17%乙二胺四乙酸冲洗液对牙本质粪肠球菌黏附的影响

目的 研究17%乙二胺四乙酸（EDTA）冲洗液对促进牙本质粪肠球菌黏附的影响。方法 将48例半劈样本及12例牙本质片样本随机分为3个实验组和1个对照组,实验组分别经17%EDTA处理1、3、5 min,对照组经生理盐水处理5 min,接种粪肠球菌后通过扫描电子显微镜、激光共聚焦电子显微镜（CLSM）、菌落形成单位及组织学方法进行细菌黏附量的评价。结果 除组织学革兰染色显示,1 min组与对照组、3 min组的细菌侵入深度无明显差异（P>0.05）外,其余结果显示,实验组细菌生物膜厚度、侵入牙本质小管深度（CLSM测量）及菌落计数均大于对照组,差异具有统计学意义（P<0.05）;3个实验组之间比较,差异也具有统计学意义（P< 0.05）。结论 17%EDTA冲洗液可促进粪肠球菌对牙本质的黏附,且黏附量随处理时间的延长而增加。     

17％乙二胺四乙酸冲洗液对牙本质粪肠球菌黏附的影响

目的 研究17%乙二胺四乙酸（EDTA）冲洗液对促进牙本质粪肠球菌黏附的影响。方法 将48例半劈样本及12例牙本质片样本随机分为3个实验组和1个对照组,实验组分别经17%EDTA处理1、3、5 min,对照组经生理盐水处理5 min,接种粪肠球菌后通过扫描电子显微镜、激光共聚焦电子显微镜（CLSM）、菌落形成单位及组织学方法进行细菌黏附量的评价。结果 除组织学革兰染色显示,1 min组与对照组、3 min组的细菌侵入深度无明显差异（P>0.05）外,其余结果显示,实验组细菌生物膜厚度、侵入牙本质小管深度（CLSM测量）及菌落计数均大于对照组,差异具有统计学意义（P<0.05）;3个实验组之间比较,差异也具有统计学意义（P< 0.05）。结论 17%EDTA冲洗液可促进粪肠球菌对牙本质的黏附,且黏附量随处理时间的延长而增加。     

氢氧化钙和洗必泰抑制粪肠球菌的抗菌性检测

目的:检测氢氧化钙和洗必泰凝胶根管封药对粪肠球菌的抑制作用。方法:将成功建立粪肠球菌感染的60个根管模型,随机分成4组,每组15个样本,分别用氢氧化钙、20 g/L洗必泰凝胶、氢氧化钙和20 g/L洗必泰凝胶联合、生理盐水(空白对照组)进行根管内封药处理,14 d后,分别用5号和6号GG钻钻取根管内壁层深0～100μm和100～200μm的牙本质,收集碎屑,用BHI梯度稀释后进行细菌培养和计数。结果:无论在根管壁深0～100μm处,还是100～200μm处,空白组均有大量的细菌生长,而3个实验组根管内粪肠球菌明显减少,分别与空白对照组相比,差异均具有统计学意义(P<0.05);各实验组之间相比,氢氧化钙组活菌数最高,与其他两组相比,差异均有统计学意义(P<0.05),其次为20 g/L洗必泰凝胶组,氢氧化钙联合洗必泰组最低,但与单独使用洗必泰组相比,差异无统计学意义(P>0.05)。结论:在根管封药中,氢氧化钙和洗必泰均对粪肠球菌有较好的抗菌性,其中洗必泰凝胶的抑菌作用更强。     

Effect of smear layer and direction of dentinal tubules on osteoblast adhesion to human dentin tissue

The purpose of this study was to investigate the effect of smear layer and direction of dentinal tubules on osteoblast adhesion to human dentin tissue in vitro. Dentin disks were made from human premolars extracted for orthodontic reasons. Dentin disks were cut either perpendicularly to the long axis of the tooth or at 45 degrees to the long axis of tooth. The smear layer was removed by 34% phosphoric acid gel from half of the dentin disk surface. Human osteoblast-like Saos-2 cells were grown in RPMI medium with 10% fetal bovine serum and 1% antibiotic/antimycotic cocktail under standard cell culture conditions. Cells were seeded into Nunc four-well culture plates at 1.5 x 10(5) cells per well with dentin disks in the bottom of each well. After 1 day in culture the dentin disks along with cells grown on their surface were examined with a scanning electron microscopy. Osteoblasts attached and spread on the dentin surface and formed a monolayer in the presence and absence of a smear layer. Cells spread over the dentinal tubules despite their direction. These results suggest that cell adhesion and spreading of osteoblasts is not influenced either by the existence of a smear layer or the direction of the dentinal tubules on the dentin surface.     

