "HDL quality... could protect LDL" or "lowering hypertension risk". These must be softened to "associated with" or "correlated with.

Author Response

Thank you for your insightful comments of round 2.

Following the reviewer’s suggestion, we made point-to-point response and reflected on our second revision.

Please find attached doc as our response.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Thank you for the point-wise response to the concerns raised over the manuscript. The manuscript in the present revised form is much clearer in its framing and presentation. Revised and expanded methodologies would be helpful for the readers to understand the rationale of the study and ensure reproducibility. The revised schematic appropriately presents your model of study. With these revisions in place, I consider the manuscript can be accepted for publication.

Thank you!

Round 3

Reviewer 1 Report

Comments and Suggestions for Authors

The revised manuscript adequately addresses the previous concerns and is suitable for publication.