Comparative Evaluation of Three Different Pulpotomy Agents in Primary Molars

1. Background and Rationale Dental caries in primary molars often progresses to pulp involvement. Pulpotomy - removal of infected coronal pulp while preserving radicular vitality - is the standard treatment. Despite many available medicaments, no consensus exists on the optimal agent. Mineral trioxide aggregate (MTA) is highly biocompatible and bioactive, releasing calcium and hydroxyl ions to promote an alkaline, antibacterial environment and stimulate hard-tissue bridge formation. However, its high cost and handling difficulties limit universal use. Ferric sulfate (FS) is inexpensive and hemostatic, acting by forming ferric ion-protein complexes that control bleeding. It lacks long-term bioactivity; histological studies have demonstrated that FS-induced complexes can penetrate pulp tissue, causing persistent chronic inflammation and internal resorption over time. Chitosan is a natural polysaccharide derived from crustacean chitin. It has FDA approval for general medical use as a topical hemostatic and antimicrobial agent. Chitosan possesses intrinsic hemostatic capacity, broad-spectrum antimicrobial activity, biocompatibility, and biodegradability. In vitro studies suggest it may also enhance fibroblast proliferation and angiogenesis. Despite these properties, clinical data on chitosan as a pulpotomy medicament in primary teeth are limited to short-term, small-sample studies. No long-term randomized controlled trial has directly compared chitosan against MTA and FS. This study is designed to fill that gap by providing 18-month clinical and radiographic outcomes. 2. Specific Objectives * Primary objective: To compare the 18-month overall success rate (defined as simultaneous clinical and radiographic success) of MTA, FS, and a 2% chitosan hydrogel when used as pulpotomy agents in primary molars. * To test the null hypothesis that there is no statistically significant difference in treatment outcome among the three pulpotomy materials at 18 months. * To estimate risk differences, relative risks, and odds ratios for all pairwise comparisons, with Bonferroni correction applied to account for multiple testing (adjusted threshold α = 0.0167). * To document the clinical and radiographic success rates at each follow-up interval (3, 6, 12, and 18 months) for each material. 3. Trial Design and Key Methodological Details 3.1 Study Design and Setting * Design: Single-center, parallel-group, three-arm randomized controlled trial (RCT), adhering to the CONSORT 2010 statement. * Setting: Pediatric dentistry clinic, Al-Mustansiriyah University College of Dentistry, Baghdad, Iraq. * Ethics and registration: Approved by the institutional ethics committee (REC206, study number MUPEDO6); registered in the Thai Clinical Trials Registry (TCTR20260309003). 3.2 Randomization and Allocation Concealment * A computer-generated sequence with variable block sizes (allocation ratio 1:1:1) was prepared by an independent statistician not involved in clinical procedures. * Allocation concealment was maintained using sequentially numbered, opaque, sealed envelopes opened only after coronal pulp amputation and confirmation of hemostasis - ensuring that teeth with uncontrolled bleeding were excluded before group assignment. 3.3 Blinding * Operators could not be blinded due to the different physical forms of the materials (liquid, powder, gel). * All participants and parents/guardians were blinded to group assignment throughout the study. * The clinical outcome assessor (a calibrated pediatric dentist not involved in treatment) was blinded to treatment group at all follow-up visits. * The radiographic assessor (an experienced endodontist not involved in treatment) was blinded to group assignment. * Data analysts remained blinded to group assignment until analysis was complete. * Two pediatric dentists performed all procedures; cases were distributed approximately equally across operators and balanced across treatment groups. Inter-operator reliability was assessed on pilot cases (κ = 0.91). Intra-examiner consistency of the blinded clinical assessor: κ = 0.89. 3.4 Sample Size * Sample size was calculated based on published MTA and FS data, assuming an 18% absolute difference between groups (success rates: 98% for MTA and 80% for FS), with 80% power and α = 0.05. * This yielded 44 teeth per group. Accounting for an anticipated 20% attrition, the final sample was increased to 55 teeth per group (total N = 165). 3.5 Participants - Inclusion and Exclusion Criteria Inclusion criteria: * Healthy children aged 5 to 8 years. * At least one primary molar with carious exposure of the pulp indicated for pulpotomy. * Pulp assessed as clinically vital at the time of treatment (controlled bleeding within 5 minutes after coronal amputation). * Absence of spontaneous pain, swelling, abscess, fistula, or sinus tract. * Absence of pathological root resorption, furcal or periapical radiolucency on preoperative digital radiograph. * Tooth amenable to final restoration with a preformed stainless-steel crown. Exclusion criteria: * Necrotic pulp or irreversible pulpitis (failure to achieve hemostasis within 5 minutes). * Radiographic evidence of internal or external pathological root resorption, or more than one-third physiological root resorption. * Periapical or furcal radiolucency on preoperative radiograph. * Known allergy to shellfish or crustaceans (relevant to chitosan arm). * Systemic disease or medical condition contraindicating routine dental treatment. 