Composition Vendor Newgen KnowledgeWorks (P) Ltd. PLoS One plos plosone PLOS One 1932-6203 Public Library of Science San Francisco, CA USA 10.1371/journal.pone.0352702 PONE-D-25-63924 Study Protocol Medicine and health sciencesClinical medicineSigns and symptomsPain Engineering and technologyEquipmentOptical equipmentLasers Medicine and health sciencesOral medicineOrthodontics Computer and information sciencesArtificial intelligence Research and analysis methodsDatabase and informatics methodsDatabase searching Medicine and health sciencesEpidemiologyMedical risk factors Research and analysis methodsMathematical and statistical techniquesStatistical methodsMetaanalysis Physical sciencesMathematicsStatisticsStatistical methodsMetaanalysis Research and analysis methodsResearch assessmentSystematic reviews Efficacy of low-level laser therapy in pain relief during orthodontic treatment: Protocol for a systematic review and meta-analysis Efficacy of low-level laser therapy in pain relief during orthodontic treatment https://orcid.org/0009-0003-1982-6929 Ronchi Lemos Cintia Conceptualization Data curation Formal analysis Methodology Validation Visualization Writing – original draft Writing – review & editing 1 2 Bianchini Orlandi Daniela Data curation Formal analysis Investigation Methodology Writing – review & editing 1 2 Pereira Thimoteo Anna Carolina Data curation Formal analysis Investigation Methodology Validation Writing – review & editing 1 2 Polmann Helena Data curation Formal analysis Methodology Supervision Validation Writing – review & editing 1 2 ‡ https://orcid.org/0000-0002-1762-7059 Pauletto Patrícia Conceptualization Formal analysis Methodology Project administration Supervision Writing – review & editing 3 ‡ ¤ * Miron Stefani Cristine Conceptualization Formal analysis Methodology Supervision Validation Writing – review & editing 1 4 ‡ https://orcid.org/0000-0002-7622-1550 Neves Silva Guerra Eliete Methodology Validation Writing – review & editing 4 ‡ Flores-Mir Carlos Supervision Writing – review & editing 5 ‡ De Luca Canto Graziela Conceptualization Methodology Project administration Supervision Validation Writing – review & editing 1 2 6 ‡ Brazilian Centre for Evidence-Based Research (COBE), Brazil Federal University of Santa Catarina (UFSC), Department of Dentistry, Florianópolis, Brazil School of Dentistry, Universidad De Las Americas (UDLA), Quito, Ecuador University of Brasília (UnB), School of Health Sciences, Brasília, Brazil Mike Petryk School of Dentistry, University of Alberta, Edmonton, Canada Postgraduate Program in Evidence-Based Health, Federal University of São Paulo, São Paulo, Brazil Papageorgiou Spyridon N. Editor University of Zurich, SWITZERLAND * E-mail: patricia.pauletto@udla.edu.ec

The authors declare no commercial or noncommercial conflicts of interest.

Current address: School of Dentistry, Universidad De Las Americas (UDLA), Cristóbal Colón Avenue, E9-241 Quito, Ecuador

☯ These authors contributed equally to this work.

‡ HP,PP,CMS, ENSG, CFM and GDLG also contributed equally to this work.

 972026 2026 21 7 e0352702 4122025 1262026 2026 Ronchi Lemos et al This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Introduction

Patient adherence to orthodontic treatment depends on comfort, absence of pain, and improved quality of life. Pain and discomfort from procedures like interproximal separation or appliance activation are often underestimated by professionals. Low-level laser therapy, with minimal side effects, shows potential for pain relief in orthodontics, though its efficacy remains uncertain.

 Objectives

To investigate the efficacy of low-level laser therapy in controlling pain in orthodontic patients.

 Methods

A systematic review of randomized controlled trials with orthodontic patients assessing low-level laser therapy will be done. Studies with specific treatments, such as hybrid orthodontics with skeletal anchorage or intermaxillary elastics, will be excluded. The intervention will compare low-level laser therapy alone versus sham therapy, no treatment, or other interventions. No restrictions will apply regarding publication date, language, or patient age. Primary outcomes will be pain; secondary outcomes will be quality of life and adverse effects. Searches will be performed in Cochrane, Embase, LILACS, MEDLINE (via PubMed), and Scopus, plus gray literature (Google Scholar, ProQuest), reference lists, expert consultations, and ClinicalTrials.gov. Retrieved records will be merged in a reference manager, with duplicates removed, and imported into LASER AI software. Calibrated authors will independently select studies in two phases: title/abstract screening and full-text screening. Included studies will undergo narrative synthesis, and, if sufficiently homogeneous, meta-analysis. Risk of bias will be assessed with RoB 2, and certainty of evidence graded with GRADE.

 Author Anna Carolina Pereira Thimoteo received a PIBIC scholarship from CNPq (Brazilian National Council for Scientific and Technological Development). Data Availability All data generated from this review, including extracted datasets, data extraction forms, AI-assisted annotations, and statistical code, will be made publicly available in an open repository (Open Science Framework, OSF) upon study completion. Introduction

Patient adherence to orthodontic treatment can be influenced by pain and changes in quality of life, making it essential to address patients’ concerns and provide adequate management [1–3]. Cooperation may decrease after appliance placement, as pressure and intermittent pain are common, especially with fixed appliances [1]. During the first three months of treatment, pain peaks between 24 h and 3 days, and analgesic use reflects moderate pain perception [2]. Appliance type also affects discomfort, with fixed and functional appliances causing more pain than simple removable plates [3]. Psychological factors strongly influence adaptation, and managing expectations helps maintain adherence [1–3].

Low-Level Laser Therapy (LLLT) is a complementary dental treatment that benefits hard and soft oral tissues with few side effects [4]. It delivers low-intensity photons that modulate cellular activity without generating heat, promoting tissue repair and analgesia [5,6]. The main challenge is defining optimal parameters to achieve consistent clinical outcomes [6].

