互联网时代,知识重装分类,获取知识的路径和学识标准已完全改变,这对高校教师是机遇也是挑战.探讨了当前医学高等教育的一些问题.从教育以学生为中心、智慧为中心、终身为中心等视角,提出了一点个人浅见.

Quorum-sensing signalling molecules such as N-acyl homoserine lactones (AHLs) enable certain Gram-negative bacteria to respond to environmental changes through behaviours, such as biofilm formation and flagellar movement. The present study aimed to identify Acinetobacter baumannii AHLs and assess their influence on antibiotic resistance. A clinical isolate of A. baumannii strain S (AbS) was collected from the wound of a burn patient and high-performance liquid chromatography and tandem quadrupole or quadrupole time-of-flight high-resolution mass spectrometry was used to identify AbS AHLs. Antibiotic sensitivity was assessed in an AHL-deficient AbS mutant (AbS-M), and the expression of drug-resistance genes in the presence of meropenem in AbS, AbS-M and AbS-M treated with the AHL N-3-hydroxy-dodecanoyl-homoserine lactone (N-3-OH-C-12-HSL). AbS-M was more sensitive to meropenem and piperacillin than wild-type AbS, but resistance was restored by supplementation with N-3-OH-C-12-HSL. In addition, meropenem-treated AbS-M expressed lower levels of the drug-resistance genes oxacillinase 51, AmpC, AdeA and AdeB; treatment with N-3-OH-C-12-HSL also restored the expression of these genes. Overall, the results of the present study indicate that N-3-OH-C-12-HSL may be involved in regulating the expression of drug-resistance genes in A. baumannii. Therefore, this quorum-sensing signalling molecule may be an important target for treating multidrug-resistant A. baumannii infections.

Aim"Perioceutics" including antimicrobial therapy and host modulatory therapy has emerged as a vital adjunctive treatment of periodontal disease. Melatonin level was significantly reduced in patients with periodontal diseases suggesting melatonin could be applied as a potential "perioceutics" treatment of periodontal diseases. This study aims to investigate the effects of melatonin receptor agonists (melatonin and ramelteon) on Porphyromonas gingivalis virulence and Porphyromonas gingivalis-derived lipopolysaccharide (Pg-LPS)-induced inflammation.MethodsEffects of melatonin receptor agonists on Porphyromonas gingivalis planktonic cultures were determined by microplate dilution assays. Formation, reduction, and viability of Porphyromonas gingivalis biofilms were detected by crystal violet staining and MTT assays, respectively. Meanwhile, biofilms formation was also observed by confocal laser scanning microscopy (CLSM). The effects on gingipains and hemolytic activities of Porphyromonas gingivalis were evaluated using chromogenic peptides and sheep erythrocytes. The mRNA expression of virulence and iron/heme utilization was assessed using RT-PCR. In addition, cell viability of melatonin receptor agonists on human gingival fibroblasts (HGFs) was evaluated by MTT assays. After pretreatment of melatonin receptor agonists, HGFs were stimulated with Pg-LPS and then release of cytokines (IL-6 and lL-8) was measured by enzyme-linked immunosorbent assay (ELISA).ResultsMelatonin and ramelteon did exhibit antimicrobial effects against planktonic culture. Importantly, they inhibited biofilm formation, reduced the established biofilms, and decreased biofilm viability of Porphyromonas gingivalis. Furthermore, they at sub-minimum inhibitory concentration (sub-MIC) concentrations markedly inhibited the proteinase activities of gingipains and hemolysis in a dose-dependent manner. They at sub-MIC concentrations significantly inhibited the mRNA expression of virulence factors (kgp, rgpA, rgpB, hagA, and ragA), while increasing the mRNA expression of ferritin (ftn) or hemolysin (hem). They did not show obvious cytotoxicity toward HGFs. They inhibited Pg-LPS-induced IL-6 and IL-8 secretion, which was reversed by luzindole, the melatonin receptor antagonist.ConclusionMelatonin receptor agonists can inhibit planktonic and biofilm growth of Porphyromonas gingivalis by affecting the virulent properties, as well as Pg-LPS-induced inflammatory response. Our study provides new evidence that melatonin receptor agonists might be useful as novel "perioceutics" agents to prevent and treat Porphyromonas gingivalis-associated periodontal diseases.

