Novel LRAP-binding partner revealing the plasminogen activation system as a regulator of cementoblast differentiation and mineral nodule formation in vitro.

Amelogenin isoforms, including full-length amelogenin (AMEL) and leucine-rich amelogenin peptide (LRAP), are major components of the enamel matrix, and are considered as signaling molecules in epithelial-mesenchymal interactions regulating tooth development and periodontal regeneration. Nevertheless, the molecular mechanisms involved are still poorly understood. The aim of the present study was to identify novel binding partners for amelogenin isoforms in the cementoblast (OCCM-30), using an affinity purification assay (GST pull-down) followed by mass spectrometry and immunoblotting. Protein-protein interaction analysis for AMEL and LRAP evidenced the plasminogen activation system (PAS) as a potential player regulating OCCM-30 response to amelogenin isoforms. For functional assays, PAS was either activated (plasmin) or inhibited (ε-aminocaproic acid [aminocaproic]) in OCCM-30 cells and the cell morphology, mineral nodule formation, and gene expression were assessed. PAS inhibition (EACA 100 mM) dramatically decreased mineral nodule formation and expression of OCCM-30 differentiation markers, including osteocalcin (Bglap), bone sialoprotein (Ibsp), osteopontin (Spp1), tissue-nonspecific alkaline phosphatase (Alpl) and collagen type I (Col1a1), and had no effect on runt-related transcription factor 2 (Runx2) and Osterix (Osx) mRNA levels. PAS activation (plasmin 5 µg/µl) significantly increased Col1a1 and decreased Bglap mRNA levels (p < .05). Together, our findings shed new light on the potential role of plasminogen signaling pathway in the control of the amelogenin isoform-mediated response in cementoblasts and provide new insights into the development of targeted therapies.

Does enamel matrix derivative application improve clinical outcomes after semilunar flap surgery? A randomized clinical trial.

OBJECTIVES: To evaluate the treatment of gingival recessions by semilunar coronally positioned flap plus enamel matrix derivative (SCPF + EMD). MATERIALS AND METHODS: Thirty patients with class I localized gingival recession were included. They were randomly allocated in two groups: SCPF + EMD and SCPF. Recession height (RH), recession width (RW), width of keratinized tissue (WKT), thickness of keratinized tissue (TKT), probing depth (PD), and clinical attachment level (CAL) were measured at baseline, 6 and 12 months post-surgery. Patient/professional evaluation of esthetics and root sensitivity was performed. RESULTS: After 12 months, mean root coverage was 1.98 ± 0.33 mm for SCPF + EMD (90.86 ± 14.69%) and 1.85 ± 0.41 mm (79.76 ± 17.44%) for SCPF (p > 0.05). The esthetic evaluation by the patient showed preference for SCPF + EMD. According to the professional evaluation (QCE), the use of EMD decreases the appearance of postoperative scar tissue line. There was a significant reduction in root hypersensitivity with no further complaints by the patients. CONCLUSIONS: The addition of EMD provides significantly better esthetics to SCPF, according to patient and professional assessments. SCPF + EMD is effective but not superior to SCPF for root coverage, after 12 months. CLINICAL RELEVANCE: Previous clinical trials showed that the combination of EMD with coronally advanced flaps may enhance the outcome of root coverage. There is a lack of studies testing the combination of EMD with SCPF. The combination SCPF + EMD provides better esthetics when compared to the SCPF and is effective, but not superior, to SCPF for root coverage, after 12 months. TRIAL REGISTRATION: NCT02459704.

Leucine-Rich Amelogenin Peptide (LRAP) Uptake by Cementoblast Requires Flotillin-1 Mediated Endocytosis.

