Relationship Apical Periodontitis and the Main Bacterial Species in the Oral Microbiota

Relationship of MMP-9 With the Clinical Course of Apical Periodontitis and the Main Bacterial Species in the Oral Microbiota

Aim: Apical periodontitis(AP) caused by root canal infection is the most frequent pathological lesion in the jaws. Bacterial products, host immune cells and biologically active factors called locally produced cytokines(such as IL-1, TNF-α) have been reported to play an important role in the pathogenesis of AP. Metalloproteinases(MMP), is a measurable biomarker that plays an important role in the degradation and regeneration of collagen and is an indicator of collagen. This study aimed to determine the main bacterial species in the microbiota as Gr(+) and Gr(-) and to compare the relationship between MMP-9 and TNF-α with controlled patient groups. Methodology:60 patients with AP and extraction indication were included in the study. 30 systemically and orally healthy volunteers were selected as the control group. After access cavity preparation, an initial microbiologic sample(S1) was taken from the root canal. After atraumatic extraction of the tooth, second microbial sample(S2) was taken from the external root surface. After bacterial DNA extraction, 16S rRNA gene primer was designed for sequence analysis. Bacterial community profiling was made by Sanger sequencing of the PCR products. Besides, blood samples were collected from all of the patients. Enzyme-linked immunosorbent assay was used to measure levels of MMP-9 and TNF-α.

Study Overview

• Status: Completed
• Conditions: Periapical Periodontitis; MMP-9
• Intervention / Treatment: Other: apical periodontitis

Detailed Description

60 patients with AP and extraction indication were included in the study. 30 systemically and orally healthy volunteers were selected as the control group. After access cavity preparation, an initial microbiologic sample(S1) was taken from the root canal. After atraumatic extraction of the tooth, second microbial sample(S2) was taken from the external root surface. After bacterial DNA extraction, 16S rRNA gene primer was designed for sequence analysis. Bacterial community profiling was made by Sanger sequencing of the PCR products. Besides, blood samples were collected from all of the patients. Enzyme-linked immunosorbent assay was used to measure levels of MMP-9 and TNF-α Approximately 450 patients who applied to xxxx University, Faculty of Dentistry, Endodontics Department for routine examination due to apical periodontitis complaints were examined and their demographic characteristics were recorded. Patients who had any acute or chronic inflammatory disease, known infection status, history of trauma or surgical operation within 6 months that may affect MMP-9 levels, pregnant and/or lactating, malignity, morbid obesity, arthritis, immunologic diseases, cardiovascular diseases or use of vitamins with high biotin were not included in the study. Additionally, patients who used antibiotics or anti-inflammatory drugs within the last 6 months, and patients with periodontal disease were excluded from the study. Of these patients, 60 patients with AP and extraction indication (due to restorative causes and lesion size) were included in the study. A total of 30 volunteers who were determined to be systemically and orally healthy (not having AP indication), were included in the study as the control group. The study was conducted between March 2022 and February 2023.

Demographic Characteristics & Clinical and Radiographic Evaluation Laboratory Analysis ELISA measurement procedure of MMP-9 and TNF-α levels Bacteriological Sampling from the Root Canal System and Outer Root Surface Genomic DNA Isolation and Measurement of DNA Concentration PCR (16S Universal primer) Purification and sequencing of the 16S rRNA gene Power Analysis and Sample Size Calculations

• Study Type: Interventional
• Enrollment (Actual): 60
• Phase: Not Applicable

Contacts and Locations

Study Locations

• Turkey
  • Esenler
    • Istanbul, Esenler, Turkey
      • Istanbul Medipol University, Faculty of Dentistry

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Adult; Older Adult
• Accepts Healthy Volunteers: Yes

Description

Inclusion Criteria:

• AP and extraction indication (due to restorative causes and lesion size) were included in the study

Exclusion Criteria:

• Patients who had any acute or chronic inflammatory disease, known infection status, history of trauma or surgical operation within 6 months that may affect MMP-9 levels, pregnant and/or lactating, malignity, morbid obesity, arthritis, immunologic diseases, cardiovascular diseases or use of vitamins with high biotin were not included in the study. Additionally, patients who used antibiotics or anti-inflammatory drugs within the last 6 months, and patients with periodontal disease were excluded from the study.

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Treatment
• Allocation: Randomized
• Interventional Model: Parallel Assignment
• Masking: Double

Number of Arms

2

Arms and Interventions

Participant Group / Arm	Intervention / Treatment
Experimental: apical periodontitis; 60 patients with AP and extraction indication (due to restorative causes and lesion size) were included in the study	Other: apical periodontitis; Of these patients, 60 patients with AP and extraction indication (due to restorative causes and lesion size) were included in the study
Experimental: control; A total of 30 volunteers who were determined to be systemically and orally healthy (not having AP indication), were included in the study as the control group.	Other: apical periodontitis; Of these patients, 60 patients with AP and extraction indication (due to restorative causes and lesion size) were included in the study

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Laboratory Analysis	Blood samples were collected from the patients for MMP-9 and TNF-α analysis. The blood samples taken from the patients in a tube with a gel separator were kept at room temperature for 20-30 minutes to coagulate, and then centrifuged at 1800 g for 10 minutes. T	The study was conducted between March 2022 and February 2023

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Bacteriological Sampling from the Root Canal System and Outer Root Surface	After plaque removal and rubber dam isolation, the operative field was cleaned with 3% hydrogen peroxide and disinfected with 2.5% NaOCl solution. Then, old coronal restorations if present and carious defects were removed, and an access preparation was completed when the root canal space was properly exposed. Afterwards, the tooth (including the pulp chamber), clamp and adjacent rubber dam were once again disinfected with 2.5% NaOCl, followed by inactivation with 10% sodium thiosulfate in order to avoid interference with bacteriological sampling. The working length (WL) was established 1-mm short of the apical foramen with an apex locator (Raypex6; VDW GmbH).	The study was conducted between March 2022 and February 2023
ELISA measurement procedure of MMP-9 and TNF-α levels	BT lab branded Human Matrix Metalloproteinase 9 and Human Tumor Necrosis Factor-α ELISA Kits (Shangai, China) were used to measure MMP-9 and TNF-α levels. The sensitivity of the kits is 15.12 ng/L and 1.520 pg/mL, respectively, and the measuring range of the kit is 30 - 9000 ng/L and 3 - 900 pg/mL, respectively. The within-run coefficient of variation is < 10%. In the study, measurements were made using BIO-TEK ELx500 ELISA Reader and BIO-TEK ELx50 (USA) washing devices.	The study was conducted between March 2022 and February 2023

Collaborators and Investigators

• Sponsor: Seyda Ersahan, DDS, PhD
• Investigators: Principal Investigator: Seyda Ersahan, Assoc.Prof., IstanbulMedipol University

Study record dates

Study Major Dates

• Study Start (Actual): November 15, 2021
• Primary Completion (Actual): November 15, 2022
• Study Completion (Actual): February 15, 2023

Study Registration Dates

• First Submitted: April 26, 2023
• First Submitted That Met QC Criteria: May 4, 2023
• First Posted (Actual): May 12, 2023

Study Record Updates

• Last Update Posted (Actual): May 12, 2023
• Last Update Submitted That Met QC Criteria: May 4, 2023
• Last Verified: May 1, 2023

More Information

Terms related to this study

• Additional Relevant MeSH Terms: Stomatognathic Diseases; Periodontal Diseases; Mouth Diseases; Jaw Diseases; Periapical Diseases; Periodontitis; Periapical Periodontitis
• Other Study ID Numbers: Oral Microbiota

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No