Cryotherapy, PGE2, NGF Levels and Pain After Endodontic Retreatment

Official title

Evaluation of the Relationship Between PGE2, NGF, and Postoperative Pain in Patients With and Without Intracanal Cryotherapy in Single-visit Endodontic Retreatment

Overview

This prospective, randomized, controlled, single-blind, parallel-group clinical trial evaluates whether intracanal cryotherapy reduces postoperative pain and inflammatory biomarker levels following single-visit endodontic retreatment of teeth with asymptomatic apical periodontitis. A total of 100 patients (18-65 years, systemically healthy, ASA I-II) requiring non-surgical retreatment of a single-rooted mandibular premolar tooth will be randomly assigned via computer-generated randomization to one of two equal groups (n=50 each): a cryotherapy group, receiving intracanal final irrigation with sterile saline cooled to 2.5°C for 5 minutes, and a control group, receiving the same irrigation protocol with saline at room temperature (20-22°C). Salivary nerve growth factor (NGF) and gingival crevicular fluid (GCF) prostaglandin E2 (PGE2) levels will be measured by ELISA at baseline (T0), 24 hours (T1), and 72 hours (T2). Postoperative pain will be assessed using a visual analog scale (VAS, 0-100 mm) at 6, 12, 24, 48, and 72 hours and on day 7, with the 24-hour VAS score as the primary comparison point. Rescue analgesic (paracetamol) consumption will also be recorded and compared between groups. The study aims to determine whether intracanal cryotherapy lowers postoperative pain and biomarker levels compared to standard-temperature irrigation, and to evaluate the correlation between PGE2/NGF levels and pain intensity.

Detailed description

Postoperative pain following endodontic retreatment is associated with periapical inflammation mediated by prostaglandin E2 (PGE2) and nerve growth factor (NGF). Intracanal cryotherapy has been proposed to reduce postoperative pain through local hypothermia-induced vasoconstriction, neuropraxia, and reduced metabolic and inflammatory mediator activity. While prior studies have examined cryotherapy's effect on postoperative pain, no randomized controlled trial has jointly measured PGE2 and NGF using minimally invasive sampling methods (GCF and saliva) in the context of single-visit retreatment.

Eligible patients will be systemically healthy adults (18-65 years, ASA I-II) with a single-rooted mandibular premolar tooth requiring non-surgical retreatment due to a previously inadequate root canal treatment, presenting with asymptomatic apical periodontitis and periapical radiolucency (PAI ≥ 3). Key exclusion criteria include pregnancy or lactation, uncontrolled diabetes mellitus (HbA1c \> 7%), autoimmune disease, immunosuppressive therapy, systemic corticosteroid use within the past 3 months, BMI \> 35 kg/m², acute infection (abscess/cellulitis), advanced periodontal breakdown, heavy smoking (\>10 cigarettes/day), recent analgesic/NSAID or antibiotic use, and chronic pain syndromes.

Patients will be randomly allocated (computer-generated randomization, sealed envelopes) to one of two parallel groups:

Group A (Cryotherapy): Following standard non-surgical retreatment instrumentation, final intracanal irrigation with sterile saline cooled to 2.5°C, 20 mL, over 5 minutes.

Group B (Control): Identical protocol using sterile saline at room temperature (20-22°C), 20 mL, over 5 minutes.

All treatments will be performed in a single visit by one calibrated operator following a standardized protocol; the two groups differ only in irrigation temperature. Given the nature of the intervention, the operator cannot be blinded; however, laboratory personnel analyzing saliva/GCF samples and statisticians will remain blinded to group allocation (single-blind design).

Outcome measures:

Primary: Postoperative pain intensity measured by VAS (0-100 mm) at 6, 12, 24, 48, and 72 hours and on day 7, with the 24-hour time point as the primary comparison.

Secondary: Salivary NGF levels (ELISA) and GCF PGE2 levels (ELISA) at baseline (T0), 24 hours (T1), and 72 hours (T2); rescue analgesic (paracetamol 500 mg, oral, as needed) consumption over 7 days; and correlation between biomarker levels and VAS pain scores.

Each patient will be followed for a total of 7 days. Saliva and GCF samples will be collected between 08:00-10:00 to minimize circadian variation. Sample size was calculated using G\*Power 3.1.9.7 (repeated-measures ANOVA, 80% power, α=0.05, f=0.25), yielding a minimum of 34 participants per group; accounting for an estimated 20% dropout rate, 50 patients per group (100 total) will be enrolled. Statistical significance will be set at p\<0.05. The study is sponsored and funded internally by Alanya Alaaddin Keykubat University, with no external funding or industry sponsorship.

