Identification of a glucan-binding protein C gene homologue in Streptococcus macacae

BACKGROUND/AIMS: The past few decades have seen the isolation of certain glucosyltransferases and a number of proteins from mutans streptococci. Some of these proteins have been shown to possess glucan-binding capabilities which confer an important virulence property on mutans streptococci for the role of these bacteria play in dental caries. Among these proteins is glucan-binding protein C, which is encoded by the gbpC gene, and which we have identified as being involved in the dextran-dependent aggregation of Streptococcus mutans. However, gbpC homologues have yet to be identified in other mutans streptococci. METHODS: We carried out polymerase chain reaction amplification of Streptococcus macacae using primers that were designed based on conserved sequences of S. mutans gbpC and identified a gbpC gene homologue. The gene of that homologue was then characterized. RESULTS: Nucleotide sequencing of the S. macacae gbpC homologue revealed a 1854 bp open reading frame encoding a protein with an N-terminal signal peptide. The molecular mass of the processed protein was calculated to be 67 kDa. We also found an LPxTG motif, the consensus sequence for gram-positive cocci cell wall-anchored surface proteins, which was followed by a characteristic sequence at the carboxal terminal region of the putative protein. This suggests that the S. macacae GbpC homologue protein was tethered to the cell wall. CONCLUSION: Based on these results, together with the demonstrated glucan-binding ability of the S. macacae GbpC homologue protein, we suggest that S. macacae cells are capable of binding dextran via the GbpC homologue protein, which is similar to the S. mutans GbpC protein. In addition, Southern hybridization analysis using the S. macacae gbpC homologue as a probe showed a distribution of gbpC homologues throughout the mutans streptococci.     

The evaluation of microleakage and bond strength of a silicone-based resilient liner following denture base surface pretreatment

STATEMENT OF PROBLEM: The failure of adhesion between a silicone-based resilient liner and a denture base is a significant clinical problem. PURPOSE: The purpose of this study was to examine the effects of denture base resin surface pretreatments with different chemical etchants preceding the silicone-based resilient liner application on microleakage and bond strength. The initial effects of chemical etchants on the denture base resin in terms of microstructural changes and flexural strength were also examined. MATERIAL AND METHODS: Forty-two polymethyl methacrylate (PMMA) denture base resin (Meliodent) specimens consisting of 2 plates measuring 30 x 30 x 2 mm were prepared and divided into 7 groups (n = 6). Specimen groups were treated by immersion in acetone for 30 (A30) or 45 (A45) seconds, methyl methacrylate monomer for 180 (M180) seconds, and methylene chloride for 5 (MC5), 15 (MC15) or 30 (MC30) seconds. Group C had no surface treatment and served as the control. Subsequently, an adhesive (Mollosil) and a silicone-based resilient denture liner (Mollosil) were applied to the treated surfaces, and all specimens were immersed in the radiotracer solution (thalium-201 chloride) for 24 hours. Tracer activity (x-ray counts), as a parameter of microleakage, was measured using a gamma camera. For bond-strength measurement, 84 rectangular PMMA specimens (10 x 10 x 40 mm) were surface-smoothed for bonding and treated with the different chemical etchants using the same previously described group configurations. The adhesive and the silicone-based denture liner were applied to the treated surfaces. Tensile bond-strength (MPa) was measured in a universal testing machine. Flexural strength measurement was performed with 49 PMMA specimens (65 x 10 x 3.3 mm according to ISO standard 1567) in 7 groups (n = 7), with 1 flat surface of each treated with 1 of the chemical etchants preceding adhesive application. The flexural strength (MPa) was measured using a 3-point bending test in a universal testing machine. The data were analyzed by 1-way analysis of variance and the Tukey HSD test (alpha = .05). RESULTS: Significant differences were found between the groups in terms of microleakage (P < .0001). The lowest microleakage was observed in group M180 (30,000 x-ray counts) and the highest in the control group (44,000 x-ray counts). The mean bond strength to PMMA resin ranged from 1.44 to 2.22 MPa. All treated specimens showed significantly higher bond strength than controls (P < .01). The flexural strength values all significantly differed (P < .05). All experimental specimens that had chemical surface treatments showed lower flexural strength than controls (P < .05). CONCLUSION: Treating the denture base resin surface with chemical etchants increased the bond strength of silicone-based resilient denture liner to denture base and decreased the microleakage between the 2 materials. Considering the results of both tests together, the use of methyl methacrylate monomer for 180 seconds was found to be the most effective chemical treatment.     

