Sex- and Exercise-Dependent Modulation of Hypertrophic Remodeling by the MCT1 rs1049434 Polymorphism

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

To the Authors:

The subject of the work is interesting, but it is not performed thoroughly. Additional tests, including broad genetic and metabolic analyses, should be performed to explain the main subject.

Remarks relating to:

The Introduction

The author described the clinical significance of the monocarboxylate transporter 1 (MCT1) and provided information on the clinical relevance of the genetic rs1049434 polymorphism (T1470A; Asp490Glu) in hypertrophic cardiomyopathy (HCM), but there is no information on the gene encoding MCT1.  The background on the influence of other polymorphisms in this gene on HCM, including their relationship to gender, requires more detailed study.

Is there only one polymorphism in the SLC16A1 gene with an HCM clinical impact?

There is also no information on why the authors chose only one of the polymorphisms in the SLC16A1 gene over hypertrophic cardiomyopathy, which could explain energy metabolism and lactate utilization efficiency.

 

The Material and Methods

The material and methods are presented insufficiently; for example, no details of the methods used in this study are provided for genotyping the SLC16A1 gene. This should be provided.

There is no testing of lactate levels or any additional serum test results to evaluate acidosis in the tested subjects.

In the “Discussion” section, lactate levels in women are presented from the literature; therefore, it is valuable to present these test results.

 

General remarks

The gene abbreviations presented in this study should be written in italic type.

 

 

 

Comments for author File: Comments.pdf

Author Response

Response to the Reviewer 1

We thank the Editor and the three Reviewers for their constructive comments. We have revised the manuscript accordingly, strengthening methodological detail (genotyping and exercise definition), clarifying interpretation of lactate-related statements, improving readability of the male athletic subgroup description, adding recommended references, and ensuring consistent formatting (italicization of gene symbols). Genotype distribution data for the full cohort and the HCM subgroup are provided in Supplementary Tables S1–S2.

REVIEWER 1

Introduction

Comment 1: The author described the clinical significance of the monocarboxylate transporter 1 (MCT1) and provided information on the clinical relevance of the genetic rs1049434 polymorphism (T1470A; Asp490Glu) in hypertrophic cardiomyopathy (HCM), but there is no information on the gene encoding MCT1.  The background on the influence of other polymorphisms in this gene on HCM, including their relationship to gender, requires more detailed study.

Response: We thank the reviewer for this important comment. We have revised the Introduction to explicitly state that MCT1 is encoded by the SLC16A1 gene. In addition, we have expanded the rationale for prioritizing the rs1049434 (T1470A; Asp490Glu) polymorphism by emphasizing that, among the multiple variants described within SLC16A1, rs1049434 is the most extensively functionally characterized and the one most consistently associated with altered lactate transport, substrate utilization, and glycolytic strain across independent cohorts and experimental settings.

We also briefly acknowledge the existence of additional SLC16A1 polymorphisms that have been linked to interindividual variability in lactate metabolism, particularly in athletic populations, while noting that many of these variants show context-dependent effects or limited functional annotation. A representative reference has been added to support this point (Maculewicz et al., Genes, 2025).

These additions clarify both the genetic background of MCT1 and the biological justification for focusing on rs1049434 without extending beyond the scope of the present study

 

Comment 2: Is there only one polymorphism in the SLC16A1 gene with an HCM clinical impact?

Response: We thank the reviewer for this question. At present, rs1049434 is the only SLC16A1 polymorphism for which reproducible functional data and a plausible link to myocardial energetic stress exist, supporting its evaluation as a potential disease modifier in hypertrophic cardiomyopathy. Although several additional SLC16A1 variants have been reported—mainly in the context of exercise physiology and athletic performance—there is currently no evidence supporting a direct clinical impact of these variants on HCM phenotype or penetrance. For this reason, and to avoid speculative interpretation, the present study focused on rs1049434, while briefly acknowledging other polymorphisms with limited or context-dependent effects.

 

Comment 3: There is also no information on why the authors chose only one of the polymorphisms in the SLC16A1 gene over hypertrophic cardiomyopathy, which could explain energy metabolism and lactate utilization efficiency.

