Saccharibacteria (TM7), but not other bacterial taxa, are associated with childhood caries regardless of age in a South China population.

Background: Human microbiome dysbiosis is related to various human diseases, and identifying robust and consistent biomarkers that apply in different populations is a key challenge. This challenge arises when identifying key microbial markers of childhood caries. Methods: We analyzed unstimulated saliva and supragingival plaque samples from children of different ages and sexes, performed 16S rRNA gene sequencing, and sought to identify whether consistent markers exist among subpopulations by using a multivariate linear regression model. Results: We found that Acinetobacter and Clostridiales bacterial taxa were associated with caries in plaque and saliva, respectively, while Firmicutes and Clostridia were found in plaque isolated from children of different ages in preschool and school. These identified bacterial markers largely differ between different populations, leaving only Saccharibacteria as a significant caries-associated phylum in children. Saccharibacteria is a newly identified phylum, and our taxonomic assignment database could not be used to identify its specific genus. Conclusion: Our data indicated that, in a South China population, oral microbial signatures for dental caries show age and sex differences, but Saccharibacteria might be a consistent signal and worth further investigation, considering the lack of research on this microbe.

Pulp revascularization on an adult mandibular right second premolar: A case report.

BACKGROUND: Pulp revascularization has become a new method for the treatment of periapical diseases in young permanent teeth in recent years. Through root canal flushing and disinfection, avoiding mechanical preparation, guiding apical stem cells into the root canal and promoting the continuous development of tooth roots, it has achieved good clinical curative effects. But in adult patients with chronic periapical periodontitis with immature roots and open apices, apical barrier technology is often used to treat these teeth. CASE SUMMARY: Pulp revascularization of a 26-year-old patient's tooth was performed using cefaclor instead of minocycline and iRoot BP instead of mineral trioxide aggregate as intracanal medication. The case was followed up for 36 mo. Observations showed evidence of regression of clinical signs and symptoms, resolution of apical periodontitis and no discolouration of affected teeth. CONCLUSION: For adult patients with chronic periapical periodontitis with immature roots and open apices, pulp revascularisation showed favourable results in treating these teeth.

Overview of noncoding RNAs involved in the osteogenic differentiation of periodontal ligament stem cells.

Periodontal diseases are infectious diseases that are characterized by progressive damage to dental support tissue. The major goal of periodontal therapy is to regenerate the periodontium destroyed by periodontal diseases. Human periodontal ligament (PDL) tissue possesses periodontal regenerative properties, and periodontal ligament stem cells (PDLSCs) with the capacity for osteogenic differentiation show strong potential in clinical application for periodontium repair and regeneration. Noncoding RNAs (ncRNAs), which include a substantial portion of poly-A tail mature RNAs, are considered "transcriptional noise." Recent studies show that ncRNAs play a major role in PDLSC differentiation; therefore, exploring how ncRNAs participate in the osteogenic differentiation of PDLSCs may help to elucidate the underlying mechanism of the osteogenic differentiation of PDLSCs and further shed light on the potential of stem cell transplantation for periodontium regeneration. In this review paper, we discuss the history of PDLSC research and highlight the regulatory mechanism of ncRNAs in the osteogenic differentiation of PDLSCs.

Plantamajoside attenuates inflammatory response in LPS-stimulated human gingival fibroblasts by inhibiting PI3K/AKT signaling pathway.

Periodontitis is an important inflammatory disease that often causes by periodontopathic bacteria. The present study, we tested the anti-inflammatory effects of plantamajoside on LPS-stimulated human gingival fibroblasts. Human gingival fibroblasts (HGFs) were stimulated with LPS from Porphyromonas gingivalis. Plantamajoside was administrated 1 h before LPS treatment. The results demonstrated that plantamajoside decreased the production of PGE2, NO, IL-6, and IL-8 in LPS-stimulated HGFs. LPS-induced NF-κB p65 and IκB phosphorylation were also suppressed by plantamajoside. Furthermore, plantamajoside inhibited LPS-induced PI3K and AKT phosphorylation. In conclusion, these results suggested that the mechanism of plantamajoside was through inhibiting PI3K/AKT signaling pathway, which lead to the inhibition of NF-κB activation and inflammatory response.