Influence of Enterococcus faecalis proteases and the collagen-binding protein,Ace, on adhesion to dentin

Enterococcus faecalis is a pathogen that persists in medicated root canals. Here, we tested the hypothesis that the E. faecalis proteases, serine protease and gelatinase, and the collagen-binding protein (Ace) contribute to adhesion to the root canal. Scanning electron microscopy was used to examine dentin binding by four E. faecalis strains: OG1RF, the wild type, and three mutant derivatives of OG1RF, TX5128, TX5243 and TX5256 deficient in serine protease and gelatinase, serine protease, and Ace, respectively. For each strain, 20 root halves were exposed to 3 x 10(9) to 5 x 10(9) cells/ml for 6 h, and 50 fields per root half were examined for adherent bacteria. Statistical analysis revealed that adherence of OG1RF was significantly greater than the mutant strains (P < 0.001), while significant differences were not detected between the protease mutants. The data indicate that serine protease and Ace aid E. faecalis binding to dentin, while the role of gelatinase is uncertain.     

Effect of tissue fluids on hydrophobicity and adherence of Enterococcus faecalis to dentin

This in vitro study was carried out to determine (1) the hydrophobicity of selected oral bacteria, (2) the influence of growth media (saliva and serum) and mode of growth (planktonic or biofilm) on the hydrophobicity of Enterococcus faecalis, and (3) the influence of growth media and conditioning fluids on the adherence of E. faecalis to dentin. The ability to bind to a hydrocarbon phase (xylene) was used as an index of relative hydrophobicity of cells. Fluorescent microscopy-based technique was used to assay the bacterial adherence to dentin. Results showed that bacteria involved in the primary stage of oral biofilm formation such as Streptococcus mutans, Fusobacterium nucleatum, and Porphyromonas gingivalis are relatively more hydrophobic than E. faecalis. The hydrophobicity of E. faecalis was significantly increased during starvation and biofilm mode of growth (p < .05). The adherence of E. faecalis to dentin was appreciably increased after starvation and when dentin was conditioned with saliva. It was observed that surface conditioning of dentin with saliva and starvation can enhance the adherence of E. faecalis to dentin. The findings from this study indicated that the coronal leakage of saliva and the physiologic state of microbes might play an important role in the adherence and biofilm formation of bacteria to root canal dentin.     

Influence of Enterococcus faecalis proteases and the collagen‐binding protein,Ace, on adhesion to dentin

Enterococcus faecalis is a pathogen that persists in medicated root canals. Here, we tested the hypothesis that the E. faecalis proteases, serine protease and gelatinase, and the collagen‐binding protein (Ace) contribute to adhesion to the root canal. Scanning electron microscopy was used to examine dentin binding by four E. faecalis strains: OG1RF, the wild type, and three mutant derivatives of OG1RF, TX5128, TX5243 and TX5256 deficient in serine protease and gelatinase, serine protease, and Ace, respectively. For each strain, 20 root halves were exposed to 3 × 10⁹ to 5 × 10⁹ cells/ml for 6 h, and 50 fields per root half were examined for adherent bacteria. Statistical analysis revealed that adherence of OG1RF was significantly greater than the mutant strains (P < 0.001), while significant differences were not detected between the protease mutants. The data indicate that serine protease and Ace aid E. faecalis binding to dentin, while the role of gelatinase is uncertain.     

Effect of smear layer removal agents on the microhardness and roughness of radicular dentin

PurposeTo evaluate the effect of phytic acid (IP6) on the surface roughness and microhardness of human root canal dentin and compare it to other smear layer removal agents.Materials and methodsFifty extracted human maxillary incisors were sectioned longitudinally into a total of 100 specimens followed by embedding in auto-polymerizing acrylic resin. The specimens were polished and then randomly divided into five groups (n = 20) according to the test solution used to condition root canal dentin: 17% ethylenediaminetetraacetic acid (EDTA); 10% citric acid (CA); 1% IP6; 37% phosphoric acid (PA); or distilled water (control group). Each specimen was treated with a total volume of 1 ml of each solution for 1 min with agitation. Each group was then divided into two subgroups of 10 specimens each. The specimens of the first subgroup were used to determine microhardness, using Vickers hardness tester, and the specimens of the second subgroup were used to measure surface roughness, using a confocal laser scanning microscope. The results were analyzed statistically using one-way ANOVA and Tukey tests, α = 0.05.ResultsAll the tested groups exhibited microhardness and surface roughness values that were statistically significantly different when compared with the control group (P < 0.05). The microhardness value obtained with IP6 was significantly lower when compared to EDTA, CA, and the control group, whereas its roughness value was significantly higher compared to the aforementioned groups. However, there was no significant difference between IP6 and PA (P > 0.05).ConclusionsIP6 and PA showed the lowest microhardness and the highest surface roughness values.     