Evidence suggests that LLLT can alleviate pain after orthodontic activation, but variability in protocols, study quality, and possible placebo effects limit certainty [7–10]. For example, irradiation at 980 nm, 2.5 W/cm² and 600 J has shown significant pain reduction [11]. Nevertheless, the overall efficacy of LLLT for orthodontic pain remains uncertain, and standardized protocols are lacking [12].

Orthodontic pain is a frequent adverse effect of fixed appliances, aligners, separators, and orthopedic forces, with peak intensity 24–48 h after activation [13,14]. Up to 95% of patients experience pain from the inflammatory response involving cytokines, prostaglandins, and neuropeptides [15–18]. Since this mechanism is well established, identifying effective interventions is critical to improve adherence and comfort [1–3]. LLLT is a promising, non-invasive option with minimal adverse effects, yet studies show conflicting results and practical aspects such as cost and treatment time require consideration [4–8,19–21].

A recent randomized clinical trial (RCT) comparing low-level laser therapy with paracetamol–caffeine demonstrated clinically meaningful pain reductions, highlighting the need for a comprehensive synthesis of the available evidence [22]. Although two recent systematic reviews [20,23] examined orthodontic pain control, important gaps remain. One of them pooled several physical interventions without isolating LLLT, did not stratify by appliance type or laser parameters, and did not present certainty of the evidence [20], while another one focused only on separator-related pain and provided low-to-moderate certainty of evidence [23]. Neither incorporated a prospectively registered, peer-reviewed search with broad database and grey literature coverage, justifying a more comprehensive and methodologically rigorous review [20,23]. This review will also encompass different orthodontic treatment phases (such as separator placement, fixed appliance activation, and clear aligner insertion) to ensure a comprehensive synthesis of pain control across clinical scenarios.

Therefore, a new systematic review is needed to address these gaps. This review will evaluate the efficacy of LLLT for pain relief and postoperative sensitivity in orthodontic patients.

 Methods

The protocol for this review was developed following the PRISMA-P (Preferred Reporting Items for Systematic Review and Meta-Analysis Protocols) guidelines [24]. It was registered in the International Prospective Register of Systematic Reviews (PROSPERO) under the number CRD420251033037. A systematic review of intervention studies will be conducted by the methodology proposed in the Cochrane Handbook [25].

Study status and timeline

A preliminary search strategy was developed and pilot-tested to refine keywords, indexing terms, and database-specific adaptations. However, the full electronic search has not yet been conducted. The complete and final search is planned for June 2026, after which study selection, data extraction, and subsequent stages of the review will proceed according to the predefined protocol. The protocol is being submitted before the final search to ensure methodological transparency and avoid selective reporting, in accordance with PRISMA-P recommendations.

 Research question

“What is the efficacy of low-level laser therapy in alleviating pain, compared to other interventions, sham therapy, or no treatment, in patients undergoing orthodontic treatment?”

The research question was formulated based on the PICOS acronym (Population, Intervention, Comparison, Outcome, and Study Design), as outlined in Table 1.

10.1371/journal.pone.0352702.t001PICOS Framework for the Research Question.
Population Orthodontic patients undergoing
Intervention Low-Level Laser Therapy (LLLT)
Comparison Other interventions, sham therapy or no treatment
Outcome Primary outcomes: duration and intensity of painSecondary outcomes: quality of life, adverse effects
Study Design Randomized controlled trials
Eligibility Criteria Inclusion criteria.

Orthodontic patients of all ages will be included, whether undergoing treatment with fixed appliances or clear aligners. The intervention of interest is LLLT, encompassing various types and wavelengths, applied to the alveolar region. LLLT is considered within photobiomodulation therapy. However, this review will include only laser-based interventions and exclude LED-based PBMT due to differences in dosimetry and coherence. Eligible comparators will include other interventions, sham therapy, and no treatment. The primary outcome is pain intensity. Secondary outcomes include quality of life and patient satisfaction, and adverse effects of LLLT. Randomized controlled trials, including split-mouth and crossover designs (considering only data from the first assessment), will be included. No restrictions will be applied regarding language or publication date. If multiple tools are used to measure an outcome, the one most commonly adopted in the literature will be prioritized.

 Exclusion criteria

Studies involving specific treatments such as hybrid orthodontics or skeletal anchorage. These exclusions aim to reduce clinical heterogeneity, as skeletal anchorage and intermaxillary elastics involve different pain mechanisms that may confound the isolated effect of LLLT.

Studies involving patients under continuous use of medications (e.g., analgesics, anti-inflammatory drugs, corticosteroids) that could influence pain perception or confound the effect of the laser.

Studies in which the effect of the laser cannot be isolated from other interventions.

Reviews, letters, books, conference abstracts, case reports, opinion articles, technique articles, posters, and clinical practice guidelines;

Full text is unavailable, even after contacting the corresponding authors (three attempts in 3 weeks). Additional efforts will include contacting co-authors and searching repositories; if unavailable, data may be described narratively.

 Search Methods for Study Identification Information sources and search strategy.

A preliminary search strategy for PubMed was developed by the first author (C.R.L.) and improved with the assistance of a librarian. This strategy was peer-reviewed using the PRESS (Peer Review of Electronic Search Strategies) guideline to ensure accuracy and comprehensiveness by a second librarian. The peer-reviewed PubMed search strategy, provided verbatim (APPENDIX 1), will serve as the basis for and be adapted to the other databases. The search will be applied to the Cochrane, Embase, LILACS (in Spanish: Literatura Latinoamericana y del Caribe en Ciencias de la Salud), MEDLINE (via PubMed), and Scopus databases. Additionally, searches will be conducted in gray literature via ProQuest, Google Scholar, and Clinical Trials. Experts will be consulted, and references from included articles will be manually checked.

 Study records Data management.

The first author (C.R.L.) will compile all references into a single file and remove duplicates into a reference software manager (EndNote-WebTM, ClarivateTM, Jersey, USA).

 Methods for study selection

The selected studies will be imported into a screening platform, and the selection process will be conducted in two phases using LASER AI software (Evidence Prime Inc., https://www.evidenceprime.com/laser). In the first phase, three reviewers (C.R.L., D.B.O., and A.C.P.T.) will independently select titles and abstracts; in the second phase, they will apply the eligibility criteria to the full texts. A fourth reviewer (H.P.) will address discrepancies through consensus. The process will be sufficiently detailed to complete the PRISMA flow diagram [26].

