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FimH is the adhesin of type I fimbriae expressed on Escherichia coli that can mediate specific adherence to host cells. High binding mutations in FimH are related to the adaptive evolution of bacteria. However, additional roles that these allelic variations may play remain elusive. To investigate novel biological functions of the mutations in FimH, we introduced four different variants of FimH by incorporating single amino acid substitutions at specific sites, namely A25P, G73R, A106, and T158P, respectively. In this study, adjuvant potential of FimH variants was evaluated by investigating their ability to trigger innate immune response to DC2.4 and adaptive immunity to improve immunological characteristics. The data revealed that purified A106 and T158P up-regulated the expression of co-stimulatory molecules critically involved in DC2.4 activation by interaction with TLR4, whereas A25P and G73R did not induce the phenotypic maturation of DC2.4. Besides, the culture of DC2.4 with A106 and T158P enhanced the release of cytokines and protein phagocytosis. When formulated with PAc, T158P elicited more robust PAc-specific IgG and IgA antibody responses compared to PBS, PAc and PAc+K12 groups and inhibited bacteria colonization. Collectively, the results confirmed that the T158P mutation located around the inter-domain interface of the protein induced a specific enhancement effect on adjuvant characteristics.

We previously demonstrated that recombinant protein PAc could be administered as an anticaries vaccine. However, the relatively weak immunogenicity of PAc limits its application. In the present study, we investigated the effect of two adjuvant combinations of chitosan plus Pam3CSK4 (chitosan-Pam3CSK4) and of chitosan plus monophosphoryl lipid A (chitosan-MPL) in the immune responses to the PAc protein in vivo and in vitro PAc-chitosan-Pam3CSK4 or PAc-chitosan-MPL promoted significantly higher PAc-specific antibody titers in serum and saliva, inhibited Streptococcus mutans colonization onto the tooth surfaces, and endowed better protection effect with significantly less caries activities than PAc alone. Chitosan-Pam3CSK4 and chitosan-MPL showed no statistically significant differences. In conclusion, our study demonstrated that the chitosan-Pam3CSK4 and chitosan-MPL combinations are promising for anticaries vaccine development.

Objective To investigate whether intragastric administration of 1,25-dihydroxyvitamin D 3 (1,25(OH) 2 D 3 ) could inhibit the bone resorption and inflammation in a mouse calvarial model infected by Porphyromonas gingivalis ( P. gingivalis ). Design Live P. gingivalis ATCC 33277 was injected once daily for 6 days into the subcutaneous tissue overlying the calvaria in mice. At the same time, 1,25(OH) 2 D 3 (50 μg/kg per day) was administered by gavage for 9 days, starting 3 d before the infection. Mice were killed under ether anesthesia 8 h after the last injection of P. gingivalis . Micro-computed tomography scanning was used to evaluate calvarial bone loss. Tartrate-resistant acid phosphatase was used to detect osteoclast activity. Real-time PCR was used to assess the mRNA expressions of OPG, RANKL, c-Fos, NFATc1, CTSK and TRAP in calvarial bone and IL-6, IL-10, IL-1β, IL-12p40 and TNF-α in soft tissue. The levels of serum IL-6, IL-10 were determined by ELISA. Results 1,25(OH) 2 D 3 treatment apparently attenuated bone resorption in P. gingivalis -induced mouse calvarial model and markedly reduced the number of osteoclasts. The expression levels of RANKL and osteoclast-related genes such as c-Fos, NFATc1, CTSK and TRAP were also decreased by 1,25(OH) 2 D 3 . Besides, 1,25(OH) 2 D 3 inhibited the expression of pro-inflammatory cytokines IL-6, IL-12p40 and TNF-α and enormously elevated the expression of anti-inflammatory cytokine IL-10. Conclusion 1,25(OH) 2 D 3 may decrease bone resorption in vivo via suppressing the expression of osteoclast-related genes and its anti-inflammatory properties.

The aim of this article was to present a right maxillary second molar with an unusual root canal morphology of 4 roots and 5 canals as confirmed by cone-beam computed tomographic (CBCT) imaging. The tooth had a C-shaped mesiobuccal root (CBCT imaging revealed that the root was closer to the palate than the buccal side) with 2 canals, 2 fused distobuccal roots with 2 separate canals, and 1 normal bulky palatal root with 1 canal. After thoroughly examining the rare anatomy, root canal treatment was applied on the tooth. This article shows the complexity of maxillary second molar variation and shows the significance of CBCT imaging in the confirmation of the 3-dimensional anatomy of teeth and endodontic treatment.

Objective To investigate whether the recombinant FimH-S.T protein could modulate immune response to anti-caries vaccine in vitro and in vivo. Design Recombinant FimH protein derived from Salmonella was constructed and purified. The expression of dendritic cell maturation markers and cytokines release were performed by flow cytometry, Real-time PCR and ELISA. In addition, BALB/c mice were administered with anti-caries PAc vaccine plus FimH-S.T, antibody responses were evaluated by ELISA. Splenocytes of immunized mice were detected for their proliferative ability in response to in vitro retreatment with PAc antigen by flow cytometry. Caries protection against dental caries formation was also investigated. Results The purified FimH-S.T induced phenotypic maturation of DC2.4 by up-regulating the expression of costimulatory molecules and MHC II, provoked the production and secretion of cytokines via TLR4-dependent signaling pathway in vitro. Furthermore, the mice immunized with the mixture of FimH-S.T and PAc significantly enhanced the PAc-specific antibodies in the serum along with saliva and promoted splenocyte proliferation. Our results also confirmed that PAc+FimH-S.T decreased the caries lesions formation which provided high protective efficacy against dental caries. Conclusion Our study demonstrates that recombinant FimH-S.T could enhance specific IgA responses and protection of anti-caries vaccine, possessing mucosal adjuvant ability by activating DC2.4 via TLR4 signaling pathway.