3.6 Interventions - Material Application Protocols Ferric Sulfate (FS): * Product: Astringedent® 15.5% ferric sulfate solution (Ultradent Products Inc., South Jordan, UT, USA). * A cotton pellet saturated with FS solution was applied to the pulp stumps for 15 seconds, then gently rinsed with sterile saline. * A thick mix of reinforced zinc oxide-eugenol cement (IRM®, Dentsply Sirona) was then placed over the pulp chamber floor. Mineral Trioxide Aggregate (MTA): * Product: White ProRoot® MTA (Dentsply Maillefer, Tulsa Dental Specialties, Switzerland). * Mixed according to the manufacturer's instructions. * Placed as a 2-3 mm layer directly over the pulp stumps. Chitosan Hydrogel (CHI): * Preparation: Medical-grade chitosan powder (Sigma-Aldrich, Product No. 448877; degree of deacetylation: 85%; molecular weight: 300-350 kDa) was dissolved in 1% (v/v) acetic acid at 2% (w/v) concentration under constant magnetic stirring at room temperature for 24 hours. * Neutralization: Solution was neutralized to pH 7.0 ± 0.1 by dropwise addition of 1 M sodium hydroxide (NaOH) with continuous stirring; pH was monitored with a calibrated pH meter. * Sterilization: The hydrogel was aseptically dispensed into sterile single-use glass vials and sealed. Sealed vials were sterilized by gamma irradiation at 25-30 kGy (Cobalt-60 source; Steris Nordion, Ottawa, Canada) per ISO 11137 standards. Post-sterilization quality control confirmed sterility per USP \<71\>, viscosity 850 ± 50 cP at 25°C, and molecular weight 290 ± 20 kDa. * Storage: Vials were stored at 4°C in the dark and used within 3 months of manufacture. * Application: Sterile hydrogel was drawn into a sterile 3 mL syringe and applied 2-3 mm thick directly over the pulp stumps. Common restoration protocol (all three groups): * A reinforced glass ionomer base (Fuji IX GP, GC Corporation) was placed over the pulp chamber floor. * A preformed stainless-steel crown (3M™ ESPE™) was cemented and occlusion was adjusted. * A postoperative periapical digital radiograph was taken immediately to serve as baseline. 3.7 Follow-Up Schedule * Follow-up visits at: 3 months, 6 months, 12 months, and 18 months (primary endpoint). * Each visit included clinical examination and periapical digital radiograph. * Clinical examination performed by a blinded, calibrated pediatric dentist. * Radiographic assessment performed by a blinded endodontist not involved in treatment. 3.8 Outcome Definitions Clinical success (all criteria must be met): * Absence of spontaneous or provoked pain. * Absence of soft-tissue swelling or fistula. * Absence of sinus tract. * Absence of pathological tooth mobility. Radiographic success (all criteria must be met): * Absence of pathological internal root resorption. * Absence of pathological external root resorption (beyond normal physiological resorption). * Absence of furcal radiolucency. * Absence of periapical radiolucency. Overall success (primary endpoint): • Simultaneous fulfillment of all clinical AND radiographic success criteria at the 18-month visit. Failure handling: * Any tooth meeting any failure criterion at any recall visit was recorded as failed from that time point onward. * Missing follow-up data (if any) would have been classified as treatment failure (intention-to-treat principle). 3.9 Statistical Analysis Plan * All analyses performed using IBM SPSS Statistics v26.0, following an intention-to-treat approach. * Baseline characteristics compared using chi-square tests or one-way ANOVA as appropriate. * Success rates compared using chi-square tests or Fisher's exact tests. * Bonferroni correction applied for three pairwise comparisons: adjusted significance threshold α = 0.0167. * Effect measures: Risk differences (RD) with 95% confidence intervals (Newcombe's method), relative risks (RR), and odds ratios (OR) with exact confidence intervals. * Number needed to treat (NNT) calculated as 1/RD for statistically significant comparisons only. 4. Anticipated Limitations The following limitations were identified during study design and are acknowledged prospectively: * Operators cannot be blinded due to the different physical forms of the three materials; this is an inherent limitation of comparative pulpotomy trials. * Histological confirmation of pulpal healing is not feasible for ethical reasons in a clinical trial involving children. * The chitosan formulation used is prepared in-house; results may not be directly generalizable to commercially available chitosan products. * The study is conducted at a single center, which may limit generalizability. * Data on pulp exposure size, depth of carious lesion, and exact hemostasis time are not collected as separate variables. * The 18-month follow-up period, while robust, may not capture very late failures occurring prior to natural tooth exfoliation. * No data on cost-effectiveness, patient-reported pain outcomes, or quality of life are collected. 5. Relation to Existing Evidence Systematic reviews and meta-analyses consistently show that calcium-silicate cements, including MTA, achieve high pulpotomy success rates in primary molars, generally outperforming hemostatic agents. Ferric sulfate provides hemostasis through a chemical reaction with blood proteins but offers no bioactive stimulation; histological studies have linked FS to persistent chronic inflammation and internal resorption. Prior clinical studies of chitosan as a pulpotomy agent are limited to short follow-up periods (6-12 months) and small sample sizes. No previous randomized trial has compared chitosan directly to both MTA and FS over 18 months with simultaneous clinical and radiographic success criteria. This trial is designed to provide a more rigorous and longer-term assessment of chitosan's role in pediatric pulp therapy.