 Data collection process

Data extraction using LASER AI will be conducted semi-automatically, integrating machine learning with human validation. Initially, the AI will be trained using initial manual annotations. Subsequently, the AI will process the articles and suggest extracted data. One reviewer (C.R.L.) will verify and adjust the extracted information, while two other reviewers (D.B.O. and A.C.P.T.) will reevaluate the corrections and identify any discrepancies. In cases of conflicts, a fourth reviewer (H.P.) will act as an arbitrator to facilitate consensus. Artificial intelligence (AI) tools (LASER AI, Evidence Prime Inc., and ChatGPT, OpenAI, model version Plus) will be used exclusively as assistive tools. All AI outputs will be independently verified by human reviewers. Prompts, outputs, and corrections will be documented to ensure auditability and reproducibility. AI will not be used for final decision-making. Once validated, the final data will be exported in Microsoft Excel spreadsheet format (Microsoft Corporation, Redmond, WA, USA) for statistical analysis (APPENDIX 2), thereby enhancing efficiency and standardization. If data are missing, the study authors will be contacted. Data from graphs can be extracted using WebPlotDigitizer (https://automeris.io/WebPlotDigitizer/) [27]. The data will be presented in tables in the article. Reports originating from the same study will be analyzed collectively.

 Data items

The variables to be extracted will be predefined, including study characteristics (author and year), study design, sample size (with sex distribution), and age (mean and standard deviation). Intervention groups will be categorized as laser, sham therapy, or control. Laser parameters such as wavelength (nm), power, irradiance, energy density, spot size, pulse mode, application time, number os sessions, irradiation sites, and dose (Joules) will also be extracted. Additionally, outcome measurement instruments, pain assessment time points, and results (mean and standard deviation) will be recorded. The results will specifically address the placement of separator elastics and the activation of orthodontic mechanics, with pain intensity evaluated at four predefined time points: T1 (up to 24 hours post-intervention), T2 (more than 24–48 hours), T3 (more than 48 hours to 7 days), and T4 (after 7 days). Meta-analyses will be conducted separately for each of these time points. If data for T3 or T4 are not reported, only T1 and T2 will be included.

 Outcomes and prioritization

The primary outcome is pain intensity, measured using the Visual Analog Scale (VAS) at the specified time points. Secondary outcomes include quality of life and patient satisfaction, assessed with the OHIP-14 (Oral Health Impact Profile) [26,27], analgesic consumption (when reported), and LLLT adverse effects. The latter will be recorded using a systematic checklist that categorizes events by type (e.g., headache, irritation) and intensity (mild, moderate, severe).

 Risk of bias analysis

The risk of bias will be independently assessed by two reviewers (C.R.L. and D.B.O.) using the Cochrane RoB 2 tool [28]. Given that the use of AI in evidence synthesis is evolving and requires caution, with evidence suggesting variability in accuracy compared to human reviewers (Taneri PE, 2025), AI such as ChatGPT (ChatGPT, OpenAI, 2024) [29] will be used only as an assistive tool. Specifically, ChatGPT will aid the reviewers by processing specific commands to answer questions for each domain of the tool. All AI outputs will be verified independently by three authors (C.R.L., D.B.O., and A.C.P.T.). Disagreements will be resolved through consensus meetings or adjudication by a fourth reviewer (H.P.). The evaluation will cover biases related to randomization, deviations from intended interventions, incomplete outcome data, outcome measurement, and reporting results, with the risk categorized as high, low, or “some concerns.” Judgments will be presented in tables using RevMan (RevMan 5 (Review Manager) Version 5.4 [Software] The Cochrane Collaboration, 2020) and included in the forest plot of the meta-analyses. Studies will be classified as having a low risk of bias if all key domains are rated as low risk, as having some concerns if any domain is rated as some concerns, and as high risk if any domain is rated as high risk.

 Data synthesis

A narrative synthesis of the included studies will be conducted, with an additional quantitative synthesis through meta-analyses if sufficient clinical and methodological homogeneity is identified. Meta-analyses will be performed using RevMan 5.4 (Review Manager, The Cochrane Collaboration, 2020), and a random-effects model will be applied. The specific effect measures (Relative Risks, Mean Differences, and Standartized Mean Differences) are detailed in the Effect Measures sub-section. Statistical heterogeneity will be assessed using the Chi² test (P < 0.10) and quantified with the I² statistic; between-study variance (τ²) will also be reported when available. For multi-arm trials, relevant intervention groups will be combined when appropriate, or comparator groups will be split to avoid double-counting. When multiple timepoints are reported, outcomes will be grouped according to predefined time windows, and a single timepoint per study will be selected for each window based on proximity to the defined interval.

If additional analyses beyond the capabilities of RevMan are required, statistical analyses may be supplemented using R software (e.g., meta or metafor packages).

 Effect measures

For dichotomous outcomes, results will be reported as RR with 95% confidence intervals (CI). For continuous outcomes, results will be expressed as mean differences (MD) when the same scale is used or as standardized mean differences (SMD) for different scales, with 95% CI. When scales differ, standardized mean differences will be used. VAS 0–100 mm scores will be converted to a 0–10 scale. The mean and standard deviation for each group will be calculated relative to the baseline and follow-up measurements using RevMan (RevMan 5 (Review Manager) Version 5.4 [Software] The Cochrane Collaboration, 2020). The SMD will be interpreted as a small effect (0.2), moderate effect (0.5), or large effect (0.8) [30].

 Unit of analysis

The unit of analysis will be the patient for randomized controlled trials and each side of the patient for split-mouth studies. The split-mouth studies will be grouped separately in different meta-analyses and will use paired analyses when possible. If the correlation is unavailable, it will be imputed and tested in sensitivity analyses. For crossover trials, only first-period data will be used.