Influence of irrigation regimens on the adherence of Enterococcus faecalis to root canal dentin

Enterococcus faecalis is frequently associated with post-treatment endodontic infections. Because adherence of bacteria to a substrate is the earliest stage in biofilm formation, eliciting the factors that links adherence of this bacterium to dentin would help in understanding its association with treatment-failed root canals. This investigation aimed to study the effects of endodontic irrigants on the adherence of E. faecalis to dentin. The bacteria adherence assay was conducted by using fluorescence microscopy, and the adhesion force was measured by using atomic force microscopy. There were significant increases in adherence and adhesion force after irrigation of dentin with ethylenediaminetetraacetic acid (EDTA), whereas sodium hypochlorite (NaOCl) reduced it. With the use of chlorhexidine (CHX), the force of adhesion increased, but the adherence assay showed a reduction in the number of adhering bacteria. The irrigation regimen of EDTA, NaOCl, and CHX resulted in the least number of adhering E. faecalis cells. This study highlighted that chemicals that alter the physicochemical properties of dentin will influence the nature of adherence, adhesion force, and subsequent biofilm formation of E. faecalis to dentin.     

Enterococcus faecalis adhesin, Ace, mediates attachment to particulate dentin

An enzyme linked immunosorbent assay was developed to assess E. faecalis adhesion to particulate dentin. E. faecalis, OG1RF, which expresses the collagen binding protein (Ace+), and a derivative of OG1RF, TX5256, deficient in the collagen binding protein (Ace-) were grown at 46 degrees C, necessary for in vitro expression of Ace, and at 37 degrees C. E. faecalis binding to dentin was measured at 0, 15, 30, 60, 120, and 360 minutes. Compared to TX5256 and OG1RF grown at 37 degrees C, OG1RF grown at 46 degrees C adhered significantly better at all time points except 15 minutes (p < 0.001) exhibiting maximum binding at 120 minutes (17.4% of a positive control). Type I collagen at 100 microg/ml inhibited dentin binding by OG1RF grown at 46 degrees C in both competition (p < 0.005) and displacement assays (p < 0.046). Immunoaffinity purified anti-Ace IgG at 200 microg of protein inhibited adhesion of OG1RF grown at 46 degrees C to dentin.     

Adhesion of sealer cements to dentin with and without the smear layer

The influence of a smear layer on the adhesion of sealer cements to dentin was assessed in recently extracted human anterior teeth. A total of 120 samples was tested, 40 per sealer; 20 each with and without the smear layer. The teeth were split longitudinally, and the internal surfaces were ground flat. One-half of each tooth was left with the smear layer intact, while the other half had the smear removed by washing for 3 min with 17% EDTA followed by 5.25% NaOCl. Evidence of the ability to remove the smear layer was verified by scanning electron microscopy. Using a specially designed jig, the sealer was placed into a 4-mm wide x 4-mm deep well which was then set onto the tooth at a 90-degree angle and allowed to set for 7 days in 100% humidity at 37 degrees C. This set-up was then placed into a mounting jig which was designed for the Instron Universal Testing Machine so that only a tensile load was applied without shearing or applying preloading forces. The set-up was subjected to a tensile load at a crosshead speed of 1 mm per min. The results show significant differences (p less than 0.001) among AH26, Sultan, and Sealapex, with AH26 being the strongest and Sealapex being the weakest. The only significant difference with regard to the presence or absence of the smear layer was found with AH26, which had a stronger bond when the smear layer was removed.     