Objective: To analyze the genetic diversity of Prevotella intermedia in primary endodontic infections?Methods:Seventeen microbial samples were taken from the root canals of teeth with primary endodontic infec?tions?DNA extracted from the samples was amplified by nested PCR to detect P?intermedia?It was conducted with P?intermedia ?specific primers based on 16 S rDNA V3?V6 region,and several DGGE bands were excised for further sequen?cing?Results:The prevalence of P?intermediain primary endodontic infections was 47?06%?There were 14 DGGE bands present at different position?It was founded 3 genetic according to sequencing and blasting?Conclusion:Genetic diversity of P?intermedia was presented by PCR?DGGE.

目的 分析原花青素对变形链球菌生物膜的体外抑制作用.方法 采用96孔微量板液体微量稀释法进行抑菌试验,观察对变形链球菌生长的抑制情况.盖玻片上形成变形链球菌生物膜模型,用共聚焦显微镜观察原花青素和对照试剂N-乙酰半胱氨酸对变形链球菌生物膜的影响.结果 原花青素对变形链球菌的抑制作用比较强.原花青素在终浓度为2～5 mg/mL,N-乙酰半胱氨酸在终浓度为7.5～75 mg/mL对变形链球菌生物膜均有明显的影响.N-乙酰半胱氨酸明显减少了生物膜的多糖,而原花青素明显减少了生物膜的蛋白.结论 原花青素和对照药物N-乙酰半胱氨酸具有不同的抗生物膜机制,原花青素对生物膜蛋白影响更多一些,而N-乙酰半胱氨酸对生物膜多糖的影响更多一些.

ObjectiveThe aim of this study was to examine the influence of various time intervals on the composition of the supragingival plaque microbiome, especially the dynamic core microbiome, and to find a suitable observation interval for further studies on oral microbiota.Methods and MaterialsEight qualified volunteers whose respective age ranges from 25 to 28 years participated in the present study. The supragingival plaque was collected from the buccogingival surface of the maxillary first molar at eight time slots with different intervals (day 0, 1 day, 3 days, 1 week, 2 weeks, 3 weeks, 1 month, and 3 months). Bioinformatic analyses was performed based on 16S rDNA pyrosequencing (454 sequencing platform) targeting at the hypervariable V4-V5 region, in order to assess the diversity and variation of the supragingival plaque microbiome.ResultsA total of 359,565 qualified reads for 64 samples were generated for subsequent analyses, which represents 8,452 operational taxonomic units identified at 3% dissimilarity. The dynamic core microbiome detected in the current study included five phyla, 12 genera and 13 species. At the genus level, the relative abundance of bacterial communities under the "1 day," "1 month," and "3 months" intervals was clustered into sub-category. At the species level, the number of overlapping species remained stable between the "1 month" and "3 months" intervals, whereas the number of dynamic core species became stable within only 1 week.ConclusionsThis study emphasized the impact of different time intervals (days, weeks and months) on the composition, commonality and diversity of the supragingival microbiome. The analyses found that for various types of studies, the time interval of a month is more suitable for observing the general composition of the supragingival microbiome, and that a week is better for observing the dynamic core microbiome.

Dental caries and periodontal diseases have a close relationship with microbes such as Streptococcus mutans, Porphyromonas gingivalis and Fusobacterium nucleatum. Graphene oxide (GO), as the derivative of graphene, plays an important role in many areas including biology and medicine. In particular, it has been known as a promising antimicrobial nanomaterial. In this study, we focused on the antimicrobial property of GO against dental pathogens. With the utilization of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) reduced test, colony forming units (CFU) counting, growth curve observation, live/dead fluorescent staining, and confocal laser scanning microscopy (CLSM), we found GO nanosheets were highly effective in inhibiting the growth of dental pathogens. Transmission electron microscopy (TEM) images revealed that the cell wall and membrane of bacteria lost their integrity and the intracellular contents leaked out after they were treated by GO. Therefore, GO nanosheets would be an effective antibacterial material against dental pathogens and the potential applications in dental care and therapy are promising.

