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Updated: Aug 5, 2026
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Isolation and Culture of Primary Human Gingival Epithelial Cells using Y-27632
Published on: November 6, 2021
Enhancing the Outgrowth of Oral Mucosal Cell Sheets for Cultivated Rabbit Oral Mucosal Epithelial Transplantation
Wei-Lun Huang1,2,3, Shang-Chih Yang3, Vladlen Klochkov3
1Department of Ophthalmology, National Taiwan University Hospital Hsinchu Branch, Hsinchu, Taiwan.
Current Eye Research
|July 29, 2026
Summary
Rho-associated protein kinase (ROCK) inhibitor Y-27632 enhances oral mucosal epithelial cell sheet growth for limbal stem cell deficiency (LSCD) treatment. This optimization speeds up cultivated oral mucosal epithelial transplantation (COMET) procedures.
Area of Science:
- Ophthalmology
- Regenerative Medicine
- Tissue Engineering
Background:
- Cultivated oral mucosal epithelial transplantation (COMET) treats limbal stem cell deficiency (LSCD).
- Current COMET methods face limitations due to long cultivation times and complexity.
- Rho-associated protein kinase (ROCK) inhibitors promote cell proliferation, migration, and cytoskeleton stability.
Purpose of the Study:
- To investigate the efficacy of ROCK inhibitor Y-27632 in enhancing oral mucosal epithelial cell sheet growth.
- To determine if Y-27632 maintains the characteristics of oral mucosal epithelial cells for transplantation.
- To optimize COMET for LSCD treatment by accelerating cell sheet production.
Main Methods:
- Oral mucosal epithelial cells were cultured from rabbits using a collagenase-based technique.
- Cells were cultured with or without amniotic membrane (AM) and Y-27632 in transwell plates.
- Cell sheet outgrowth, density, and expression of stem cell (p63), differentiation (K3, K4, K13), and junctional markers (ZO-1, occludin, connexin 43) were analyzed.
Main Results:
- Y-27632 and AM significantly accelerated cell sheet outgrowth and expansion from day 7 to 15 (p < 0.05).
- Y-27632 treatment dose-dependently increased cell density and inhibited squamous differentiation.
- Y-27632 increased p63-positive cells, potentially preserving stem cell properties and inhibiting differentiation.
Conclusions:
- Y-27632 combined with AM creates an efficient environment for producing oral mucosal epithelial cell sheets.
- These agents may accelerate oral epithelial cell-based therapy for LSCD.
- Y-27632 offers a promising adjunctive strategy to optimize COMET for LSCD treatment.