Effect of S-PRG Eluate on Biofilm Formation and Enzyme Activity of Oral Bacteria

Abstract

Recently, the antibacterial activity of a composite resin containing prereacted glass ionomer (S-PRG) filler was revealed. We examined the effect of an S-PRG eluate on various biologic activities of Streptococcus mutans and Porphyromonas gingivalis. Adherence ability of S. mutans was evaluated by microtiter plate assay; protease and gelatinase activities of P. gingivalis were examined by synthetic substrate hydrolysis and gelatin film spot assay, respectively. Coaggregation of P. gingivalis with Fusobacterium nucleatum was also examined. S-PRG eluate was found to suppress streptococcal adherence. S-PRG eluate inhibited the protease and gelatinase activities of P. gingivalis and the coaggregation between P. gingivalis and F. nucleatum. These results indicate that S-PRG eluate suppresses streptococcal adherence and inhibits the protease and coaggregation activities of P. gingivalis. These findings may prompt research into novel strategies for preventing caries and periodontitis.

Figure 1

Adherence of Streptococcus mutans JCM 5705 in the presence and absence of PBS or S-PRG eluate. To evaluate the effect of S-PRG eluate on adherence, different percentages of S-PRG were substituted for culture medium (circle). For the control, phosphate-buffered saline was used instead of S-PRG (triangle). The plates were incubated at 37°C for 16 h under anaerobic conditions, stained with 0.25% safranin, and the amount of stain in an ethanol-solubilized sample quantified by measuring absorbance at 492 nm. Data are representative of three independent experiments and the bars indicate the standard deviation.

Figure 2

BAPNA hydrolysis activity (as an indicator of protease activity) in the presence and absence of S-PRG eluate. P.-gingivalis-sonicated extract was added to the reaction mixture. To analyze the effect of S-PRG, distilled water in control mixture (triangle) was replaced with S-PRG solution (circle). Colored metabolites were quantified using a colorimetric absorbance assay at 405 nm. Data are representative of three independent experiments and the bars indicate the standard deviation.

Figure 3

Gelatinase spot assay. Different amounts of S-PRG eluate were added to the reaction mixture. P.-gingivalis-sonicated extract was then added and the mixture was serially diluted with distilled water. A 15 μL spot of each mixture was placed onto gelatin-coated X-ray film and incubated at 37°C for 2 h in a humidified atmosphere before being washed with tap water. Removal of the gelatin surface beneath the spotted area revealed the base color of the film and indicated the presence of gelatinase activity. (a) Photograph of X-ray film after assay. (b) Results of gelatinase spot assay.

Figure 4

Coaggregation between P. gingivalis and F. nucleatum in the presence and absence of S-PRG eluate. (a): photographs of coaggregation. (b): results of coaggregation assay.