[Enhancer of zeste homolog 2 affects dental pulp inflammation by regulating macrophage chemotaxis].

OBJECTIVE: To investigate the expression changes of the epigenetic regulator enhancer of zeste homolog 2 (EZH2) during pulp inflammation and the effect of EZH2 on macrophages migration. METHODS: Rat dental pulp was stimulated with 10 g/L lipopolysaccharide (LPS) to establish a model of rat pulpitis at different stages of inflammation. Immunohistochemical staining was used to detect the expression changes of EZH2 during the progression of pulp inflammation. Immunofluorescence double staining was used to detect the expression of EZH2, CD68 and their colocalization. To screen the appropriate concentration of EZH2 recombinant protein to stimulate hDPCs and human leukaemia-derived monocytic cell line (THP-1) cells, the effects of different concentrations (1, 10, 20, 40, and 100 µg/L) of EZH2 recombinant protein on proliferation of human dental pulp cells (hDPCs) and human monocyte cell line THP-1 were detected by cell counting kit-8 (CCK-8). Transwell migration assay was used to detect the effect of supernatants of hDPCs treated with EZH2 recombinant protein on the migration of THP-1 cells. RESULTS: HE staining results showed that in the model of rat pulp inflammation induced by LPS, with the prolongation of LPS stimulation, the inflammation response of pulp gradually increased. Immunohistochemical results showed that EZH2 expression decreased within 8 h of LPS-induced dental pulp inflammation; but after 1, 3, and 7 d of stimulation, EZH2 expression gradually increased with the extension of the stimulation time. As for the normal rat dental pulp tissue, the positive expression of EZH2 was scattered in the odontoblast cell layer and the pulp proper. Compared with the control group, LPS stimulated the expression of EZH2 and CD68 in the infected dental pulp, and the colocalization of EZH2 and CD68 could be detected in macrophages. The results of CCK-8 suggested that the appropriate concentration of EZH2 recombinant protein to stimulate hDPCs and THP-1 cells was 20 µg/L. Transwell cell migration assay confirmed that compared with the supernatant of EZH2 untreated HDPCs group, the supernatant of EZH2treated hDPCs significantly promoted macrophage chemotaxis. CONCLUSION: EZH2 is involved in the development of pulpitis and promotes the chemotaxis of macrophages, which suggests that EZH2 may play an important regulatory role in the development of pulp inflammation.

[Feasibility study of cone-beam CT for the analysis of cortical bone thickness in the posterior area of the jaw bone].

Objective: To investigate the feasibility of cone-beam CT (CBCT) in evaluating the thickness of cortical bone in jaw bone. Methods: Sixty patients [twenty-three for males and forty-seven for females, at an average age of (43.8±1.7) years] from Center of Stomatology, The First Affiliated Hospital of USTC & Anhui Provincial Hospital with 63 operational regions were included in the present study. Totally 63 bone sections from these areas were all selected at last. Case Viewer and oral dynamic system were used for the measurements in sections and CBCT graphs of the cortical bone thicknesses at alveolar ridges. Paired samples t test was performed to compare the difference between CBCT measurement and Case Viewer measurement. Results: The cortical bone thicknesses measured by Case Viewer were (1.20±0.75), (0.68±0.46) and (1.48±0.77) mm in the posterior, maxillary posterior and mandibular posterior areas, respectively. The cortical bone thicknesses measured by dynamic navigation software were (1.14±0.77), (0.64±0.24) and (1.41±0.83) mm in the posterior, maxillary posterior and mandibular posterior areas, respectively. There were no significant differences between either the two methods or the different areas (P>0.05). Conclusions: CBCT would be a useful equipment for the analysis of cortical bone thickness with a reliable and convincible accuracy.

Remineralization and fluoride uptake of white spot lesions under dental varnishes.

INTRODUCTION: The aim of this study was to evaluate white spot lesion (WSL) remineralization and fluoride uptake by the application of fluoride varnishes directly onto artificial WSLs in vitro. METHODS: MI varnish containing casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) and 2.26% fluoride and Duraphat varnish containing 2.26% fluoride (no added calcium) were compared with a placebo varnish (no added calcium or fluoride). Two WSLs were prepared in enamel slabs and varnish applied to cover one of the two lesions. Each slab was immersed in artificial saliva for 14 days at 37°C. Mineral content was determined using transverse microradiography and fluoride uptake using electron probe microanalysis. The data were statistically analysed using a linear mixed model. RESULTS: Both MI and Duraphat varnishes significantly remineralized the covered and uncovered WSLs when compared with the placebo varnish (P < 0.001). The WSLs covered with varnish showed greater remineralization than those uncovered. MI varnish produced the highest level of remineralization and significantly greater fluoride uptake (0.44 ± 0.08 wt%) compared with Duraphat (0.24 ± 0.03 wt%) and the placebo varnish (0.06 ± 0.05 wt%). CONCLUSION: Varnish containing fluoride and CPP-ACP was superior to varnish containing fluoride alone in promoting WSL remineralization and fluoride uptake.

