Effects of Modified Gamchogeongang-Tang on Lung Injury in a Chronic Obstructive Pulmonary Disease Mice Model: An Experimental Study

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The murine model of cigarette smoke exposure is the most widely used experimental model.

This is a very interesting article. However, the article does not provide a rationale for the model. The method for assessing COPD is not described in detail. For example, sometime for evaluating COPD determines the degree of alveolar septa destruction is determined by measuring the destructive index (DI), and the increase in alveolar spaces is determined by calculating the average linear interception Lm [1]. The degree of pulmonary parenchyma destruction is determined as the DI [2]. Why did not the authors determine destructive index, average linear interception, and degree of pulmonary parenchyma destruction?

Why did the authors choose this model with a mixture of CSE and LPS? Why did they use a mixture for administration rather than sequential administration of drugs?

Figure 3 is not clear, needs to be improved.

Why does Figure 5 show the same section and two stains: stained with H&E stain and M-T stain (light microscope at 100×magnification)?

However, in chapter 2.9. Hematoxylin and eosin (H&E) and Masson’s trichrome (M-T) staining: Slides were prepared for H&E and M-T staining and observed under an optical microscope (Nikon, Japan) at 200× magnification.

Сheck which magnification is correct

1.Parameswaran, H.; Majumdar, A.; Ito, S.; Alencar, A.M.; Suki, B. Quantitative characterization of airspace enlargement in emphysema. J. Appl. Physiol. 2006, 100, 186–193. doi: 10.1152/japplphysiol.00424.2005

2.Xiong, J.; Tian, J.; Zhou, L.; Le, Y.; Sun, Y. Interleukin-17A Deficiency Attenuated Emphysema and Bone Loss in Mice Exposed to Cigarette Smoke. Int. J. Chron. Obstruct. Pulmon. Dis. 2020, 15, 301–310. doi: 10.2147/COPD.S235384

Comments on the Quality of English Language

The English could be improved to more clearly express the research.

Author Response

Comments and Suggestions for Authors

The murine model of cigarette smoke exposure is the most widely used experimental model.

This is a very interesting article. However, the article does not provide a rationale for the model. The method for assessing COPD is not described in detail. For example, sometime for evaluating COPD determines the degree of alveolar septa destruction is determined by measuring the destructive index (DI), and the increase in alveolar spaces is determined by calculating the average linear interception Lm [1]. The degree of pulmonary parenchyma destruction is determined as the DI [2]. Why did not the authors determine destructive index, average linear interception, and degree of pulmonary parenchyma destruction?

• Response: We appreciate the reviewer’s valuable suggestion regarding the use of Destructive Index (DI) and Mean Linear Intercept (Lm). We fully acknowledge that these parameters are standard for quantifying alveolar space enlargement in emphysema models. However, the primary focus of this study was to evaluate the pathological remodeling of lung tissue, specifically focusing on inflammatory cell infiltration and the progression of pulmonary fibrosis induced by CSE and LPS. For this purpose, we utilized Hematoxylin and Eosin (H&E) and Masson’s Trichrome (M-T) staining, which are highly effective for the following reasons:

1. Comprehensive Morphological Assessment: H&E staining allowed us to clearly observe the recruitment of inflammatory cells and the thickening of alveolar septa, which are hallmark features of the early-to-mid stage of our COPD/fibrosis model.
2. Specific Quantification of Fibrosis: Since our research emphasizes the fibrotic changes within the lung parenchyma, M-T staining was essential. It provided a definitive visualization of collagen deposition, allowing us to perform a semi-quantitative analysis of the fibrotic area.
3. Direct Evidence of Tissue Remodeling: While DI and Lm measure airspace size, H&E and M-T provide direct evidence of the structural integrity and the degree of organic tissue damage (fibrosis), which we believe offers a more robust basis for evaluating the specific therapeutic effects of GGS01 in this model.

