Hydroxy decenoic acid down regulates gtfB and gtfC expression and prevents Streptococcus mutans adherence to the cell surfaces

Abstract

Background: 10-Hydroxy-2-decenoic acid, an unsaturated fatty acid is the most active and unique component to the royal jelly that has antimicrobial properties. Streptococcus mutans is associated with pathogenesis of oral cavity, gingivoperiodontal diseases and bacteremia following dental manipulations. In the oral cavity, S. mutans colonize the soft tissues including tongue, palate, and buccal mucosa. When considering the role of supragingival dental plaque in caries, the proportion of acid producing bacteria (particularly S. mutans), has direct relevance to the pathogenicity of the plaque. The genes that encode glucosyltransferases (gtfs) especially gtfB and gtfC are important in S. mutans colonization and pathogenesis. This study investigated the hydroxy-decenoic acid (HDA) effects on gtfB and gtfC expression and S. mutans adherence to cells surfaces.

Methods: Streptococcus mutans was treated by different concentrations of HPLC purified HDA supplied by Iran Beekeeping and Veterinary Association. Real time RT-PCR and western blot assays were conducted to evaluate gtfB and gtfC genes transcription and translation before and after HDA treatment. The bacterial attachment to the cell surfaces was evaluated microscopically.

Results: 500 μg ml-1 of HDA inhibited gtfB and gtfC mRNA transcription and its expression. The same concentration of HDA decreased 60% the adherence of S. mutans to the surface of P19 cells.

Conclusion: Hydroxy-decenoic acid prevents gtfB and gtfC expression efficiently in the bactericide sub-concentrations and it could effectively reduce S. mutans adherence to the cell surfaces. In the future, therapeutic approaches to affecting S. mutans could be selective and it's not necessary to put down the oral flora completely.

Figure 1

The structure of Trans 10-Hydroxy-2-Decenoic acid (HDA) from Royal jelly.

Figure 2

Time-kill curve for S. mutans strain by hydroxy-decenoic acid at 4 times of the MIC (data not shown). Symbols: ▴: Test group and Δ: control group of S. mutans ATCC 25175.

Figure 3

Transcription of gtfB/gtfC from S. mutans following HDA treatment. gtfB/gtfC expression is completely abrogated by exposure to HDA. gtfB/gtfC levels are also reduced with lower concentrations of HDA. The error bars represent mean and standard deviations of experiments performed in triplicate. gtf genes were more abundantly expressed in cultures that treated with 0.0, 100 and 200 μg ml^-1 of HDA but missing in the cells treated with 1000 μg ml^-1 of HDA.

Figure 4

Western blot analysis of GtfB and GtfC enzymes. GtfB and GtfC proteins that are present in S. mutans treated with 0.0, 100, and 200 μg ml^-1 concentrations of HDA but are missing in the cells treated with 500 and 1000 μg ml^-1 or higher concentration of HDA. The 16SrRNA protein was used as control.

Figure 5

Cultures f P19 embryonal carcinoma monolayer cells: after treatment with HDA disrupted the organization of the monolayer of P19 cells (A); and addition of S. mutans to monolayer of P19 cells that disrupted the organization of monolayer to a greater extension (B); S. mutans treated with HDA in P19 cultures (C) and not treated culture (D). Magnification in part A is 600X and in the all other images is X400.