PURPOSE:To establish an immortalized human dental pulp stem cell line used for basic and clinical research of oral science.METHODS:Human telomerase reverse transcriptase (hTERT) cDNA was transferred into human dental pulp stem cells (hDPSCs) by lentivirus. The resultant stable clones reproduced successively and the expression of hTERT was identified.RESULTS:The hTERT gene was transferred into human dental pulp stem cells successfully. The transformed cells expressed telomerase activity and divided vigorously. p35 had been obtained so far.CONCLUSIONS:The hDPSCs can be immortalized by transferring exogenous hTERT gene to constitute telomerase activity.

The aim of this study was to investigate the effect of silver-supported material (Novaron (N)) in acrylic resin (poly(methyl methacrylate) (PMMA)) composites, which reinforced with zirconium dioxide nanoparticles (nano-ZrO2) and aluminum borate whiskers (ABWs), on the mechanical behavior, antibacterial properties and cytotoxicity. Silanized ABWs (4 wt%) and nano-ZrO2 (2 wt%) were mixed with PMMA powder to obtain nano-ZrO2-ABW/PMMA matrices. Various amounts of Novaron particles were incorporated into the matrices and the pure PMMA to test the flexural strength. In addition, Streptococcus mutans (S. mutans) and Canidia albicans (C. albicans) biofilms on the specimen surface and in the culture medium were investigated for metabolic activity and colony-forming units (CFUs). Extracts taken in the cell culture medium of the specimens were used to evaluate cell viability. Results showed that the silanized nano-ZrO2 and ABWs could improve the flexural strength of composites compared with the pure PMMA. Novaron itself had no mechanical function for composites while it had synergistic effect when it mixed with silanized nano-ZrO2 and ABWs. And when 4 wt% (N-4) Novaron mixed in nano-ZrO2-ABW/PMMA composites, flexural strength achieved an increase of 44%, getting the maximum value. For the antibacterial properties, the values of MTT and CFUs of S. mutans and C. albicans biofilms on the composites surface were greatly reduced (p < 0.05) with the higher proportion of Novaron, and no significant difference was found in the culture medium. The composites did not have an adverse affect on fibroblast growth in this study (p > 0.05). (C) 2014 Elsevier Ltd. All rights reserved.

Objective: The aim of this article was to study the variation in oral microflora of the subgingival plaque during and after radiotherapy.Design: During and after radiotherapy, microbial samples were collected at seven time points (early stage, medium stage, and later stage of radiotherapy, and 1 month, 3 months, 6 months, and 1 year after radiotherapy) in three subjects for a total of 21 samples. Polymerase chain reaction (PCR) amplification was carried out on the 16S rDNA hypervariable V1-V3 region, and then the PCR products were determined by high-throughput pyrosequencing.Results: The rarefaction curve indicating the richness of the microflora demonstrated that the number of operational taxonomic units (OTUs) was in decline from the early stage of radiotherapy to the time point 1 month after radiotherapy and then trended upward. The Shannon diversity index declined during radiotherapy (ranging from 4.59 to 3.73), and generally rose after radiotherapy, with the lowest value of 3.5 (1 month after radiotherapy) and highest value of 4.75 (6 months after radiotherapy). A total of 120 genera were found; five genera (Actinomyces, Veillonella, Prevotella, Streptococcus, Campylobacter) were found in all subjects across all time points.Conclusion: The richness and diversity of oral ecology decreased with increased radiation dose, and it was gradually restored with time. (C) 2015 Elsevier Ltd. All rights reserved.