Interventions

• Procedure Intracanal cryotherapy
  Final intracanal irrigation with sterile 0.9% saline solution cooled to 2.5°C (using an ice-water bath), 20 mL administered over 5 minutes, performed after canal instrumentation during single-visit non-surgical endodontic retreatment.
• Procedure Room-temperature saline irrigation
  Final intracanal irrigation with sterile 0.9% saline solution at room temperature (20-22°C), 20 mL administered over 5 minutes, performed after canal instrumentation during single-visit non-surgical endodontic retreatment - identical protocol to the cryotherapy group except for irrigant temperature.

Primary outcome measures

• Postoperative pain intensity (VAS score) [Time frame: Baseline, then at 6, 12, 24, 48, and 72 hours and on Day 7 postoperatively (primary comparison at 24 hours)]

Secondary outcome measures (2)

• Salivary nerve growth factor (NGF) levels [Time frame: Baseline (T0), 24 hours (T1), and 72 hours (T2) postoperatively]
• Gingival crevicular fluid (GCF) prostaglandin E2 (PGE2) levels [Time frame: Baseline (T0), 24 hours (T1), and 72 hours (T2) postoperatively]

Eligibility criteria

Inclusion criteria

• Age 18-65 years, systemically healthy individuals (ASA I-II)
• Single-rooted mandibular premolar teeth with a previous root canal treatment, requiring non-surgical retreatment due to asymptomatic apical periodontitis
• Presence of periapical radiolucency (Periapical Index \[PAI\] ≥ 3)
• No antibiotic use within the past 1 month
• Willing to participate and has signed the informed consent form

Exclusion criteria

• Pregnancy or lactation
• Uncontrolled diabetes mellitus (HbA1c > 7%), diagnosed autoimmune disease, or receiving immunosuppressive therapy
• Systemic corticosteroid use within the past 3 months
• Body mass index (BMI) greater than 35 kg/m²
• Presence of acute infection (e.g., acute apical abscess or cellulitis)
• Conditions precluding retreatment, including advanced periodontal breakdown (clinical attachment loss > 5 mm), root resorption, perforation, or advanced canal calcification
• Absence of an opposing (antagonist) tooth resulting in lack of functional occlusal contact
• Diagnosed chronic pain syndrome or currently receiving active psychiatric treatment
• Smoking more than 10 cigarettes per day
• Use of analgesics or NSAIDs within the past 24 hours
• Antibiotic use within the past 1 month

Criteria are shown verbatim from the registry (in English). Final eligibility is always assessed by the study center.

Healthy volunteers: No

Study design

Allocation

Randomized

Model

Parallel assignment

Masking

Double blind

Primary purpose

Treatment

Study locations

Turkey (Türkiye) · 1 center

• Alanya Alaaddin Keykubat University, Faculty of Dentistry — Antalya

Publications

• Kaya Mumcu A, Tuzcu E, Kurnaz S, Kiraz G. Evaluation of the effect of intracanal cryotherapy on postoperative pain after retreatment: a randomized controlled clinical trial. Clin Oral Investig. 2025 Jun 25;29(7):355. doi: 10.1007/s00784-025-06435-w. PMID 40560202
• Keskin C, Aksoy A, Kalyoncuoglu E, Keles A, Ilik AA, Komec O, Yuzgulec E, Akgun H, Alak SG, Tokur O. Effect of intracanal cryotherapy on the inflammatory cytokine, proteolytic enzyme levels and post-operative pain in teeth with asymptomatic apical periodontitis: A randomized clinical trial. Int Endod J. 2023 Aug;56(8):932-942. doi: 10.1111/iej.13937. Epub 2023 May 30. PMID 37222468
• Vera J, Ochoa J, Romero M, Vazquez-Carcano M, Ramos-Gregorio CO, Aguilar RR, Cruz A, Sleiman P, Arias A. Intracanal Cryotherapy Reduces Postoperative Pain in Teeth with Symptomatic Apical Periodontitis: A Randomized Multicenter Clinical Trial. J Endod. 2018 Jan;44(1):4-8. doi: 10.1016/j.joen.2017.08.038. Epub 2017 Nov 1. PMID 29079057

Identifiers

NCT: NCT07733700 · ALKU-EC-2026-44

Primary sources (government registries)

• ClinicalTrials.gov (US NIH)

View this study on ClinicalTrials.gov ↗