2020 Volume 63 Issue 1 Pages 61-72
Purpose: Sulfated vizantin (Viz-S) possesses anti-biofilm activity against Streptococcus mutans as well as immune-stimulating properties. However, the mechanism associated with this activity and cellular toxicity remained unclear. The purpose of this study was to answer these questions utilizing an in vitro biofilm model. In addition, we also analyzed whether a high concentration of sucrose in the growth media would degrade anti-biofilm activity.
Methods: Human gingival epithelial cells were incubated in the presence of Viz-S at the concentrations of 0, 5, 10, 50, 75, 100 and 500 μmol/l for 3 h at 37℃. The cellular viability was assessed using an MTT assay. S. mutans biofilm was developed in a brain-heart infusion broth (BHI) supplemented with 0.2, 0.4, 0.8 and 1.6% sucrose using a chambered coverglass containing Viz-S at the concentrations of 0, 10 and 50 μmol/l for 24 h at 37℃ under anaerobic conditions. The wells were washed two times with phosphate-buffered saline and the amount of residual biofilms was measured by the method of crystal violet staining. The expression of genes related to biofilm formation in the biofilm cells in the absence and presence of 50 μmol/l Viz-S was investigated using a real-time polymerase chain reaction. The expression of proteins of cell-associated glucosyltransferase (CA-GTF) and cell-free glucosyltransferase (CF-GTF) was investigated by western blot analysis.
Results: Supplementation with Viz-S at a concentration of less than 75 μmol/l did not affect the gingival cells. Viz-S at the concentration of 50 μmol/l significantly upregulated the expression of gtfB and gtfC in the biofilm cells developed in the BHI supplemented with more than 0.4% sucrose, whereas the expression of gtfD was significantly downregulated by 0.44- to 0.66-fold in all culture conditions. CA-GTF, corresponding to GTFB and GTFC, showed reduced expression in the presence of a low concentration of sucrose, however, expression in the presence of 1.6% sucrose was elevated. Surprisingly, CF-GTF, corresponding to GTFD, mostly was not detected.
Conclusion: Viz-S disrupted the localization balance between CA-GTF and CF-GTF, resulting in detachment of S. mutans biofilm.