Response:  We thank the reviewer for this comment. The selection of rs1049434 was based on its unique combination of functional characterization, reproducibility, and biological plausibility in relation to myocardial energy metabolism. Among the polymorphisms described within SLC16A1, rs1049434 is the only variant for which consistent evidence demonstrates a direct effect on MCT1-mediated lactate transport, substrate utilization, and glycolytic strain—mechanisms directly relevant to energetic inefficiency in hypertrophic cardiomyopathy.

Other SLC16A1 polymorphisms have been reported, mainly in exercise physiology studies, but their effects are often context-dependent, lack functional validation at the cellular level, and have not been linked to myocardial remodeling or HCM phenotype. For this reason, and to avoid speculative associations, we focused on rs1049434 as the most biologically justified candidate to explore a potential metabolic modifier role in sarcomere-positive HCM. This rationale has now been clarified in the Introduction.

Changes in manuscript (Introduction): Monocarboxylate transporter 1 (MCT1), encoded by the SLC16A1 gene, facilitates the transmembrane transport of lactate and other monocarboxylates, enabling metabolic coupling between glycolytic and oxidative tissues through the lactate shuttle [6,7]. Among the multiple single-nucleotide polymorphisms described within SLC16A1, the common rs1049434 variant (T1470A; Asp490Glu) has been the most extensively characterized from a functional standpoint. This polymorphism affects MCT1-mediated lactate transport and has been consistently associated with altered lactate kinetics, substrate utilization, and increased glycolytic strain during exhaustive exercise [8–11].

Although several additional MCT1 variants have been linked to interindividual differences in lactate production and clearance in athletic populations, most show limited functional annotation or context-dependent effects, often emerging only in large, performance-based cohorts or under specific experimental conditions (e.g., high-intensity anaerobic protocols) [12]. In contrast, rs1049434 has repeatedly demonstrated robust as-sociations across independent cohorts and experimental settings, supporting its prioritization as a biologically relevant modifier of lactate handling. While these effects have been characterized primarily in skeletal muscle, their relevance to myocardial remodeling and energetic stress in genetically determined cardiomyopathies remains uncertain.

 

Material and Methods

Comment 1: The material and methods are presented insufficiently; for example, no details of the methods used in this study are provided for genotyping the SLC16A1 gene. This should be provided.

Response: We thank the reviewer for this comment and agree that additional methodological detail was required. We have revised the Materials and Methods section to include a clear description of the genotyping procedure. Specifically, we now report the use of dried blood spot sampling obtained by finger prick, DNA extraction using the QIAamp® DNA Mini Kit, and genotyping of the SLC16A1 rs1049434 polymorphism by real-time PCR using a TaqMan® SNP Genotyping Assay on a StepOne™ Real-Time PCR System, including appropriate quality controls. These additions improve methodological transparency and reproducibility.

Changes in manuscript (Materials and Methods;  Section 2.3.: Genetic analysis): Genomic DNA was extracted from dried blood spot samples obtained by finger prick using the QIAamp® DNA Mini Kit (Qiagen, Hilden, Germany), following the manufacturer’s instructions. Genotyping of the SLC16A1 (MCT1) rs1049434 polymorphism (T1470A; Asp490Glu) was performed by real-time polymerase chain reaction using a TaqMan® SNP Genotyping Assay (Applied Biosystems, Foster City, CA, USA) on a StepOne™ Real-Time PCR System (Applied Biosystems). Appropriate positive and negative controls were included on each plate to ensure genotyping accuracy. Genotype and allele frequencies were tested for Hardy–Weinberg equilibrium.

 

Comment 2: There is no testing of lactate levels or any additional serum test results to evaluate acidosis in the tested subjects.

Response: We thank the reviewer for this important observation. We acknowledge that direct measurements of circulating lactate levels or systemic acid–base status were not performed in this study. This was a cross-sectional observational investigation conducted in a clinical setting, and the assessment of dynamic lactate responses would have required standardized exercise testing or metabolic challenge protocols, which were beyond the scope of the present work and not routinely feasible in patients with hypertrophic cardiomyopathy.

