Clinical and Histological Analysis of Human Gingival Phenotypes

Clinical, Histological and Immunohistochemical Characterization of Human Gingival Phenotypes

The goal of this observational study is to compare the composition of the human gingiva in different gingival phenotypes. The main questions to answer are:

• Is there any difference in the cellular composition of the gingiva between thin and thick gingival phenotype?
• Is there any difference in the molecular composition of the gingiva between thin and thick gingival phenotype?

The participants were divided in two groups (thin and thick phenotype) and a biopsy of healthy gingiva was obtained from each one them. The biopsies were analyzed histologically and the collected data will be analyzed statistically in order to identify possible differences between the gingival phenotypes.

Study Overview

• Status: Completed
• Conditions: Healthy
• Intervention / Treatment: Diagnostic test: Histological analysis; Diagnostic test: Immunohistochemical analysis

Detailed Description

Healthy volunteers were assigned in one of two groups:

• Group 1: Thin gingiva, when the free gingiva was evaluated as transparent, after the insertion of a periodontal probe (Hu-Friedy XP-23/QW, Hu-Friedy,Chicago,IL,USA) in the middle of the facial dentogingival sulcus of a maxillary central incisor
• Group 2: Thick gingiva: when the free gingiva was evaluated as non-transparent, using the same methodology.

A full thickness sample of the oral mucosa of each one of the participants was collected under local anaesthesia. The sample had a rectangular shape, with a length of 4 millimeters and a width of 1 millimeter. It had a vertical orientation and it was expanding at the both sides of the mucogingival junction.

The oral mucosa samples were processed for histological and immunohistochemical analysis. Staining with haematoxylin-eosin was performed in order to describe the tissue histologically and also calculate the total number of the cells it contained. Immunohistochemical staining with anti-Vimentin,anti-Cluster of Differentiation 68, anti-Ki-67 and anti-Smooth Muscle Actin antibodies was applied for the calculation of the numbers of fibroblasts, macrophages, Ki-67-positive cells and Smooth muscle actin positive cells respectively. Immunohistochemical staining was also performed in order to determine the levels of expression of Collagen I, Collagen V, Elastin and Hyaluronic acid.

Whole slide images of the specimens were acquired with the use of the NanoZoomer 2.0HT (Hamamatsu Pho-tonics K.K., Hamamatsu, Japan). Cell counting will be performed automatically using an image analysis software (QuPath 3.0). The levels of molecular expression will be assessed with the use of the same software and will be expressed as a percentage of the area of the connective tissue that is occupied by the investigated molecules.

• Study Type: Observational
• Enrollment (Actual): 45

Contacts and Locations

Study Locations

• Greece
  • Macedonia
    • Thessaloniki, Macedonia, Greece, 54124
      • Aristotle University of Thessaloniki

Participation Criteria

Eligibility Criteria

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

• Sampling Method: Probability Sample

Study Population

Healthy volunteers referred to the Department of Preventive Dentistry, Periodontology and Implant Biology of the Dental School of the Aristotle University of Thessaloniki.

Description

Inclusion Criteria:

• Healthy adults

Exclusion Criteria:

• Attachment loss or Probing depths greater than 2 millimeters at the maxillary central incisors
• Altered passive eruption of the maxillary incisors
• Previous surgery or orthodontic treatment in the anterior maxilla
• Medical conditions or medication affecting soft tissue metabolism
• Tooth crowding or abnormal angulation or abrasion of the maxillary incisors
• Restorations at the buccal surface of the maxillary central incisors
• Smoking
• Pregnancy or lactation

Study Plan

How is the study designed?

Number of groups / cohorts

2

Cohorts and Interventions

Group / Cohort	Intervention / Treatment
Thin Phenotype; Participants with transparent free gingiva of the maxillary central incisor	Diagnostic test: Histological analysis; Histological staining of gingival biopsies with haematoxylin-eosin.; Diagnostic test: Immunohistochemical analysis; Immunohistochemical staining of gingival biopsies using anti-Vimentin, anti-Collagen I, anti-Collagen V, anti-Elastin, anti-Hyaluronic acid, anti-Smooth muscle actin, anti-Cluster of Differentiation 68 and anti-ki67 antibodies.
Thick Phenotype; Participants with non-transparent free gingiva of the maxillary central incisor	Diagnostic test: Histological analysis; Histological staining of gingival biopsies with haematoxylin-eosin.; Diagnostic test: Immunohistochemical analysis; Immunohistochemical staining of gingival biopsies using anti-Vimentin, anti-Collagen I, anti-Collagen V, anti-Elastin, anti-Hyaluronic acid, anti-Smooth muscle actin, anti-Cluster of Differentiation 68 and anti-ki67 antibodies.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Cell density	The number of all types of cells per square millimeter of the connective tissue	Baseline

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Fibroblast Density	The number of fibroblasts per square millimeter of the connective tissue	Baseline
Macrophage Density	The number of macrophages per square millimeter of the connective tissue	Baseline
Density of Ki-67-positive cells	The number of Ki-67-positive cells per square millimeter of the connective tissue	Baseline
Density of Smooth Muscle Actin positive cells	The number of Smooth Muscle Actin positive cells per square millimeter of the connective tissue	Baseline
Collagen I area ratio	The percentage of the area of the connective tissue which is occupied by Collagen I	Baseline
Collagen V area ratio	The percentage of the area of the connective tissue which is occupied by Collagen V	Baseline
Elastin area ratio	The percentage of the area of the connective tissue which is occupied by Elastin	Baseline
Hyaluronic acid area ratio	The percentage of the area of the connective tissue which is occupied by Hyaluronic acid	Baseline

Other Outcome Measures

Outcome Measure	Measure Description	Time Frame
Keratinized tissue width	The length (millimeters) of the line that connects the mucogingival junction and the gingival margin, at the middle of the facial surface of the maxillary central incisor	Baseline
Gingival thickness	The thickness (millimeters) of the gingiva, measured with an ultrasonic biometer (PIROP Biometric scanner, ECHO-SON S.A., Poland) at the middle of the line that connects the mucogingival junction and the gingival margin, at the middle of the facial surface of the maxillary central incisor	Baseline

Collaborators and Investigators

• Sponsor: Aristotle University Of Thessaloniki
• Collaborators: Umeå University
• Investigators: Study Director: Sotiris Kalfas, Professor, Aristotle University of Thessaloniki

Study record dates

Study Major Dates

• Study Start (Actual): March 5, 2020
• Primary Completion (Actual): December 1, 2022
• Study Completion (Actual): December 1, 2022

Study Registration Dates

• First Submitted: March 22, 2023
• First Submitted That Met QC Criteria: June 27, 2023
• First Posted (Actual): June 28, 2023

Study Record Updates

• Last Update Posted (Actual): June 28, 2023
• Last Update Submitted That Met QC Criteria: June 27, 2023
• Last Verified: June 1, 2023

More Information

Terms related to this study

• Keywords: phenotype; gingiva; periodontal; immunohistochemical; histological; biotype
• Other Study ID Numbers: 1399

Drug and device information, study documents

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