A 3-year clinical evaluation of endodontically treated posterior teeth restored with two different materials using the CEREC AC chair-side system.

STATEMENT OF PROBLEM: The introduction of polymer-infiltrated ceramic network (PICN) materials may provide more options for dentists in restoring short clinical crowns and extensively damaged posterior teeth, but clinical data for their performance are lacking. PURPOSE: The purpose of this clinical study was to compare the 3-year performance and survival rates of PICN material with those of conservative ceramic onlay restorations for endodontically treated posterior teeth using the CEREC AC chair-side system. MATERIAL AND METHODS: A total of 101 onlay restorations of endodontically treated posterior teeth using the CEREC AC chair-side system were provided in 93 participants. The 101 teeth were divided into 2 groups: Vita Enamic group and Vitablocs Mark II group. Using the modified US Public Health Service quality evaluation system, 2 calibrated evaluators examined the performance of the onlay restorations over 3 years. The Kaplan-Meier method was adopted to analyze the survival rate of restorations (α=.05). The log rank test was used to compare the survival rates of the 2 groups. The Fisher exact test was performed to detect differences in the success rates for extensively damaged teeth and short clinical crown restorations between the 2 groups. The Silness and Löe gingival index was also recorded. RESULTS: The restoration survival rates in the 2 groups were 97.0% (Vita Enamic) and 90.7% (Vitablocs Mark II) (P>.05). Five failures were recorded (4.95%). These failures were caused by restoration debonding (60%), ceramic fractures (20%), and tooth fractures (20%). There were no significant differences between the success rates of restoring extensively damaged teeth and short clinical crowns between the 2 groups (P>.05). The periodontal condition of 25% of participants was improved 3 years after the onlay restorations. CONCLUSIONS: Onlay restorations of endodontically treated posterior teeth with Vita Enamic using the CEREC AC chair-side system are clinically promising prosthodontic alternatives, with a survival rate of 97.0% after 3 years. More research is needed to verify the results of this study.

[Gelatin/alginate hydrogel scaffolds prepared by 3D bioprinting promotes cell adhesion and proliferation of human dental pulp cells in vitro].

OBJECTIVE: To evaluate the cytotoxicity of gelatin/alginate hydrogel scaffolds prepared by 3D bioprinting in human dental pulp cells (HDPCs) and compare the cell adhesion and proliferation of the cells seeded in the biomaterial using two different methods. METHODS: HDPCs isolated by tissue block culture and enzyme digestion were cultured and passaged. Gelatin/alginate hydrogel scaffolds were printed using a bioplotter, and the cytotoxicity of the aqueous extracts of the scaffold material was tested in the third passage of HDPCs using cell counting kit-8. Scanning electron microscopy and trypan blue were used to assess the adhesion and proliferation of the cells seeded in the scaffold material at a low or high concentration. RESULTS: The aqueous extract of the scaffolds at different concentrations showed no obvious cytotoxicity and promoted the proliferation of HDPCs. The scaffolds had a good biocompatibility and HDPCs seeded in the scaffold showed good cell growth. Cell seeding at a high concentration in the scaffold better promoted the adhesion of HDPCs and resulted in a greater cell number on the scaffold surface compared with low-concentration cell seeding after a 5-day culture (P<0.05). CONCLUSION: Gelatin

Periostin is essential for periodontal ligament remodeling during orthodontic treatment.