 Handling of missing data

The corresponding author of the original studies will be contacted via email to obtain any missing data. If the data are not provided, they will be considered as missing. Only the available data will be analyzed. Missing SDs will be estimated from available statistics, and medians will be converted when appropriate. Sensitivity analyses will be performed to assess how sensitive the results will be to reasonable changes in the assumptions made [31]. All correspondence with the authors will be documented.

 Assessment of heterogeneity

Random-effects models will be used, with I² and τ² reported when available. Statistical heterogeneity will be assessed across the results of different trials using the Chi-squared (Chi²) test, with significance defined as P < 0.1. The I² statistics will quantify heterogeneity among the studies in each analysis. Heterogeneity will be categorized as “not important” (0% to 40%), moderate (30% to 60%), substantial (50% to 90%), and considerable (75% to 100%) [32]. Additionally, heterogeneity will be visually assessed by inspecting the overlap of CIs and may be explored through subgroup analyses.

 Subgroup analysis

To minimize multiplicity and focus on clinically relevant heterogeneity, subgroup analyses will be limited to three prespecified factors: [1] orthodontic appliance type (separator elastics, fixed appliance activation, removable appliance activation, or new aligner insertion); [2] laser type (low-level laser therapy vs. diode laser); and [3] wavelength (e.g., 660, 808, 940 nm). Additional subgroup analyses may include energy density, number of sessions, and comparator type if data allow. Analyses will be conducted as reported by the included studies.

Regarding comparator handling, each distinct active comparator will be assessed in its own meta-analysis. Different non-pharmacologic interventions will not be pooled in a single model, except for sham/placebo, which may be combined. Pharmacologic comparators may be pooled only if they belong to the same therapeutic class (e.g., different nonsteroidal anti-inflammatory drugs); otherwise, they will be analyzed separately.

 Sensitivity analysis

A sensitivity analysis will be performed to assess the robustness of the overall effect estimate. This analysis will examine the influence of studies with a high risk of bias and small sample sizes on the overall effect estimate. This will be done by removing studies with these characteristics from each meta-analysis to assess their influence on the results. Sensitivity analyses will include exclusion of high-risk studies, imputed data, and outliers.

 Assessment of publication bias

If more than 10 studies are included in any comparison, publication bias will be assessed through funnel plot analysis and using Egger’s test [33].

 Certainty of evidence assessment

The certainty of the evidence will be evaluated according to the Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) approach [34]. The assessment process will be conducted by the first author (C.R.L.) and verified by the second and third authors (A.C.P.T. and D.B.O.). A summary of the findings table will be created using the GRADEpro GDT software (GRADEpro: Guideline Development Tool, 2015) [35]. Five domains for downgrading the certainty will be evaluated: risk of bias, inconsistency, indirect evidence, imprecision, and publication bias. These domains will be classified as “not serious,” “serious,” or “very serious.” The certainty of the evidence will be categorized as high, moderate, low, or very low.

 Expected results and impacts

Pain is a common concern among patients undergoing orthodontic treatment, affecting their quality of life and treatment adherence. The LLLT presents itself as a non-invasive approach to pain relief and healing improvement, offering a promising alternative without the side effects of conventional medications. This study has the potential to assist healthcare professionals in clinical decision-making, enhancing treatment protocols and outcomes. A systematic review can identify knowledge gaps, guide future research, and develop evidence-based guidelines, leading to more effective treatments and greater patient satisfaction.

This systematic review aims to examine the evidence on the efficacy of laser therapy in reducing pain caused by orthodontic treatment [4]. The study stands out for its robust methodology, including a comprehensive search of five databases, grey literature, reference lists, and collaboration with experts, ensuring the reliability of the results. The transparent methodology facilitates replication, allowing independent validation by other researchers [36]. The review employs three blinded reviewers (C.R.L., D.B.O., and A.C.P.T.), supported by a fourth (H.P.) to avoid selection bias. The analysis provides valuable insights for professionals and patients considering laser therapy, potentially influencing therapeutic decisions and improving informed consent. Despite potential limitations, such as the risk of bias, this review can serve as a valuable resource, contributing to a comprehensive understanding of the efficacy of laser therapy and guiding informed decisions on pain relief in orthodontic treatment.

 Strengths and limitations of this study

This protocol is based on a comprehensive and peer-reviewed search strategy, including grey literature sources and expert consultation.

It incorporates artificial intelligence tools (Laser AI and ChatGPT) combined with double human cross-checking to enhance efficiency and accuracy in study selection and data extraction.

The variability in laser parameters and treatment protocols is expected to introduce heterogeneity, which may limit the feasibility of meta-analysis.

Exclusion of studies without available full texts may introduce a risk of publication bias.

 Supporting information PRISMA-P_completed.

(DOCX)

 Abbreviations PRESS

Peer Review of Electronic Search Strategies

LILACS

Latin American and Caribbean Health Sciences Literature

LLLT

Low-Level Laser Therapy

nm

nanometers

J

Joules

RCTs

Randomized Controlled Trials

VAS

Visual Analog Scale

ODs

Odds Ratio

RRs

Relative Risk

CI

Confidence Intervals

MD

Mean Differences

SMD

Standardized Mean Differences

GRADE

Grading of Recommendation, Assessment, Development, and Evaluation

We are grateful to the librarians Karyn Lehmkuhl from the Federal University of Santa Catarina, Brazil (email: karyn.lehmkuhl@ufsc.br), and Camila Belo Tavares Ferreira from National Cancer Institute (INCA), Brazil, for support in the development of the search strategies.

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Revisões sistemáticas da literatura: guia prático. 2020. 10.1371/journal.pone.0352702.r001 Decision Letter 0 Papageorgiou Spyridon Academic Editor 2026 Spyridon Papageorgiou This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Submission Version 0

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Reviewers' comments:

Reviewer's Responses to Questions

-->Comments to the Author

1. Does the manuscript provide a valid rationale for the proposed study, with clearly identified and justified research questions?

The research question outlined is expected to address a valid academic problem or topic and contribute to the base of knowledge in the field.-->

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

-->2. Is the protocol technically sound and planned in a manner that will lead to a meaningful outcome and allow testing the stated hypotheses?