Basic, pre-clinical, and clinical studies have documented the potential of amelogenin, and its variants, to affect cell response and tissue regeneration. However, the mechanisms are unclear. Thus, the aim of the present study was to identify, in cementoblasts, novel binding partners for an alternatively spliced amelogenin form (Leucine-Rich Amelogenin Peptide-LRAP), which is supposed to act as a signaling molecule in epithelial-mesenchymal interactions. LRAP-binding protein complexes from immortalized murine cementoblasts (OCCM-30) were achieved by capture affinity assay (GST pull down) and proteins present in these complexes were identified by mass spectrometry and immunoblotting. Flotillin-1, which functions as a platform for signal transduction, vesicle trafficking, endocytosis, and exocytosis, was identified and confirmed by co-precipitation and co-localization assays as a protein-binding partner for LRAP in OCCM-30 cells. In addition, we found that exogenously added GST-LRAP recombinant protein was internalized by OCCM-30 cells, predominantly localized in the perinuclear region and, that inhibition of flotillin1-dependent functions by small interference RNA (siRNA) methodology significantly affected LRAP uptake and its biological properties on OCCM-30 cells, including LRAP effect on the expression of genes encoding osteocalcin (Ocn), bone sialoprotein (Bsp), and runt-related transcription factor 2 (RunX2). In conclusion, LRAP uptake by cementoblast involves flotillin-assisted endocytosis, which suggests an involvement of LRAP in lipid-raft-dependent signaling pathways which are mediated by flotillin-1. J. Cell. Physiol. 232: 556-565, 2017. © 2016 Wiley Periodicals, Inc.

Full-mouth ultrasonic debridement associated with povidone iodine rinsing in GAgP treatment: a randomised clinical trial.

OBJECTIVE: This study evaluated the clinical, immunological and microbiological results of full-mouth ultrasonic debridement (FMUD) with 10 % povidone iodine (PVPI) as the cooling liquid in the treatment of generalised aggressive periodontitis (GAgP). MATERIAL AND METHODS: Twenty-eight patients presenting GAgP were randomly assigned to one of the following groups for evaluation: FMUD + SS (n = 14)--single session of FMUD with 0.9 % saline solution as cooling agent and FMUD + PVPI (n = 14)--single session of FMUD with PVPI solution as cooling agent. Probing depth (PD), relative clinical attachment level (RCAL), relative position of gingival margin, plaque index (FMPI) and bleeding score (FMBS), immunological (interleukin-10 and interleukin-1ß concentrations in gingival crevicular fluid) and microbiological (Aa and Pg amounts) parameters were evaluated at baseline, first, third and sixth months after treatment. RESULTS: The two groups presented reduction of FMPI and FMBS and had statistically significant PD reductions, RCAL gains and gingival recession (p < 0.05). Both therapies reduced Pg levels in deep and in moderate pockets (p < 0.05). FMUD + PVPI reduced Aa levels in deep pockets. However, no inter-group differences in clinical, immunological and microbiological parameters were observed (p > 0.05). CONCLUSIONS: It could be concluded that 10 % PVPI used as an irrigant solution in FMUD decreased Aa levels in deep pockets but had no additional benefits when compared with saline solution irrigation in terms of clinical, microbiological and immunological results. CLINICAL RELEVANCE: The FMUD is a valid option for the treatment of GAgP, but the use of 10 % PVPI did not improve the results of the periodontal therapy.

Periodontal clinical and microbiological characteristics in healthy versus generalized aggressive periodontitis families.

AIM: Generalized aggressive periodontitis (GAP) is a severe and multifactorial disease in which a familial aggregation and a specific microbiological profile have been suggested. Thus, this case-control study evaluated the clinical and subgingival microbial profile of GAP subjects and their families compared to healthy families. METHODS: Fifteen families with parents presenting periodontal health and 15 with parents with a history of GAP were selected. Each family should have at least one child between 6 and 12 years old. Plaque index (PI), gingival index (GI), and periodontal probing depth (PPD), as well as Porphyromonas gingivalis, Tannerella forsythia, and Aggregatibacter actinomycetemcomitans (Aa) amounts (by qPCR), were assessed from all subjects. RESULTS: Children of GAP families showed a higher PI, GI, and PPD when compared to children of healthy families (p ≤ 0.05). A higher frequency of detection and amounts of Aa was observed in GAP children compared to children of healthy families (p ≤ 0.05). Moreover, a significant association between Aa amounts and gingival bleeding was observed in children (p ≤ 0.05, r = 0.37). CONCLUSION: Children from GAP families have worst clinical conditions, i.e. higher levels of PI, GI, and PPD, a more pathogenic microbiological profile, and the amount of Aa are associated with a higher marginal inflammation.