PURPOSE:To evaluate the efficacy of Beyond cold-light tooth bleaching on the formation of main cariogenic bacteria biofilm on enamel surfaces. METHODS:Twenty enamel discs with the size of 4 mm×4 mm×1 mm in size, were made. The enamel discs were divided into 4 groups randomly: cold-light bleaching group, bleaching gel group, cold-light group and control group. Five discs were in each group. Cold-light bleaching group was whitened 3 times with bleaching gel and cold-light, and 12 min per session. Bleaching gel was smeared on the surface of enamel in bleaching gel group for 3 times and 12 min per session. Enamel discs of cold-light group were treated with cold-light for 12 min and 3 sessions. Control group was treated without any processing. The 4 groups were incubated in mixed bacteria liquid, including Streptococcus mutans(SM), Actinomyces viscosus (Av) and Fusobacterium nucleatum (Fn), within the artificial oral cavity model. After 36 h, the samples were observed under confocal laser scanning microscopy(CLSM). The data was analyzed with SAS8.2 software package. RESULTS:The biofilms in 3 experimental groups were sparser than the control group under CLSM, and the thickness significantly decreased after treatment (P<0.05), while no significant difference was found among 3 experimental groups (P>0.05).Compared with the control group, the percentage of vital bacteria in biofilm of the experimental groups decreased significantly after treatment (P<0.001). CONCLUSIONS:Cold-light tooth bleaching can inhibit the formation of mixed bacteria biofilm, damage the structure of biofilm and reduce the number of vital bacteria. Supported by Research Fund of Ninth People's Hospital of Shanghai Jiao Tong University School of Medicine (2013-06).

Enterococcus faecalis is the species most frequently isolated from failed endodontic treatments because it can survive under stress conditions imposed by root canal treatment. The objective of this study was to determine the ability of E. faecalis to invade dentine tubules under alkaline and energy-starvation stress and to explore the potential mechanisms. Roots from single-rooted human teeth were infected with E. faecalis under alkaline and energy-starvation stress conditions. After 4 wk of culture, samples were processed to establish the tubule-penetration distance. In addition, the hydrophobicity of E. faecalis cells under these conditions was analysed and the expression of genes involved in adhesion was quantified by real-time quantitative PCR. Culture of E. faecalis under alkaline and energy-starvation stress conditions resulted in a marked reduction of tubule-penetration distance, a significant increase in hydrophobicity of the bacterial surface, and marked down-regulation of most adhesin genes compared with E. faecalis cultured in tryptic soy broth. The results indicate that the dentine tubule invasion ability of E. faecalis was markedly decreased under alkaline and glucose-starvation stress conditions, possibly because of the increase of hydrophobicity and down-regulation of some adhesion genes.

PURPOSE:To explore the stem cell surface markers expressed in human dental pulp stem cells which were selected and isolated by magnetic beads.METHODS:Human dental pulp cells (hDPCs) were separated and cultured from dental pulp of healthy third molars for orthodontic purpose. HDPSCs were isolated from cultured hDPCs by magnetic-activated cell sorting's (MACS) indirect magnetic cell labeling and positive selection strategy with antibody STRO-1 in the 2nd generation. Then the stem cell surface markers (CD73, CD90, CD105, CD166 and STRO-1) were respectively detected in 3, 4, 5, 6, 7 and 8 generation of dental pulp stem cells. HDPSCs were induced to differentiation by adipogenic medium and osteogenic medium in the 3rd generation. Adipogenic differentiation was assessed by oil red O staining in day 21, and osteogenic differentiation was assessed by alizarin red staining in day 21.RESULTS:HDPSCs could differentiate into adipocyte and osteoblasts. Oil red O staining and alizarin red staining were positively expressed after induction of HDPSCs. STRO-1's expression was decreased with the increase of generation. The expressions of CD73, CD90, CD105 and CD166 were relatively stable.CONCLUSIONS:The expression of STRO-1 is declined with the increase of generation, and the expressions of CD73, CD90, CD105 and CD166 are relatively stable with the changes of generation. Supported by National Natural Science Foundation of China (81070826/81371143) and Shanghai Rising-Star Program (12QH1401400).

PURPOSE:To complement the activated methyl cycle (AMC) pathway at an AI-2 defect background in Streptococcus mutans (S. mutans) luxS null strain.METHODS:A sahH gene was amplified from Pseudomonas aeruginosa and introduced into the S. mutans luxS null strain to complement the methyl-metabolic disruption at an AI-2 defect background. Western blot, reverse-transcription PCR and AI-2 bioassay were performed to confirm the heterogenous expression of SahH in S. mutans luxS null strain. The data was statistically analyzed by SAS8.0 software package.RESULTS:LuxS and SahH were detected to express in Escherichia coli BL21 as well as their mRNA were confirmed to be successfully transcribed in S. mutans luxS null strain. AI-2 production was found in wide type S. mutans and its luxS-introduced luxS null strain but not found in the luxS null strain and its sahH and empty plasmid-introduced strains.CONCLUSIONS:A new S. mutans derivative with the AMC pathway complements while the AI-2 defect is constructed.