Orthodontic tooth movement is a process stimulated and maintained by external tensile stress; periodontal ligament remodeling serves an important role during this process. However, the function and underlying mechanism of periostin (PN) during orthodontic periodontal ligament remodeling remain unclear. The present study established in vitro and in vivo models of orthodontic treatment to investigate the expression levels of PN under conditions of external tensile stress load. These results indicated that tensile stress load increased the expression levels of PN in mouse peridontal ligaments and human periodontal ligament cells (hPDLCs), during orthodontic tooth movement. Furthermore, the present study demonstrated that the expression levels of PN were regulated by transforming grown factor ß, and that PN promotes type I collagen and α­smooth muscle actin expression levels in hPDLCs. Therefore, PN may be essential for periodontal ligament remodeling during orthodontic treatment, and therefore may represent a potential therapeutic target.

Minimally invasive root canal therapy / 口腔疾病防治

@#Minimally invasive root canal therapy should be defined as a discipline which adhere to a concept of preserving the healthy tooth structure as much as possible during all the root canal therapy procedure. In the past 15 years, the concept of minimally invasive has spread and developed fast throughout the diagnosis and treatment of endodontics, which made the root canal therapy (RCT) procedure safer, more accurate and efficient. Minimally invasive endodontics rely on the development of various kinds of therapeutic devices and materials, including the 3D image auxiliary equipment, operation microscope, the NiTi instrument systems and the disinfection and obturation material. Minimally invasive endodontics is a therapeutic concept of the modern root canal therapy which redefined the standard of what a successful RCT is.

The impact of different DNA extraction methods on the analysis of microbial diversity of oral saliva from healthy youths by polymerase chain reaction-denaturing gradient gel electrophoresis.

BACKGROUND/PURPOSE: Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), as a conventional molecular technique, was utilized to analyze the diversity of oral microbiota. However, studies found that the results of PCR-DGGE were affected by the DNA isolation method. This study compared QIAamp DNA Micro Kit extraction method with the phenol and chloroform extraction method for DNA isolation of saliva of healthy youths and analyzed PCR-DGGE fingerprints. MATERIALS AND METHODS: In the first stage, samples were divided into two after collection from eight health youths. Two methods were used to isolate the DNA for PCR-DGGE analysis. In the second stage, another 16 samples were collected from 14 youths. The better method, QIAamp DNA Micro Kit, was used to isolate the DNA for PCR-DGGE analysis. The cluster analysis was performed with unweighted pair-group method with arithmetic means. RESULTS: The results in the first stage showed that the QIAamp DNA Micro Kit extraction method was more suitable for DNA extraction of saliva than the phenol-chloroform extraction method. In the second stage, the bands were changed into numbers "0", "1", and "2" to analyze the similarity of samples according to the bands' lightness. The similarity indices of different periods from the same individual showed that the microbiological composition was very similar (>0.95), while those from different individuals varied greatly (<0.90). CONCLUSION: PCR-DGGE was more accurate in assessing oral microbial diversity by QIAamp DNA Micro Kit. Different individuals had large differences in oral microbial diversity but also had some common microbial dominant communities.

A novel small deletion mutation in RUNX2 gene in one Chinese family with cleidocranial dysplasia.

Cleidocranial dysplasia (CCD) is a skeletal dysplasia with autosomal-dominant inheritance. The runt related transcription factor 2 (RUNX2) gene is the only gene in which mutations are known to cause CCD. We report identification of a novel small deletions mutation in the RUNX2 gene in a Chinese family with CCD. A 29-year-old female was diagnosed as proband of CCD based on the clinical findings, which show delayed closure of the fontanels, hypoplastic or aplastic clavicles and dental anomalies. Similar dental and skeletal symptoms were also observed in the other three affected individuals. We prepared genomic DNA from all four affected individuals, unaffected individual from her family members, as well as 100 unrelated healthy controls. PCR was conducted using the above genomic DNA as template and the RUNX2 gene-specific primers. The PCR product was subjected to direct sequencing and the sequence was compared to that of RUNX2 gene within the NCBI database. We detected a small deletion CCTA from nucleotide 635 to nucleotide 638 in exon 3 of RUNX2 gene of the proband. This will lead to the introduction of a translational stop codon at codon 220, resulting in a truncated RUNX2 protein, and therefore within the runt domain of the RUNX2 protein. We detected the same mutation in the the other three affected individuals, and did not detect any mutation in the unaffected family members or the 100 unrelated healthy controls, demonstrating that this is a novel missense mutation in RUNX2 gene and therefore, contributes to the molecular diagnosis of CCD.