[Water sorption and hygroscopic changes of five core buildup composite resins].

Objective: To evaluate water sorption and hygroscopic dimensional changes of five core buildup composite resins, and to provide references for material modification on water sorption. Methods: Five commercial core buildup materials (group A: Smart Dentin Replacement(TM); group B: NanoFil; group C: ParaCore(TM); group D: LuxaCore Z; group E: EmbraceCore(TM) Resin Cement) were fabricated to disk-shaped specimens: (15.0±0.1) mm diameter, (2.0±0.1) mm thickness (n=10). Specimens were thoroughly irradiated with curing lights. The initial mass in air was recorded, and the initial mass in deionized water was recorded. Five specimens of each group were immersed in deionized water for 28 d. They were weighed as a function of different immersion time (1, 2, 3, 4, 5, 6, 7, 14, 28 d). The mass in air was recorded, and the displayed mass in deionized water was recorded. Archimedes' principle was applied to calculate the dimensional changes. The other five specimens of each group were stored in artificial saliva and were tested by the same methods. Results: All specimens gained weight and hygroscopic changes during 28 d immersion. When stored in deionized water, the apparent mass change of group A [(10.6±0.9) µg/mm(3)] and the dimensional change of group A [(0.39±0.10)%] were the lowest, while the mass change of group E [(48.0±0.2) µg/mm(3)] and the dimensional change of group E [(3.16±0.13)%] were the highest (P<0.05). In artificial saliva, the lowest apparent mass change was found in group A [(11.8±1.0) µg/mm(3)] while the highest change was found in group E [(47.4±3.5) µg/mm(3)] (P<0.05). The lowest dimensional change was found in group C [(0.37±0.09)%] and the highest was found in group E [(3.07±0.19)%] in artificial saliva (P<0.05). Conclusions: Water sorption and dimensional changes vary in immersion fluids with different osmotic pressure. Water sorption and dimensional changes of core buildup composite resins are highly correlated with test materials in both of the deionized water and artificial saliva.

Spheres of influence: Porphyromonas gingivalis outer membrane vesicles.

Outer membrane vesicles (OMVs) are asymmetrical single bilayer membranous nanostructures produced by Gram-negative bacteria important for bacterial interaction with the environment. Porphyromonas gingivalis, a keystone pathogen associated with chronic periodontitis, produces OMVs that act as a virulence factor secretion system contributing to its pathogenicity. Despite their biological importance, the mechanisms of OMV biogenesis have not been fully elucidated. The ~14 times more curvature of the OMV membrane than cell outer membrane (OM) indicates that OMV biogenesis requires energy expenditure for significant curvature of the OMV membrane. In P. gingivalis, we propose that this may be achieved by upregulating the production of certain inner or outer leaflet lipids, which causes localized outward curvature of the OM. This results in selection of anionic lipopolysaccharide (A-LPS) and associated C-terminal domain (CTD) -family proteins on the outer surface due to their ability to accommodate the curvature. Deacylation of A-LPS may further enable increased curvature leading to OMV formation. Porphyromonas gingivalis OMVs that are selectively enriched in CTD-family proteins, largely the gingipains, can support bacterial coaggregation, promote biofilm development and act as an intercessor for the transport of non-motile bacteria by motile bacteria. The P. gingivalis OMVs are also believed to contribute to host interaction and colonization, evasion of immune defense mechanisms, and destruction of periodontal tissues. They may be crucial for both micro- and macronutrient capture, especially heme and probably other assimilable compounds for its own benefit and that of the wider biofilm community.

Cariogenicity of Streptococcus mutans strains with defects in fructan metabolism assessed in a program-fed specific-pathogen-free rat model.

