The protocol was reviewed and accepted by the Research Ethics Committee, Faculty of Dentistry, Cairo University.
A) Samples, Intervention and Outcomes
1. Cell Isolation and Culture:
Create the gingival fibroblast-derived cell sheets in the stem cell center, Faculty of Dentistry, Cairo University, as described by Diar-Bakirly and El-Bialy (2021). Briefly:
I) Obtain gingival tissue from patients undergoing gingivectomy or gingivoplasty, after assigning the informed consent.
II) Cut the gingival tissues into small pieces, plate over tissue culture dishes, and incubate at 37°C in a humidified incubator with 5% CO2.
III) Expand the cells to passages until the number of cells is sufficient for experimentation.
2. Characterization of Gingival Fibroblasts:
Examine the morphological characterization of gingival fibroblast using an inverted microscope. (Gamal et al., 2017).
3. Intervention for Each Group, Cell Sheet Fabrication:
After passages of expansion, seed cells on culture dishes and culture in a complete medium containing ascorbic acid (Wei et al., 2012) with and without rapidly resorbable scaffold. Harvest and evaluate autologous gingival fibroblasts-derived cell sheets.
4. Examine physicochemical properties of the produced cell sheets:
Cell Sheet Thickness (Histomorphometric Evaluation).
1- Number of Cells and Cell Survival Rate using Trypan Blue Staining.
2- Laminin and Fibronectin Production using Enzyme-Linked Immunosorbent Assay (ELISA).
3-Resistance to Mechanical Forces using Universal Testing Machine.
4- Cell Morphology (Scanning Electron Microscope (SEM) Evaluation).
5. Test data statistically after checking its normality by a blinded statistician.
B) Assignment to Intervention
1- Sequence Generation
Cell culture dishes will be randomly assigned to intervention and control groups.
Random allocation sequence will be generated using computer-assisted randomization.
2- Allocation Concealment
The generated random allocation sequence will be sealed using numbered opaque sealed envelopes. Opaque envelopes will be numbered and each envelope will receive the group number according to the generated random allocation sequence, then will be adequately sealed to the time of applying the procedures of cell sheet formation.
Sequence generation and allocation concealment will be performed by a person not involved in the study.
Outcome assessors and statistician will be blinded and will not be informed about the samples assignment to which study group.