The manuscript should describe the methods in sufficient detail to prevent undisclosed flexibility in the experimental procedure or analysis pipeline, including sufficient outcome-neutral conditions (e.g. necessary controls, absence of floor or ceiling effects) to test the proposed hypotheses and a statistical power analysis where applicable. As there may be aspects of the methodology and analysis which can only be refined once the work is undertaken, authors should outline potential assumptions and explicitly describe what aspects of the proposed analyses, if any, are exploratory.-->

Reviewer #1: Yes

Reviewer #2: Partly

Reviewer #3: Yes

**********

-->3. Is the methodology feasible and described in sufficient detail to allow the work to be replicable?

Descriptions of methods and materials in the protocol should be reported in sufficient detail for another researcher to reproduce all experiments and analyses. The protocol should describe the appropriate controls, sample size calculations, and replication needed to ensure that the data are robust and reproducible.-->

Reviewer #1: Yes

Reviewer #2: No

Reviewer #3: Yes

**********

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Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: No

**********

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Reviewer #1: Yes

Reviewer #2: No

Reviewer #3: Yes

**********

-->6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above and, if applicable, provide comments about issues authors must address before this protocol can be accepted for publication. You may also include additional comments for the author, including concerns about research or publication ethics.

You may also provide optional suggestions and comments to authors that they might find helpful in planning their study.

(Please upload your review as an attachment if it exceeds 20,000 characters)-->

Reviewer #1: This protocol outlines a planned systematic review and (conditional) meta-analysis to assess the efficacy of low-level laser therapy (LLLT; photobiomodulation) in reducing orthodontic pain across different treatment scenarios (e.g., separators, fixed appliance activation, aligner insertion). The authors will include randomized controlled trials, compare LLLT against sham/no treatment/other interventions, and evaluate primary (pain intensity/duration) and secondary outcomes (quality of life, adverse effects), using RoB 2 and GRADE to appraise risk of bias and certainty of evidence.

Technical novelty and innovation

The protocol focuses specifically on LLLT for orthodontic pain (rather than pooling heterogeneous physical modalities), aiming to stratify by orthodontic phase and laser parameters—an important refinement over prior reviews.

Explicit plan to use GRADE and RoB 2 should enhance methodological transparency and utility for clinicians.

Inclusion of gray literature and expert consultation increases comprehensiveness and mitigates publication bias.

Experimental rigor and validation

PROSPERO registration and adherence to PRISMA-P indicate prospective methodological planning and reporting.

Two-phase independent screening with calibration and duplicate processes is appropriate for minimizing selection bias.

Clarity of presentation

The clinical problem and rationale for a focused, up-to-date synthesis are well motivated; the target population, interventions, comparators, and outcomes are clearly defined at a high level.

Significance of contributions

Orthodontic pain is a common, clinically significant issue; clarifying the effectiveness and parameters of LLLT has direct implications for patient comfort, adherence, and clinical decision-making.

Technical limitations or concerns

Important statistical/analytical details are missing: prespecified timepoints for pain outcomes; choice of effect size metrics; handling of split-mouth/crossover designs; statistical model (random- vs fixed-effects); subgroup and sensitivity analyses; small-study effects assessment.

Exclusion criteria omit justification for excluding skeletal anchorage and intermaxillary elastics broadly despite their prevalence and clinical relevance; this may limit applicability.

Ambiguity about whether LED-based PBMT is included/excluded (modern PBMT often uses LEDs in addition to lasers).

Experimental gaps or methodological issues

The planned use of AI tools (LASER AI and ChatGPT) is not sufficiently detailed (versioning, tasks, safeguards, human oversight, reproducibility), raising transparency and bias concerns.

No explicit plan for data extraction items (laser parameters, dosimetry details, analgesic consumption), missing data handling, or contacting authors for outcome data.

No clear plan for dealing with multiple arms/timepoints (e.g., several wavelengths/doses or repeated measurements) and to prevent unit-of-analysis errors.

Clarity or presentation issues

Minor inconsistencies/typos (e.g., software name “LASER Al,” parameter notation “2.5 W/cm^2 and 600 J”), and PROSPERO ID formatting should be corrected; some sections are truncated (e.g., search strategy details).

Technical soundness evaluation

The PICOS framework is appropriate, but the protocol lacks essential pre-specifications:

Outcomes: Define primary timepoints (e.g., 6h, 24h, 48h, 72h, 7 days) and whether a composite (e.g., area under the pain-time curve) will be used. Predefine acceptable pain scales and how to convert heterogeneous scales; consider minimally important difference thresholds where available.

Effect measures: State the primary effect measure (MD for homogeneous scales like 0–100 mm VAS; SMD if mixed scales). Specify handling of skewed data, medians/IQR, and imputation of SD.

Design issues: For split-mouth trials, specify paired analyses and the approach to within-person correlation (use reported correlations, calculate from paired data, or impute with sensitivity analyses). For crossover trials, confirm use of first-period data only and explain how carryover will be managed.

Multiple arms/timepoints: Predefine how to select among multiple laser doses/wavelengths or multiple comparator arms (avoid double counting by appropriate arm combination or comparator splitting). For multiple post-intervention timepoints, either prespecify primary timepoint(s) or meta-analyze time windows separately.

Heterogeneity and meta-analysis model: State that a random-effects model (e.g., REML with Hartung-Knapp adjustment) will be used given expected clinical/methodological heterogeneity; report I² and τ²; plan meta-regression only if k is sufficient and covariates are prespecified (e.g., wavelength, irradiance, energy density, pulsing, number of sessions).

Subgroups/sensitivity analyses: Prespecify subgroups by treatment phase (separators vs first archwire activation vs aligners), wavelength (e.g., red vs NIR), energy density, irradiance, pulsing vs continuous-wave, application site (buccal vs buccal+palatal), number of sessions, comparator type (sham vs usual care), age group (adolescents vs adults), and risk of bias strata. Sensitivity analyses: exclude high RoB trials, imputed correlations/SDs, non-peer-reviewed sources, and outliers/influential studies.

Small-study effects/publication bias: Plan funnel plots and Egger’s regression (or Peters/Macaskill) where ≥10 studies per analysis; consider contour-enhanced plots.