To define the role of dental plaque fructans and the enzymes involved in their metabolism in the initiation and progression of dental caries, we constructed otherwise-isogenic mutants of Streptococcus mutans defective in the ability to synthesize fructans, to degrade fructans, or to do both. The cariogenic potential of these organisms was evaluated in a specific-pathogen-free rat model in which the feeding patterns of the animals were controlled by means of a König-Hofer programed feeder. Specifically, rats were infected with wild-type S. mutans UA159 or derivatives of this strain which contained an insertionally-inactivated fructanase (fruA) gene, fructosyltransferase (ftf) gene, or which had both genes inactivated. The animals were fed 17 meals per day of Diet 2000 containing 56% sucrose at 70-minute intervals for five weeks, and caries experience was evaluated. Animals infected with S. mutans with a mutated fruA gene only had statistically significant decreases in sulcal caries severity. Such a decrease was not observed in previous studies with ad libitum-fed animals (Wexler et al., 1992). The manifestation of diminished virulence in the programmed feeding model, but not in ad libitum-fed animals, supports the concept that the primary contribution of FruA to virulence is through the utilization of fructans storage polysaccharides. Animals infected with strains carrying the ftf mutation or simultaneous mutations in ftf and fruA did not display decreased virulence, perhaps indicating that sucrose utilization pathways may compete for substrate in vivo, or that accumulation of fructans may affect the ecology or the physicochemical characteristics of dental plaque in such a way as to reduce its cariogenic potential. The results of this study also emphasize that the contribution of a particular virulence determinant to the caries process may be highly dependent on the experimental design, feeding regimen and diet, and the presence or absence of other enzymatic activities.

Synthesis and characteristics of [60]fullerene polysiloxane stationary phase for capillary gas chromatography.

A new type of fullerene-containing polysiloxane was synthesized by reacting [60]fullerene with azidopropyl polysiloxane directly. The polysiloxanes have been used successfully as stationary phases for capillary gas chromatography. They displayed high column efficiency, wide operational temperature and high thermostability, and exhibited unique selectivity for many organic substances, such as alkanes, alcohols, ketones and aromatic compounds. The stationary phase was especially suitable for separation of high boiling-point compounds like polycyclic aromatic hydrocarbons and phthalic esters, etc. It was also found that some alcoholic or aromatic positional isomers could be well separated on the column. The influence of the fullerene content on the separation was also investigated.

Cardiac troponin I: a reliable marker and early myocardial involvement with meningoencephalitis after fatal enterovirus-71 infection.

OBJECTIVES: A major outbreak of enterovirus 71 (EV71) in Taiwan in 1998 caused many severe cases and 78 deaths. Our purpose was to find reliable markers and early indicators of fatal EV71 central nervous system (CNS) infection. METHODS: From June 2000 to November 2001, 21 patients with hand foot mouth disease or herpangina with CNS infection were admitted to Kaohsiung Veterans General Hospital. All 21 had culture-confirmed EV71 infection or were EV71 IgM positive. Patients were divided into two groups: group I included the five fatalities at our institution and group II, the 16 surviving patients. RESULTS: Of the 21 infants and children with EV71 infection with CNS involvement, MR imaging studies were completed on 17, and 15 showed hyperintensity in the posterior portions of brain stem. All patients received intravenous immunoglobulin (IVIG) 1 g/day for two days and supportive care. Five patients rapidly deteriorated owing to irreversible hypotension and died. The other 16 patients recovered completely without sequel. In group I patients, the decrease of cardiac ejection function is significant and laboratory findings showed lower platelet count (P=0.0192). The mean of initial cTnI level for groups I and II was 10.6+/-11.6 and 0.48+/-0.55 ng/dl, respectively, higher in group I than in II (P=0.0019). CONCLUSION: We hypothesized that like patients with severe burns, those with severe EV-71 CNS meningoencephalitis have varying degrees of non-ischemic cardiac injury, manifesting as leakage of cTnI from myocytes into the circulation. EV-71 CNS meningoencephalitis likely to die with an early myocardial involvement evidenced by reduced ejection fraction and release of cTnI. We conclude that fatal EV71 CNS infection quickly leads to death due to severe encephalopathy associated with cardiomyopathy.

The effect of bonding agent and fissure sealant on microleakage of composite resin restorations.

The purpose of this study was to assess, in vitro, the microleakage of Class V composite resin restorations following sealing of the margins with a fissure sealant or the resin component of a dentinal bonding agent, and to compare it to the microleakage of an unsealed control group. All groups of teeth showed some degree of leakage at the gingival margin. Statistical analysis showed that there was significantly less leakage when the fissure sealant (P less than .01) and bonding agent (P less than .001) were used than there was in the control group, in which the margins had not been sealed. There was no statistically significant difference between the two experimental groups. There was no leakage at the enamel margin in any of the teeth.

Cytotoxicity of release products from magnetic nanocomposites in targeted drug delivery.

