EVALUATION OF SALIVA AND SERUM HEME OXYGENASE, ARYLESTERASE AND NUCLEAR FACTOR ERYTHROID 2-RELATED FACTOR 2 LEVELS IN PATIENTS WITH STAGE III PERIODONTITIS
Trial Snapshot
- Phase
- Not Applicable
- Status
- Completed
- Enrollment
- 74
- Locations
- 1
- Primary Endpoint
- serum and saliva OSI levels
Study Overview
Brief Summary
Periodontal disease is a chronic and progressive inflammatory disease in which the hard and soft tissues that support the teeth are damaged. It is caused by the interaction between harmful bacteria and the body's immune responses, and most periodontal tissues are damaged by the body's abnormal response to these microorganisms and their products. When bacteria enter the body, immune cells (neutrophils) produce reactive oxygen species (ROS) in a process called the "respiratory burst". These ROS damage cells, causing tissue destruction through a variety of mechanisms, including DNA damage, fat oxidation and protein damage. Studies have shown that neutrophils from individuals with periodontal disease produce more ROS than neutrophils from healthy individuals. High amounts of ROS lead to oxidative damage to gum tissue, periodontal ligament and alveolar bone. Oxidative stress occurs when antioxidants in the body are insufficient or when high levels of ROS are present. Therefore, disruption of the balance between oxidant and antioxidant activities is considered an important cause of oxidative damage in periodontal tissues. Parameters such as total antioxidant status (TAS), total oxidant status (TOS) and oxidative stress index (OSI) are used to determine oxidative stress.
Furthermore, some enzymes such as arylesterase (ARE), heme oxygenase (HO) and nuclear factor erythroid 2-related factor 2 (NRF-2) are involved in defense mechanisms against oxidative stress. Many recent studies have shown a strong association between oxidative stress and periodontal disease.
Detailed Description
This study was divided into 2 groups, 37 systemically and periodontally health individuals and 37 systemically healthy individuals with stage III grade B periodontitis were included. Clinical periodontal parameters plaque index (PI), gingival index (GI), probing pocket depth (SCD), bleeding on probing (BOP) and clinical attachment level (CAL) were recorded and saliva and serum samples were collected. ELISA method was used for analyses of TOS, TAS, OSI, ARE, HO-1, NRF-2 levels.
Study Design
- Study Type
- Observational
- Observational Model
- Case Control
- Time Perspective
- Cross Sectional
Eligibility Criteria
- Ages
- 18 Years to 65 Years (Adult, Older Adult)
- Sex
- All
- Accepts Healthy Volunteers
- Yes
Inclusion Criteria
- •systemically healthy
- •between the ages of 18-65
Exclusion Criteria
- •Patients with systemic diseases and/or conditions that may affect periodontal tissues (cardiovascular diseases, immune system diseases, rheumatoid arthritis, diabetes mellitus, chemotherapy/radiotherapy).
- •taking any medication that may have an effect on periodontal tissues,
- •less than 20 natural teeth excluding third molars
- •used antibiotics, anti-inflammatory or immunosuppressive drugs
- •periodontal treatment in the last 6 months
- •pregnant or lactating women.
Outcomes
Primary Outcomes
serum and saliva OSI levels
Time Frame: 6 months
TAS values (initially in mmol Trolox Eq/L) were converted to µmol Trolox Eq/L. OSI was then computed using the following formula: OSI = \[(TOS, µmol/L) / (TAS, µmol Trolox Eq/L)\] × 100. The oxidative stress index is significantly higher in periodontitis patients compared to healthy individuals, reflecting an imbalance between oxidants and antioxidants.
serum and saliva TAS (mmol Trolox Eq/L) levels
Time Frame: 6 months
TAS concentrations were evaluated utilizing an innovative automated colorimetric assay, originally developed by Erel. The outcomes were reported as millimoles of Trolox equivalent per liter (mmol Trolox Eq/L). Total antioxidant levels are significantly lower in periodontitis patients compared to healthy individuals, suggesting increased oxidative stress associated with periodontal inflammation.
serum and saliva TOS (µmol H₂O₂ Eq/L) levels
Time Frame: 6 months
TOS concentrations were determined through a distinct automated colorimetric technique, also described by Erel. Results were expressed as micromoles of hydrogen peroxide equivalent per liter (µmol H₂O₂ Eq/L). Compared to healthy controls, periodontitis patients show higher total oxidant levels, which may contribute to tissue destruction and disease progression.
serum and saliva ARE (U/L) levels
Time Frame: 6 months
Arylesterase activity was assessed using commercially available assay kits (Rel Assay Diagnostics, Turkey). For the measurement of arylesterase activity, phenylacetate was used as the substrate. The enzymatic activity was calculated based on the molar extinction coefficient of the phenol formed (1,310 M-¹cm-¹). One unit of arylesterase activity was defined as the amount of enzyme required to hydrolyze 1 µmol of phenol per minute under the assay conditions and was also expressed as U/L. Arylesterase activity is significantly decreased in periodontitis patients compared to healthy controls, suggesting impaired antioxidant enzyme function.
serum and saliva NRF-2 (ng/ml) levels
Time Frame: 6 months
The level of NRF-2 was quantified using an enzyme-linked immunosorbent assay (ELISA) kit (BT lab.). In this assay, wells were pre-coated with human NRF-2-specific antibodies. Samples containing NRF2 were added to the wells, allowing target binding. Subsequently, a biotin-conjugated anti-NRF-2 antibody was applied, followed by streptavidin-horseradish peroxidase (HRP) conjugate. After washing to remove unbound components, a substrate solution was added, producing a colorimetric reaction proportional to the amount of NRF-2 present. The reaction was terminated by adding an acidic stop solution, and absorbance was measured at 450 nm. It is suggested that the reduced expression of NRF-2 in periodontitis patients, relative to healthy controls, contributes to increased oxidative stress and tissue damage.
serum and saliva HO-1(ng/ml) levels
Time Frame: 6 months
The concentration of HO-1 was measured using an ELISA kit (BT Lab.) Wells were pre-coated with human HO-1-specific antibodies. Following the addition of samples, human HO-1 proteins bound to the immobilized antibodies. A biotinylated anti-HO-1 antibody and subsequently streptavidin-HRP conjugate were introduced. After removal of unbound components via washing, a substrate solution was added to initiate color development, which was proportional to the HO-1 concentration. The reaction was stopped using an acidic solution, and absorbance was recorded at 450 nm. Compared to healthy subjects, HO-1 is considered to be less expressed in periodontitis patients, potentially compromising antioxidant and anti-inflammatory defenses
Secondary Outcomes
- Gingival Index (GI)(6 months)
- Plaque Index (PI)(6 months)
- Bleeding on Probing Index (BoP)(6 months)
- Clinical Attachment Loss (CAL) (millimeter)(6 months)
- Pocket Depth (PD) (millimeter)(6 months)
Investigators
Zeynep Hazan YILDIZ
Doctor
Saglik Bilimleri Universitesi