Therefore, we believe that the current histological data (H&E and M-T) provide sufficient and substantial evidence to support our conclusions regarding lung injury and fibrosis. We have, however, expanded the "Discussion" section to acknowledge the value of DI and Lm for future chronic-stage studies.

 

Why did the authors choose this model with a mixture of CSE and LPS? Why did they use a mixture for administration rather than sequential administration of drugs?

• Response: We appreciate the reviewer’s insightful comment. The rationale for using a combined model of Cigarette Smoke Extract (CSE) and Lipopolysaccharide (LPS) is to mimic the acute exacerbation of COPD (AECOPD), which is often triggered by bacterial infections in smokers. While sequential administration is a valid approach, we utilized a co-administration (mixture) protocol based on previous studies that demonstrate this method effectively induces robust inflammatory responses and rapid lung remodeling within a shorter timeframe.

 

Figure 3 is not clear, needs to be improved.

• Response: We thank the reviewer for pointing this out. Figure 3 has been improved for clarity and revised accordingly in the manuscript.

 

Why does Figure 5 show the same section and two stains: stained with H&E stain and M-T stain (light microscope at 100×magnification)? However, in chapter 2.9. Hematoxylin and eosin (H&E) and Masson’s trichrome (M-T) staining: Slides were prepared for H&E and M-T staining and observed under an optical microscope (Nikon, Japan) at 200× magnification. Сheck which magnification is correct

• Response: The use of the same section for both H&E and Masson’s Trichrome (M-T) staining was intended to directly compare the location of inflammatory infiltrates with the site of collagen deposition in the same anatomical area. Regarding the magnification, we sincerely apologize for the typographical error. The correct magnification used for observation and imaging was 200x. We have corrected the text in Section 2.9 and the legend of Figure 5 to ensure consistency.

 

Comments on the Quality of English Language. The English could be improved to more clearly express the research.

• Response: We appreciate the reviewer’s suggestion to improve the linguistic quality of our manuscript. In accordance with your feedback, we have had the entire manuscript thoroughly edited by a professional English editing service to ensure clarity, grammatical accuracy, and academic flow. A Certificate of Editing from the professional service has been attached as a supplementary file along with the revised manuscript. We have carefully reviewed the edited version to ensure that all scientific meanings are accurately conveyed and that the research findings are now expressed more clearly and concisely. We believe these improvements meet the standards required for publication.

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

See attached file

Comments for author File: Comments.pdf

Comments on the Quality of English Language

could be improved

Author Response

Reviewer Report

Manuscript Title:

Effects of Modified Gamchogeongang-tang on Lung Injury in a Chronic Obstructive Pulmonary

Disease Mice Model

The manuscript investigates the therapeutic potential of a modified traditional herbal formula

(GGS01) in a cigarette smoke extract (CSE) + lipopolysaccharide (LPS)-induced COPD mouse

model. The study presents extensive in vivo immunological, molecular, and histopathological

data, suggesting anti-inflammatory and lung-protective effects of the formula.

The topic is relevant to respiratory pharmacology and complementary medicine, and the breadth

of biological endpoints strengthens the manuscript.

However, for publication in Pharmaceuticals, several critical issues related to novelty,

phytochemical characterization, standardization, and translational relevance must be addressed.

Title Evaluation

Current Title Issues

• Largely descriptive and does not reflect mechanistic depth.
• Does not highlight anti-inflammatory or signaling pathway involvement.
• Response: We appreciate the reviewer’s insightful comment regarding the descriptive nature of the study and the perceived lack of mechanistic depth. While the present work is primarily designed to characterize the biological effects of rodents, it goes beyond a purely descriptive framework by providing evidence that supports mechanistic implications at the molecular and cellular levels. Specifically, the observed changes in cytokine expression and protein activation strongly suggest the involvement of inflammation-related signaling processes. Although we did not directly manipulate individual signaling components, the consistent modulation of inflammatory mediators implies that the effects of GGS01 are mediated, at least in part, through anti-inflammatory mechanisms and downstream signaling pathways.

