IADR Abstract Archives
Establishing a Clinically Relevant in-Vitro Cytotoxicity Model for Denture Adhesives
Objectives: Methods to assess the in-vitro cytotoxicity of medical devices, including ISO10993-5, utilizes murine fibroblasts for evaluation of cytotoxicity instead of human fibroblasts. Previous studies have reported mice to have several cell death domains that are not present in human orthologous. Hence, in order to establish a clinically relevant cytotoxicity model for denture adhesives, our objective was to investigate the differences in responses to denture adhesives between the two cell lines.
Methods: Twelve commercially available denture adhesives were diluted in a cell culture medium (Dulbecco's Modified Eagle Medium-DMEM with 10% Fetal Bovine Serum-FBS with antibiotics) with concentrations ranging from 0.015%- 2.5%. Human gingival fibroblasts (ATCC HGF-1) and Murine fibroblasts (ATCC, L929) were co-cultured with denture adhesives for 24 hours in 96 well plates under 5% CO2 condition. After 24 hours of culture, the supernatant was collected for LDH assay to measure cytotoxicity. MTT solution was added to the cell culture to detect cell viability. Mean cytotoxicity was compared using 2-tailed t-tests, with alpha=0.05 and Bonferroni correction.
Results: For both cell types, 41.7% of denture adhesives exhibited cytotoxicity and suppressed cell viability. However, L929 showed 15 times higher IC50 in LDH cytotoxicity when compared to HGF-1 (p-values ≤0.033 in each product %). Furthermore, L929 cells were more sensitive to MTT assay when compared to HGF-1 Microscopic analysis showed cell membrane rupture as the main cause of cell death.
Conclusions: HGF-1 and L929 respond differently when tested for cytotoxicity against denture adhesives. L929, the common choice of cell line for those tests, presented 15 times higher sensitivity in cytotoxicity compared to HGF-1 in LDH assay. Also, L929 cells indicate high sensitivity to MTT cell viability assay compared to HGF-1.To test the oral cytotoxicity of denture adhesives, human gingival fibroblasts may be the best choice for a cell line.
Methods: Twelve commercially available denture adhesives were diluted in a cell culture medium (Dulbecco's Modified Eagle Medium-DMEM with 10% Fetal Bovine Serum-FBS with antibiotics) with concentrations ranging from 0.015%- 2.5%. Human gingival fibroblasts (ATCC HGF-1) and Murine fibroblasts (ATCC, L929) were co-cultured with denture adhesives for 24 hours in 96 well plates under 5% CO2 condition. After 24 hours of culture, the supernatant was collected for LDH assay to measure cytotoxicity. MTT solution was added to the cell culture to detect cell viability. Mean cytotoxicity was compared using 2-tailed t-tests, with alpha=0.05 and Bonferroni correction.
Results: For both cell types, 41.7% of denture adhesives exhibited cytotoxicity and suppressed cell viability. However, L929 showed 15 times higher IC50 in LDH cytotoxicity when compared to HGF-1 (p-values ≤0.033 in each product %). Furthermore, L929 cells were more sensitive to MTT assay when compared to HGF-1 Microscopic analysis showed cell membrane rupture as the main cause of cell death.
Conclusions: HGF-1 and L929 respond differently when tested for cytotoxicity against denture adhesives. L929, the common choice of cell line for those tests, presented 15 times higher sensitivity in cytotoxicity compared to HGF-1 in LDH assay. Also, L929 cells indicate high sensitivity to MTT cell viability assay compared to HGF-1.To test the oral cytotoxicity of denture adhesives, human gingival fibroblasts may be the best choice for a cell line.