Objective: To develop an in vitro three-dimensional model of human sulcular epithelium.
Materials and Methods: Primary normal human gingival keratinocytes were grown three-dimensionally on top of collagen type I gels populated with primary normal human gingival fibroblasts (GF) or periodontal ligament fibroblasts (PLF). The cultures were grown in serum-free medium for 7 and 9 days at air-liquid interface. Experiments were repeated 3 times in duplicate. Tissue architecture (histomorphometry), cell proliferation (Ki67) and cell differentiation (cytokeratins CK19, CK13, CK10) were assessed in cultured tissues and 5 biopsies of human sulcular epithelium and gingiva.
Results: PLF models had thinner epithelium and less epithelial cell layers than GF models at both 7 and 9 days. Basal cell proliferation was significantly higher in PLF models than in GF models (p=0,025) at day 7, but comparable at day 9. A superficial cell layer with flattened, terminally differentiated keratinocytes was observed in GF models only at both time points. Strong expression of CK19, with only few scattered cells expressing CK10, were observed in PLF models in contrast to the GF models showing weaker CK19 expression and stronger CK10 expression. CK13 was strongly expressed suprabasally in both models. Changes in the cytokeratin expression were more proeminent at day 7.
Conclusion: Tissue morphology, cell proliferation, cytokeratin expression and their distribution pattern in the models showed major differences dependent on the type of fibroblasts used and the harvesting time. The models constructed with PLF harvested at 7 days showed similar tissue morphology and cytokeratin expression to the in vivo sulcular epithelium.
Funding: This study was partially supported by Colgate-Palmolive scholarship.