Mingwen Fan

The performance of dental pulp stem cells on nanofibrous PCL/gelatin/nHA scaffolds

The aim of current study is to investigate the in vitro and in vivo behavior of dental pulp stem cells (DPSCs) seeded on electrospun poly(epsilon-caprolactone) (PCL)/gelatin scaffolds with or without the addition of nano-hydroxyapatite (nHA). For the in vitro evaluation, DNA content, alkaline phosphatase (ALP) activity and osteocalcin (OC) measurement showed that the scaffolds supported DPSC adhesion, proliferation, and odontoblastic differentiation. Moreover, the presence of nHA upregulated ALP activity and promoted OC expression. Real-time PCR data confirmed these results. SEM micrographs qualitatively confirmed the proliferation and mineralization characteristics of DPSCs on both scaffolds. Subsequently, both scaffolds seeded with DPSCs were subcutaneously implanted into immunocompromised nude mice. Scaffolds with nHA but without cells were implanted as control. Histological evaluation revealed that all implants were surrounded by a thin fibrous tissue capsule without any adverse effects. The cell/scaffold composites showed obvious in vivo hard tissue formation, but there was no sign of tissue ingrowth. Further, the combination of nHA in scaffolds did upregulate the expression of specific odontogenic genes. In conclusion, the incorporation of nHA in nanofibers indeed enhanced DPSCs differentiation towards an odontoblast-like phenotype in vitro and in vivo.

Fibrous scaffolds loaded with protein prepared by blend or coaxial electrospinning

The aim of the present study was to fabricate polycaprolactone-based nanofibrous scaffolds with incorporated protein via either the blend or coaxial electrospinning technique. Both techniques were compared with respect to processing set-up and scaffold characteristics as well as the release kinetics and biological activity of the loaded protein. Bovine serum albumin was used as a model protein to determine release profiles, while alkaline phosphatase was used to determine protein activity after the electrospinning process. Coaxial electrospinning resulted in a uniform fiber morphology with a core-shell structure, and a homogeneous protein distribution throughout the core of the fibers. In contrast, blend electrospinning formed bead-like fibers with a heterogeneous protein distribution in the fibers. The coaxial scaffold exhibited more sustained release profiles than the comparative blend scaffold, and the additive poly(ethylene glycol) (PEG) in the coaxial scaffold accelerated protein release. Both electrospinning processes decreased the biological activity of the incorporated protein, but coaxial electrospinning with PEG as an additive showed up to 75% preservation of the initial biological activity. Thus, coaxial electrospinning was demonstrated to be superior to blend electrospinning for the preparation of nanofibrous scaffolds with a uniform fibrous structure and protein distribution and sustained protein release kinetics as well as high preservation of the protein activity.

Untreated Cavitated Dentine Lesions: Impact on Children’s Quality of Life

The aim of the present investigation was to assess the impact of dental caries prevalence and the consequences of untreated cavitated dentine lesions on quality of life of 6- and 7-year-old Brazilian children. A total of 826 schoolchildren were assessed using ICDAS and pufa (to score consequences of dental caries on soft tissues) indices. History of extraction and toothache was recorded. Oral health-related quality of life was assessed using the Brazilian version of the Early Childhood Oral Health Impact Scale (B-ECOHIS). A multiple logistic regression model was used to analyze the relationship between the prevalence of dentine carious lesions, pufa, history of extraction and toothache with the B-ECOHIS scores. A total of 587 questionnaires were analyzed. The prevalence of cavitated dentine lesions and pufa was 74.8 and 26.2%, respectively. Some 21.8% of children reported toothache and 9.2% had had at least one tooth extraction. The chance (OR) for children with cavitated dentine lesions, pufa ≥1, history of extraction and toothache of having higher B-ECOHIS scores than those not affected was 1.90 (95% CI: 1.18–3.06), 6.26 (95% CI: 3.63–10.83), 6.87 (95% CI: 2.75–17.16) and 3.68 (95% CI: 2.12–6.39), respectively. Children’s quality of life was negatively influenced by untreated cavitated dentine lesions and their consequences.