RCCS enhances EOE cell proliferation and their differentiation into ameloblasts.

In this article we report on the culturing of dental enamel organ epithelia (EOE) using a rotary cell culture system (RCCS) bioreactor associated with a cytodex-3 microcarrier. This culture system enhanced the proliferation and differentiation of the EOE into ameloblasts. Primary dental EOE trypsinized from 4-day old post-natal rat pups were cultured in the RCCS associated with Cytodex-3. The results were analyzed in comparison to a conventional plate system (control). Cells grown in RCCS have shown higher viabilities (above 90%) and final cell densities in terms of cells/ml than in the control system. In the case of RCCS, 46±2 manifold increases were obtained, while significantly lower yields of 10.8±2.5 manifod were obtained for control plates. Throughout the experiments, glucose levels were maintained within the accepted physiological range. In this case, LDH levels are kept low (below 150 mmol/ml), which is in accordance with the low cell death observed in the RCCS. Scanning electron microscopy revealed cells that were spread and forming three dimensional aggregates on the surface of cytodex-3. Cells cultured in the RCCS exhibited a stronger positive immunofluorescence staining for ameloblastin than those in control plates. RT-PCR results revealed that cells cultured in RCCS have higher amelogenin mRNA levels compared to controls. We have done an exploratory study on biological characteristics and self-assembling of epithelium cellula intersitialis, which demonstrated that the special 3D environment enhanced the rat dental EOE cell proliferation and differentiation into ameloblasts. The study has revealed that RCCS could be used to study the reaction of the EOE cells, tooth enamel organ cells and mesenchymal cells under the spacial 3D culture system, which will also provide a novel hypothesis for dental regeneration.

[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.

[Application of three-dimensional laser scanning-based maxillofacial soft tissue reconstruction in orthodontic treatment].

OBJECTIVE: To establish a convenient and rapid method for constructing a digital model of the maxillofacial soft tissue based on three-dimensional laser surface scanning to allow direct and accurate observation of the soft tissue changes in the course of orthodontic treatment. METHODS: The point cloud data of three-dimensional laser scanning of the maxillofacial region were acquired from a healthy woman with Angle Class I occlusion, who maintained a horizontal Frankfort plane during scanning with the scanner placed at a distance of 80 cm. The scanning was repeated twice after wearing the dental cast for an Angle Class I occlusion. The three-dimensional digital model of the maxillofacial soft tissue was constructed based on the point cloud using GeoMagic10.0 software. RESULTS: The high-resolution three-dimensional model of the maxillofacial soft tissue reconstructed allowed accurate observation of the distinct facial anatomical landmarks and represented directly the soft tissue changes in the process of orthodontic treatment by merging the models. Using the analytic tool provided by the software, this model also allowed direct quantitative measurement of the nasolabial angle and the distances from the esthetic plane to the upper lip, labral inferior, and mentolabial sulcus, which were 111.86°, -3.57 mm, -2.54 mm, and 3.95 mm before orthodontic treatment as compared to 114.31°, -2.73 mm, -1.06 mm, and 3.46 mm during treatment, and 116.53°, -0.15 mm, 0.64 mm, and 3.11 mm after the treatment, respectively. CONCLUSION: Three-dimensional laser surface scanning enables accurate and rapid construction of the digital model of the facial soft tissues, which may provide valuable assistance in orthodontic treatment.

Disturbed tooth germ development in the absence of MINT in the cultured mouse mandibular explants.