The efficacy of chemotherapy can be significantly improved if the therapeutic agent remains localized at the afflicted area and released at controlled rates. Such a targeted drug delivery can be achieved using magnetic nanocomposite (MNC), which incorporates drug and magnetic nanoparticles in biodegradable polymer microspheres. Reported here are results from an in vitro study on drug release rate and cytotoxicity of other release products from MNC. The model system contains an anti-cancer chemotherapy agent 5-flurouracil (5-FU) and (Co(0.5)Zn(0.5))Fe(2)O(4) in poly(lactic-co-glycolic acid) (PLGA) matrix produced by an oil/oil emulsion technique. Cell proliferation data indicate a sustained release of 5-FU for mouse macrophage cell eradication, whereas other microsphere components of magnetic nanoparticles and PLGA have little cytotoxic effects.

The relationship between dental caries status and dental plaque urease activity.

INTRODUCTION: Ammonia production from the metabolism of urea by urease enzymes of oral bacteria moderates plaque acidification and may inhibit dental caries, as suggested by in vitro studies and indirect clinical observations. The objective of this study was to examine the relationship of urease activity with dental caries at the clinical level. METHODS: Urease activity was measured in dental plaque and saliva samples from 25 caries-free subjects (CF) and in eight subjects with six or more open caries lesions (CA). Plaque and saliva collection was repeated for each subject 1 week later using identical procedures. RESULTS: Urease-specific activity in the dental plaque of CF subjects was significantly higher compared to that in the subjects with caries. The association of low plaque urease levels with increased caries was further supported by odds ratio analysis using different plaque urease cut-off points. Using a receiver operating characteristic curve it was estimated that there was an approximately 85% probability of correctly classifying the subjects as CA or CF based on the relative ordering of their plaque urease activity levels. No statistically significant differences were observed in salivary urease activity. CONCLUSION: This study suggests that loss of alkali-generating potential of tooth biofilms via the urease pathway has a positive relationship to dental caries.

Comparison of bond strength between lingual and buccal surfaces on young premolars.

The purpose of this experiment was to compare the bond strength and the failure interfaces on lingual and buccal surfaces of young human premolars. Premolar buccal brackets were adapted and bonded on the lingual or buccal surfaces of crowns after etching with a 37% phosphoric acid solution for 15 seconds. The bond strengths were 7.2 MPa and 7.0 MPa for lingual and buccal surfaces, respectively. The bond failure interfaces were located between the bracket and the resin, within the resin itself, or between the resin and the enamel. Tooth fragmentation was not found. There were no statistically significant differences (p > 0.05). This indicates that it is not necessary to specially treat a lingual surface for increasing the bond strength.

Analysis of gene expression in Streptococcus mutans in biofilms in vitro.

The purpose of this study was to develop methods for the consistent production of biofilms of S. mutans containing reporter gene fusions, and to examine the expression of genes involved in sucrose metabolism in adherent populations of this organism. Three strains of S. mutans harboring reporter gene fusions to the gene promoter regions of the gtfBC genes, ftf, and scrA were grown in a Rototorque biofilm fermenter in a tryptone-yeast extract-sucrose medium. Quasi-steady-state levels of reporter gene activity were measured after the biofilms were grown for either 48 hrs of 7 days. Also, induction of gene expression by the addition of sucrose to biofilm cells was monitored. Reporter gene activity was measurable from all gene fusion strains. This study (i) establishes the feasibility of doing detailed molecular and physiologic studies on immobilized populations of S. mutans, (ii) demonstrates that the polysaccharide synthesis machinery of S. mutans is differentially expressed in biofilms, and (iii) opens the way for a more detailed analysis of the environmental signals and signal transduction pathways governing the regulation of gene expression by S. mutans cells that are immobilized on a solid surface.

Streptococcus salivarius urease: genetic and biochemical characterization and expression in a dental plaque streptococcus.

