Cytotoxic effects of hydrogen peroxide on periodontal cells

Abstract

To evaluate the in vitro cytotoxic effects of hydrogen peroxide (H 2 O 2) on periodontal cells, based on the cellular viability and morphology of immortalized osteoblast and gingival fibroblast cultures, with different exposure times and concentrations. Methods: Immortalized human gingival fibroblast and human fetal osteoblast cell lines were cultured, separately, in 96-well plates. After reaching confluency, they were exposed to H 2 O 2 solutions at 16 different concentrations ranging between 0.05 µg/ml and 10 µg/ml for 1 h, 24 h or 72 h in triplicate assays (n=24), using culture media alone as the control. Cell viability was measured by previously established fluorometric methods, using a resazurin-based assay, and cell morphology by using an inverted microscope with integrated phase-contrast optics. Data were statistically analyzed using a one-way ANOVA with Tukey's and Dunnet's post hoc tests and Pearson correlation coefficient (r), as appropriate (α=0.05). Results: H 2 O 2 induced a decrease in cell viability to below 50% in fibroblasts and around 50% in osteoblasts, in all tested concentrations after 1h exposure, and a decrease in cell viability to above 70% after 24 h and 72 h (P<0.05). A significant negative correlation was detected between H 2 O 2 and cell viability at 1 h and 72 h for osteoblast (r=-0.471) and HGF (r=-0.12) cells, respectively. The cell morphology analysis showed cell detachment and lower cell density, in agreement with these findings. Conclusions: H 2 O 2 induced cell alterations with moderate to severe cytotoxic effects in osteoblast and gingival fibroblasts.