Review Reports - The PATH Protocol for Integrated Facial Rejuvenation: A Preliminary Three-Patient Case Report Series and Narrative Review of the Literature

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript addresses a clinically relevant topic; however, several major methodological, ethical, and interpretative concerns should be resolved.

Abstract

The statement regarding a “favorable safety profile” is too strong for a study involving only three patients with 60 days of follow-up. It should be limited to the observation that no serious adverse events were reported during the follow-up period.

Introduction

Lines 69–75: The manuscript discusses procedures supported by different levels of scientific evidence. In particular, clinical evidence concerning succinic acid-containing products remains limited. Please clearly distinguish well-established applications from approaches for which the evidence is still preliminary.

Lines 76–80: The PATH protocol is presented as a coherent clinical method, but the manuscript does not state who developed it, whether it has been previously published or validated, or whether it is associated with a specific manufacturer. Please provide this information and cite the original source of the protocol.

Lines 78–80: The claims that the protocol affects the extracellular matrix, restores soft tissues, and maintains the skin barrier are presented as established effects. These statements require appropriate references or should be reformulated as theoretical assumptions.

Lines 81–83: The stated aim combines a case series with a narrative literature review. Please clearly define the article type.

Literature Review and Rationale

Lines 85–96: The description of the narrative review is too general to assess how the literature was selected. Please explain how publications were chosen from the retrieved records.

Lines 207–210: Please specify who developed the PATH protocol, whether it has been previously published or validated, and whether it is linked to a specific manufacturer or product. This is relevant to the assessment of novelty and potential conflicts of interest.

3.1. Study Design and Objectives

Lines 225–232: The methods describe planned treatment, predefined eligibility criteria, scheduled follow-up visits, and prospective outcome collection. This appears inconsistent with the statement that the study was a retrospective analysis of routine clinical practice. Please clearly define the actual study design and revise the ethics statement accordingly.

In addition, full-face photographs with only the eyes covered cannot generally be considered anonymous. Please confirm that the patients provided written informed consent for the publication of potentially identifiable images, rather than only for the use of “anonymized” images.

Lines 228–244: A sample of three patients and a short follow-up period do not permit a meaningful assessment of the safety of the protocol.

Line 238: The assessments of “facial harmony” and “skin quality” are subjective. Please state who performed these evaluations, whether validated scales were used, and whether the assessor was independent.

Line 242: Please clarify whether the GAIS was completed by the patients, the physician, or both, and describe exactly what the scale assessed.

Lines 245–263: The method used to select the three patients is not reported. Please specify whether they represented all consecutive eligible patients treated during a defined period or whether they were retrospectively selected cases. This is essential for assessing the risk of selective presentation of favorable outcomes.

PATH Protocol and Treatment Products

Lines 265–288: The protocol is not described in sufficient detail to allow reproduction. Please provide the names and compositions of the products, doses, administered volumes, type of peel, number of sessions, and the exact homecare regimen.

Lines 282–285: Only approximate treatment intervals of 15–21 days are reported. Since the protocol was individualized and only three patients were included, the exact treatment schedule should be presented separately for each patient.

Lines 320–328: Please clarify whether mixing mesohyal™ Organic Silicon and mesohyal™ X-DNA in a single injectable solution is consistent with the manufacturers’ instructions and whether compatibility data are available.

The authors declare no external funding and no conflicts of interest, but they do not state whether the Mesoestetic products used throughout the study were purchased routinely or supplied free of charge by the manufacturer. This should be explicitly disclosed.

The complete composition of all products used in the study, preferably according to INCI nomenclature where applicable, should be provided at the beginning of the Materials and Methods section.

Tables

Tables 1 and 2: The tables are not formatted in accordance with the journal’s requirements and should be revised for clarity and consistency.

Discussion

Lines 653–666: The authors attribute the observed effects to the PATH protocol, although several active interventions were used simultaneously, including a topical retinoid with an independent effect on skin texture and quality, and no control group was included. It is therefore not possible to determine which component was responsible for the observed improvement.

Throughout the manuscript, conclusions regarding safety and efficacy should be formulated with considerable caution because of the extremely small sample size and short follow-up period.

Recommendation

I believe that the manuscript should be rejected because of substantial ethical concerns, the inconsistency between the declared retrospective nature of the study and the apparently planned treatment and assessment protocol, and the inability to draw reliable conclusions regarding efficacy and safety from three selected cases without a control group.