The Msx2-interacting nuclear target protein (MINT) is a nuclear matrix protein that regulates the development of many tissues. However, little is known regarding the role of MINT in tooth development. In this study, we prepared polyclonal antibodies against MINT, and found that that MINT was expressed in different cells at each stage of tooth germ development by immunohistochemistry. The role of MINT in tooth development was further illustrated by the misshapen and severely hypoplastic tooth organ in the cultured mandibular explants of MINT deficient mice. From the initiation to cap stage, the differences between mutants and wild-type molars were more and more distinguished histologically. In the MINT-deficient mandibular explants, the tooth germ was reduced in the overall size and lacked enamel knot, with abnormal dental lamina and collapsed stellate reticulum. Furthermore, the BrdU incorporation experiment showed that the proliferation activity was significantly reduced in MINT-deficient dental epithelium. Our results suggest that MINT plays an important role in tooth development, in particular, epithelial morphogenesis.

Inhibition of RANK/RANKL signal transduction pathway: a promising approach for osteoporosis treatment.

Osteoporosis is a bone disease causing impaired bone strength. It is characterized by increased osteoclast formation or enhanced bone resorption, leading to an increased risk of fragility fractures. Its prevalence increases with age. The advent of an aging population suggests that progressively more individuals will develop this disease in the aging population. A number of drugs for the prevention and treatment of osteoporosis act by inhibiting bone resorption. However, the effectiveness of osteoporosis treatment in clinical practice is limited. Since the osteoclast is the only cell in the body that is capable of resorbing bone, understanding its biology will be necessary for developing a new therapeutic approach for osteoporosis. Recently, it was discovered that the receptor activator of nuclear factor kappaB (RANK)/RANK ligand (RANKL)/osteoprotegerin (OPG) system is an important signal transduction pathway that regulates osteoclast formation. The binding of OPG to RANKL inhibits the binding between RANKL and RANK; this, in turn, prevents osteoclast precursors from differentiating and fusing to form mature osteoclasts. Therefore, the inhibition of the RANK/RANKL pathway inhibits osteoclast formation, differentiation, activation, and bone resorption. A potential clinical antiresorptive therapy can be developed by using an anti-RANKL monoclonal antibody, such as denosumab, that binds to RANKL with high affinity and specificity and blocks RANKL-RANK interactions.

[Detection of Helicobacter pylori in oral cavity of patients with recurrent aphthous ulcer].

OBJECTIVE: To investigate the association of recurrent aphthous ulcer (RAU) with Helicobacter pylori (Hp) infection and digestive diseases. METHODS: Saliva samples were collected from 82 patients with RAU and 74 healthy volunteers for Hp detection with PCR. RESULTS: The positivity rates of HP differed significantly between RAU patients and healthy volunteers (43.9% vs 16.2%, P<0.001). In the 82 RAU patients, 22 (26.82%) were identified to have gastritis and peptic ulcer, whereas only 7 out of the 74 healthy volunteers (10.45%) had such digestive diseases, showing significant difference between them (P<0.01). CONCLUSION: Hp might in some way associate with RAU, which in turn is associated with an increased incidence of digestive diseases.

[Generation of transgenic mouse of dentin sialoprotein and transgene expression analysis].

PURPOSE: To generate the transgenic mouse model of DSP and perform transgene expression analysis by RT-PCR. METHODS: Plasmid pcDNA3.1-CX was constructed by substituting promoter cbeta-actin for CMV promoter of pcDNA3.1, and the ultimate transgenic vector, pcDNA3.1-CX-dsp, was constructed by cloning DSP coding sequence into pcDNA3.1-CX. The pcDNA3.1-CX-dsp plasmid was linearized and microinjected into the male pronucleus of the zygotes. The tail DNA of pups was tested by PCR and Southern blot. A member of F1 generation of one positive mouse was used to perform transgene expression analysis by RT-PCR. RESULTS: 717 embryos were implanted to 29 recipient pseudopregnant mice, 4 of the 67 pups carrying the transgene. Expression of DSP was detected in a member of F1 generation of one positive mouse by RT-PCR. CONCLUSION: Founders of the DSP transgenic mouse were obtained successfully, and the expression of DSP was primarily confirmed.