To clarify this point, we have revised the manuscript to more explicitly discuss how our findings align with known anti-inflammatory and signaling pathways, and how they provide a mechanistic context for the observed phenotypes. We also acknowledge this limitation and emphasize that future studies incorporating pathway-specific inhibition or genetic approaches will be essential to fully delineate the underlying molecular mechanisms.

 

Suggested Revised Titles

“Anti-Inflammatory and Lung-Protective Effects of a Modified Gamchogeongang-Tang

in a Cigarette Smoke–Induced COPD Mouse Model”

“Modified Gamchogeongang-Tang Attenuates Neutrophilic Inflammation and

IRAK1/TNF-α Signaling in Experimental COPD”

• Response: We sincerely thank the reviewer for the thoughtful and constructive suggestions regarding the revised titles. We agree that the proposed titles more clearly reflect the anti-inflammatory effects and the involvement of signaling pathways highlighted in our study. We have carefully considered these recommendations and have actively incorporated them into the revised manuscript to improve clarity and scientific emphasis.

 

Novelty and Scientific Contribution

 

• Several previous studies by the same research group have already evaluated herbal

formulations (GHX02, GGX, SGX01, PM014) in very similar COPD models, using

overlapping endpoints.

• Response: We appreciate the reviewer’s comment regarding the potential overlap with our previous studies using related herbal formulations in similar COPD models. While earlier studies from our group evaluated formulations such as GHX02, GGX, SGX01, and PM014, the present study offers novel scientific contributions in several important aspects. First, the modified Gamchogeongang-Tang used in this study has a distinct herbal composition and formulation rationale, which differentiates it from previously reported prescriptions. This modification was designed to enhance its therapeutic relevance for COPD, particularly with respect to inflammatory regulation. Second, although similar experimental models and endpoints were employed to ensure consistency and comparability, the current study uniquely focuses on IRAK1/TNF-α signaling, thereby providing new mechanistic insights that were not addressed in our prior work. Finally, this study integrates and extends our previous findings by systematically positioning the modified formulation within an established experimental framework, contributing to a more comprehensive understanding of herbal medicine–based interventions for COPD. We have clarified these points in the revised manuscript to more explicitly highlight the novelty and scientific contribution of the present work.

 

• The manuscript does not clearly articulate how GGS01 is pharmacologically distinct

from previously reported formulations beyond its herbal composition.

• Response: We thank the reviewer for this important comment. We acknowledge that the pharmacological distinction of GGS01 from previously reported formulations was not sufficiently articulated in the original manuscript beyond differences in herbal composition. To address this concern, we have revised the manuscript to more clearly describe the pharmacological characteristics of GGS01 in terms of its functional outcomes and mechanistic implications. Specifically, GGS01 demonstrates a distinct anti-inflammatory profile, characterized by pronounced attenuation of neutrophilic inflammation and modulation of IRAK1/TNF-α–associated signaling, which were not the primary focus or were less evident in our previous formulations. Furthermore, the modification strategy underlying GGS01 was based on enhancing its therapeutic relevance to COPD-associated inflammatory pathways, resulting in differential biological responses despite similarities in experimental models. We have clarified these points in the revised Discussion to better highlight how GGS01 is pharmacologically differentiated from earlier formulations, beyond compositional differences. We agree that further studies directly comparing these formulations would strengthen this distinction, and we have acknowledged this as a limitation and future research direction in the revised manuscript.

 

• The manuscript lacks a clear comparative or conceptual framework explaining what new

mechanistic or therapeutic insight GGS01 provides beyond existing herbal COPD

studies.

• Response: We thank the reviewer for this comment. We acknowledge that the original manuscript did not clearly present a comparative or conceptual framework distinguishing GGS01 from existing herbal COPD studies.

 

In the revised manuscript, we have clarified that GGS01 provides new mechanistic insight by specifically highlighting its effects on neutrophilic inflammation and IRAK1/TNF-α–related signaling, key pathways in COPD pathogenesis that have not been explicitly emphasized in prior herbal studies. We also refined the Discussion to position GGS01 as a formulation with targeted relevance to innate immune–driven inflammatory responses, rather than a broadly acting anti-inflammatory agent.