Importantly, the rationale of the study was not to quantify lactate concentrations per se, but to evaluate the clinical and structural impact of a well-characterized functional polymorphism in the SLC16A1 (MCT1) gene, whose effects on lactate transport and glycolytic strain have been consistently demonstrated in experimental and human exercise studies. Accordingly, our findings should be interpreted as evidence of a clinical association supporting a metabolic modifier role, rather than as direct proof of altered myocardial lactate levels.

We have clarified this limitation and the interpretative scope of the study in the Discussion.

Changes in manuscript: Before drawing final conclusions, several methodological aspects should be acknowledged when interpreting these results. First, the exploratory nature of this study, together with the limited sample size, warrants cautious interpretation of the findings, which should be confirmed in larger, independent cohorts. In addition, no direct measurements of circulating lactate levels, systemic acid–base status, or myocardial metabolic flux were available. Consequently, mechanistic interpretations regarding lactate handling are inferred from established functional data on the MCT1 rs1049434 polymorphism and should be considered hypothesis-generating. Finally, the absence of direct myocardial metabolic or functional validation further constrains mechanistic in-ference and should be addressed in future investigations.

 

Discussion

Comment 1: In the Discussion section, lactate levels in women are presented from the literature; therefore, it is valuable to present these test results.

Response: We thank the reviewer for this comment. We would like to clarify that lactate levels were not directly measured in our cohort. The statement in the Discussion refers to previously published experimental and human exercise studies describing altered lactate transport and increased glycolytic strain in carriers of the MCT1 rs1049434 T allele. To avoid any potential misunderstanding, we have revised the wording of this paragraph to explicitly attribute the lactate-related evidence to the literature and to frame our findings in terms of genetic association rather than direct biochemical measurements.

Changes in manuscript: In women, the presence of the T allele of the MCT1 rs1049434 polymorphism, previously associated with reduced lactate transport efficiency, is related to greater septal hypertrophy. This observation is consistent with experimental models and human exercise studies showing increased glycolytic strain in carriers of this variant [9,11].

 

General remarks

Comment 1: The gene abbreviations presented in this study should be written in italic type.

Response:  We thank the reviewer for this observation. Gene abbreviations have now been formatted in italic type throughout the manuscript in accordance with standard nomenclature guidelines.

Reviewer 2 Report

Comments and Suggestions for Authors

This is a well-conceived and timely translational study that investigates an important and underexplored area: the role of metabolic genotype in modifying the phenotypic expression of sarcomeric hypertrophic cardiomyopathy (HCM).

Suggestions:

1. The primary limitation is the small sample size, particularly within subgroups (e.g., 10 women with HCM, subdivided by genotype). This severely limits statistical power and increases the risk of Type II errors. The non-significant p-values in the male exercise analysis (e.g., LVOT obstruction p=0.136) may represent meaningful biological trends obscured by low power.

2. The authors correctly note the absence of multivariable adjustment due to the exploratory nature and sample size. However, potential confounders such as age, specific sarcomeric gene mutation (MYBPC3 vs. MYH7), variant pathogenicity level, and medication use (e.g., beta-blockers) could influence septal thickness. The lack of adjustment weakens the claim of an independent modifying effect of MCT1 genotype. In the Discussion, this should be framed as a key limitation for future studies to address. 

3. Definition and Quantification of "Vigorous Exercise": The definition provided is qualitative ("competitive or high-intensity endurance sports... over multiple years"). This lacks objective quantification (e.g., MET-hours/week, VO2 max data, training logs) and is subject to recall bias. Mechanistic Speculation vs. Evidence:

4. The discussion justifiably links the findings to known biology (glycolytic strain, lactate shuttle). However, the study provides no direct evidence of altered myocardial metabolism, lactate levels, or mitochondrial function in the patients. The title of the Discussion section or a key paragraph should more explicitly state that the study demonstrates a clinical association which supports the hypothesis of metabolic modulation. 

5.The use of a well-characterized familial HCM cohort with comprehensive clinical, echocardiographic, and cardiac MRI data strengthens the phenotypic assessment and helps distinguish pathological hypertrophy from other causes. However I suggest to consider to cite the article "Cardiac imaging in athlete’s heart: The role of the radiologist" by Fogante et al. 