[Comparison and analysis of human dentin matrix protein 1 promoter activity in three different cells].

OBJECTIVE: To observe and compare the luciferase activities of different length segments of human dentin matrix protein 1 promoter in human dental pulp stem cells (HDPSC), osteoblasts (OC) and Hela cells. METHODS: The differentlength desired DNA segments were obtained from 2 195 bp Dmp1 promoter cloned by PCR method. The amplified promoter segments with different length were cloned into luciferase report gene vector pGL3-Basic, the correct orientation of those inserts was verified by cutting with two different restrict enzymes. The luciferase activity was observed after different pGL3-PDmp1 vectors were transfected transiently into those three different-type cells. RESULTS: 6 Dmp1 promoter segments with different-length were obtained successfully, and luciferase report gene vectors with different promoter segments were successfully constructed after identified by restriction enzymes cutting. They had different luciferase activities when they were transfected transiently into HDPSC, and the region of -505(-)-193 bp and -935(-)-505 bp could be regarded as the specific promoters of Dmp1 promoter for HDPSC and OC respectively, which could include the basic regulatory elements. CONCLUSION: The correct clone of the upstream of human Dmp1 promoter segments with different length had been obtained, and they had strong luciferase activities in HDPSC and OC, but very low in Hela cell. These results will make an important basis for studying mineralized tissue-specific transcriptional regulation mechanisms of Dmp1.

[Reconstruction of dentin-pulp complex structure by tissue engineering technology].

OBJECTIVE: To investigate the possibility of reconstruction of dentin-pulp complex by tissue engineering technology. METHODS: Rat dental pulp stem cells were seeded into HA-TCP scaffold and incubated for 20 hours in vitro. Then the cell-scaffold complex was implanted subcutaneously into the dorsal side of nude mice. 8 weeks postimplantation, the samples were extracted for histological and immunohistochemical examinations. RESULTS: Three strata of tissue were observed in the hole of HA-TCP scaffold. They were dentin-like tissue, predentin-like tissue and pulp-like tissue respectively from the inner surface of the pore to the center. Dentin tubules were obvious in predentin-like and dentin-like tissue lining from the pulp-like tissue through predentin-like tissue and dentin-like tissue. Cells localized along the edge of pulp-like tissue were dense and polarized, resembling odontoblasts. Immunohistochemical study demonstrated DSP and DMP1 expression in these odontoblast-like cells and in the area of predentin-like tissue. CONCLUSIONS: Tissue-engineered rat dentin-pulp complex was reconstructed by seeding HA-TCP scaffold with rat dental pulp stem cells.

[Cloning and transcriptional regulation analysis of amelogenin promoter in different cells].

PURPOSE: To clone different promoter region of amelogenin gene and analyze their transcriptional activity. METHODS: The upstream regulation sequences of amelogenin gene were retrieved and analyzed. Amplified by PCR from the genomic DNA of mouse C57BL/6J and digested with restriction endonucleases enzyme, different transcriptional regulation sequences of 5'flanking of amelogenin gene including the basal promoter were cloned and ligated with luciferase gene in PGL3-Basic vector. These report vectors were transiently transfected into CHO, Hela and UMR-106 cells and luciferase assay was performed to analyse the transcription activation of these promoters. RESULTS: 6 promoters different in length were cloned. The activity of luciferase was very strong in Hela cells. On the contrary, the CHO and UMR-106 cells showed weak fluorescence. Luciferase activity fluctuated with the different promoter lengths in Hela cell as well as in CHO and UMR-106 cells. 975 bp and 532 bp of amelogenin 5'flanking DNA had a strong transcriptional activation, but 285 bp of amelogenin 5'flanking DNA had a weaker transcriptional activation. CONCLUSION: The sequence of amelogenin promoter can be activated in Hela cell. Hela cell can be used as a good model to study the transcriptional regulation of amelogenin promoter. According to the different activities of different lengths, it is suggested that there were some potential siliencer located between -975 and -532, and some potential activator in the region between -532 and -285.