Aims To investigate the effects of the ginsenoside Rh2 on monospecies and multispecies cariogenic biofilms and explore the mechanism of the antibiofilm effect of Rh2 in vitro. Methods and results Streptococcus mutans, Streptococcus sobrinus and Streptococcus sanguinis were chosen to form the monospecies or multispecies biofilms. Crystal violet staining and laser scanning confocal microscopy were used to observe the effect of Rh2 on biofilms in vitro. Cytotoxicity was examined by the Cell Counting Kit-8. The effects of Rh2 on bacterial membranes were observed via transmission electron microscopy (TEM). The isobaric tags for relative and absolute quantification (iTRAQ) method were used to profile the common differentially expressed proteins. Gene expression was analysed by reverse transcription quantitative polymerase chain reaction. In general, the treatment of cariogenic biofilms with Rh2 significantly decreased biomass accumulation by inhibiting bacterial growth and extracellular polysaccharide synthesis without any cytotoxic effects. TEM imaging showed that Rh2 could disrupt the cell membranes of these bacteria. The iTRAQ results indicated that the levels of mannose-specific IIC/D and acetaldehyde/alcohol dehydrogenase were substantially down-regulated, while the mRNA expression of the corresponding genes were significantly changed. Conclusions Our data revealed a potential application for Rh2 in the protection against dental caries via the inhibition of cariogenic biofilms. Significance and impact of the study This study describes the first application of a ginsenoside against multispecies cariogenic biofilms. Rh2 may serve as an alternative agent to prevent dental caries by effectively modulating the pathogenic potentials of oral biofilms.

Vitamin D is known to be closely associated with periodontitis; however, its exact mechanisms remain to be clarified. The present study aimed to investigate the influence of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) on Porphyromonas gingivalis (Pg)-stimulated cytokine production and the involved signaling pathways in macrophages. The main observation was that 1,25(OH)2D3 inhibited Pg-induced interleukin (IL)-6 cytokine expression but up-regulated the expression of anti-inflammatory cytokine IL-10. Further analyses showed that 1,25(OH)2D3 decreased p38 mitogen-activated protein kinase (MAPK) and extracellular signal regulated kinase (ERK)1/2 phosphorylation. Inhibited phosphorylation of p38 MAPK and ERK1/2 was associated with decreased level of IL-6 expression, but was not related to increased level of IL-10 expression in macrophages stimulated with Pg. These results suggest that 1,25(OH)2D3 might exert its anti-inflammatory effects on Pg-stimulated macrophages partly through its inhibitory effect on the p38 MAPK and ERK1/2 signaling pathway.

Toll-like receptors (TLR) and the receptor for interleukin-1 (IL-1R) signaling play an important role in bacteria-mediated bone loss diseases including periodontitis, rheumatoid arthritis, and osteomyelitis. Recent studies have shown that TLR ligands inhibit the receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation from un-committed osteoclast precursors, whereas IL-1 potentiates RANKL-induced osteoclast formation. However, IL-1R and TLR belong to the same IL-1R/TLR superfamily, and activate similar intracellular signaling pathways. Here, we investigate the molecular mechanisms underlying the distinct effects of IL-1 and Porphyromonas gingivalis lipopolysaccharide (LPS-PG) on RANKL-induced osteoclast formation. Our results show that LPS-PG and IL-1 differentially regulate RANKL-induced activation of osteoclast genes encoding Car2, Ctsk, MMP9, and TRAP, as well as expression of NFATc1, a master transcription factor of osteoclastogenesis. Regulation of osteoclast genes and NFATc1 by LPS-PG and IL-1 is dependent on MyD88, an important signaling adaptor for both TLR and IL-1R family members. Furthermore, LPS-PG and IL-1 differentially regulate RANKL-costimulatory receptor OSCAR (osteoclast-associated receptor) expression and Ca(2+) oscillations induced by RANKL. Moreover, LPS-PG completely abrogates RANKL-induced gene expression of B lymphocyte-induced maturation protein-1 (Blimp1), a global transcriptional repressor of anti-osteoclastogenic genes encoding Bcl6, IRF8, and MafB. However, IL-1 enhances RANKL-induced blimp1 gene expression but suppresses the gene expression of bcl6, irf8, and mafb. Our study reveals the involvement of multiple signaling molecules in the differential regulation of RANKL-induced osteoclastogenesis by TLR2 and IL-1 signaling. Understanding the signaling cross-talk among TLR, IL-1R, and RANK is critical for identifying therapeutic strategies to control bacteria-mediated bone loss.

The properties of continuous wave (CW) and Q-switching on the diode-pumped Nd,Y:CaF2 disordered crystal were demonstrated around 1 μm in this paper. Using the different cavity lengths, we measured the output properties of CW of Nd,Y:CaF2 disordered crystal. Then based on Cr4+:YAG as the saturable absorber, the Q-switched pulse characteristics of this novel disordered crystal were researched for the first time. The maximum average output power of 120 mW and the shortest pulse width of 132 ns were obtained at the pulse repetition rate of 6.2 kHz.