The hydrolysis of urea by urease enzyme of oral bacteria is believed to have a major impact on oral microbial ecology and to be intimately involved in oral health and diseases. To begin to understand the biochemistry and genetics of oral ureolysis, a study of the urease of Streptococcus salivarius, a highly ureolytic organism which is present in large numbers on the soft tissues of the oral cavity, has been initiated. By using as a probe a 0.6-kpb internal fragment of the S. salivarius 57.I ureC gene, two clones from subgenomic libraries of S. salivarius 57.I in an Escherichia coli plasmid vector were identified. Nucleotide sequence analysis revealed the presence of one partial and six complete open reading frames which were most homologous to ureIAB-CEFGD of other ureolytic bacteria. Plasmid clones were generated to construct a complete gene cluster and used to transform E. coli and Streptococcus gordonii DL1, a nonureolytic, dental plaque microorganism. The recombinant organisms expressed high levels of urease activity when the growth medium was supplemented with NiCl2. The urease enzyme was purified from E. coli, and its biochemical properties were compared with those of the urease produced by S. salivarius and those of the urease produced by S. gordonii carrying the plasmid-borne ure genes. In all cases, the enzyme had a Km of 3.5 to 4.1 mM, a pH optimum near 7.0, and a temperature optimum near 60 degrees C. S. gordonii carrying the urease genes was then demonstrated to have a significant capacity to temper glycolytic acidification in vitro in the presence of concentrations of urea commonly found in the oral cavity. The ability to genetically engineer plaque bacteria that can modulate environmental pH through ureolysis will open the way to using recombinant ureolytic organisms to test hypotheses regarding the role of oral ureolysis in dental caries, calculus formation, and periodontal diseases. Such recombinant organisms may eventually prove useful for controlling dental caries by replacement therapy.

Phenotypic and functional analysis of activated B cells of autoimmune NZB x NZW F1 mice.

Polyclonal B-cell activation is the central theme in the production of autoantibodies and possible activation of autoreactive T cells in both human and murine lupus. The abnormal expansion of CD5+ B cells in murine lupus has been suggested, in particular, to be one of the most characteristic findings in these mice. Activated B cells can be separated from the B cells of resting stage by the difference in cell density. The aim of this study was to investigate the characteristics of different densities of the spleen cells separated by gradient density. Furthermore, the ability of anti-DNA antibody secretion in each percoll gradient fraction of B cells was also analysed. The results showed: a higher percentage of CD5+ B cells, which corresponded to the activated B-cell population, in percoll gradient 1 and 2 fractions; that splenic B cells of NZB/W F1 mice had proliferative response to interleukin (IL)-4 or IL-5 but not to IL-10 or interferon-gamma (IFN-gamma); and that B cells isolated by percoll gradient produced anti-DNA antibody after stimulation with lipopolysaccharide (LPS) plus IL-5 and IFN-gamma, but not IL-4 and IL-10. These data suggest that B cells at different stages of activation express differential characteristics and functions.

[Preparation and chromatographic characteristics of linear [60] fullerene polysiloxane stationary phase for capillary gas chromatography].

Hydrosilylation of polymethylhydrosiloxane (active hydrogen content was 0.05%) and reactions with allylbromide (or undecenyl bromide), sodium azide and [60] fullerene in their given orders is described. Two kinds of fullerene-containing polysiloxanes were obtained [polymer(3)]. Their [60] fullerene contents were 30 mg (C60-PSO-1) and 60 mg (C60-PSO-2) respectively. These polymers (3) were characterized by FT-IR, 1H-NMR, UV-V, 13C-NMR and used as stationary phase for capillary chromatography. Two fused-silica capillary columns (0.25 mm i.d.) were coated with a 5 g/L solution of polymer (3) in dichloromethane by the static method. The column efficiencies were 3,450 plates/m (C60-PSO-1) and 3,020 plates/m (C60-PSO-2), for naphthalene at 120 degrees C. The average polarities of these columns, represented by the McReynold's Constant, were 79 and 93 respectively. The polarity of C60-PSO-2 was a little higher than that of C60-PSO-1, which was in turn a little higher than that of SE-30(43). These columns showed wide operation temperature range (100-360 degrees C) and outstanding thermostability. It could be used at 360 degrees C with a baseline drift of 30-40 fA. Grob test mixtures eluted on these columns in the ordre of 1,3-butanediol, n-decane, 1-octanol, n-undecane, 2,6-dimethylphenol, n-dodecane, 2,4-dimethylaniline, naphthalene, indicating that the strong pi-pi interaction of [60] fullerene with aromatic moieties played an important role in separation mechanism. The columns exhibited unique selectivity for many organic compounds, such as alkanes, alcohols, ketones and anilines. It can well separate benzene homologues, naphthalene homologues and polyaromatic hydrocarbons(PAHs) due to the pi-pi interaction of three-dimensional pi system of C60 and aromatic hydrocabons. And the columns are especially good for the separation of positional isomers(nonanol, butanediol, nitrotoluene, nitrochlorobenzene, nitroethylbenzene, dinitrobenzene, chlorophenol). The columns also have excellent thermostability and are particularly suitable for the separation of high boiling compounds. For example, polycyclic aromatic hydrocarbons and fatty acid methyl esters are well separated on them.