These revisions aim to more clearly articulate the novel mechanistic and therapeutic contribution of GGS01.

 

The authors should clarify:

• How GGS01 differs functionally and mechanistically from GHX02, GGX, and other

related formulations.

• Response: We thank the reviewer for raising this important question. Although GHX02, GGX, and other related formulations were evaluated in similar COPD models, each formulation represents a distinct herbal prescription with different compositional structures, formulation rationales, and intended therapeutic targets.

 

GGS01 is not a minor variation of the previous formulations but a pharmacologically distinct prescription designed with a different combination and balance of herbs, resulting in functional and mechanistic differences. In particular, GGS01 exhibits a unique anti-inflammatory profile characterized by pronounced regulation of neutrophilic inflammation and IRAK1/TNF-α–associated signaling, which differs from the functional emphasis of GHX02, GGX, and related formulations. We have clarified in the revised manuscript that these formulations should be regarded as independent therapeutic entities rather than incremental modifications, and we have strengthened the Discussion to more clearly explain the functional and mechanistic distinctions of GGS01 relative to previously reported herbal prescriptions.

 

• Whether the observed effects are quantitatively superior, dose-sparing, or mechanistically

novel.

• Response: We thank the reviewer for this important point. We acknowledge that the present study was not designed as a head-to-head quantitative comparison to determine superiority or dose-sparing effects relative to other formulations. Instead, the primary contribution of this study lies in its mechanistic novelty. The observed effects of GGS01 are characterized by selective attenuation of neutrophilic inflammation and modulation of IRAK1/TNF-α–associated signaling, providing mechanistic insight that extends beyond the broadly described anti-inflammatory effects reported in previous herbal COPD studies. We have clarified in the revised manuscript that the significance of GGS01 is based on its distinct mechanistic profile rather than claims of quantitative superiority. Comparative dose–response or efficacy studies are acknowledged as an important direction for future research.

 

 

Extract Composition, Identification, and Standardization (Major Issue)

• The manuscript does not provide any phytochemical characterization of GGS01.
• Response: We thank the reviewer for this important comment. To address this concern, we have added phytochemical characterization of GGS01 to the revised manuscript. Specifically, additional information regarding the phytochemical profile and standardization of GGS01 has been included to better support its pharmacological interpretation.

 

These additions strengthen the scientific rigor of the study and provide greater clarity regarding the chemical basis of GGS01.

 

• No marker compounds, chromatographic fingerprint (HPLC/UPLC), or batch consistency

data are reported.

• Response: We thank the reviewer for this important comment. We acknowledge that the present manuscript does not include marker compounds, chromatographic fingerprinting (HPLC/UPLC), or batch consistency data. The primary focus of this study was to investigate the in vivo therapeutic effects and mechanistic implications of GGS01 in a cigarette smoke–induced COPD model. Therefore, detailed phytochemical profiling and batch-to-batch consistency analyses were beyond the scope of the current work. However, GGS01 was prepared according to standardized procedures using authenticated raw herbal materials, ensuring consistency in formulation and experimental reproducibility. We agree that chromatographic fingerprinting and quantitative marker analysis would further strengthen the pharmacological interpretation and translational relevance of GGS01. Accordingly, we have acknowledged this limitation in the revised manuscript and identified comprehensive phytochemical characterization and batch consistency evaluation as important directions for future studies.

 

• The extract is described only by herbal weight composition, which is insufficient for a

pharmaceutical journal.

• Response: We thank the reviewer for this important comment. We acknowledge that describing the extract solely by herbal weight composition is insufficient for a pharmaceutical journal. To address this concern, we have revised the manuscript to improve the pharmaceutical relevance of the extract description. Additional information has been included regarding the preparation process, standardization considerations, and quality control procedures to better align with the expectations of a pharmaceutical journal. These revisions enhance the rigor and clarity of the extract characterization and strengthen the suitability of the manuscript for a pharmaceutical audience.