Author Response

Response to the Reviewer 2

We thank the Editor and the three Reviewers for their constructive comments. We have revised the manuscript accordingly, strengthening methodological detail (genotyping and exercise definition), clarifying interpretation of lactate-related statements, improving readability of the male athletic subgroup description, adding recommended references, and ensuring consistent formatting (italicization of gene symbols). Genotype distribution data for the full cohort and the HCM subgroup are provided in Supplementary Tables S1–S2.

REVIEWER 2

Comments and Suggestions for Authors:  This is a well-conceived and timely translational study that investigates an important and underexplored area: the role of metabolic genotype in modifying the phenotypic expression of sarcomeric hypertrophic cardiomyopathy (HCM).

Response:  We sincerely thank the reviewer for these positive and encouraging comments. We appreciate the recognition of the translational relevance of our work and the focus on metabolic genotype as a modifier of phenotypic expression in sarcomeric hypertrophic cardiomyopathy.

 

Suggestion 1: The primary limitation is the small sample size, particularly within subgroups (e.g., 10 women with HCM, subdivided by genotype). This severely limits statistical power and increases the risk of Type II errors. The non-significant p-values in the male exercise analysis (e.g., LVOT obstruction p=0.136) may represent meaningful biological trends obscured by low power.

Response: We fully agree with the reviewer. As acknowledged in the Discussion, the limited sample size, particularly within sex- and genotype-stratified subgroups, substantially reduces statistical power and increases the risk of Type II error. Consequently, non-significant findings, especially in the analysis of exercise-related phenotypic differences in men, should be interpreted cautiously and may reflect biologically meaningful trends that could emerge as significant in larger cohorts. We have emphasized this point in the Discussion.

 

Suggestion 2: The authors correctly note the absence of multivariable adjustment due to the exploratory nature and sample size. However, potential confounders such as age, specific sarcomeric gene mutation (MYBPC3 vs. MYH7), variant pathogenicity level, and medication use (e.g., beta-blockers) could influence septal thickness. The lack of adjustment weakens the claim of an independent modifying effect of MCT1 genotype. In the Discussion, this should be framed as a key limitation for future studies to address.

Response: We agree with the reviewer that the absence of multivariable adjustment represents an important limitation. As discussed, potential confounders such as age, sarcomeric gene involved, variant pathogenicity, and medication use could influence septal thickness and overall phenotype. Owing to the exploratory design and limited sample size, multivariable modeling was not feasible. We have framed this explicitly in the Discussion as a key limitation that future, larger studies should address.

 

Suggestion 3: Definition and Quantification of "Vigorous Exercise": The definition provided is qualitative ("competitive or high-intensity endurance sports... over multiple years"). This lacks objective quantification (e.g., MET-hours/week, VO2 max data, training logs) and is subject to recall bias. Mechanistic Speculation vs. Evidence:

Response: We thank the reviewer for this comment and acknowledge that the definition of vigorous exercise in our study does not rely on direct physiological quantification such as MET-hours, VO₂max measurements, or training logs, which may introduce recall bias.

However, we would like to clarify that the classification used was not purely qualitative. Participants were categorized based on long-term engagement in competitive or high-intensity endurance sports, following objective and standardized criteria aligned with the Participant Classification Framework proposed by McKay et al. (Int J Sports Physiol Perform, 2022). This framework was specifically developed to avoid subjective or poorly defined terminology in exercise science by emphasizing training history, training volume, and performance-related characteristics rather than point estimates of physical activity.

By prioritizing chronic training exposure, this approach captures the sustained physiological state most relevant to cardiac adaptation and is readily applicable in routine clinical settings where detailed exercise testing is often impractical. Nonetheless, we agree that future studies should incorporate quantitative exercise metrics to further refine exercise exposure assessment.

 

Suggestion 4: The discussion justifiably links the findings to known biology (glycolytic strain, lactate shuttle). However, the study provides no direct evidence of altered myocardial metabolism, lactate levels, or mitochondrial function in the patients. The title of the Discussion section or a key paragraph should more explicitly state that the study demonstrates a clinical association which supports the hypothesis of metabolic modulation.