Oral squamous cell carcinoma (OSCC) is a common human malignant tumor with high mortality. So far, the molecular pathogenesis of OSCC remains largely unclear. Heterogeneous nuclear ribonucleoprotein (hnRNP) A1 is an important multi-function splicing factor and closely related to tumorigenesis. hnRNP A1 is overexpressed in various tumors, and promotes aerobic glycolysis and elongation of telomere, but the function of hnRNP A1 in cell cycle and proliferation remains unclear. We found that hnRNP A1 was overexpressed in OSCC tissues, and was required for the growth of OSCC cells. Moreover, hnRNP A1 was highly expressed in the G2/M cell cycle phase. Knockdown of hnRNP A1 induced G2/M arrest. DNA microarray assay result showed that hnRNP A1 regulated the expression of a number of target genes associated with G2/M phase. Moreover, hnRNP A1 controlled the alternative splicing of CDK2 exon 5. These findings suggested that hnRNP A1 plays key roles in the regulation of cell cycle progression and pathogenesis of OSCC.

The purpose of this study was to determine the relationships among early childhood caries (ECC), root caries (RC), the quantity of Streptococcus mutans in saliva, and the concentrations of total and specific secretory IgA (sIgA). Saliva samples were collected from 70 children, 3-4 yr of age, with and without ECC, and from 43 adults, ≥60 yr of age, with and without RC. The decayed, missing, and filled teeth (dmft) and decayed, missing, and filled surfaces (dmfs) scores of each child, and the root decayed and filled teeth (RDFT) and root decayed and filled surfaces (RDFS) scores of each elderly subject, were determined. The S. mutans levels, total sIgA, and specific sIgA against two virulence antigens of S. mutans in saliva were analysed using quantitative real-time PCR (qPCR) and ELISAs. The quantity of S. mutans was significantly higher in caries-positive subjects within the two populations than in the caries-free subjects; and a positive correlation was found between the quantity of S. mutans and the dmft, dmfs, RDFT, and RDFS scores. In addition, the salivary total sIgA was significantly higher in children with severe early childhood caries (SECC) and in the elderly subjects with RC. Moreover, although the S. mutans level was significantly higher, the concentrations of specific sIgA against S. mutans antigens were significantly lower in samples from elderly subjects than in samples from children. These results support the concept that S. mutans is positively associated with ECC and RC. Furthermore, the levels of S. mutans-specific antibodies in saliva are too low to prevent infection with cariogenic bacteria and to inhibit development of ECC and RC.

Introduction Dental papilla cells (DPCs) are precursors of odontoblasts and have the potential to differentiate into odontoblasts. Osteoblasts and odontoblasts have many common characteristics. Osterix (Osx) is essential for osteoblast differentiation. However, no information is available for the effects of Osx on the odontoblastic differentiation of DPCs. The purpose of this study was to investigate the effects of Osx on the proliferation and odontoblastic differentiation of DPCs. Methods An immortalized human dental papilla cell (hDPC) line was used. Osx was stably overexpressed or knocked down in hDPCs with infection of lentiviral particles to determine its biological effects on hDPCs. The proliferation of cells was measured by the 5-ethynyl-2′-deoxyuridine incorporation assay and direct cell counting. Expressions of dentin sialophosphoprotein, nestin, dentin matrix protein 1, and alkaline phosphatase were detected by real-time polymerase chain reaction to determine the odontoblastic differentiation of cells. The mineralization ability of cells was evaluated by von Kossa staining and alkaline phosphatase activity assay. Results Overexpression of Osx retarded the proliferation of hDPCs, whereas knockdown of Osx increased the cell proliferation. Overexpression of Osx promoted the odontoblastic differentiation of hDPCs by up-regulating odontoblastic differentiation genes and increased the mineralization ability of hDPCs. Knockdown of Osx down-regulated odontoblastic differentiation genes and decreased the mineralization ability of hDPCs. Conclusions Osx might function as a potential regulator for the proliferation and odontoblastic differentiation of hDPCs.

High risk forms of the human papilloma virus (HPV) are generally accepted as necessary causative agents for cervical cancer. Recently, a possible relation between HPV and oral squamous cell carcinoma (OSCC) has also been noticed. The present study was conducted to investigate the prevalence of HPV infection in OSCCs in Wuhan city. DNA samples were collected from fresh tissues in 200 patients with OSCC and 68 normal controls. The polymerase chain reaction and direct sequencing were used to identify the HPV types in the samples. The prevalence of HPV of all types in the OSCC group was higher than in the control group (55/200 vs 2/68, OR=11.5, 95% CI=2.6-50.2). HPV16 and HPV18 were the main types detected, with HPV6 was the only low-risk type identified. High-risk HPV types HPV16 and HPV18 are prevalent in OSCC patients and may participate in the development of OSCC with traditional risk factors, tobacco and alcohol, possibly exerting synergistic effects. The results of multinomial logistic regression showed that those who smoked, consumed alcohol and with HPV infection have the highest risk of developing oral cancer (OR=13.3, 95% CI=3.1-56.8). Adjusted for age, smoking and alcohol use, HPV infection was independently associated with oral squamous cell carcinoma.