Molecular analysis of bacteria in asymptomatic and symptomatic endodontic infections

The purpose of the present study was to use terminal restriction fragment length polymorphism analysis and the 16S rRNA gene clone library to investigate the diversity of the microbiota associated with asymptomatic and symptomatic endodontic infections and to compare the bacterial community structure in these two clinical conditions. Samples were taken from asymptomatic endodontic infections associated with chronic periradicular lesions and from symptomatic infections clinically diagnosed as acute abscesses. 16S rRNA genes from DNA isolated from clinical samples were used to construct clone libraries or were subjected to terminal restriction fragment length polymorphism analysis. Sequence analysis of 186 clones revealed 42 taxa; 23 (55%) were uncultivated phylotypes, of which seven were unique to endodontic infections. Clone sequencing and terminal restriction fragment length polymorphism analysis revealed that the most commonly detected taxa were Fusobacterium nucleatum (including terminal restriction fragment types 1 and 2), Peptostreptococcus micros/Peptostreptococcus sp. oral clone AJ062/BS044/FG014, Prevotella species, Dialister species, Mogibacterium species, Lachnospiraceae oral clone 55A-34, Filifactor alocis, Megasphaera sp. oral clone CS025/BS073, and Veillonella sp. oral clone BP1-85/Veillonella dispar/V. parvula. Bacteroides-like sp. oral clone X083/Bacteroidales oral clone MCE7_20 and Dialister sp. oral clone BS016/MCE7_134 were detected only in asymptomatic teeth. On the other hand, F. nucleatum terminal restriction fragment type 2, Prevotella intermedia, Dialister pneumosintes, and some phylotypes were exclusively detected in symptomatic samples. Bacterial profiles of symptomatic endodontic infections generated by terminal restriction fragment length polymorphism analysis were clearly different from those of asymptomatic infections. Overall, the average number of terminal restriction fragments in symptomatic samples was significantly larger than in asymptomatic samples. Molecular analysis of the microbiota associated with symptomatic or asymptomatic endodontic infections indicates that the endodontic bacterial diversity is greater than previously described by culture methods and that the structure of the microbiota differ significantly between asymptomatic and symptomatic infections.     

Immunodominant region of Actinobacillus actinomycetemcomitans 40-kilodalton heat shock protein in patients with rheumatoid arthritis

Bacterial heat shock proteins have been implicated in the pathogenesis of several diseases, and the immunological relationship between rheumatoid arthritis (RA) and Escherichia coli DnaJ has been reported. Since there are similarities in the tissue destruction process of RA and periodontitis, we examined the reactivities of antibodies in sera from RA patients to the DnaJ protein from Actinobacillus actinomycetemcomitans. An enzyme-linked immunosorbent assay showed that IgG titers to the N-terminal conservative region of the DnaJ are significantly higher in RA patients compared with the healthy controls (p < 0.05). Furthermore, we examined IgG titers of disease controls to determine the specificity of the immune responses to this region in RA patients. The difference between RA and infectious disease patients was also significant (p < 0.05). These results suggest that the N-terminal region of DnaJ from A. actinomycetemcomitans may contribute to the etiologic analysis of RA.     

Induction of dental pulp fibroblast matrix metalloproteinase-1 and tissue inhibitor of metalloproteinase-1 gene expression by interleukin-1alpha and tumor necrosis factor-alpha through a prostaglandin-dependent pathway

Matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) are involved in the degradation of extracellular matrix in many inflammatory diseases. Little is known regarding the expression of these mediators in dental pulp fibroblasts. The effects of proinflammatory cytokines (interleukin (IL)-1alpha and tumor necrosis factor-alpha (TNF-alpha)) and prostaglandin E2 (PGE2) on pulp fibroblast MMP-1 and TIMP-1 gene expression were investigated. Northern hybridization showed that IL-1alpha and TNF-alpha induced significant MMP-1 gene expression, with only little effect on TIMP-1 gene. Exogenous PGE2, however, upregulated TIMP-1 mRNA synthesis but not MMP-1. Concomitant addition of IL-1alpha and PGE2 or TNF-alpha and PGE2 suppressed MMP-1 mRNA production, compared with the groups treated with IL-1alpha or TNF-alpha alone. In contrast, PGE2 enhanced the upregulatory effects of TIMP-1 mRNA by IL-1alpha or TNF-alpha. Furthermore, cytokine stimulation of MMP-1 and TIMP-1 gene expressions can be enhanced or blocked by indomethacin, respectively, and reversed by exogenous PGE2. These results suggested that cytokine-stimulated MMP-1 and TIMP-1 gene expression in dental pulp fibroblasts was mediated, at least in part, by a prostaglandin-dependent pathway. The differential regulation of IL-1alpha or TNF-alpha-induced MMP-1 and TIMP-1 mRNA synthesis, as well as the direct upregulation of TIMP-1 gene expression by PGE2, also implied that prostaglandin may serve as a protective mechanism from excessive tissue breakdown during pulpitis.     

Vertical distraction of a free vascularized osteocutaneous scapular flap in the reconstructed mandible for implant therapy

This paper describes a case of vertical distraction osteogenesis of a free vascularized osteocutaneous scapular flap in the reconstructed mandible before implant therapy. The patient was a 67-year-old woman with squamous cell carcinoma of the right lower gingiva. She underwent segmental mandibulectomy for tumor ablation and reconstruction with an osteocutaneous scapular flap. The distraction protocol, clinical course and implant therapy are presented. Through this procedure, the bone height of the scapular graft increased by 10mm. Implants with adequate length could be placed in the distracted area. Two years after masticatory loading, the condition of these implants was stable. Vertical distraction osteogenesis of the scapular flap was considered effective when performed before implant therapy, to facilitate postoperative functional and esthetic restoration after tumor resection.