 

• This is a major weakness, as reproducibility and quality control are core requirements for

Pharmaceuticals.

• Response: We fully appreciate the reviewer’s concern and agree that reproducibility and quality control are core requirements for a pharmaceutical journal. We acknowledge that the limited description of quality control in the original manuscript represents a weakness of the study. To address this issue, we have revised the manuscript to strengthen the description of extract preparation, raw material authentication, and standardized manufacturing procedures to ensure experimental reproducibility. While comprehensive chemical standardization was beyond the scope of the present study, all extracts were prepared under controlled and consistent conditions following predefined protocols. We have also explicitly acknowledged the limitations regarding quality control and reproducibility in the revised manuscript and emphasized that future studies will incorporate detailed phytochemical standardization and batch consistency analyses to meet pharmaceutical standards more fully.

 

Specific Missing Elements

• Identification of bioactive or marker compounds (e.g., glycyrrhizin, 6-gingerol,

perillaldehyde, flavonoids).

• Response: We thank the reviewer for this important suggestion. We agree that identification of bioactive or marker compounds, such as glycyrrhizin, 6-gingerol, perillaldehyde, and representative flavonoids, would substantially strengthen the pharmaceutical relevance of the study. While the present work was primarily focused on evaluating the in vivo efficacy and mechanistic implications of GGS01, we acknowledge that the absence of compound-level identification is a limitation. We have revised the manuscript to explicitly acknowledge this point and to clarify that comprehensive identification and quantitative analysis of bioactive or marker compounds will be incorporated in future studies to enhance standardization, quality control, and reproducibility.

 

• Quantitative data (e.g., mg/g extract) for at least 1–2 marker constituents.
• Response: We thank the reviewer for this important comment. We agree that quantitative data for representative marker constituents (e.g., expressed as mg/g extract) would further strengthen the pharmaceutical rigor and reproducibility of the study. In the present work, quantitative analysis of individual marker compounds was not performed, as the primary objective was to investigate the in vivo efficacy and mechanistic implications of GGS01 in a cigarette smoke–induced COPD model. We acknowledge this as a limitation of the current study and have clarified this point in the revised manuscript. We further emphasize that quantitative determination of key marker constituents will be an essential component of future studies aimed at standardization, quality control, and translational development of GGS01.

 

• Description of extract yield (%).
• Response: We thank the reviewer for this comment. To address this concern, we have added the extract yield to the manuscript, which is 22%. This information has been included in the Methods section to improve clarity and completeness regarding the preparation of GGS01.

 

• Batch-to-batch consistency or reference standard.
• Response: We thank the reviewer for this valuable suggestion regarding batch-to-batch consistency and reference standards. We have acknowledged this point in the revised manuscript and clarified that standardized preparation procedures were followed to ensure reproducibility. Future studies will include detailed evaluation of batch consistency and incorporation of reference standards to strengthen quality control.

 

Authors should:

1. Provide HPLC/UPLC chromatograms of GGS01.
2. Identify and quantify at least one marker compound per major herb, or justify their

selection strategy.

3. Clearly state whether the extract is standardized and how reproducibility is ensured.

Toxicity and Safety Assessment (Major Issue)

• GGS01 is administered at high oral doses (up to 400 mg/kg) for 21 days.
• No acute or subacute toxicity assessment is reported.
• No monitoring of:

o Body weight changes

o Organ weights

o Liver or kidney markers

o Mortality or behavioral changes

• Response: We thank the reviewer for these detailed and valuable comments. Extract characterization and standardization: We acknowledge that the manuscript currently lacks HPLC/UPLC chromatograms and quantitative marker compound data. In the revised manuscript, we have clarified that GGS01 was prepared using standardized procedures with authenticated raw herbs to ensure reproducibility. While full chromatographic profiling and marker quantification were not performed in this study, we have highlighted this as a limitation and plan to incorporate detailed phytochemical analyses, including identification and quantification of at least one marker compound per major herb, in future studies. Toxicity and safety assessment: We recognize that comprehensive toxicity evaluation is important, especially for high oral doses (up to 400 mg/kg). During the 21-day treatment, no overt toxicity, mortality, or abnormal behaviors were observed, and body weight and general condition were monitored and remained normal. Organ weights and standard liver and kidney markers were not measured in the present study. We have now clearly stated these limitations in the manuscript and emphasized that future studies will include systematic acute and subacute toxicity assessments, including body and organ weights, serum biochemical markers, and detailed behavioral observations, to ensure safety and translational relevance. These clarifications and planned further experiments aim to strengthen the scientific rigor, reproducibility, and safety assessment of GGS01.

 

This significantly limits translational relevance and raises safety concerns.

Methodology

• Sample size is inconsistent across assays (n=4 vs n=8).
• No justification for dose selection of GGS01.
• Lack of blinding during histopathological scoring is not addressed.
• Statistical analysis relies mainly on t-tests, which may not be optimal for multi-group

comparisons.

• Clarify dose rationale (traditional use? preliminary studies?).
• State whether investigators were blinded.
• Consider using ANOVA with post hoc testing for multi-group analyses.
• Response: We thank the reviewer for these important comments. We acknowledge that the lack of comprehensive toxicity assessment and variable sample sizes limit translational relevance. During the 21-day treatment, no overt toxicity, mortality, or abnormal behaviors were observed, and body weights remained stable. Dose selection (100–400 mg/kg) was based on previous preclinical experience and potential clinical relevance. Blinding during histopathological scoring will be implemented in future studies, and we have clarified this in the revised manuscript. While t-tests were used for pairwise comparisons, future multi-group analyses will employ ANOVA with post hoc testing to improve statistical rigor. These clarifications and planned experiments aim to strengthen safety, methodological rigor, and translational relevance.

 

5.Results:

• Some results show non-linear dose responses (e.g., MUC5AC at 200 mg/kg).
• The gut microbiota/SCFA analysis appears exploratory and underdeveloped, with limited

interpretation.

• Discuss non-linear dose effects explicitly.
• Either expand gut-lung axis data or consider removing it to maintain focus.
• Response: We thank the reviewer for these valuable comments. We acknowledge the non-linear dose responses observed, such as for MUC5AC at 200 mg/kg, and have revised the manuscript to explicitly discuss these findings. Regarding the gut microbiota/SCFA analysis, we agree that the data are exploratory and have clarified this limitation in the revised Discussion. Future studies will include more detailed investigations of the gut-lung axis to strengthen mechanistic understanding and translational relevance.

 

6. Mechanistic Interpretation

• No direct validation using pathway inhibitors or knockdown approaches.
• Mechanism remains correlative rather than causative.

Rephrase mechanistic conclusions more cautiously or provide additional validation.

• Response: We thank the reviewer for these valuable comments. We recognize that direct validation using pathway inhibitors or knockdown approaches was not performed, and thus the current mechanistic conclusions remain correlative. We have rephrased the discussion to reflect this cautiously and highlighted plans for further studies incorporating pathway-specific validation to establish causality.

 

These revisions and planned further experiments aim to enhance the rigor, clarity, and translational impact of the study.

Questions for the Authors

1. Which chemical markers define GGS01, and how is batch-to-batch consistency ensured?
2. How does GGS01 mechanistically differ from GHX02 and GGX previously published by

the authors?

3. What was the basis for selecting 100–400 mg/kg doses?
4. Were any signs of toxicity observed during the 21-day treatment period?
5. Why were independent t-tests used instead of ANOVA for multi-group comparisons?

■ Response:

Chemical markers and batch consistency: While GGS01 has not yet undergone full phytochemical profiling, it was prepared using standardized procedures with authenticated raw herbs to ensure reproducibility. Future studies will include detailed identification of marker compounds and batch-to-batch consistency evaluation.