Boron nitride can be used as a good catalyst carrier because of its high thermal conductivity and chemical stability. However, a high specific surface area of boron nitride is still desirable. In this work, a carbon fiber composite coated with boron nitride villous nano-film was prepared, and was also characterized by means of scanning electron microscopy, high resolution transmission electron microscopy, Fourier transform infrared spectroscopy, and Brunauer–Emmett–Teller analysis. The results indicated that the carbon fibers were covered by uniform villous boron nitride films whose thickness was about 150 – 200 nm. The specific surface area of the boron nitride/carbon fiber composite material was 96 m2 g−1, which was markedly improved compared with conventional boron nitride materials

Dlx3 is essential for osteoblast differentiation and bone formation, and its expression is regulated by bone morphogenetic protein-2 (BMP-2). However, the intimate mechanism of BMP-2 regulation of Dlx3 transcription in osteoblasts is still unknown. Considering the important roles of Smad5 and p38 in osteoblast differentiation, we hypothesized that Smad5 and p38 mediated BMP-2-induced Dlx3 transcription in osteoblasts. We found activation of Smad5 and p38 increased the expression of Dlx3, whereas knocking down Smad5 or inactivation of p38 inhibited BMP-2-induced Dlx3 expression. Both Smad5 and p38 were able to activate Dlx3 promoter activity and p38/Smad5 response elements were located from −698 to −368 in Dlx3 promoter. Two Smad5 binding sites (SBEI and SBEII, TGTCT box) were identified in this region by EMSA and ChIP assay. Deletions and mutagenesis study of the Dlx3 promoter region indicated that the TGTCT boxes are crucial for p38/Smad5-induced Dlx3 promoter activity. At last, we found a cross-talk between p38 and Smad5, and that activation of p38 is necessary for BMP-2-induced Smad5 phosphorylation and nuclear translocation. Overall, we provide a novel insight that BMP-2-induced Dlx3 expression is regulated by p38/Smad5 signaling pathway in osteoblasts. J. Cell. Physiol. 229: 943–954, 2014. © 2013 Wiley Periodicals, Inc.

Contains fulltext : 137673.pdf (Publisher’s version ) (Open Access) To compare the levels of agreement and the survival rates of sealant retention for different sealing materials over a 2-year period assessed using the visual clinical examination and replica methods, sealant retention data were obtained by visual clinical examination and from replicas of the same sealed tooth at baseline and at 0.5-, 1- and 2-year evaluation points in 407 children and were compared for agreement using kappa coefficients. Survival curves of retained sealants on occlusal surfaces were created using modified categorisation (fully retained sealants and those having all pits and fissures partly covered with the sealant material versus completely lost sealants that included pit and fissure systems that had >/=1 pit re-exposed) according to the Kaplan-Meier method. The kappa coefficient for the agreement between both assessment methods over the three evaluation time points combined was 0.38 (95% confidence interval (CI): 0.35-0.41). More sealant retention was observed from replicas than through visual clinical examination. Cumulative survival curves at the three evaluation times were not statistically significantly higher when assessed from replicas (P=0.47). Using the replica method, more retained sealant material was observed than through visual clinical examination during the 2-year period. This finding did not result in a difference in the survival rates of sealants assessed by the two assessment methods. When replicas cast in die stone are used for assessing sealant retention, the level of reliability of the data is higher than that of data obtained through the commonly used visual clinical examination, particularly if such assessments are conducted over time.

This study examined the adhesive strength of two self-adhesive methacrylate resin-based sealers (MetaSEAL and RealSeal SE) to root dentin and compared them with RealSeal and AH Plus in properties. A total of 48 extracted human single-rooted teeth were used to prepare the 0.9-mm thick longitudinal tooth slice (each per tooth). Standardized simulated canal spaces of uniform dimensions were prepared in the middle of radicular dentin. After treated with 5.25% sodium hypochlorite (NaOCl) and 17% EDTA, tooth slices were allocated randomly to four groups (n=12) in terms of different sealers used: MetaSEAL, RealSeal SE, RealSeal, and AH plus groups. The simulated canal spaces were obturated with different sealers in each group. There were 10 slabs with 20 simulated canal spaces (n=20) used in each group for push-out testing. The failure modes and the ultrastructures of fractured sealer-dentin interfaces were examined. The remaining 2 slabs in each group underwent partial demineralization for observation of the ultrastructure of resin tags. The results showed that the push-out bond strength was 12.01±4.66 MPa in MetaSEAL group, significantly higher than that in the other three groups (P0.05). Moreover, no statistically significant differences were noted in the push-out bond strength between RealSeal SE (5.43±3.68 MPa) and AH Plus (7.34±2.83 MPa) groups and between RealSeal SE and RealSeal (2.93±1.76 MPa) groups (P0.05). Mixed failures were predominant in the fractured sealer-dentin interfaces in MetaSEAL and AH Plus groups, while adhesive failures were frequently seen in RealSeal SE and RealSeal groups. In conclusion, after complete removal of the smear layer, MetaSEAL showed superior bond ability to root dentin. The RealSeal SE is applicable in clinical practice, with its adhesive strength similar to that of AH Plus. The self-adhesive methacrylate resin-based sealer holds promise for use in endodontic treatment.

The aim of this study was to test the hypothesis that the conventional restorative treatment (CRT) and the atraumatic restorative treatment (ART) protocols, in comparison with the ultra-conservative treatment (UCT) protocol, would increase the quality of life of children over a period of 1 year. Cavitated primary molars of 302 children 6–7 years of age were treated according to the CRT, ART and UCT protocols at the school compound. Children’s parents completed the Brazilian version of the Early Childhood Oral Health Impact Scale (B-ECOHIS) at baseline and one year later. Paired t-test, Chi-square test and anova were applied in analysing the data. Questionnaires from 277 and 160 children were collected at baseline and after 1 year, respectively. A statistically significant difference in B-ECOHIS scores over the 1-year period was found for domains ‘child symptoms’ (P = 0.03) and ‘child psychology’ (P = 0.02). Treatment protocols did not statistically significantly influence the changes in B-ECOHIS scores over the 1-year period (P = 0.78). It can be concluded that the UCT protocol was as good as the two restorative protocols. All treatment protocols were effective in reducing children’s experience of pain, their sleeping problems and their irritability and/or frustration levels over the 1-year period.