Mineralized Tissue Formation by BMP2-transfected Pulp Stem Cells

Previously, in vitro differentiation of odontoblasts was shown for dental pulp stem cells (DPSCs) transfected with bone morphogenetic protein-2 (Bmp2). For this study, we hypothesized that such cells also show potential for mineralized tissue formation in vivo. DPSCs were transfected with Bmp2 and seeded onto a ceramic scaffold. These complexes were cultured in medium without dexamethasone, and thereafter placed subcutaneously in nude mice for 1, 4, and 12 weeks. Samples were evaluated by histology and real-time PCR for osteocalcin, bone sialoprotein, dentin sialophosphoprotein, and dentin matrix protein 1. Results indicated that only the transfected DPSCs showed obvious mineralized tissue generation, and 12 weeks of implantation gave the highest percentage of mineralized tissue formation (33 ± 7.3% of implant pore area). Real-time PCR confirmed these results. In conclusion, Bmp2-transfected DPSCs effectively show mineralized tissue formation upon ectopic implantation.

Root Canal Systems in Mandibular First Premolars with C-shaped Root Configurations. Part I: Microcomputed Tomography Mapping of the Radicular Groove and Associated Root Canal Cross-sections

The purpose of this study was to investigate the morphology of the radicular grooves and root canal cross-sections in mandibular first premolars that exhibit a C-shaped root. Eighty-six mandibular first premolars with C-shaped roots were scanned from the coronal to anatomical apex using microCT80. The location, length, and depth of the radicular grooves were measured. The cross-sectional morphology of root canal systems was evaluated at 11 levels. Results were subject to the Kruskal-Wallis test. Of the 86 C-shaped roots, a total of 93 radicular grooves were noted. Most of the grooves (78.5%) were located on the mesiolingual surface of the root. The C-shaped canal could vary considerably in shape at different levels, and the incidence of two canals at the middle and apical was 21% and 80%, respectively. This study shows that radicular grooves may present relevant morphologic variations in the root canal system when the C-shaped root is present.

Anti-Inflammatory Activity of Curcumin in Macrophages Stimulated by Lipopolysaccharides from Porphyromonas gingivalis

Background:Porphyromonas gingivalis, a major periodontopathic bacterium, is necessary for periodontitis to take place. The lipopolysaccharide (LPS) of P. gingivalis stimulates cytokine secretion in immune cells, and thereby initiates the inflammation related to periodontitis. Macrophages are the important ones of the immune cells that are prominent at inflammatory periodontal sites. Curcumin, a major curcumanoid found in the spice turmeric, exhibits anti-inflammatory properties. The aim of this study was to investigate the anti-inflammatory effect and the mechanism of action of curcumin in macrophages stimulated by P. gingivalis LPS. Methods: RAW264.7 cells pre-treated with various concentrations of curcumin were stimulated by P. gingivalis LPS. TNF-α and IL-1β expressions were separately detected by RT-PCR and ELISA. Next, activation of NF-ĸB-dependent transcription was examined by luciferase assay. Results: Curcumin dose-dependently inhibited TNF-α and IL-1β gene expression and protein synthesis in RAW264.7 cells stimulated with P. gingivalis LPS. P. gingivalis LPS activated NF-ĸB-dependent transcription in RAW264.7 cells, which were down-regulated by pre-treatment with curcumin as well. Conclusion: Our data suggest that curcumin can inhibit P. gingivalis LPS-induced cytokine expression, and that this could be due to the inhibition of the NF-ĸB pathway.

Non-viral bone morphogenetic protein 2 transfection of rat dental pulp stem cells using calcium phosphate nanoparticles as carriers.