Salivary carriage of periodontal pathogens in generalized aggressive periodontitis families.

BACKGROUND: Generalized aggressive periodontitis (GAP) is a multifactorial disease that shows a specific microbial profile and a familial aggregation. AIM: This study evaluated the salivary microbial profile of families with a history of GAP and compared them with healthy families. DESIGN: Fifteen families with parents presenting periodontal health and 15 with parents with a history of GAP were selected. Each family had a child aged 6-12 years. Stimulated saliva was collected from all subjects, and Porphyromonas gingivalis (Pg), Tannerella forsythia (Tf), and Aggregatibacter actinomycetemcomitans (Aa) amounts were determined. RESULTS: Children of GAP families showed higher detection of Aa (90%) than children of healthy families (45%) (P < 0.05). Parents with GAP showed a Pg salivary concentration statistically higher than that of healthy parents (P < 0.05).Children of GAP families, however, exhibited similar Pg concentration than healthy children (P > 0.05). Tf amounts did not differ either in parents or in children (P > 0.05) The infection risk calculation indicates that children who have one parent who is positive for Aa have 16.3 times (95% CI 3.1-87.2) more risk of being infected with Aa (P < 0.05) than children from an Aa-negative family. CONCLUSION: It may be concluded that children of parents with aggressive periodontitis have higher levels and higher risk of Aa infection.

Parathyroid hormone (1-34) compensates the negative effect of smoking around implants.

OBJECTIVES: This study aimed to investigate the effect of Recombinant Human Parathyroid Hormone (PTH 1-34) on attenuating the influence of cigarette smoke on bone around titanium implants. MATERIAL AND METHODS: Forty-eight female Wistar rats were used. At the beginning of the study, 15 animals were randomly assigned to Group 1 (control) and received subcutaneous injections of saline solution, three-times/week, after implant placement. The other animals received intermittent cigarette smoke inhalation (CSI), 60 days prior and 60 days after implant placement ( Al 2 O 3 -blasted titanium implants - 4.0 × 2.2 mm). After surgery, these animals were randomly assigned to: Group 2 - subcutaneous injections of saline solution, three-times/week (n = 16) and Group 3 - intermittent doses of PTH (1-34) (40 µg/Kg), three-times/week (n = 17). Animals were sacrificed 60 days after surgery, and degree of bone-to-implant contact (BIC), bone area (BA) within the limits of the threads and proportion of mineralized tissue (PMT) adjacent to the implants (500 µm wide zone) were separately obtained in cortical and cancellous bone. RESULTS: Data analysis confirmed that CSI negatively affects bone around implants, as observed for BIC in cortical zone (Cohen's d (d) = -1.26) and for PMT in both zones (d = -6.09 and d = -4.46 for cortical and cancellous zones, respectively). In addition, in the presence of CSI, PTH (1-34) promoted the highest BIC in both regions and BA and PMT in cancellous bone (P < 0.05). The histometric parameter that was not influenced by both PTH and CSI (1-34) was BA in cortical bone (P > 0.05). CONCLUSION: In the presence of cigarette smoke, a factor related to poor bone healing and low bone density, PTH (1-34) increased bone volume around implants.

One stage, full-mouth, ultrasonic debridement in the treatment of severe chronic periodontitis in smokers: a preliminary, blind and randomized clinical trial.