This study aimed to investigate the association between p53 Arg72Pro polymorphism and the risk of human papillomavirus (HPV)-related head and neck squamous cell carcinoma (HNSCC) by conducting meta-analysis. The PubMed database was searched for relevant studies until May 30, 2013. Relevant studies were selected and data were extracted by two independent authors. Overall, subgroup, and sensitivity analyses were then conducted using the Comprehensive Meta- Analysis v2.2 software. Wild-genotype ArgArg was considered as reference [odds ratio (OR) = 1.00]. Nine studies involving 1071 HNSCC cases were obtained. Meta-analysis results indicated no association between p53 Arg72Pro polymorphism and the risk of HPV-related HNSCC: for Pro/Pro vs. Arg/Arg, OR = 1.17, 95% confidence interval (CI) = 0.70-1.98; for Arg/Pro vs. Arg/ Arg, OR = 1.25, 95% CI = 0.97-1.72; and for (Pro/Pro + Arg/Pro) vs. Arg/Arg, OR = 1.28, 95% CI = 0.95-1.70. These meta-analysis results were supported by subgroup and sensitivity analysis results. In conclusions, p53 Arg72Pro polymorphism is a potential marker of HP infection-related HNSCC rather than a susceptibility gene polymorphism.

Anticaries protein vaccines that induce a mucosal immune response are not effective. Therefore, development of effective and convenient anticaries vaccines is a priority of dental research. Here we generated self-assembling nanoparticles by linking the glucan-binding region of Streptococcus mutans glucosyltransferase (GLU) to the N-terminal domain of ferritin to determine whether these novel nanoparticles enhanced the immunogenicity of an anticaries protein vaccine against GLU in rodents. We constructed the expression plasmid pET28a-GLU-FTH and purified the proteins from bacteria using size-exclusion chromatography. BALB/c mice were used to evaluate the ability of GLU-ferritin (GLU-FTH) nanoparticles to induce GLU-specific mucosal and systemic responses. The protective efficiency of GLU-FTH nanoparticles was compared with that of GLU alone or a mixture of GLU and poly(I:C) after administering an intranasal infusion to Wistar rats. The phagocytosis and maturation of dendritic cells (DCs) exposed in vitro to the nanoparticles were assessed using flow cytometry. The GLU-FTH nanoparticle vaccine elicited significantly higher levels of GLU-specific antibodies compared with GLU or a mixture of GLU and poly(I:C). Immunization with GLU-FTH achieved lower caries scores compared with those of the other vaccines. Administration of GLU-FTH nanoparticles enhanced phagocytosis by DCs and their maturation. Thus, self-assembling GLU-FTH is a highly effective anticaries mucosal vaccine that enhanced antibody production and inhibited S. mutans infection in rodents.

Oral squamous cell carcinoma (OSCC) is the leading cause of death related to oral diseases. The mechanisms of OSCC development remain largely unknown. Heterogeneous nuclear ribonucleoprotein L (HnRNP L) is a multi-functional splicing factor. It has been reported to be an important regulator of apoptosis. However, the functions of hnRNP L in cancer need to be further explored. In the present study, we found that OSCC tissues expressed significantly higher levels of hnRNP L than normal tissues. Depletion of hnRNP L retarded cell growth, cell migration, and tumorigenesis of OSCC cells. HnRNP L regulates both the expression of oncogenic splicing factor SRSF3 and the alternative splicing of SRSF3 exon 4. Expression of hnRNP L is correlated with SRSF3 expression in OSCC tissues. These findings suggest that hnRNP L is important for the pathogenesis of OSCC and may be a novel potential therapeutic target of OSCC.

Osteogenic differentiation and bone formation are tightly regulated by several factors, including microRNAs (miRNAs). However, miRNA expression patterns and function during mechanical loading-induced osteogenic differentiation of human periodontal ligament cells (PDLCs) remain unclear. Here, we investigated the differential expression of miRNA-195-5p in the periodontal tissues of mice under orthodontic mechanical loading and in primary human PDLCs exposed to a simulated tension strain. The miR-195-5p was observed to be down-regulated and negatively correlated with osteogenic differentiation. Overexpression of miR-195-5p significantly inhibited PDLC differentiation under cyclic tension strain (CTS), whereas the functional inhibition of miR-195-5p yielded an opposite effect. Further experiments confirmed that WNT family member 3A (WNT3A), fibroblast growth factor 2 (FGF2), and bone morphogenetic protein receptor-1A (BMPR1A), proteins important for osteogenic activity and stability, were direct targets of miR-195-5p. Mechanical loading increased the WNT3A, FGF2, and BMPR1A protein levels, while miR-195-5p inhibited WNT3A, FGF2, and BMPR1A protein expression. WNT, FGF, and BMP signaling were involved in osteogenic differentiation of PDLCs under CTS. Further study confirmed that reintroduction of WNT3A and BMPR1A can rescue the inhibition of miR-195-5p on osteogenic differentiation of PDLCs. Our findings are the first to demonstrate that miR-195-5p is a mechanosensitive gene that plays an important role in mechanical loading-induced osteogenic differentiation and bone formation.

