A Membrane-Targeted Peptide Inhibiting PtxA of Phosphotransferase System Blocks Streptococcus mutans.

Streptococcus mutans, the primary cause of dental caries, takes up carbohydrates through the phosphoenolpyruvate sugar phosphotransferase system (PTS). This study aimed to identify a novel membrane-targeted antimicrobial peptide (AMP) that could also target the L-ascorbate-specific PtxA component of the S. mutans PTS system. C10-KKWW was identified and selected using virtual screening of a lipopeptide library, a minimum inhibiting concentration (MIC) assay, cytotoxicity assays and a hemolysis assay. Surface plasmon resonance confirmed that C10-KKWW had a high binding affinity for PtxA. Combining with scanning electron microscopy and cell permeability assay, it was shown that the effects of C10-KKWW could be attributed to both membrane and PtxA. Wild type (WT) S. mutans, a ptxA deletion mutant (ΔptxA), and a mutant-complemented strain (CptxA), were cultured consistently in brain heart infusion (BHI) medium, tryptone-vitamin medium supplemented with 15 mM L-ascorbate (TVL), or for 5 h in BHI supplemented with 7.4 mM sodium L-ascorbate. Compared to ∆ptxA, in WT S. mutans and CptxA, C10-KKWW had a stronger MIC (3.9 µg/mL), and distinctively decreased biofilm viability. The extracellular concentrations of L-ascorbate/sodium L-ascorbate were not changed before and after WT treated with C10-KKWW. L-ascorbate-induced operon genes, or other PTS genes, were significantly suppressed by C10-KKWW. In conclusion, C10-KKWW has been developed; it acts through interaction with the bacterial membrane and interferes with L-ascorbate translocation to inhibit S. mutans growth and eradicate its biofilm. C10-KKWW may be especially effective at optimal oral ascorbate levels. A combination of C10-KKWW with sodium L-ascorbate might also be a novel strategy for dental caries treatment.

[Role of SMU.2055 gene in cariogenic capacity of Streptococcus mutans].

OBJECTIVE: To construct a SMU.2055-dificient mutant strain of Streptococcus mutans (S. mutans) and evaluate its cariogenic capacity in comparison with wild-type S. mutans. METHODS: The SMU.2055-dificient mutant strain of S. mutans was constructed using homologous recombination technique and observed with scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The absorbance at 600 nm and pH values of the wild-type and mutant strains were monitored to evaluate their growth and acid production. After acid adaption, the two strains were challenged with acid shock and their survival rates were determined. RESULTS: PCR and sequence analyses verified the successful construction of the SMU.2055-dificient mutant strain. Observation with SEM revealed obvious changes in the morphology of the mutant strain, which showed reduced irregular substances between the individual bacteria as compared with the wild-type strain. TEM revealed major alterations in the cellular architecture of the mutant strain with blurry cell membrane and disruption of the membrane integrity. The growth capacity of the mutant strain decreased in both normal and acidic conditions but its acid production capacity remained unaffected. CONCLUSION: SMU.2055 gene is associated with morphology maintenance, growth capacity and acid resistance of S. mutans but is not related to the acid production capacity of the bacterium.

[Effects of intracellular Porphyromonas gingivalis on proliferation and osteogenic differentiation of human periodontal ligament cells in vitro].

OBJECTIVE: To investigate the ability of Porphyromonas gingivalis to invade human periodontal ligament cells (hPDLCs) and the effect of intracellular P. gingivalis on cell proliferation and osteogenic differentiation in vitro. METHODS: The invasion ability of P. gingivalis in hPDLCs was tested using an antibiotic protection assay at the multiplicity of infection (MOI) of 10 and 100. The proliferation of the infected cells was detected using a CFDA-SE kit, and the cells were sorted by fluorescence-activated cell sorting (FACS) followed by alizarin red staining for detecting mineralization nodules deposition; real-time PCR was used to examine the expression of Runx2 mRNA in the cells. RESULTS: P. gingivalis actively invaded hPDLCs, and the internalized P. gingivalis was able to resist antibiotic treatment. The cells infected by P. gingivalis exhibited no significant suppression of cell proliferation, but showed significantly lowered capacity for osteogenic differentiation, down-regulated RUNX2 mRNA expression, and reduced mineral deposition. CONCLUSION: Intracellular P. gingivalis does not significantly affect the proliferation of hPDLCs but inhibits osteogenic differentiation of the cells.

[Six degree-of-freedom acquisition and analysis of jaw opening and closing with motion capture system].

OBJECTIVE: To explore the six degrees of freedom of jaw opening and closing movement with motion capture and analysis system to establish a quantitative method for studying mandibular movement and a digital basis for virtual reality study of mandibular movement. METHODS: In a male adult with normal dentition without temporomandibular joint disorders, 3 fluorescent markers were pasted in the upper dentition and 4 in the lower dentition. Six cameras of the motion capture system were arranged in a semi-circular fashion. The subject sat in front of the camera at an 80-cm distance with the Frankfort plane kept parallel to the horizontal plane. The degree-of-freedom (3 linear displacement and 3 angular displacement) of jaw opening and closing movement was obtained by collecting the marker motion. RESULTS: Six degrees of freedom of jaw opening and closing were obtained using the motion capture system. The maximum linear displacements of X, Y and Z axes were 5.888 089 cm, 0.782 269 cm, and 0.138 931 cm, and the minimum linear displacements were -3.649 83 cm, -35.961 2 cm, -5.818 63 cm, respectively. The maximum angular displacements of X, Y and Z axes were 0.760 088°, 2.803 753°, and 0.786 493°, with the minimum angular displacements of -2.526 18°, -0.625 94°, and -25.429 8°, respectively. Variations of linear displacements during jaw opening and closing occurred mainly in the Y axis, and those of angular displacement occurred mainly in the Z axis. CONCLUSION: The six degree-of-freedom of mandibular movement can be accurately obtained with the motion capture system to allow quantitative examination of the mandibular movement.