This study was conducted at the Oral and Maxillofacial Radiology Department of School of Dentistry, Shahid Beheshti University of Medical Sciences within a 6-month period in 2022. The study was approved by the ethics committee of the university (IR.SBMU.DRC.REC.1400.138) and registered in the Iranian Registry of Clinical Trials (IRCT20211229053566N1).

Trial design

A clinical trial was conducted in which GT of patients was measured at 40 points first with the intraoral probe of an ultrasound and then by transgingival probing. The results were reported in accordance with the Consolidated Standards of Reporting Trials.

Participants, eligibility criteria, and settings

The inclusion criteria were candidates with healthy periodontium and presence of all anterior and premolar teeth in the oral cavity. Due to the use of local anesthetic injection, it was crucial for ethical reasons that this study would have a gain for the patients thus candidates for crown lengthening surgery with a healthy gingiva were selected.

The exclusion criteria were tobacco use, intake of medications affecting gingival health such as phenytoin, pregnancy, presence of periodontal pockets, gingival inflammation, and gingival recession at the measurement sites.

The sample consisted of 34 patients including 17 eligible patients with a thick gingival biotype and 17 with a thin gingival biotype (as determined by visual inspection by a periodontist) that were selected after assessment of patient records and clinical periodontal examination of patients.

Interventions

The records of patients presenting to the School of Dentistry of Shahid Beheshti University of Medical Sciences and a private radiology clinic in Tehran city who required crown lengthening surgery were assessed. Eligible patients underwent clinical periodontal examination, and after ensuring their gingival health and no inflammation or periodontal pockets around their anterior teeth and premolars, all teeth were probed at three points in the buccal surface using a UNC-15 periodontal probe (Premier Dental, USA). Written informed consent was obtained from all patients prior to their enrollment. All GT measurements were made by one operator with high reliability under the supervision of an experienced radiologist.

For measurement of GT by ultrasonography, the patients were asked to have an upright position on dental chair, with their head in natural head position. The intraoral probe of ultrasound (B-Scan, E-CUBE 7, ALPINION, Korea) with 2 cm length and 12 MHz frequency was disinfected, and lubricant gel was applied over the probe detector as a conducting medium. The probe was positioned vertically at the mid-buccal region of anterior teeth and premolars of both jaws such that part of the probe was on the tooth and part of it was on the mid-buccal gingiva (Fig. 1). GT was measured at all 40 points by the measurement tool of ultrasound.

Fig. 1

Positioning of intraoral probe of ultrasound for measurement of gingival thickness

Real-time images displayed on the monitor indicated the buccal surface of the tooth, cementoenamel junction, and buccal bone covering the root surface in the sagittal plane (Fig. 2). The entire hypoechoic tissue from the superior image margin (indicating the probe surface on the tooth and gingiva) to the hyperechoic area related to tooth or bone indicated the GT at the mid-buccal region. The measurement tool of the software was used to mark 2- and 4-mm distances from the free gingival margin (FGM), and then measure the GT from the superior margin of the image to the bone surface using the same measurement tool (Fig. 3). The measured values were recorded in Microsoft Excel 2019.

Fig. 2

Real-time images showing the buccal surface of the tooth, cementoenamel junction, and buccal bone covering the root surface in the sagittal plane. In the right side, a hyperechoic area at the top (A-arrow) indicates the clinical tooth crown (buccal surface) on which, the probe is placed. The next hyperechoic line (C-arrow) indicates the buccal cortical plate. The intersection of the above mentioned two hyperechoic lines indicates the cementoenamel junction of the tooth (B-arrow). The entire hypoechoic tissue from the superior image margin (indicating the probe surface on the tooth and gingiva) to the hyperechoic area related to tooth or bone indicates the gingival thickness at the mid-buccal region

Fig. 3

Measuring the gingival thickness (GT) at 4 mm distance from the free gingival margin. Points A and B indicate the gingival margin, and 4 mm distance from the free gingival margin, respectively. As shown, the GT at point B was 0.8 mm

For measurement of GT by the transgingival probing method, infiltration anesthesia was induced in the gingiva adjacent to the respective teeth by local injection of a few drops of 2% lidocaine plus 1:80,000 epinephrine (3% Citanest for patients with contraindications for lidocaine such as cardiovascular patients). After 10 min, a periodontal probe was used to mark 2- and 4-mm distances from the FGM at the mid-buccal region of the respective tooth (Fig. 4). Next, a #25 endodontic finger spreader (25 mm, 2% convergence; Mani, Japan) was vertically inserted into the gingival tissue until it reached the underlying bone for measurement of GT at the marked points. Upon contact with bone, the rubber stop of the spreader was adjusted and the spreader was removed. The length indicated by the rubber stop was measured by a digital caliper (Tech instruments, China) with 0.01 mm accuracy. The measured values were recorded in Microsoft Excel 2019.

Fig. 4

Marking a point at 2 mm distance from the free gingival margin at the mid-buccal of right central incisor by a periodontal probe and insertion of an endodontic spreader into the gingival tissue at this point for measurement of gingival thickness

After conventional measurement of the GT at 4 mm distance from the FGM, the patients were accordingly assigned to two groups of thick (> 1.5 mm) and thin (< 1.5 mm) gingival biotype [5].

Outcomes (primary and secondary)

The main objective of this study was to compare GT measured by the intraoral probe of ultrasound with GT measured by transgingival probing. Comparison of GT measured by the two methods in the maxilla and mandible, anterior and premolar regions of the jaw, and at different levels from the FGM were the secondary outcomes.

Sample size calculation

The sample size was calculated to be 17 according to a study by Savitha and Vandana [9] assuming α = 0.05, study power of 0.90, standard deviation of 0.25, and minimum significant difference of 0.2 between the two methods.

To increase the sample size compared with similar previous studies [9, 11], comparison of anterior and premolar regions of the jaws, and apico-coronal assessment of teeth, 40 points were assessed in each individual including two points at 2- and 4-mm distances from the FGM at the mid-buccal of the teeth from the left to the right second premolars in both the maxilla and mandible.

Interim analyses and stopping guidelines

No interim analyses were performed, and no stopping guidelines were established.

Randomization

Not applicable.

Blinding

Blinding of the operator was not possible in the present study. However, the statistician who analyzed the data was blinded to the group allocation of participants.

Statistical analysis

Data were analyzed by SPSS 26 (SPSS Inc., IL, USA). Paired t-test was used to compare the two methods, maxilla and mandible, and GT at 2- and 4-mm distances. Independent t-test was used to compare the values between the two jaws, anterior and premolar areas, and males and females. The intraclass correlation coefficient (ICC) was applied to assess the agreement of data for the mandibular right central incisor and second premolar, and right maxillary canine teeth between the two methods in each patient. Values < 0.4 indicated poor agreement, values between 0.4 and 0.59 indicated fair agreement, values between 0.6 and 0.75 indicated good agreement, and values > 0.75 indicated excellent agreement [8]. In addition to paired t-test, generalized estimating equation was also used for comparison of different areas in the same patient, which yielded results similar to t-test. To assess the clinical significance of the obtained difference between the two measurement methods, the formula of Power and Effect Size for independent means suggested by Kellar [16] was applied. Wherever significant differences were found between the measurements made by the two methods, the mean difference was divided by the standard deviation using the following formula and the obtained value was compared with the ranges reported by Kellar [16] (values up to 0.2 indicated very small difference, 0.2 to 0.5 indicated small difference, 0.5 to 0.8 indicated moderate difference, and 0.8 and higher indicated high difference) to find out the clinical significance of the difference between the ultrasonography and the transgingival probing measurements.