Effectiveness of selected materials against Enterococcus faecalis: part 3. The antibacterial effect of calcium hydroxide and chlorhexidine on Enterococcus faecalis

It has been found that Enterococcus faecalis is most commonly isolated in failed endodontic treatment. Irrigation with chlorhexidine gluconate has been suggested based on its antimicrobial effect and substantivity. Calcium hydroxide also is an effective antimicrobial agent because of its high alkalinity. The purpose of this study was to test the individual and combined effect of calcium hydroxide and chlorhexidine against E. faecalis. The agar-diffusion test was performed on Mueller-Hinton plates. Paper disks were impregnated with: (a) CaOH powder with sterile water; (b) Pulpdent; (c) 0.12% Peridex; (d) CaOH powder with Peridex; and (e) Pulpdent with Peridex. Ampicillin served as a control. The plates were incubated at 37 degrees C for 72 h. Peridex showed significantly larger zones of inhibition compared with CaOH. No statistically significant difference was found between Peridex and the combination of CaOH and Peridex.     

Growth at high pH increases Enterococcus faecalis adhesion to collagen

AIM: To evaluate the effect of growth at pH levels from 7.1 to 9.5 on the adherence of Enterococcus faecalis to bovine serum albumin (BSA) and collagen type I. METHODOLOGY: Enterococcus faecalis strain A197A was grown in broth of adjusted pHs varying between 7.1 and 9.5. Aliquots of bacterial suspensions were added to wells coated either with BSA or with collagen type I. Bacteria adhering to the surfaces were stained with crystal violet. Spectrophotometric measurements of the dissolved stain were used to assess the number of bacteria adhering to the surfaces. The data obtained were analysed using the Kolmogorov-Smirnov test, Levene's test and Student's t-test, with alpha = 0.05 as the level for statistical significance. RESULTS: The adhesion of E. faecalis to BSA-coated surfaces decreased inversely with alkalinity of the growth medium. The pH 7.1-grown bacteria bound to BSA significantly more than the other BSA groups. On the contrary, the adhesion to collagen type I-coated surfaces of bacteria grown at pH 8.0 and 8.5 was significantly greater than for those grown at pH 7.1. CONCLUSIONS: A minor increase in pH up to 8.5, which may be a consequence of insufficient treatment with alkaline medicaments such as calcium hydroxide, increases the collagen-binding ability of E. faecalis, in vitro. This can be a critical mechanism by which E. faecalis predominates in persistent endodontic infections.     

Effectiveness of 2% chlorhexidine gel and calcium hydroxide against Enterococcus faecalis in bovine root dentine in vitro

AIM: To evaluate the effectiveness of 2% chlorhexidine gluconate gel and calcium hydroxide (Ca(OH)2) as intracanal medicaments against Enterococcus faecalis. METHODOLOGY: One hundred and eighty dentine tubes prepared from intact freshly extracted bovine maxillary central incisors were infected in vitro for 7 days with E. faecalis. The specimens were divided into four groups, according to the intracanal medicament used, as follows: Group 1: 2% chlorhexidine gluconate gel; Group 2: calcium hydroxide in a viscous vehicle (polyethyleneglycol 400); Group 3: 2% chlorhexidine gluconate gel + calcium hydroxide and Group 4: Brain Heart Infusion (BHI) broth (control group). The medicaments were placed into the canal lumen and left there for experimental times of 1, 2, 7, 15 and 30 days. After each period, irrigation with sterile saline to remove the medicament was performed and the canals were dried with sterile paper points. Dentine chips were removed from the canals with sequential sterile round burs at low speed. The samples obtained with each bur were immediately collected in separate test tubes containing BHI broth. The tubes were incubated at 37 degrees C and daily observed for microbial growth, visualized by the medium turbidity. RESULTS: Chlorhexidine gel alone completely inhibited the growth of E. faecalis after 1, 2, 7 and 15 days. Calcium hydroxide allowed microbial growth at all experimental times. The combination of chlorhexidine and Ca(OH)2 was effective after 1 and 2 days demonstrating 100% antibacterial action; however, its antibacterial activity reduced between 7 and 15 days. CONCLUSION: Under the conditions of this study, it can be concluded that 2% chlorhexidine gel alone was more effective against E. faecalis than calcium hydroxide (P < 0.05). However, its antibacterial activity depended on how long it remained inside the root canal.     