OBJECTIVE: The aim of this clinical trial was to assess the performance of a full-mouth ultrasonic debridement protocol in the treatment of severe chronic periodontitis in comparison with scaling and root planing in a quadrant-wise procedure in smokers. MATERIALS AND METHODS: The trial consisted of 30 participants presenting with periodontitis divided into 3 groups: Group FMUD - full-mouth ultrasonic debridement, i.e., one session of 45 minutes of ultrasonic instrumentation for smokers (n = 10), Group SRP- scaling and root planing performed in a quadrant-wise manner for smokers (n = 10), and Group Control - SRP for nonsmokers (n = 10), treated following the same protocol as the SRP group. The parameters evaluated were: plaque/bleeding on probing indices, probing pocket depth, relative recession, and relative probing attachment level at baseline, 45, 90 and 180 days after therapy. RESULTS: Full-mouth ultrasonic debridement and scaling and root planing resulted in comparable gain of attachment 6 months after therapy. Both groups exhibited probing pocket depth reduction at all experimental periods as compared to baseline. Smokers, however, had less probing pocket depth reduction and relative probing attachment level gain compared to non-smokers, despite the mechanical protocol used (p < 0.05). Moreover, at 180 days, nonsmokers presented with fewer sites requiring re-treatment (probing pocket depth > 5 mm and bleeding on probing) than smokers (p < 0.05). CONCLUSIONS: Full-mouth ultrasonic debridement and scaling and root planing result in comparable clinical outcomes for the treatment of smokers with severe chronic periodontitis. Despite the non-surgical technique used, smokers had a less favorable clinical response than non-smokers.

Connective tissue graft plus resin-modified glass ionomer restoration for the treatment of gingival recession associated with non-carious cervical lesions: microbiological and immunological results.

OBJECTIVES: It was previously reported the clinical results of placing subgingival resin-modified glass ionomer restoration for treatment of gingival recession associated with non-carious cervical lesions. The aim of this study was to evaluate the influence of this treatment on the subgingival biofilm and gingival crevicular fluid (GCF) inflammatory markers. MATERIALS AND METHODS: Thirty-four patients presenting the combined defect were selected. The defects were treated with either connective tissue graft plus modified glass ionomer restoration (CTG+R) or with connective tissue graft only (CTG). Evaluation included bleeding on probing and probing depth, 5 different bacteria targets in the subgingival plaque assessed at baseline, 45, and 180 days post treatments, and 9 inflammatory mediators were also assessed in the GCF. RESULTS: The levels of each target bacterium were similar during the entire period of evaluation (p > 0.05), both within and between groups. The highest levels among the studied species were observed for the bacterium associated with periodontal health. Additionally, the levels of all cyto/chemokines analyzed were not statistically different between groups (p > 0.05). CONCLUSION: Within the limits of the present study, it can be concluded that the presence of subgingival restoration may not interfere with the subgingival microflora and with GCF inflammatory markers analyzed. CLINICAL RELEVANCE: This approach usually leads to the placement of a subgingival restoration. There is a lack of information about the microbiological and immunological effects of this procedure. The results suggest that this combined approach may be considered as a treatment option for the lesion included in this study.

Autologous periodontal ligament cells in the treatment of Class III furcation defects: a study in dogs.

AIM: This study aimed to evaluate, histomorphometrically, the use of periodontal ligament cells (PDL cells) in the treatment of class III furcation defects. MATERIAL AND METHODS: PDL cells were obtained from the mandibular tooth extracted from each dog (7), cultured in vitro and phenotypically characterized. Bilateral class III furcation defects were created at mandibular 3rd and 4th premolars and were randomly assigned to: CONTROL GROUP: coronally positioned flap, GTR Group: GTR, Sponge Group: carrier + GTR, Cell Group: carrier + PDL cells + GTR. RESULTS: After 3 months of healing, data analysis demonstrated that the Cell Group presented a superior length of new cementum (4.82 ± 0.61 mm; 3.66 ± 0.95 mm; 2.87 ± 0.74 mm and 1.70 ± 0.60 mm, p < 0.001), a greater extension of periodontal regeneration (3.43 ± 1.44 mm; 2.33 ± 0.95 mm; 1.52 ± 0.39 mm and 0.69 ± 0.59 mm, p = 0.001) and a larger area of new bone (5.45 ± 1.58 mm(2) ; 3.94 ± 1.52 mm(2) ; 2.91 ± 0.56 mm(2) and 1.89 ± 0.95 mm(2) , p = 0.0012), when compared with Sponge, GTR and CONTROL GROUP, respectively. CONCLUSION: The PDL cells in association with GTR may significantly promote periodontal regeneration in class III furcation defects surgically created in dogs.