Author Response

Reviewer

The manuscript addresses a clinically relevant topic; however, several major methodological, ethical, and interpretative concerns should be resolved.

Abstract

Response:
We thank the Reviewer for this important observation. We agree that the expression “favorable safety profile” was too strong considering the limited sample size and the 60-day follow-up period. Accordingly, we have revised the Abstract to state that no serious adverse events or systemic complications were reported during the follow-up period. We have also removed the reference to a “favorable safety profile” from the Conclusions and clarified that the small number of patients and the short follow-up period do not allow definitive conclusions regarding the safety of the protocol.

Introduction

Lines 81–83: The stated aim combines a case series with a narrative literature review. Please clearly define the article type.

Response:

We thank the Reviewer for these important and constructive comments. The Introduction has been extensively revised accordingly.

First, we have distinguished well-established aesthetic procedures, such as chemical peeling and cross-linked hyaluronic acid fillers, from approaches supported by a more limited and heterogeneous body of evidence, including skin boosters, non-cross-linked hyaluronic acid formulations, and products combining hyaluronic and succinic acids. We have also clarified that, although preliminary findings suggest potential benefits in terms of hydration, firmness, elasticity, and overall skin quality, the specific clinical contribution of succinic acid has not yet been clearly established and requires further investigation.

Second, we have specified that the PATH protocol, an acronym for Profundity, Action, Time, and Homecare, was developed by mesoestetic® and is therefore associated with a specific manufacturer. We have clarified that the protocol has already been described and used clinically, including for treatment of the periocular region, and have added the available original source. However, because the currently available evidence remains limited and predominantly manufacturer-associated, we have explicitly stated that its previous clinical use should not be interpreted as formal or independent validation.

Third, statements concerning extracellular matrix remodeling, soft-tissue support, and skin-barrier function have been reformulated. These effects are now presented as component-specific biological rationales or theoretical assumptions rather than as established outcomes of the combined PATH protocol.

Finally, we have clarified that the present article is a descriptive case series involving three patients. The focused overview of the literature is included only to contextualize the biological and clinical rationale of the individual treatments and is not intended to constitute a standalone narrative review. The aim of the study has been revised accordingly.

Literature Review and Rationale

Lines 85–96: The description of the narrative review is too general to assess how the literature was selected. Please explain how publications were chosen from the retrieved records.

Response:

We thank the Reviewer for these important comments. The literature-review methodology has been expanded to clarify how publications were selected from the retrieved records. We now specify that titles and abstracts were screened by two authors (E.F. and L.S.), followed by full-text assessment of potentially relevant publications. The revised manuscript also reports the inclusion and exclusion criteria, the types of evidence prioritized, the procedure used to resolve uncertainties, and the final number of publications included in the narrative synthesis (n = 32).

We have also clarified the origin and status of the PATH protocol. The revised manuscript now specifies that PATH was developed by mesoestetic® and is therefore manufacturer-associated. Its previous clinical description has been cited, while also clarifying that previous use does not constitute formal independent validation and that the protocol has not yet been validated through large independent controlled studies. The novelty of the present article has consequently been defined as the retrospective description of its individualized application in three routine-care cases rather than the development of the PATH concept itself.

3.1. Study Design and Objectives

Lines 228–244: A sample of three patients and a short follow-up period do not permit a meaningful assessment of the safety of the protocol.

Line 242: Please clarify whether the GAIS was completed by the patients, the physician, or both, and describe exactly what the scale assessed.

Response:

We thank the Reviewer for these important observations. We agree that the previous wording did not clearly distinguish routine clinical care from a prospectively planned research protocol. The Methods section has therefore been revised to clarify that the article is a retrospective, single-center, descriptive case series based on routinely collected clinical data.

The treatments, clinical assessments, instrumental examinations, and follow-up visits were performed as part of routine clinical practice and were not assigned specifically for research purposes. The available clinical records and documentation were subsequently reviewed retrospectively for the present report. We have also clarified that the eligibility criteria were applied retrospectively to identify patients with sufficiently complete and comparable baseline and 60-day documentation.

We agree that a sample of three patients with 60 days of follow-up cannot provide a meaningful assessment of protocol safety. Safety assessment has therefore been removed as a study objective. The revised manuscript states only that treatment-related events documented during routine follow-up were retrospectively collected and descriptively reported, without drawing conclusions regarding the safety profile of the protocol.

We have also specified who evaluated facial harmony and skin quality, whether the assessor was independent or blinded, and whether validated scales were used. In the absence of validated scales, these outcomes are now explicitly described as subjective and descriptive. The GAIS methodology has been clarified by reporting who completed the scale and that it assessed overall aesthetic improvement compared with baseline.