Dye penetration of the smear layer and fluoride application to the dentin surface

A smear layer is formed after cavity or root canal preparation. The aim of the present study was to reinforce the dental surface in order to prevent the invasion of foreign irritants, by treating the smear layer with fluoride. Dentinal samples whose surfaces had been washed with water after the formation of a smear layer, and corresponding samples without washing, were examined by the dye penetration test, and the results were compared. Although there was no significant difference between the two groups of samples, dye penetration was suppressed by about 30% in washed samples, whereas the suppression was 20% in unwashed samples. When washed samples were treated with 1.0% SnF2, 10.0% SnF2, 7.5% Na2PO2F, 15% Na2PO2F, APF, 10 mM In(NO3)3, 100 mM TiF3, 50 mM TiF3, or 10 mM TiF3, samples washed after treatment with 1.0% SnF2 showed a dye penetration suppression of about 60% as a whole, in comparison with samples having no smear layer. Hardly any suppression of dye penetration was observed after treatment with other fluorides.     

The influence of the dentin smear layer on adhesion: a self-etching primer vs. a total-etch system

OBJECTIVE: To determine the effect of dentin smear layers created by various abrasives on the adhesion of a self-etching primer (SE) and total-etch (SB) bonding systems. METHODS: Polished human dentin disks were further abraded with 0.05 micro m alumina slurry, 240-, 320- or 600-grit abrasive papers, # 245 carbide, # 250.9 F diamond or # 250.9 C diamond burs. Shear bond strength (SBS) was evaluated by single-plane lap shear, after bonding with SE or SB and with a restorative composite. Smear layers were characterized by thickness, using SEM; surface roughness using AFM; and reaction to the conditioners, based on the percentage of open tubules, using SEM. RESULTS: Overall, SBS was lower when SB was used than when SE was used. SBS decreased with increasing coarseness of the abrasive in the SE group. Among burs, the carbide group had the highest SBS, and 320- and 240-grit papers had SBS close to the carbide group. Surface roughness and smear layer thickness varied strongly with coarseness. After conditioning with SE primer, the tubule openness of specimens abraded by carbide bur did not differ from 240- or 320-grit paper, but did differ from the 600-grit. SIGNIFICANCE: Even though affected by different surface preparation methods, SE yielded higher SBS than SB. The higher SBS and thin smear layer of the carbide bur group, suggests its use when self-etching materials are used in vivo. Overall, the 320-grit abrasive paper surface finish yielded results closer to that of the carbide bur and its use is recommended in vitro as a clinical simulator when using the SE material.     

Antimicrobial efficacy of chlorhexidine and two calcium hydroxide formulations against Enterococcus faecalis

The purpose of this study was to investigate the efficacy of chlorhexidine (CHX) and calcium hydroxide (Ca(OH2) against Enterococcus faecalis in vitro. Extracted single-rooted human teeth were instrumented up to size 40. After removal of the smear layer, an inoculum of E. faecalis was inserted into the root canals. After incubation, the inoculum was removed and the root canals were filled with one of three different disinfectants: Ca(OH2 paste, CHX 2%, and a mixture of CHX and Ca(OH2 paste (n = 10 in each group). Control teeth were filled with water of standardized hardness (n = 10). The teeth were then incubated for 3 days. After incubation, each root canal was instrumented, and the removed dentin was examined microbiologically. CHX was significantly more effective against E. faecalis than was Ca(OH2 paste or a mixture of CHX with Ca(OH2 paste (p < 0.05). There was no increase in the efficiency of Ca(OH2 paste when CHX was added (p > 0.05). The results suggest that CHX is effective in the elimination of E. faecalis from dentinal tubules under the conditions of this study.     

A new method for studying the adhesion of Candida albicans to dentin in the presence or absence of smear layer

OBJECTIVES: The purpose of this study was to develop a reproducible, quantitative model of Candida albicans adhesion to human dentin through the use of a colorimetric method and to evaluate the effect of smear layer on candidal adhesion. STUDY DESIGN: Dentin disks with or without smear layer were incubated with C albicans (10(8) cells/mL) for 4 hours. After incubation, the disks were exposed to an (2,3)-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)-carbonyl]-2H-tetrazolium hydroxide-coenzyme Q solution for 2 hours. The color of (2,3)-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)-carbonyl]-2H-tetrazolium hydroxide formazan in the supernatant was determined spectrophotometrically at 492 nm. To relate formazan formation to cell numbers, standard curves were generated with known numbers of yeast cells without dentin. The number of adherent cells per square millimeter was then calculated. RESULTS: The number of attached C albicans cells was 2.4 x 10(4) per square millimeter in dentin with smear layer and 1.5 x 10(4) in dentin without smear layer (P <.05). CONCLUSION: (2,3)-Bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)-carbonyl]-2H-tetrazolium hydroxide assay is a potential microbiologic tool for the quantitative determination of Candida adhesion to human dentin.