Response: We agree with the reviewer and have been careful to frame our findings as clinical associations rather than direct mechanistic evidence. The Discussion explicitly states that the proposed metabolic mechanisms are hypothesis-generating and supported by existing experimental and human literature, while acknowledging the absence of direct myocardial metabolic or lactate measurements in our cohort.

 

Suggestion 5: The use of a well-characterized familial HCM cohort with comprehensive clinical, echocardiographic, and cardiac MRI data strengthens the phenotypic assessment and helps distinguish pathological hypertrophy from other causes. However, I suggest to consider to cite the article "Cardiac imaging in athlete’s heart: The role of the radiologist" by Fogante et al.

Response: We thank the reviewer for this valuable suggestion. We have added the reference by Fogante et al. to the manuscript to further support the role of advanced cardiac imaging in differentiating physiological athletic remodeling from pathological hypertrophy in HCM.

Reviewer 3 Report

Comments and Suggestions for Authors

This study investigated how the MCT1 rs1049434 polymorphism influences the clinical expression of hypertrophic cardiomyopathy (HCM) in patients carrying sarcomeric gene variants. Among women with HCM, carriers of the T allele showed more pronounced septal hypertrophy, whereas in men, engagement in vigorous physical activity was associated with a less severe phenotype. Overall, these results point to a complex interaction between genotype, sex, and environmental factors. Specifically, altered lactate transport may contribute to enhanced hypertrophic remodeling in women, while exercise-related metabolic adaptations appear to mitigate disease severity in men. I have few comments: 

- Introduction: The introduction would benefit from a more detailed discussion of the role of MCT1 in cardiac energy metabolism, as well as a clearer rationale for focusing on this specific polymorphism in the context of HCM.

- Line 87: Please specify whether the study received approval from a single ethics committee or from multiple committees.

- Line 101: The definition of “vigorous physical activity” should be clarified in greater detail, given its central importance to the analysis.

- Results : Presenting genotype distribution data for both the overall cohort and the HCM subgroup in a table could improve clarity and readability.

- Lines 134–144: The description of phenotypic differences observed in men could be streamlined to better highlight the main findings.

- Discussion : Consider expanding the discussion of potential biological mechanisms underlying the observed genotype–sex–environment interaction, drawing on existing literature related to cardiac metabolism and exercise physiology.

- Lines 186–197: This section could be reorganized to improve flow, clearly separating methodological limitations from the interpretation of the results.

Author Response

Response to the Reviewer 3

We thank the Editor and the three Reviewers for their constructive comments. We have revised the manuscript accordingly, strengthening methodological detail (genotyping and exercise definition), clarifying interpretation of lactate-related statements, improving readability of the male athletic subgroup description, adding recommended references, and ensuring consistent formatting (italicization of gene symbols). Genotype distribution data for the full cohort and the HCM subgroup are provided in Supplementary Tables S1–S2.

 

REVIEWER 3

Comments and Suggestions for Authors: This study investigated how the MCT1 rs1049434 polymorphism influences the clinical expression of hypertrophic cardiomyopathy (HCM) in patients carrying sarcomeric gene variants. Among women with HCM, carriers of the T allele showed more pronounced septal hypertrophy, whereas in men, engagement in vigorous physical activity was associated with a less severe phenotype. Overall, these results point to a complex interaction between genotype, sex, and environmental factors. Specifically, altered lactate transport may contribute to enhanced hypertrophic remodeling in women, while exercise-related metabolic adaptations appear to mitigate disease severity in men.

Response:  We thank the reviewer for this accurate and thoughtful summary of our findings. We appreciate the recognition of the genotype–sex–environment interaction highlighted in our study and the emphasis on metabolic mechanisms potentially underlying the observed differences in hypertrophic remodeling between women and men.

 

Introduction

Comment 1: The introduction would benefit from a more detailed discussion of the role of MCT1 in cardiac energy metabolism, as well as a clearer rationale for focusing on this specific polymorphism in the context of HCM.