Objective The objective of the present in vitro study was to investigate the antibacterial effect of three kinds of root canal sealers, AH-Plus, Vitapex and zinc oxide eugenol root canal sealer on mixed bacteria in root canals by a root canal filling model. Methods 48 freshly extracted human mandibular premolars for orthodontic treatment were collected for root canal preparation, and were divided into four groups randomly (n=12). Three experiment groups were filled using the three root canal sealers respectively, leaving a group of blank root canal as negative control, and following by thermal cycling treatment. The international reference strains including Streptococcus mutans, Actinomyces viscosus, Lactobacillus acidophilus, Enterococcus faecalis, Veillonella dispar, Porphyromonas gingivalis and Fusobacterium nucleatum were incubated. Aliquots of mixed bacterial suspension were sealed in the root canal. After incubation, the samples were cross cut to collect bacteria in the apical 1/3, middle 1/3 and coronal 1/3, and then to observe the bacterial colonies. Using the ANOVA (P=0.05) software to analyze the differences between the groups. Results Bacteria were found in all samples. P. gingivalis and F. nucleatum were detected in root canals of the control group, but not in the experiment groups. Lactobacillus and Actinomycetes were missing in the apical 1/3, while the detection rate of E. faecalis was higher in all parts of the canals. Comparing the numbers of residual bacteria in root canal, Vitapex was found to have more effective antimicrobial activity then AH-Plus in coronal 1/3, however, AH-Plus was superior to Vitapex in the apical 1/3.Conclusion Based on the inhibitory effects on bacteria in the apical 1/3 of root canals, AH-Plus has an advantage over Vitapex and conventional zinc oxide eugenol sealer.

The aim of this study was to develop new Prevotella intermedia-specific PCR primers based on the 16S rRNA. The new primer set, Pi-192 and Pi-468, increased the accuracy of PCR-based P. intermedia identification and could be useful in the detection of P. intermedia as well as epidemiological studies on periodontal disease.

OBJECTIVETo investigate the predominant contribution of methyl-metabolism pathway to the regulation of LuxS of Strecptococcus mutans.METHODSThe differences in biofilm formation and aciduricity of Strecptococcus mutans among the methyl-metabolism-complementation strain (KO-S), the parental wide-type strain (WT) and the luxS null strain (KO) were observed by real-time PCR for monitoring the transcriptional level of genes related to biofilm formation (smu.238, gtfD) and aciduricity (smu.44, smu.46) of the studied strains, methyl thiazolyl tetrazolium (MTT) for quantifying the biofilm of the exhibited strains and confocal laser scanning microscopy for estimating the structure of the biofilm.RESULTSThe transcriptional level of smu.44, smu.46, smu.238, gtfD in WT were 1.289 ± 0.051, 1.694 ± 0.140, 1.565 ± 0.107, 1.667 ± 0.196 respectively; in KO were 1.001 ± 0.045, 1.007 ± 0.151, 1.000 ± 0.021, 1.012 ± 0.196 respectively, downregulated compared with WT (P < 0.05); in KO-S were 4.662 ± 0.091, 5.019 ± 0.258, 3.462±0.029, 3.071 ± 0.136 respectively, upregulated compared both with KO and with WT (P < 0.05). The quantity of biofilms formed by the studied strains were WT (1.592 ± 0.213), KO (0.939 ± 0.029), KO- S (2.177 ± 0.226), KO- P (1.020 ± 0.093), respectively, representing a less quantity by KO and KO-P than WT (P < 0.05) and a more quantity by KO-S than other three stains (P < 0.05). According to the observation of biofilms texture by confocal laser scanning microscopy, the WT biofilm was condensed and even. In contrast, fissures and gaps were found scattered in biofilms of KO, KO-P while lessened in that of KO-S, in which high-density bacterial aggregates were observed. The acid assay indicated a smaller biofilm decrease by WT and KO-S than that by KO and KO- P(P < 0.05).CONCLUSIONSThe methyl- metabolism pathway contributes to LuxS regulation on biofilm formation and auiduricity of Strecptococcus mutans.