Calcium phosphate nanoparticles have shown potential as non-viral vectors for gene delivery. The aim of this study was to induce bone morphogenetic protein (Bmp)2 transfection in rat dental pulp stem cells using calcium phosphate nanoparticles as a gene vector and then to evaluate the efficiency and bioactivity of the transfection. We also intended to investigate the behavior of transfected cells when seeded on 3-dimensional titanium fiber mesh scaffolds. Nanoparticles of calcium phosphate encapsulating plasmid deoxyribonucleic acid (DNA) (plasmid enhanced green fluorescent protein-BMP2) were prepared. Then, STRO-1-selected rat dental pulp stem cells were transfected using these nanoparticles. Transfection and bioactivity of the secreted BMP2 were examined. Thereafter, the transfected cells were cultured on a fibrous titanium mesh. The cultures were investigated using scanning electron microscipy and evaluated for cell proliferation, alkaline phosphatase activity and calcium content. Finally, real-time polymerase chain reaction was performed for odontogenesis-related gene expression. The results showed that the size of the DNA-loaded particles was approximately 100 nm in diameter. Nanoparticles could protect the DNA encapsulated inside from external DNase and release the loaded DNA in a low-acid environment (pH 3.0). In vitro, nanoparticle transfection was shown to be effective and to accelerate or promote the odontogenic differentiation of rat dental pulp stem cells when cultured in the 3-dimensional scaffolds. Based on our results, plasmid DNA-loaded calcium phosphate nanoparticles appear to be an effective non-viral vector for gene delivery and functioned well for odontogenic differentiation through Bmp2 transfection.

Dentin phosphoprotein frameshift mutations in hereditary dentin disorders and their variation patterns in normal human population

Background: Dentin phosphoprotein (DPP) is the most abundant non-collagenous protein in dentin, which is highly phosphorylated and plays key roles in dentin biomineralisation. The aetiology of isolated hereditary dentin disorders in most affected families is largely unknown and the association between DPP and dentin disorders has not been well established. This study aims to determine whether there are some involvements for DPP mutations in inherited dentin disorders and to clarify the sequence variation patterns of DPP in normal population. Methods: Genomic DNA was analysed in eight families with hereditary dentin disorders and 110 individuals in the normal population. The full coding sequence of DPP was amplified by polymerase chain reaction (PCR) and screened for mutations and variations by direct sequencing and TOPO TA-cloning sequencing. Results: Five frameshift mutations in DPP coding region were identified in five of the eight families. The mutations co-segregated with the disease phenotypes in affected families and were not found in 220 control chromosomes. In the normal population, we revealed 14 in-frame indels (insertion/deletion), six non-synonymous single nucleotide polymorphisms (SNPs), and five synonymous SNPs in the DPP coding region. These variants display extensive linkage disequilibrium and constitute a total of 15 haplotypes with three predominant haplotypes in the investigated normal population. Conclusions: Our data provide the first evidence that DPP mutations can cause hereditary dentin disorders and suggest that in-frame length variations and missense SNPs in DPP have no obvious pathogenetic effects on dentin formation.

Human Papillomavirus as an Independent Predictor in Oral Squamous Cell Cancer

AimThere is an increasing evidence for the role of high risk human papillomavirus (HPV) in the pathogenesis of oral squamous cell carcinoma (OSCC). The purpose of this study is to evaluate the relevance of HPV infection to the survival and prognosis of OSCC.MethodologyFifty‐two patients with OSCC were followed from 4 to 88 months with a median of 50.7 months. HPV DNA was identified in formalin‐fixed, paraffin‐embedded tumor specimens by nested PCR with MY09/MY11 and GP5+/GP6+ primer pairs and the HPV genotype was determined by direct DNA sequencing. Association between the HPV status and risk factors for cancer as well as tumor‐host characteristics were analyzed. Survival curves were calculated by the Kaplan‐Meier method and analyzed using the log‐rank test.ResultsHPV was found in 40.4% of the tumors with HPV16 accounting for 63.5%, HPV18 for 30.8%, HPV6 for 3.9% and HPV11 for 1.8%. No infection with more than one HPV genotype was detected. HPV infection was significantly associated with poor histological grade, TNM stage I–II, alcohol usage and no smoking status. Multi‐variate analysis showed that HPV had an independent prognostic effect on the overall survival after adjusting other confounding factors such as histological grade, TNM stage and tobacco usage. The presence of HPV was significantly correlated with a better survival in patients with OSCC.ConclusionHPV infection can act as an independent predictor for the survival and prognosis of OSCC.

Caries-preventive effect of sealants produced with altered glass-ionomer materials, after 2 years

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.