Eligibility criteria:

Clarify and justify exclusions of skeletal anchorage, hybrid orthodontics, and intermaxillary elastics; these are common and clinically relevant. If the aim is to maintain homogeneity, consider including them but analyzing in prespecified subgroups—or justify exclusion with respect to differential pain mechanisms.

Clarify whether LED-based PBMT will be included (contemporary PBMT includes lasers and LEDs). Align terminology: LLLT vs PBMT.

Experimental evaluation assessment

Study selection and data extraction: Provide full details on calibration (pilot rounds and target kappa), number of reviewers for extraction, conflict resolution, and standardized extraction forms. Explicitly list data items: participant characteristics, appliance type, orthodontic procedure, comparator(s), pain measurement timing/instruments, analgesic use, LLLT parameters (device type, wavelength, power, power density, spot size/beam area, energy, energy density, pulse parameters/duty cycle, application points, time per point, sessions, contact mode), and adverse effects definitions/collection method.

Missing/incomplete data: State procedures for contacting authors (attempts, time windows) not only for full texts but also for missing outcome or parameter data; describe imputation rules where applicable.

Certainty of evidence: Identify which outcomes/timepoints will populate Summary of Findings tables; specify GRADE domains and how imprecision thresholds will be judged (e.g., optimal information size).

Comparison with related work (using the summaries provided)

Prior reviews have combined heterogeneous physical modalities or limited scope (e.g., separator pain only) and often did not grade certainty. Your plan to focus on LLLT across treatment phases and apply GRADE is appropriate. Additionally, contemporary PBMT trials in orthodontics and related dental applications exist (e.g., mini-implant peri-implant soft tissue outcomes), highlighting parameter sensitivity and the need for careful dosimetry capture—your planned parameter-level extraction/subgrouping will be valuable if explicitly prespecified.

Mechanistic reviews of LLLT/PBMT emphasize dose-response and wavelength dependence; bringing these into prespecified subgroup analyses may yield clinically actionable insights (e.g., red vs NIR wavelengths, pulsed vs continuous delivery).

Discussion of broader impact and significance

If executed with the suggested methodological refinements, this review can meaningfully guide clinical protocols (dosimetry, timing, and application points) and patient counseling about expected pain trajectories and adverse effects. The inclusion of quality-of-life and adverse event outcomes enhances translational value. Clear reporting of laser parameters will also support harmonization of future trials.

Questions for Authors

Will LED-based photobiomodulation trials (PBMT) be included, or are you restricting the review strictly to laser devices? Please clarify terminology and inclusion criteria (LLLT vs PBMT).

What are the prespecified primary timepoints for pain assessment (e.g., 24h, 48h, 72h)? Will you analyze time windows separately or use area-under-the-curve metrics?

Which effect size metrics will you use (MD vs SMD), and how will you handle heterogeneous pain scales, medians/IQR, and missing SDs?

How will you analyze split-mouth RCTs (paired analyses and correlation coefficients) and manage crossover trials (beyond first-period-only inclusion)? Please specify correlation imputation and planned sensitivity analyses.

What is your planned meta-analytic model (random-effects vs fixed) and heterogeneity metrics (I², τ²)? Will you use Hartung-Knapp adjustments?

Which subgroup and sensitivity analyses will be prespecified (e.g., wavelength, energy density, irradiance, pulsing, number of sessions, orthodontic procedure, comparator, age, RoB strata)?

How will you manage multi-arm trials (multiple doses/wavelengths) and multiple timepoints to avoid double counting and multiplicity?

Will analgesic consumption be included as a secondary outcome, given its frequent reporting and clinical relevance?

Please provide more detail on the AI tools: versions, specific tasks (screening, extraction), thresholds for human override, validation steps, data privacy safeguards, and how you will ensure reproducibility and auditability of AI-assisted decisions.

The protocol excludes studies involving skeletal anchorage and intermaxillary elastics. Could you justify these exclusions, or consider including them with planned subgroup analyses?

Reviewer #2: Manuscript title (revise to reflect content):

===========

Low‑Level Laser Therapy for Orthodontic Pain Relief: Protocol for a Systematic Review and Meta‑analysis

Small summary

==========

This manuscript describes a PRISMA‑P–based protocol (PROSPERO CRD420251033037) to systematically review randomized trials assessing low‑level laser therapy (LLLT) for pain control during orthodontic treatment. The authors plan broad database and grey‑literature searches, AI‑assisted screening/extraction (LASER AI and ChatGPT), RoB 2 risk‑of‑bias assessment, and GRADE certainty appraisal. The protocol outlines eligibility, outcomes (pain, QoL, adverse effects), and planned meta‑analyses by predefined time points.

Major points

========

(1) Title clarity: The current short title understates the planned meta‑analytic intent; change to include “systematic review and meta‑analysis” for accuracy.

(2) AI methods transparency: The protocol relies heavily on LASER AI and ChatGPT for screening/extraction but lacks sufficient detail on training, validation, reproducibility, and how AI decisions will be audited and reported. Provide explicit performance metrics and a reproducible workflow.

(3) Search timing and completeness: The full electronic search is deferred to January 2026; the protocol should clarify why submission precedes the planned search and ensure the peer‑reviewed search strategy (Appendix) is complete and reproducible.

(4) eligibility/exclusion bias risk: Excluding studies with unavailable full texts after only three contact attempts risks bias; justify this threshold and describe alternative approaches (e.g., contacting co‑authors, using interlibrary loan, or including abstracts with caution).

(5) Heterogeneity handling: Laser parameter variability is acknowledged but the statistical plan lacks detail on subgroup/meta‑regression strategies (wavelength, dose, application timing, appliance type). Predefine subgroup analyses and thresholds for pooling.

(6) Outcome measurement harmonization: Prioritize and justify the pain scales and time points; specify methods for converting different scales (e.g., VAS to standardized mean difference) and handling crossover/split‑mouth designs.

(7) Funding and competing interests: Financial disclosure fields appear incomplete and inconsistent with the competing‑interests statement; fully disclose all funding sources, grant numbers, and any role of funders.