Finally, we agree that full-face photographs with only the eyes covered cannot be considered completely anonymous. The revised manuscript now states that all patients provided written informed consent for publication of potentially identifiable clinical images and were informed that complete anonymity could not be guaranteed. The ethics statement has also been revised to reflect the retrospective, routine-care nature of the case series.

PATH Protocol and Treatment Products

The complete composition of all products used in the study, preferably according to INCI nomenclature where applicable, should be provided at the beginning of the Materials and Methods section.

Response:

We thank the Reviewer for these important comments. The Materials and Methods section has been extensively revised to improve the transparency and reproducibility of the treatment protocol.

First, we added a table at the beginning of the Materials and Methods section reporting the commercial name, manufacturer, quantitative composition of the injectable products, and complete INCI composition of the topical and homecare products, according to the respective product labels, instructions for use, and technical documentation.

The description of the PATH treatment procedures has also been expanded. For each product and procedure, we now report the concentrations, doses, administered volumes, anatomical areas treated, injection planes, injection devices and techniques, type of chemical peel, amount applied, number of layers, exposure time, removal or neutralization procedure, and total number of sessions. The post-procedural and homecare regimen has also been described in detail, including the amount applied, frequency, starting time, duration, retinoid suspension intervals, photoprotection, and behavioral recommendations.

Because these patients were treated as part of routine outpatient clinical care and the treatment pathway was individualized, the previously reported approximate intervals of 15–21 days have been removed. A new patient-specific table now reports, separately for each of the three patients, the exact date or day from baseline of every session, the procedures performed, products and volumes administered, anatomical areas treated, injection planes and techniques, homecare instructions, and timing of the final follow-up.

We have also clarified the administration of mesohyal™ Organic Silicon and mesohyal™ X-DNA. and the exact volume of each product administered during each session is now reported.

The combined administration was consistent with the manufacturer’s written instructions or technical confirmation, and the relevant supporting documentation has been cited or provided as Supplementary Material.

All products used during treatment were purchased through the clinic’s routine procurement process. No products, financial support, or editorial assistance were provided by Mesoestetic Pharma Group.

We have also clarified that Mesoestetic Pharma Group had no role in the clinical decision to treat the patients, treatment planning, retrospective case selection, clinical documentation, data analysis or interpretation, manuscript preparation, or the decision to publish.

Tables

Tables 1 and 2: The tables are not formatted in accordance with the journal’s requirements and should be revised for clarity and consistency.

We revised the tables according to journal’s requirements

Discussion

Throughout the manuscript, conclusions regarding safety and efficacy should be formulated with considerable caution because of the extremely small sample size and short follow-up period.

Response:

We thank the Reviewer for this important comment. The Discussion has been revised to clarify that, because several active interventions were used within the same treatment pathway and no control group was included, the observed improvements cannot be attributed to the PATH protocol as a whole or to any individual component. The possible independent contribution of chemical peeling, biorevitalization, topical retinoid therapy, and other homecare products is now explicitly acknowledged.

We have also substantially tempered all statements regarding efficacy and safety. The findings are now presented as short-term descriptive observations only, and we explicitly state that the three-patient sample and approximately 60-day follow-up do not permit conclusions regarding efficacy or a favorable safety profile. These limitations, together with the inability to isolate the contribution of individual treatment components, are now emphasized in the revised Discussion.

Recommendation

We thank the reviewer for raising this important point and agree that the absence of classification as a clinical trial of a medicinal product or as a clinical investigation of a medical device would not, by itself, be sufficient to determine the applicable ethical requirements.

The present manuscript, however, was not based on a prospectively initiated research study. It is a retrospective descriptive case series prepared from pre-existing clinical records of three patients who had already undergone treatment and routine clinical follow-up. The decision to select, analyse, and report these cases was made only after completion of the treatments and follow-up assessments.

Under Article 1 of the Italian Ministerial Decree of 30 January 2023 concerning the composition and functions of Territorial Ethics Committees, mandatory and exclusive Ethics Committee competence is expressly established for clinical trials of medicinal products, clinical investigations of medical devices, and pharmacological observational studies. For other types of studies and ethical matters, the Decree provides that Ethics Committees may perform consultative functions and that clinical centres may refer such matters for ethical evaluation.