Objective To analyze the aciduricity of Streptococcus mutans,Streptococcus sanguis,Actinomyces and Lactobacillus in vitro.Methods The bacterial growth of alone or co-culture was observed through liquid culture on a 96-well microtiter plate under different pH conditions.Results The minimum aciduricity of Lactobacillus,S.mutans,Streptococcus sanguis and Actinomycetes were pH 2.0 to 3.0,pH 3.0 to 4.0,pH 4.0 to 5.0 and pH 5.0 to 6.0 repectively in the condition of alone culture.Streptococcus bacteria or Actinomycetes showed lower pH tolerance when co-cultured with Lactobacillus.The reversal changes of aciduriccity were observed in the mixed culture of Actinomycetes and Streptococcus sanguis from pH 7.0 to pH 2.0.Conclusion Different aciduricity of bacteria may be associated with the success of bacteria in plaque and caries process.Lactobacillus bacteriocin may play a role in the growth inhibition of the bacteria co-cultured with Lactobacillus under low pH conditions.

OBJECTIVE:To study the changes of growth and biofilm formation capability of Enterococcus faecalis (Ef) in different stress conditions.METHODS:The changes of growth of Ef in stress conditions were observed by measuring the A600 value with ultraviolet spectrophotometer. Ef was incubated on glass slide in stress conditions, biofilm formation capability of cells was investigated by colony-forming unit (CFU) counting of the culturable bacteria and fluorescence confocal laser scanning microscopy.RESULTS:Ef couldn't growth under the conditions of 2%, 5%NaClO, pH = 11 and 12, the A600 value was unchanged in 96 hours. But the growth curve changed at different levels in other stress conditions: under 1%NaClO, the A600 value peaked at 1.461 at 16 hour (the peaked level was 1.238 at 6 hours in control group) ; under 0,0.05%,0.15% glucose, it peaked at 0.645,0.890, 1.173, respectively, at 6 hour (it was maximized to 1.195 at 6 hours in control group); the A600 value peaked at 1.704 at 6 hours at pH = 9 and 1.225 at 10 hours at pH = 10 (the peak level was 1.732 at 6 hours at pH = 7) . Biofilm assay showed that Ef were able to form biofilm in these stress conditions except 5%NaClO and pH = 12.CONCLUSIONS:Ef could growth and form biofilms in energy starvation, low concentrations of sodium hypochlorite and weak alkaline stress.

Objective To observe the expressions of efflux pump gene cluster adeAB in acinetobacter baumannii isolated from the burn patients and the expression changes of its upstream regulatory genes adeR and adeS and determine the influence of those genes on drug resistance of acinetobacter baumannii.Methods Nine drug-resistant strains and nine sensitive strains of acinetobacter baumannii isolated from the burn patients treated between June 2012 and March 2013 were used.After strain identification using 16SrDNA sequencing,acinetobacter baumannii standard strain ATCC19606 was employed as the control.mRNA expressions of efllux pump genes adeA and adeB and their upstream regulatory genes adeR and adeS were detected by real-time quantitative PCR.Results (1) adeA and adeB genes presented higher expressions in drug-resistant strains (3.71 ±0.95,76.16 ± 8.75) than in sensitive strains (0.92 ± 0.94,0.72 ± 0.78) (F =38.71,663.65 respectively,both P ＜ 0.05).(2) adeS and adeR genes showed higher expressions in drug resistant strains (18.02 ± 6.71,3.02 ± 2.69) than in sensitive strains (1.64±1.51,0.76±0.61) (F =51.04,5.57 respectively,bothP＜0.05).Conclusions The over-expression of efflux pump gene cluster adeAB induced by the expression alteration of its upstream regulatory genes adeR and adeS is closely associated with the drug resistance of acinetobacter baumannii in the burn patient.Besides,the regulatory genes may depend on adeS to sense the nosocomial environment condition,activate or inactivate adeR and hence regulate efflux pump expression.