Response: We thank the reviewer for this comment. We have revised the Introduction to strengthen the description of the role of MCT1 in cardiac energy metabolism, integrating its function within the lactate shuttle and its relevance to myocardial energetic remodeling as supported by available experimental and human data.

In addition, we have clarified the rationale for focusing on the rs1049434 polymorphism by explicitly stating that, among the variants described in SLC16A1, rs1049434 is the most extensively functionally characterized and the one most consistently associated with altered lactate transport efficiency and substrate utilization. Other SLC16A1 polymorphisms are briefly acknowledged, but their limited functional validation and lack of evidence in hypertrophic cardiomyopathy justify the prioritization of rs1049434. These revisions aim to improve conceptual clarity without extending beyond the scope of the study.

 

Materials and Methods

Comment 1 - Line 87: Please specify whether the study received approval from a single ethics committee or from multiple committees.

Response: We thank the reviewer for this comment. We have clarified the ethics statement to specify that the study was approved by a single regional ethics committee (Cantabria Research Ethics Committee, CEIC-IDIVAL), and we have maintained the corresponding reference number.

 

Comment 2- Line 101: The definition of “vigorous physical activity” should be clarified in greater detail, given its central importance to the analysis.

Response: We thank the reviewer for this comment and acknowledge that the definition of vigorous exercise in our study does not rely on direct physiological quantification such as MET-hours, VO₂max measurements, or training logs, which may introduce recall bias.

However, we would like to clarify that the classification used was not purely qualitative. Participants were categorized based on long-term engagement in competitive or high-intensity endurance sports, following objective and standardized criteria aligned with the Participant Classification Framework proposed by McKay et al. (Int J Sports Physiol Perform, 2022). This framework was specifically developed to avoid subjective or poorly defined terminology in exercise science by emphasizing training history, training volume, and performance-related characteristics rather than point estimates of physical activity.

By prioritizing chronic training exposure, this approach captures the sustained physiological state most relevant to cardiac adaptation and is readily applicable in routine clinical settings where detailed exercise testing is often impractical. Nonetheless, we agree that future studies should incorporate quantitative exercise metrics to further refine exercise exposure assessment.

 

Results

Comment 1: Presenting genotype distribution data for both the overall cohort and the HCM subgroup in a table could improve clarity and readability.

Response:  We thank the reviewer for this helpful suggestion. Genotype distributions and allele frequencies of the SLC16A1 rs1049434 polymorphism are now presented in tabular form in the Supplementary Material for both the full cohort of sarcomeric variant carriers (Table S1) and the subgroup of patients with hypertrophic cardiomyopathy (Table S2), with stratification by sex. In addition, we now explicitly report that genotype distributions conformed to Hardy–Weinberg equilibrium in the overall cohort as well as in sex-stratified analyses for both populations.

Changes in manuscript: Genotype distributions conformed to Hardy–Weinberg equilibrium in the overall cohort and in sex-stratified analyses for both the full carrier cohort and the hypertrophic car-diomyopathy subgroup.

 

Comment 2- Lines 134–144: The description of phenotypic differences observed in men could be streamlined to better highlight the main findings.

Response: We thank the reviewer for this helpful suggestion. We have streamlined this paragraph by removing redundant descriptions and integrating the key quantitative findings into a single, concise statement. The revised text emphasizes the overall pattern of a directionally milder phenotype in trained men, while referring detailed numerical values to Table 3, thereby improving clarity and readability.

Changes in manuscript: Among male patients with HCM, those engaged in habitual vigorous exercise showed a directionally milder phenotype, characterized by lower septal thickness and a more favorable hemodynamic and tissue profile, including a lower prevalence of left ventricular outflow tract obstruction, less frequent left atrial enlargement, and a reduced burden of myocardial fibrosis on imaging (Table 3). Although these differences did not reach statistical significance (septal thickness: 18.3 ± 4.1 vs. 19.9 ± 6.9 mm; p = 0.585), the overall pattern consistently favored trained individuals.

 

Discussion

Comment 1:  Consider expanding the discussion of potential biological mechanisms underlying the observed genotype–sex–environment interaction, drawing on existing literature related to cardiac metabolism and exercise physiology.