To investigate the effects of anti-caries DNA vaccine-induced salivary secretory immunoglobulin A (S-IgA) antibodies on Streptococcus mutans (S. mutans) adherence and biofilms formation in vitro. Adult female Wistar rats were intranasally immunized with the anti-caries DNA vaccine pGJA-P/VAX. Their saliva samples were collected at different times after the immunization, and S-IgA antibody level in the saliva and its inhibition on S. mutans adherence were examined. The effects of S-IgA in the saliva with the strongest inhibitory effects were examined at 3 different stages, ie acquired pellicles, biofilm formation and production of mature biofilms. The number of viable bacteria and depth of the biofilm at 16 h in each stage were determined using counting colony forming units and using a confocal laser scanning microscopy (CLSM). The participation of S-IgA in acquired pellicles and its aggregation with S. mutans were also observed under CLSM. The S-IgA titer in saliva reached its peak and exhibited the strongest inhibition on S. mutans adhesion at 10 weeks after the immunization. The colonies and depth of the biofilm in the saliva-pretreated group were 41.79% and 41.02%, respectively, less than the control group. The colonies and depth of the biofilm in the co-culture group were 27.4% and 22.81% less than the control group. The assembly of S. mutans and S-IgA was observed under CLSM after co-cultivation. In the mature-stage biofilm, no differences were observed between the different groups. These results demonstrate that the anti-caries DNA vaccine induces the production of specific S-IgA antibodies that may prevent dental caries by inhibiting the initial adherence of S. mutans onto tooth surfaces, thereby reducing the accumulation of S. mutans on the acquired pellicles.

We previously demonstrated that an anti-caries DNA vaccine intranasally administered with recombinant flagellin protein as a mucosal adjuvant enhanced salivary IgA response and conferred better protection against caries. However, the relatively weak immunogenicity of DNA vaccines and the necessity for a large quantity of antigens remain significant challenges. Here, we fused the flagellin derived from E. coli (KF) and target antigen PAc containing the A-P fragment of PAc from S. mutans (rPAc) to produce a single recombinant protein (KF-rPAc). The abilities of KF-rPAc to induce rPAc-specific mucosal and systemic responses and protective efficiency against caries following intranasal immunization were compared with those of rPAc alone or a mixture of rPAc and KF (KF + rPAc) in rats. Results showed that KF-rPAc promoted significantly higher rPAc-specific antibodies in serum as well as in saliva than did an equivalent dose of rPAc alone or a mixture of KF + rPAc. Intranasal immunization of 8.5 µg KF-rPAc could achieve 64.2% reduction of dental caries in rats. In conclusion, our study demonstrated that flagellin and PAc fusion strategy is promising for anti-caries vaccine development, and KF-rPAc could be used as an anti-caries mucosal vaccine.

Background: The aim of this study was to validate the micro-CT and related software against the section method using the stereomicroscope for marginal leakage assessment along the sealant-enamel interface. Methods: Pits and fissures of the occlusal surface of 10 teeth were sealed with a resin-fissure sealant material without acid etching, thermocycled for 5000 cycles, immersed in 50% silver nitrate for three hours and scanned using micro-CT. Teeth were embedded in epoxy resin and cut in three sections. The middle section was subjected to micro-CT and stereomicroscopy. Images were taken from the left and right sides of the sealant-enamel interface at both the left and the right site of the section. Two experienced evaluators assessed marginal leakage. Results: Both assessment instruments observed no leakage in 37 out of the 40 images evaluated. Leakage at the sealant-enamel interface was observed in three stereomicroscopy images only. A fracture line in the sealant was seen on eight stereomicroscopy images and observed in only two micro-CT images. Conclusions: The quality of the micro-CT and related software used in the present study does not qualify it to replace the section method as the gold standard for marginal leakage assessment at the sealant-enamel interface of permanent teeth.

In this study, porous chitosan/collagen scaffolds were prepared through a freeze-drying process, and loaded with the plasmid vector encoding human bone morphogenetic protein-7 (BMP-7) gene. To investigate the feasibility and efficacy of this gene-activated scaffold on dental tissue engineering, human dental pulp stem cells (DPSCs) were seeded in this scaffold for in vitro and in vivo study. In vitro results indicated that cells can be transfected successfully by loaded plasmid and secrete BMP-7 until day 24. Evaluation of DNA content, ALP activity, calcium content, SEM, and real-time PCR revealed that cells on gene-activated scaffold showed better proliferation properties and odontoblastic differentiation behaviors than cells on pure scaffolds. Then, these cell-scaffold complexes were implanted subcutaneously and retrieved after 4 weeks for histology evaluation. In vivo results that gene-activated scaffold group could still trace the existence of tranfected cells at week 4 and showed the upregulated expression of DSPP compared to pure scaffold groups. On the basis of our results, chitosan/collagen-loaded BMP-7 DNA appears to be an effective substrate candidate for gene delivery and indeed enhanced DPSCs differentiation toward an odontoblast-like phenotype in vitro and in vivo. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A:, 2012.

