Three-Dimensional Human Cell Cultures for Cytotoxicity Testing of Dental Filling Materials.

OBJECTIVES So far, bovine immortalized pulp cells have been used as three dimensional cultures for cytotoxicity testing of filling materials in the dentin barrier test (DBT). In this study, the use of human pulp-derived cells was evaluated, which would better simulate the clinical situation, and a composite material with a new resin base was tested. MATERIALS AND METHODS SV40-transfected human pulp cells (tHPC) were cultured in hydrogels (fibrin, peptide, collagen) and mechanical properties and cell viability (MTT or WST-1) were determined. For cell cultures in collagen, a four week - proliferation assay was performed (WST-1). After 14 days of three-dimensional culture in collagen, tHPC were introduced into the DBT with 200 µm dentin disks. After a 24-hour incubation under perfusion (0.3 ml/h), the following materials were applied according to the manufacturers' instructions (1) President (Coltene): negative (non-toxic) control, (2) CaGPG14 (ISO 7405): positive (toxic) control, (3) Tetric EvoCeram (Ivoclar Vivadent) with Clearfil SEBond (Kuraray, reference material), (4) N´Durance (Sepodont, test material), (5) N´Durance with Clearfil SEBond. Cell viability was determined after 24-hour incubation (WST-1). The percentage of relative viability was calculated (negative control=100%) and statistically analyzed (Kruskal-Wallis-test, p<0.05). RESULTS Fibrin and peptide gels had insufficient mechanical properties for the DBT Collagen and appeared suitable for three-dimensional cell culture of tHPC for up to 21 days. The cultures could be transferred to the DBT device and results for controls were similar to previous tests with bovine cells. The DBT using tHPC in the collagen showed no statistically significant difference between the test material with and without the adhesive and the reference resin composite. CONCLUSIONS tHPC in collagen can replace bovine cells in the DBT. The tested filling material is not likely to cause pulp damage, if the pulp is covered by an intact dentin layer.