Accordingly, this retrospective, non-pharmacological clinical case series was not considered to fall within the categories for which prior Ethics Committee review is expressly mandated under the applicable Italian regulatory framework. No prospective research protocol was implemented, and no treatment, visit, photograph, instrumental assessment, or other procedure was performed specifically for research purposes.

The expression “sequential treatment protocol” in the original manuscript referred to the routine clinical treatment pathway followed for these patients and not to a prospectively designed research protocol. To avoid any ambiguity, this expression has been revised throughout the manuscript.

In addition, separate written informed consent was obtained from all patients specifically for the publication of their clinical information and potentially identifiable facial

Reviewer 2 Report

Comments and Suggestions for Authors

Line 18 – define abbreviation
Line 53 – is this double space? Check whole manuscript
Line 76 – space is missing
Line 163 – reference is missing
Line 210 – is there reference for this protocol? If not, explain it was proposed by the authors
Line 225 – this study not article
Line 227 – abbreviation introduced
Line 241 – elaborate differences between 3 used devices. Are they all for scientific analysis or in clinic use?
Line 287 – have all 3 patients used same products? If not, mention that in limitation section
Line 451 – age already written
Title of table 1 is missing
Line 686 repetitive
Conclusion should be written according to the results and study design (case report)

Author Response

We thank the Reviewer for the careful reading of the manuscript and for these helpful comments. The manuscript has been revised accordingly, as detailed below.

Line 18 – define abbreviation
Response: The PATH abbreviation has now been defined at its first occurrence as Profondità/Depth, Azione/Action, Tempo/Timing, and Home care.

Line 53 – is this double space? Check whole manuscript
Response: The double spacing has been corrected, and the entire manuscript has been checked for similar formatting issues.

Line 76 – space is missing
Response: The missing space has been added.

Line 163 – reference is missing
Response: The appropriate reference has now been added.

Line 210 – is there reference for this protocol? If not, explain it was proposed by the authors
Response: The manuscript has been revised to clarify that the PATH protocol was developed by mesoestetic® and was not proposed by the present authors. A reference documenting its previous description has been added, and its manufacturer-associated origin and current limited independent validation are now explicitly stated.

Line 225 – this study not article
Response: “Article” has been replaced with “study”.

Line 227 – abbreviation introduced
Response: The abbreviation has been defined at its first occurrence and its use has been standardized throughout the manuscript.

Line 241 – elaborate differences between 3 used devices. Are they all for scientific analysis or in clinic use?
Response: The Methods section has been expanded to clarify the different functions of the three devices. OBSERV 520® was used for descriptive and iconographic comparison as part of routine clinical documentation, without quantitative outcome analysis. Antera 3D PRO® was used for quantitative assessment of selected skin surface and chromatic parameters, whereas QuantifiCare LifeViz® Infinity Pro was used for three-dimensional morphological and volumetric assessment.

Line 287 – have all 3 patients used same products? If not, mention that in limitation section
Response: We have clarified that all three patients received the same treatment products within the PATH pathway, while filler volume, anatomical distribution, and treatment details were individualized according to clinical presentation.

Line 451 – age already written
Response: The redundant age information has been removed.

Title of Table 1 is missing
Response: A title has now been added: “Summary of Baseline Clinical Characteristics, Treatment Parameters, and Follow-Up of the Three Cases.”

Line 686 – repetitive
Response: The repetitive statement has been removed/rephrased to improve clarity and avoid redundancy.

Conclusion should be written according to the results and study design (case report)
Response: The Conclusions section has been substantially revised to reflect the retrospective descriptive case-series design. Claims regarding efficacy and safety have been removed or tempered, and the findings are now presented as short-term descriptive observations from three patients, explicitly acknowledging the absence of a control group, the use of multiple concomitant interventions, and the limited follow-up.

Reviewer 3 Report

Comments and Suggestions for Authors

the subject is important.

the authors covers all aspects through a good review of previous manuscript.

the protocol is accepted

Author Response

We thank the reviewer for the comments about our manuscript.

Thank you very much.

Reviewer 4 Report

Comments and Suggestions for Authors

Thank you for the opportunity to review this manuscript. This study presents a preliminary three-patient case series evaluating the PATH protocol for integrated facial rejuvenation, supported by a narrative review of its clinical and biological rationale. The following comments are intended to improve the quality and interpretation of the manuscript.

Major Comments

It will be better if youfurther clarify the origin and novelty of the PATH protocol in the beginning. Otherwise readers may wonder whether this is simply a treatment plan proposed by the author or one that is theoretically grounded.
All images are based on visual descriptions, data charts could be addedto illustrate skin improvements more clearly.
The background colors and the proportions of the people in Figures 3 and 5 are different, and the images are not sharp enough.

