This study aims to explore the dynamic changes in different cell types and their molecular regulatory mechanisms during the healing process of peri-implant soft tissues, using single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics (ST). The research will focus specifically on the healing of soft tissues around dental implants in human subjects with the use of a healing abutment. Participants in the healing abutment group will have a micro healing abutment placed immediately after implant placement. Tissue samples will be collected at various time points (Day 4, Day 14, and Day 70) to construct a detailed single-cell map of the healing process. Spatial transcriptomics will be integrated to preserve tissue architecture, enabling the identification of the spatial distribution of different cell types and their interactions within the peri-implant microenvironment. This combined approach will allow for a comprehensive characterization of cellular interactions, spatial gene expression patterns, and regulatory networks specific to healing abutments. The goal is to identify key regulatory factors that could improve peri-implant soft tissue healing and help prevent peri-implantitis.
Study Type
INTERVENTIONAL
Allocation
NA
Purpose
BASIC_SCIENCE
Masking
NONE
Enrollment
5
The healing process of peri-implant soft tissues will be studied using a micro healing abutment (2 mm), which will be placed at least three months after implant Stage I surgery. Gingival tissue samples will then be collected at Day 4, Week 2, Week 4, and Week 8-10. At each time point, a circular incision around the micro abutment will be made using a circular punch, and the gingival tissue together with the abutment will be retrieved under local anesthesia. After sampling, the micro abutment will be replaced with a formal healing abutment to maintain healing. Samples will undergo single-cell RNA sequencing (scRNA-seq) to characterize cell types, gene expression changes, and regulatory pathways involved in soft tissue repair. Additional tissues will be used for spatial transcriptomics (ST) and histological staining.
School and Hospital of Stomatology, Fujian Medical University
Fuzhou, Fujian, China
Primary Outcome 1: Differentially Expressed Genes (DEGs) in Peri-implant Soft Tissues
Number of genes significantly differentially expressed between experimental groups, determined by single-cell RNA sequencing (scRNA-seq) analysis (FDR \< 0.05). Unit of Measure: Number of genes.
Time frame: Day 4, Week 2, Week 4 and Week 8-10 post-implantation.
Cell Type Composition in Peri-implant Soft Tissues
Proportion (%) of each annotated cell type identified by scRNA-seq based on canonical marker expression. Unit of Measure: Percentage of total cells (%)
Time frame: Day 4, Week 2, Week 4, and Week 8-10 post-implantation.
Enriched Biological Pathways
Number of significantly enriched Gene Ontology (GO) terms and KEGG pathways identified from DEGs (FDR \< 0.05). Unit of Measure: Number of enriched terms/pathways
Time frame: Day 4, Week 2, Week 4, and Week 8-10 post-implantation
Ligand-Receptor Interaction Strength
Mean interaction score of ligand-receptor pairs identified among annotated cell types using CellPhoneDB (or equivalent computational tool). Unit of Measure: Interaction score (arbitrary units)
Time frame: Day 4, Week 2, Week 4, and Week 8-10 post-implantation
Spatial Gene Expression Patterns
Localization and expression intensity of selected DEGs across tissue regions determined by the 10x Genomics Visium platform.
Time frame: Day 4, Week 2, Week 4 and Week 8-10 post-implantation.
Histological Tissue Morphology
Assessment of epithelial continuity, connective tissue organization, and inflammatory cell infiltration in peri-implant soft tissues by hematoxylin and eosin (H\&E) staining. Each parameter will be semi-quantitatively scored on a standardized histological scale (0-3), where 0 = none, 1 = mild, 2 = moderate, and 3 = severe.
Time frame: Day 4, Week 2, Week 4 and Week 8-10 post-implantation.
Immune and Stromal Cell Quantification
Quantification of immune cells (e.g., CD68⁺ macrophages, CD3⁺ T lymphocytes) and stromal cells (e.g., fibroblast-related markers) by immunohistochemistry (IHC). Mean positive cell count per high-power field (HPF) will be calculated from representative sections.
Time frame: Day 4, Week 2, Week 4 and Week 8-10 post-implantation.
Junctional Epithelium Length
Measurement of junctional epithelium length along the implant surface using histological sections and digital image analysis software. Unit of Measure: Length (µm)
Time frame: Day 4, Week 2, Week 4, and Week 8-10 post-implantation.
Basement Membrane Protein Expression
Immunohistochemical detection of Laminin-332 and related basement membrane proteins indicating epithelial attachment integrity. The percentage of positive staining area will be quantified using image analysis software. Unit of Measure: Percentage of positive area (%)
Time frame: Day 4, Week 2, Week 4, and Week 8-10 post-implantation.
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