Response: We thank the reviewer for this valuable suggestion. We have expanded the Discussion by adding a dedicated mechanistic paragraph integrating relevant literature on cardiac lactate handling and metabolic remodeling. Specifically, we discuss (i) the proposed fibroblast-to-cardiomyocyte lactate shuttle and the differential expression of MCT isoforms in cardiac cell types, (ii) experimental evidence that a lactate-rich microenvironment can promote hypertrophic signaling and transcriptional remodeling, and (iii) sex-related differences in myocardial substrate utilization that may increase susceptibility to altered monocarboxylate handling in women. We also contextualize our findings in relation to exercise-induced metabolic conditioning as a potential buffering factor in men. These additions are presented cautiously as biologically plausible, hypothesis-generating mechanisms, given the absence of direct myocardial metabolic measurements in our cohort.

Changes in manuscript: A plausible mechanistic framework linking rs1049434 to sex- and exercise-dependent remodeling in HCM involves altered lactate flux within the cardiac microenvironment. Recent evidence supports a cell-to-cell lactate shuttle in the heart, in which cardiomyocytes predominantly express MCT1, whereas cardiac fibroblasts preferentially express MCT4, potentially supplying lactate to neighboring cardiomyocytes under stress conditions [18]. Perturbations that increase local lactate availability have been shown to promote hypertrophic responses and modulate cardiomyocyte gene expression programs [18,19]. In this context, rs1049434—previously associated with altered MCT1-mediated lactate transport—could contribute to an energetic mismatch in genetically predisposed individuals, thereby amplifying hypertrophic signaling.

Sex-related differences in myocardial substrate utilization may further modulate this effect. Prior human imaging studies have reported sex differences in myocardial oxidative metabolism and glucose utilization, suggesting that female myocardium may rely on distinct metabolic pathways under comparable workloads [20]. Therefore, it is biologically plausible that impaired monocarboxylate handling could have a greater phenotypic impact in women, consistent with the greater septal hypertrophy observed in female T-allele carriers in our cohort. Supporting the relevance of lactate signaling to hyper-trophic remodeling, Wei et al. reported increased lactate uptake in hypertrophied cardiomyocytes, accompanied by upregulation of MCT1 and hypertrophy-associated genes, and showed that disruption of this lactate axis attenuated hypertrophy-linked transcriptional responses [21].

Conversely, exercise-induced metabolic conditioning may partially buffer lac-tate-related energetic stress in men. Vigorous training enhances oxidative capacity and metabolic flexibility, which may mitigate the structural expression of sarcomeric energetic inefficiency, aligning with the directionally milder phenotype observed in trained men in our cohort. Importantly, these mechanistic considerations remain hypothesis-generating in the absence of direct myocardial metabolic measurements in the present study and should be tested in larger cohorts and studies incorporating cardiac metabolic profiling.

 

Comment 2 - Lines 186–197: This section could be reorganized to improve flow, clearly separating methodological limitations from the interpretation of the results.

Response: We thank the reviewer for this suggestion. The Discussion section already separates the interpretation of findings from methodological limitations, with limitations explicitly addressed in a dedicated paragraph. To further improve clarity and guide the reader, we have slightly revised the introductory sentence of the limitations paragraph to explicitly signal this transition. No substantive changes to content were required.

Changes in manuscript: Before drawing final conclusions, several methodological aspects should be acknowledged when interpreting these results. First, the exploratory nature of this study, together with the limited sample size, warrants cautious interpretation of the findings, which should be confirmed in larger, independent cohorts. In addition, no direct measurements of circulating lactate levels, systemic acid–base status, or myocardial metabolic flux were available. Consequently, mechanistic interpretations regarding lactate handling are inferred from established functional data on the MCT1 rs1049434 polymorphism and should be considered hypothesis-generating. Finally, the absence of direct myocardial metabolic or functional validation further constrains mechanistic in-ference and should be addressed in future investigations.

 

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The introduced additions to the manuscript  are correct and sufficient to make this manuscript more transparent for readers.