Minor Comments

Sections 6.2 and 6.3 of the Discussion section both discuss mechanisms; these are not findings from this clinical trial, so they can be included in the brief review at the beginning of the paper.
You can compile a list of the types and durations of adverse reactions following each treatment to make this protocol more useful as a reference.

Recommendation: minor revision

You are encouraged to address the above comments regarding protocol clarification, image presentation, treatment details, and interpretation of mechanisms to improve the quality and clinical applicability of the manuscript.

Author Response

Response to Reviewer

We sincerely thank the Reviewer for the careful evaluation of our manuscript and for the constructive comments, which have helped us improve its clarity, methodological transparency, and clinical usefulness. The manuscript has been revised accordingly, as detailed below.

Major Comment 1. It will be better if you further clarify the origin and novelty of the PATH protocol in the beginning. Otherwise readers may wonder whether this is simply a treatment plan proposed by the author or one that is theoretically grounded.

Response:
The beginning of the manuscript has been revised to clarify both the origin and the novelty of the PATH protocol. We now specify that PATH was developed by mesoestetic® as a manufacturer-associated therapeutic framework and that it has previously been described and clinically applied, including in the periocular region. The relevant previous publication has also been cited. We further clarify that the novelty of the present report does not lie in the development of the PATH concept itself, but in its independent clinical characterization as an individualized, multilayer full-face rejuvenation framework, together with detailed documentation of its practical implementation and complementary clinical, multispectral, quantitative skin, and three-dimensional facial assessments.

Major Comment 2. All images are based on visual descriptions, data charts could be added to illustrate skin improvements more clearly.

Response:
To improve the quantitative presentation of the findings, we have added a data chart summarizing the changes in selected skin parameters derived from the quantitative Antera 3D PRO® assessments between baseline and T60. Quantitative data derived from QuantifiCare LifeViz® Infinity Pro are also reported where numerical software-generated measurements were available. OBSERV 520® findings remain presented descriptively, as this device was used exclusively for standardized multispectral photographic documentation and no quantitative outcome values derived from OBSERV 520® were used in the present case series.

Major Comment 3. The background colors and the proportions of the people in Figures 3 and 5 are different, and the images are not sharp enough.

Response:
Figures 3 and 5 have been revised to improve photographic comparability and image quality. Higher-resolution images were used, and the background appearance, cropping, facial proportions, and alignment were standardized as much as possible. The revised figures now provide a clearer and more consistent comparison between baseline and T60 while preserving the original clinical content of the photographs.

Minor Comment 4. Sections 6.2 and 6.3 of the Discussion section both discuss mechanisms; these are not findings from this clinical trial, so they can be included in the brief review at the beginning of the paper.

Response:
The mechanistic discussion regarding succinate, polynucleotides, calcium hydroxylapatite, and poly-L-lactic acid has been moved to the introductory literature review, where these concepts are now presented as biological background and theoretical rationale rather than as findings of the present case series. The corresponding section of the Discussion has been substantially shortened and now focuses on the interpretation of the clinical observations, explicitly emphasizing that no histological, biochemical, or molecular analyses were performed and that no specific biological mechanism or independent contribution of the individual treatment components can be demonstrated from these three cases.

Minor Comment 5. You can compile a list of the types and durations of adverse reactions following each treatment to make this protocol more useful as a reference.

Response:
We have added a dedicated table summarizing the treatment-related local reactions documented following each component of the PATH treatment pathway, together with their approximate duration and management. The revised manuscript now distinguishes reactions following chemical peeling, HA–succinate filler injection, intradermal biorevitalization, and the homecare regimen. All documented reactions were mild and transient and resolved spontaneously without additional medical intervention. Information regarding the absence of nodules, infections, delayed inflammatory reactions, vascular complications, and systemic adverse events during the available follow-up period has also been retained.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

hank you to the Authors for the detailed clarifications and for addressing all of the questions and concerns raised. The revisions have substantially improved the clarity of the manuscript and have adequately resolved the previously identified methodological and ethical issues.

In my opinion, the manuscript in its current revised form is suitable for publication.
However, I would recommend a few minor corrections before final acceptance. For example, in Figure 8, the images are labeled as a and b, while panel c is missing, although c is referred to and described in the figure legend. Please revise the figure labeling and legend to ensure consistency.