(8) Data availability and ethics: The Data Availability statement is generic; provide a concrete plan for sharing extracted datasets, AI training annotations, and analysis code to ensure reproducibility.

Minor points

========

(1) Title punctuation and wording: Remove redundancy in the running title and ensure it matches the main title.

(2) Appendices: Ensure the full PubMed search string (PRESS‑reviewed) is included verbatim in Appendix 1 and that database‑specific adaptations are shown.

(3) LASER AI vendor details: Provide version number, date of access, and any custom settings used.

(4) Risk of bias plan: Clarify how RoB 2 judgments will be reconciled (e.g., consensus meeting, third reviewer) and how domain‑level judgments will inform sensitivity analyses.

(5) Statistical software: State the software and packages to be used for meta‑analysis (e.g., RevMan, R metafor) and thresholds for heterogeneity (I² cutoffs).

(6) Grammar/formatting: Minor typographical errors and inconsistent numbering in the Methods should be corrected for readability.

Recommended insertions of closely related work

==============================

(1) Owayda et al. — A randomized controlled trial on the effectiveness of low‑level laser therapy versus paracetamol‑caffeine for pain control during overall orthodontic treatment (Sci Rep. 2025; doi: 10.1038/s41598-025-16658-2). Place: Introduction, paragraph summarizing existing RCT evidence for LLLT. Insertion sentence: “Notably, a recent randomized trial comparing LLLT with paracetamol‑caffeine reported clinically relevant pain reductions, underscoring the need for a comprehensive synthesis of randomized evidence (Owayda et al., 2025; doi: 10.1038/s41598‑025‑16658‑2).”

(2) Al‑Hanbali et al. — The effectiveness of low‑level laser therapy and low‑intensity pulsed ultrasound in reducing pain induced by orthodontic separation: a randomized controlled trial (BMC Oral Health. 2024; doi: 10.1186/s12903-024-03926-2). Place: Introduction, when discussing separator‑related pain and prior focused reviews. Insertion sentence: “Randomized evidence addressing separator‑related pain has been produced (Al‑Hanbali et al., 2024; doi: 10.1186/s12903‑024‑03926‑2), but prior syntheses have not consistently stratified by appliance or laser parameters.”

(3) Mousa et al. — Evaluation of low‑level laser therapy and piezocision in the en‑masse retraction of upper anterior teeth (Eur J Orthod. 2025; doi: 10.1093/ejo/cjaf026). Place: Methods or Discussion, when justifying exclusion criteria and the need to isolate laser effects from adjunctive surgical interventions. Insertion sentence: “Studies combining LLLT with surgical acceleration techniques (e.g., piezocision) highlight the difficulty of isolating laser effects, supporting our exclusion of multimodal interventions where laser impact cannot be separated (Mousa et al., 2025; doi: 10.1093/ejo/cjaf026).”

(4) Alam et al. — The Effect of Low‑Level Laser Therapy on Accelerating Tooth Movement in Orthodontic Patients: A Randomized Controlled Trial (J Pharm Bioallied Sci. 2024; doi: 10.4103/jpbs.jpbs_1168_24). Place: Discussion, when addressing secondary outcomes and biological plausibility (LLLT effects on tissue/biologic response). Insertion sentence: “LLLT’s biological effects on tooth movement and tissue response have been reported in randomized trials, which informs interpretation of pain outcomes alongside potential mechanistic effects (Alam et al., 2024; doi: 10.4103/jpbs.jpbs_1168_24).”

Reviewer #3: Overall, this is a rigorous protocol for a systematic review to determine the effectiveness of low-level laser therapy in alleviating pain during orthodontic treatment. The authors provide a good rationale for conducting the study. The proposed analyses are appropriate to address the research question. The writing is clear and concise.

Major

1) The authors indicate that they will share all data generated by the study; however, there is no information about where the data will be stored and shared.

2) Although the authors indicate that they will cross check information generated by artificial intelligence tools, I recommend providing additional information and references providing evidence of the validity of using a) Laser AI to extract data from included studies and b) ChatGPT to collect data for the risk of bias assessment. These are emerging technologies without a proven track record of accurately conducting such tasks. For example, the following study examined the validity of ChatGPT’s risk of bias assessment: Taneri PE. Human versus artificial intelligence: comparing Cochrane authors' and ChatGPT's risk of bias assessments. Cochrane Evidence Synthesis and Methods. 2025 Sep;3(5):e70044.

3) The authors could include additional information about the following:

a) Why are studies with specific treatments such as hybrid orthodontics with skeletal anchorage or intermaxillary elastics being excluded?

b) Rationale for only including the first assessment from studies using cross-over or split mouth designs

Minor

4) The authors should review reference numbers to ensure that they match the study referenced. For example, on line 90, there is mention of two recent systematic reviews (20,22). It looks like 22 is not a systematic review; however, 21 is a systematic review. Also, line 209, it is unclear how references 25 and 26 relate to the information in the sentence referenced; line 214, reference 27 relates to GRADE not the Cochrane RoB 2 tool mentioned in the sentence.

5) On line, 171, the authors reference a reading platform. Consider naming the platform that will be used.

**********

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Reviewer #1: Yes:  Ricardo Ney Cobucci

Reviewer #2: No

Reviewer #3: No

**********

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 10.1371/journal.pone.0352702.r002 Author response to Decision Letter 1 Submission Version 1

9 Jun 2026

Response to Reviewers

Manuscript Title: Adverse effects of removable orthodontic aligners: a systematic review with single-arm meta-analysis

Manuscript ID: PONE-D-26-03043

Dear Reviewers,

We thank you for the careful evaluation of our manuscript and for the constructive comments provided. We have revised the manuscript thoroughly to address all concerns. Below, we provide a detailed, point-by-point response indicating how each comment was addressed and where the corresponding revisions can be found in the manuscript.

All changes are reflected in the revised manuscript and highlighted in the tracked-changes version.

GENERAL CHANGES

The manuscript title was revised to explicitly reflect the single-arm meta-analysis approach, improving methodological transparency.

The Data Availability Statement was updated to include a public repository with DOI, ensuring full reproducibility.