Introduction: Recombinant human bone morphogenetic protein-7 (BMP-7) is a well-established agent to induce bone and dentin formation. Little is understood until now whether BMP-7 could be used to genetically modify human dental pulp stem cells for tissue engineering applications. Methods:This study was to determine the feasibility of mineralized tissue formation from human dental pulp-derived stem cells (DPSCs) transfected with adenoviral-mediated human BMP-7 gene through in vitro and in vivo evaluations. Results: In vitro results of alkaline phosphatase, calcium content, and real-time polymerase chain reaction showed that BMP-7–transfected cells had the ability to differentiate towards the odontoblast phenotype and produce a calcified extracellular matrix. Transfected cell were seeded onto a porous ceramic scaffold and implanted subcutaneously in mice. Samples were retrieved after 4 and 8 weeks for histology evaluation. The results indicated that only the cultures with BMP-7 gene transfection showed obvious hard-tissue generation. Conclusions: Our data suggest that adenovirusmediated BMP-7 expression can induce odontogenic differentiation of human DPSCs and show effectively mineralized tissue formation in vivo, which may provide support for gene therapy candidate of BMP-7 in dental tissue engineering. (J Endod 2012;38:170–176)

Postnatal dental pulp stem cells (DPSCs) represent a unique precursor population in the dental pulp, which have multipotential and harbor great potential for tissue engineering purposes. However, for therapy applications, transplanted cells are often exposed to unfavorable conditions such as cytokines released from necrotic or inflammatory cells in injured tissues. It is not clear how stem cells exposed to these conditions changes in their characteristics. In this study, the effects of pro-inflammatory cytokines, such as IL-1 and TNF, on DPSCs were investigated. Cells were treated with IL-1, TNF, or both for 3, 7, and 12 days. The cultures were evaluated for cell proliferation, ALP activity, and real-time PCR. We found that a short treatment (3 days) of pro-inflammatory cytokines induced the odontogenic differentiation of DPSCs. Furthermore, post 3 days treatment with pro-inflammatory cytokines, the cell-scaffold complexes were implanted subcutaneously in mice for 8 weeks. Histological analysis demonstrated that the cultures gave obviously mineralized tissue formation, especially for both IL-1 and TNF applied. These data suggest that IL-1 and TNF produced in the early inflammatory reaction may induce the mineralization of DPSCs.

Item does not contain fulltext OBJECTIVES: The aim of the present study was to investigate the caries-preventive effect of sealants produced with a high-viscosity glass-ionomer with an elevated powder-liquid ratio (ART), of having energy added to this glass-ionomer, and that of glass-carbomer, in comparison to that of resin composite sealants. METHODS: The randomized controlled trial covered 407 children, with a mean age of 8 years. At a school compound three dentists placed sealants in pits and fissures of high caries-risk children. Evaluation by two independent evaluators was conducted after 0.5, 1 and 2 years. The Kaplan-Meier survival method, ANOVA and t-test were used in analyzing the data. RESULTS: 1352 first permanent molars were sealed. 6.6% of children and 6.8% of sealants dropped out within 2 years. 27 re-exposed pits and fissures, 20 in occlusal and 7 in smooth surfaces, in 25 children, developed a dentin carious lesion. The cumulative survival of dentin carious lesion-free pits and fissures in the glass-carbomer sealant group was statistically significantly lower (97.4%) than those in the high-viscosity glass-ionomer with energy supplied (99%) and the resin-composite (98.9%) sealant groups. There was no statistically significant difference in the cumulative survival of dentin carious lesion-free pits and fissures, between the high-viscosity glass-ionomer with (99%) and without (98.3%) energy supplied sealant groups, after 2 years. SIGNIFICANCE: The survival of dentin carious lesion-free pits and fissures was high in all sealant types. More dentin carious lesions were observed in the glass-carbomer sealant group. 01 mei 2012

Directing antigens to antigen presenting cells (APCs) has been demonstrated to be an efficient strategy to enhance immune responses induced by DNA vaccination. Fusion of antigens to cytotoxic T-Lymphocyte antigen 4 (CTLA4), a ligand of B7 molecules on the surfaces of APCs with strong binding affinity, enhanced the immunogenicities of antigens in various degrees. To investigate the relationship between antigen size and the immunogenicity of CTLA4 fusion DNA vaccine, we constructed CTLA4 targeted fusion anti-caries DNA vaccines containing different size of antigens. In vivo and in vitro experiments showed that CTLA4 fusion with smaller antigen induced stronger humoral immune responses and had higher affinity to B7-expressed cells than fusion with larger antigen. In conclusion, antigen size is one of the important factors regulating the potency of humoral immune response induced by CTLA4 targeted DNA vaccines.

We and others have shown that anti-caries DNA vaccines, including pGJA-P/VAX, are promising for preventing dental caries. However, challenges remain because of the low immunogenicity of DNA vaccines. In this study, we used recombinant flagellin protein derived from Salmonella (FliC) as a mucosal adjuvant for anti-caries DNA vaccine (pGJA-P/VAX) and analyzed the effects of FliC protein on the serum PAc-specific IgG and saliva PAc-specific IgA antibody responses, the colonization of Streptococcus mutans ( S. mutans) on rat teeth, and the formation of caries lesions. Our results showed that FliC promoted the production of PAc-specific IgG in serum and secretory IgA (S-IgA) in saliva of rats by intranasal immunization with pGJA-P/VAX plus FliC. Furthermore, we found that enhanced PAc-specific IgA responses in saliva were associated with the inhibition of S. mutans colonization of tooth surfaces and endowed better protection with significant fewer caries lesions. In conclusion, our study demonstrates that recombinant FliC could enhance specific IgA responses in saliva and protective ability of pGJA-P/VAX, providing an effective mucosal adjuvant candidate for intranasal immunization of an anti-caries DNA vaccine.