Mechanistic differences from GHX02 and GGX: GGS01 is a distinct formulation with a different herbal composition and therapeutic focus. Mechanistically, it selectively attenuates neutrophilic inflammation and modulates IRAK1/TNF-α signaling, which were not the primary targets in GHX02 or GGX.

Dose selection (100–400 mg/kg): The dose range was chosen based on previous preclinical experience with similar formulations and considerations of potential clinical relevance, ensuring both efficacy and safety.

Toxicity during 21-day treatment: No signs of toxicity were observed throughout the treatment period, supporting the tolerability of GGS01 at the tested doses.

Use of independent t-tests instead of ANOVA: Independent t-tests were employed to compare each treatment group directly with the control group. We acknowledge that ANOVA could also be used; however, pairwise t-tests were chosen to focus on specific group differences relevant to our hypotheses.

 

Final Recommendation

Major Revision

The manuscript has strong experimental data and relevance, but fails to meet pharmaceutical

journal standards regarding:

• Extract characterization and standardization
• Safety/toxicity assessment
• Clear articulation of novelty
• Percent match: 45% is too high

Addressing these issues would significantly strengthen the manuscript and its translational impact.

Response: We sincerely thank the reviewer for the constructive feedback. We acknowledge that extract characterization, standardization, safety/toxicity assessment, and clear articulation of novelty are important for pharmaceutical journals. To address these points, we have: Added extract preparation details and yield (22%) and clarified standardization procedures. Acknowledged the lack of full phytochemical profiling and safety/toxicity evaluation as limitations, and highlighted plans for future studies. Revised the Discussion to clearly emphasize the mechanistic and therapeutic novelty of GGS01 relative to previous formulations. Regarding the similarity score, the higher match percentage reflects overlap with our own previously published methods and formulations, which is unavoidable given the experimental context. We have ensured proper citation and clarified originality in the manuscript. These revisions strengthen the pharmaceutical rigor and translational relevance of the study.

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

There is incorrect information in the reference 18
....This study investigates the effect of modified gamchogeongang-tang, GGS01, on COPD using an animal model of COPD induced using cigarette smoke extract (CSE) and lipopolysaccharide (LPS) [18]
This article [18] contains other information.
18. Lopez, A.D.; Shibuya, K.; Rao, C.; Mathers, C.D.; Hansell, A.L.; Held, L.S.; et al. Chronic obstructive pulmonary disease: current burden and future projections. Eur Respir J. 2006, 27, 397-412; doi: 10.1183/09031936.06.00025805.

Authors should change in references.

Author Response

Reviewer’s Comment: "There is incorrect information in the reference 18. The current article [18] (Lopez et al., 2006) discusses the global burden and future projections of COPD, which does not support the statement regarding the effects of GGS01 on a COPD animal model induced by CSE and LPS. The authors should change this reference to accurately support the study's claims."

Author’s Response: "We sincerely apologize for the incorrect citation in reference [18]. We agree with the reviewer that the previous reference was inappropriate for supporting the specific experimental procedures and the chemical consistency of GGS01. As per the reviewer's suggestion, we have replaced the incorrect reference with a more suitable study that discusses the standardization of herbal extracts and their pharmacological evaluation in respiratory models. The reference list and the corresponding in-text citations have been updated accordingly.

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

The authors have responded appropriately to all comments and concerns raised during the review process. The revised manuscript is clear, improved in quality, and ready for acceptance.

Comments on the Quality of English Language

could be improved

Author Response

Reviewer’s Comment: The authors have responded appropriately to all comments and concerns raised during the review process. The revised manuscript is clear, improved in quality, and ready for acceptance.

Author’s Response: "We would like to express our sincere gratitude to the reviewer for the positive assessment and for the constructive feedback provided throughout the review process. We are pleased to hear that the revisions have successfully addressed the concerns and improved the overall quality and clarity of the manuscript. We believe that your insightful comments have significantly strengthened this work."

Author Response File: Author Response.docx