Effects of GLP-1 Receptor Agonists on Saliva and Oral Health

Stimulated saliva will be collected using the paraffin wax chewing method provided in the GC Saliva-Check Buffer kit (GC Corporation). Participants will chew the paraffin piece for 5 minutes, saliva will be collected into a graduated tube, and the flow rate will be calculated in milliliters per minute (mL/min).

Resting saliva pH will be determined using the GC Saliva-Check Buffer test kit (GC Corporation). A pH test strip will be immersed in the collected unstimulated saliva sample for 10 seconds and compared against the manufacturer's standardized color chart (ranges: 5.0 to 5.8 [highly acidic], 6.0 to 6.6 [moderately acidic], and 6.8 to 7.8 [healthy/neutral]). Higher pH values indicate a more neutral/healthy oral environment.

Evaluated using the GC Saliva-Check Buffer test kit (GC Corporation). One drop of stimulated saliva will be dispensed onto each of the three test pads on the strip. After 2 minutes, the color change on each pad will be scored from 0 to 4 (Green=4, Green/Blue=3, Blue=2, Red/Blue=1, Red=0). Scores are summed to give a total buffering capacity score ranging from 0 to 12 (0-5: very low; 6-9: low; 10-12: normal/high). Higher scores indicate better buffering capacity.

Assessed using the Silness and Löe Plaque Index (PI) on designated index teeth across buccal, lingual, mesial, and distal surfaces. Plaque presence is scored from 0 to 3 (0 = No plaque; 1 = A film of plaque adhering to the free gingival margin; 2 = Moderate accumulation of soft deposits; 3 = Abundance of soft matter within the gingival pocket and/or on the tooth margin). The individual mean score ranges from 0 to 3, where higher scores represent greater plaque accumulation and poorer oral hygiene.

Cumulative caries experience will be evaluated using the World Health Organization (WHO) Decayed, Missing, and Filled Teeth (DMFT) index (ranging from 0 to 28, where higher scores indicate greater cumulative caries experience). Early coronal caries lesions will be detected using the International Caries Detection and Assessment System (ICDAS-II), scoring tooth surfaces from 0 (sound) to 6 (extensive distinct cavity with visible dentin), and the highest ICDAS score per tooth will be recorded for tooth-level analysis. Higher DMFT and ICDAS-II scores represent worse caries experience and active decay.

Evaluated using the 5-item Summated Xerostomia Inventory (SXI-D) questionnaire. Participants rate xerostomia-related symptoms over the previous month using a 3-point Likert scale (1 = Never, 2 = Occasionally, 3 = Often). Total scores range from 5 to 15, where higher scores represent more severe subjective dry mouth symptoms.

Assessed using a custom, investigator-developed food frequency questionnaire. The questionnaire evaluates daily meal frequency, fluid intake, food preferences, avoidance of dry foods, and consumption frequencies (daily, weekly, monthly) of cariogenic and protein-rich foods. No overall summary score is calculated; responses will be analyzed and reported as categorical consumption frequencies and percentages.

Concentrations of secretory immunoglobulin A (sIgA) will be quantified in saliva supernatants using commercially available enzyme-linked immunosorbent assay (ELISA) kits according to the manufacturer's protocol. Values will be expressed in micrograms per milliliter (µg/mL) and normalized to total protein concentration.

Concentrations of salivary beta-defensin-3 will be quantified in saliva supernatants using commercially available enzyme-linked immunosorbent assay (ELISA) kits according to the manufacturer's protocol. Values will be expressed in picograms per milliliter (pg/mL) and normalized to total protein concentration.

Salivary total protein concentration will be determined using the Bradford protein assay with Coomassie Brilliant Blue G-250 dye and a standard bovine serum albumin calibration curve measured at 595 nm. Results will be expressed in milligrams per milliliter (mg/mL) and used to standardize salivary antimicrobial peptide levels.

Serum C-reactive protein (CRP) concentrations will be determined from fasting venous blood samples using standard clinical laboratory immunoturbidimetric assays. Values will be reported in milligrams per liter (mg/L).

Insulin resistance will be evaluated using the Homeostatic Model Assessment for Insulin Resistance (HOMA-IR), calculated with the standard formula: [fasting glucose (mg/dL) × fasting insulin (µIU/mL)] / 405. The resulting dimensionless index reflects insulin resistance, where higher values indicate greater insulin resistance.

Glycated hemoglobin (HbA1c) levels will be measured from venous blood samples using high-performance liquid chromatography (HPLC) or certified clinical laboratory immunoassays standardized to the National Glycohemoglobin Standardization Program (NGSP). Values will be reported as a percentage (%).

Body mass index (BMI) will be calculated using standardized anthropometric measurements as body weight in kilograms divided by the square of height in meters (kg/m2). Changes over the 6-month follow-up period will be evaluated and reported in kg/m2.