Funding information was standardized and aligned with journal requirements.

Supplementary materials were expanded and cross-referenced.

The manuscript underwent full linguistic and editorial revision to ensure clarity and standard academic English.

REVIEWER #1

Comment 1 – Abstract

Please add details in background regarding the type of review conducted.

Response:

We revised the Abstract to explicitly state that this is a systematic review with single-arm meta-analysis.

Location: Abstract, Background and Methods sections.

Comment 2 – Introduction

Provide more details about adverse effects.

Response:

We expanded the Introduction to include a clearer and more comprehensive description of adverse effects (e.g., periodontal changes, enamel demineralization, speech alterations, TMD symptoms).

Location: Introduction, paragraphs describing adverse outcomes.

Comment 3 – Rationale for meta-analysis

Add rationale for the meta-analysis.

Response:

We explicitly justified the use of single-arm meta-analysis due to absence of consistent comparator groups and heterogeneity in reporting.

Location: End of Introduction and Synthesis Methods section.

Comment 4 – PRISMA flowchart

Add study flow-chart.

Response:

A PRISMA 2020 flow diagram was included and referenced in the Results section.

Location: Results – Study selection (Fig 2).

Comment 5 – Grey literature

Why include grey literature?

Response:

We clarified that grey literature was included to reduce publication bias, particularly relevant in adverse effects research.

Location: Information Sources section.

Comment 6 – Reviewer calibration

Add calibration details.

Response:

We described the calibration exercise conducted prior to screening and risk-of-bias assessment.

Location: Study Selection and Risk of Bias sections.

Comment 7 – Discussion (clinical implications)

Expand discussion on dehiscence/fenestration.

Response:

We expanded this section to include clinical implications and the importance of treatment planning and monitoring.

Comment 8 – Halitosis (long-term evidence)

Add literature beyond 4 months.

Response:

We incorporated additional discussion on longer-term findings and clarified the transient nature and limitations of current evidence.

REVIEWER #2

Major Comments

1. Title accuracy

Response:

Title revised to explicitly include single-arm meta-analysis.

Location: Title page.

2. Search date and eligibility clarity

Response:

We clarified:

Search date (May 4, 2025)

Inclusion of early online/in-press studies

Location: Methods – Information Sources and Study Selection.

3. Data availability and transparency

Response:

A complete and consistent Data Availability Statement was added, including:

OSF repository

DOI

Full datasets and R scripts

Location: Results – Data Availability Statement.

4. Funding and role of funders

Response:

Funding details were standardized and clarified.

A statement on funder role was added (no involvement in study design or analysis).

5. Use of AI tools

Response:

We clarified:

AI was used only to identify text excerpts

All decisions were made by human reviewers

Prompts were included in Supplementary Material

Location: Risk of Bias section + S4 File.

6. Heterogeneity and pooling justification

Response:

We strengthened the justification for pooling under high heterogeneity, explicitly referencing:

Conceptual consistency across studies

Use of random-effects models

Sensitivity analyses

Location: Synthesis Methods.

7. GRADE transparency

Response:

We added explicit justification for each GRADE rating and provided detailed evidence profiles.

8. Outcome harmonization

Response:

We expanded the harmonization description and added:

Stepwise procedures

Worked examples in Supplementary Material (S5)

Location: Synthesis Methods + S5.

9. Selective reporting

Response:

We conducted a structured assessment of selective reporting and included:

Dedicated supplementary file (S6)

Summary paragraph in Methods

Location: Synthesis Methods.

10. Clinical interpretation

Response:

We revised conclusions to avoid unsupported comparisons with fixed appliances, clarifying that:

Only aligner arms were analyzed

Direct comparisons require dedicated trials

Minor Comments

All minor comments were addressed:

Abstract updated with study design breakdown

PRISMA details clarified

Tables and figures standardized

Terminology harmonized (e.g., ARR vs root resorption)

Statistical details expanded (r = 0.5; sensitivity r = 0.3–0.7)

Imputed data handled and sensitivity analysis described

PROSPERO included in abstract

Language edited for clarity

Supplementary materials completed and cross-referenced

EDITORIAL REQUIREMENTS (PLOS ONE)

We confirm that all journal requirements were addressed:

Data and code fully available (OSF repository)

Funding statement corrected and standardized

Ethics statement located in Methods only

Supporting Information labeled and cited appropriately

Tables and figures included and properly formatted

FINAL STATEMENT

We believe the revised manuscript now fully addresses the reviewers’ concerns and aligns with PLOS ONE methodological and reporting standards. The revisions substantially improve transparency, methodological clarity, and reproducibility.

We appreciate the reviewers’ insightful feedback and remain available for any further clarification.

Sincerely,

Patrícia Pauletto on behalf of all authors

Contact: + 593 99900 6460

Submitted filename: Response to Reviewers.docx

 10.1371/journal.pone.0352702.r003 Decision Letter 1 Papageorgiou Spyridon Academic Editor Papageorgiou Spyridon Academic Editor 2026 Spyridon Papageorgiou This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Submission Version 1

14 Jun 2026

Efficacy of Low-Level Laser Therapy in Pain Relief During Orthodontic Treatment: Protocol for a Systematic Review and Meta-analysis

PONE-D-25-63924R1

Dear Dr. Pauletto,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

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Kind regards,

Spyridon N. Papageorgiou, DDS, Dr Med Dent

Academic Editor

PLOS One

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 10.1371/journal.pone.0352702.r004 Acceptance letter Papageorgiou Spyridon Academic Editor Papageorgiou Spyridon Academic Editor 2026 Spyridon Papageorgiou This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

PONE-D-25-63924R1

PLOS One

Dear Dr. Pauletto,

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS One. Congratulations! Your manuscript is now being handed over to our production team.

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* All references, tables, and figures are properly cited

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* There are no issues that prevent the paper from being properly typeset

You will receive further instructions from the production team, including instructions on how to review your proof when it is ready. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few days to review your paper and let you know the next and final steps.

Lastly, if your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

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Kind regards,

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on behalf of

Dr. Spyridon N. Papageorgiou

Academic Editor

PLOS One