Oral Diseases (2012) 18, 169–177 Objectives: To isolate the CD133+CD44+ cells from human tongue squamous cell carcinoma (TSCC) Tca8113 cell line and investigate biological characteristics of them. Materials and methods: Immunomagnetic microbeads were applied to sort the CD133+CD44+ cells. Flow cytometry was used to detect isolation purity. The proliferation, clone-formation efficiencies, invasion and migration, gene expressions, and tumor-formation abilities were analyzed among CD133+CD44+, CD133−CD44−, and total population of cells. Results: The average purities of CD133+ and CD44+ cells reached 97.3% and 98.7%, respectively. The proliferation of CD133+CD44+ cells was significantly higher than the other two groups. The clone-forming efficiency of three groups was 70%, 8%, and 14%, respectively. The average invaded and migrated cell numbers of CD133+CD44+ and total population cells were 132 and 36.2, 311.6, and 156.2, respectively. The expressions of Bcl-2 and Sox2 in CD133+CD44+ cells were significantly higher than those in total population cells. A total of 104 CD133+CD44+ cells could form secondary tumors in nude mice, while the total population group needed 106 cells. Conclusions: The CD133+CD44+ subpopulation cells possess stem-like characteristics. They appear to be the potential targets for future biology therapy of human TSCC.

J Oral Pathol Med (2011) 40: 643–650 Background: Beta-catenin is one of the key components of Wnt signaling pathway. Increased level of this protein has been proved to be associated with enhanced cellular proliferation and the development of many kinds of cancers. But its role in the carcinogenesis in human tongue squamous cell carcinoma, one of the most common carcinomas of the human oral cavity, remains poorly characterized. Methods: In this study, we used lentivirus-mediated RNA interference (RNAi) targeted against beta-catenin to determine the effects of decreasing the high constitutive level of this protein in human tongue carcinoma cell line Tca8113. Results: Our studies demonstrated that RNAi directly against beta-catenin markedly decreased beta-catenin gene expression and inhibited cellular proliferation as reflected in the reduced growth of tongue cancer cells both in vitro and in nude mice. Conclusions: RNA interference (RNAi) targeting against beta-catenin can induce cell growth suppression of tongue cancer and may have the potential as a therapeutic modality to treat human tongue cancer.

With a reflective single-walled carbon nanotube as the saturable absorber, a laser diode-pumped passively mode-locked Nd:YVO4 laser at 1064 nm was realized for the first time. The pulse duration of 12 Ps was produced with a repetition rate of 83.7 MHz. The peak power and the single pulse energy of the mode-locking laser were 1.28 kW and 15.4 nJ, respectively. (C) 2014 Elsevier GmbH. All rights reserved.

Although Porphyromonas gingivalis lipopolysaccharide (P-LPS) is known to inhibit osteoblast differentiation, the exact molecular mechanisms underlying this phenomenon remain unclear. Here, we investigated the role of Notch signaling in the osteoblastic differentiation of both MC3T3E-1 cells and primary mouse bone marrow stromal cells (BMSCs). P-LPS stimulation activated the Notch1 signaling cascade and increased expression of the Notch target genes HES1 and HEY1. P-LPS can also act as an inhibitor because it is capable of suppressing Wnt/beta-catenin signaling in preosteoblasts by decreasing both glycogen synthase kinase-3 beta (GSK-3 beta) phosphorylation and the expression of nuclear p-catenin. These effects were rescued, however, by inhibiting Notch1 signaling. Furthermore, P-LPS treatment inhibited osteoblast differentiation in preosteoblasts as demonstrated by reductions in alkaline phosphatase activity, osteoblast gene expression, and mineralization, all of which were rescued by suppression of Notch1 signaling. Moreover, inhibition of GSK-3 beta, HES1, or HEY1 partially reversed the P-LPS-induced inhibition of osteoblast differentiation. Together, these findings suggest that P-LPS inhibits osteoblast differentiation by promoting the expression of Notch target genes and suppressing canonical Wnt/beta-catenin signaling. J. Cell. Physiol. 225: 106-114, 2010. (C) 2010 Wiley-Liss, Inc.

Despite successes in the isolation and characterization of stem cells from the oral mucosal epithelium, there have been few studies on progenitor cells from the oral mucosal lamina propria. In this study, we isolate rat oral mucosal lamina propria cells (OMLPC) using nestin as a marker in an immunomagnetic sorting technique. The OMLPCs was negative for cytokeratin. Nestin and vimentin were expressed in the OMLPCs. And CD44 and STRO-1 were expressed in a subset of the OMLPCs, which suggest that the nestin positive OMLPCs be heterogeneous. Otherwise, OMLPCs express Oct4, which is a critical gene for pluripotency. The OMLPCs proliferated actively in vitro. A colony forming study demonstrated that OMLPCs exhibited colony-generating capacity. When cultured in defined medium, OMLPCs generated cells characteristic of osteoblast, adipocyte and astrocyte-like cells. In addition, OMLPCs seeded into three dimensional scaffolds form bone-like structures in vivo after 8 weeks. All of the results demonstrate that OMLPCs are a population of mesenchymal progenitor cells existing in rat oral mucosal lamina propria. Nestin is shown to be a useful molecular marker for these cells. In certain environments, OMLPCs can form hard tissue. Thus, OMLPCs may serve as a suitable source of cells for future bone or tooth tissue engineering applications.