Autologous periodontal ligament cells in the treatment of class II furcation defects: a study in dogs.

AIM: The goal of this study was to histologically investigate the use of periodontal ligament cells (PDL cells) in tissue engineering to regenerate class II furcation defects. MATERIAL AND METHODS: PDL cells were obtained from the mandibular tooth extracted from each dog (seven), cultured in vitro and phenotypically characterized with regard to their biological properties. Following, bilateral class II furcation lesions were created at maxillary 3rd premolars and were randomly assigned to the test group [PDL cells+guided tissue regeneration (GTR)] or the control group (GTR). After 3 months, the animals were euthanized to evaluate the histometric parameters. RESULTS: In vitro, PDL cells were able to promote mineral nodule formation and to express bone sialoprotein, type I collagen and alkaline phosphatase. Histometrically, data analysis demonstrated that the cell-treated group presented a superior length of new cementum (6.00 ± 1.50 and 8.08 ± 1.08 mm), a greater extension of periodontal regeneration (3.94 ± 1.20 and 7.28 ± 1.00 mm), a lower formation of connective tissue/epithelium (2.15 ± 1.92 and 0.60 ± 0.99 mm), a larger area of new bone (7.01 ± 0.61 and 9.02 ± 2.30 mm(2)) and a smaller area of connective tissue/epithelium (5.90 ± 1.67 and 4.22 ± 0.95 mm(2)), when compared with control group. CONCLUSION: PDL cells in association with GTR may significantly promote periodontal regeneration in class II furcation defects in dog.

TLR2 and TLR4 gene promoter methylation status during chronic periodontitis.

AIM: The objective of this study was to analyse the status of DNA methylation in the promoter region of the toll-like receptor (TLR)2 and TLR4 genes in gingival tissue samples from healthy subjects, smokers and non-smokers affected by chronic periodontitis. MATERIAL AND METHODS: Genomic DNA and total RNA were purified from gingival tissue using the TRIZOL reagent protocol. Genomic DNA was then digested by methylation-sensitive restriction enzymes, amplified by polymerase chain reaction (PCR), electrophoresed on a 10% polyacrylamide gel and stained using SYBR Gold. Real-time PCR was also performed to verify the transcript levels. RESULTS: The CpG dinucleotides analysed were observed to be unmethylated in the majority of DNA samples of the three groups and statistical differences were not found among groups (p>0.05). However, a trend towards methylation was observed in the TLR2 HhaI site in the samples of the periodontitis non-smoker groups. In fact, the analysis of all CpG sites together shows which complete methylation is observed in the shortest level in the samples of periodontitis non-smoker group. The analysis of transcript levels demonstrated no difference among groups (p>0.05). CONCLUSION: The results demonstrated major unmethylation of the TLR4 gene promoter in all groups. However, the results for the TLR2 gene promoter are inconclusive; this gene was found as a mosaic of methylated and unmethylated DNA in the majority of samples of the three groups and we also observed a trend towards the DNA methylation of CpG sites recognized by the HhaI enzyme.

Influence of different enamel substrates on microtensile bond strength of sealants after cariogenic challenge.

PURPOSE: To evaluate the microtensile bond strength (µTBS) of resin sealer on enamel substrates after cariogenic challenge. MATERIALS AND METHODS: Enamel blocks were obtained from human third molars and randomly divided into 6 groups (n = 10) according to enamel substrates (S: sound, CL: caries-like lesion, or CLTF: caries-like lesion + topical fluoride application) and sealant material (F: FluroShield, or H: Helioseal Clear Chroma). Sealants were placed on enamel surfaces, stored in 100% humidity (24 h, 37°C), and longitudinally sectioned into hourglass shapes. According to the groups, pH cycling was applied and the µTBS test was performed. The fracture patterns were assessed by SEM. RESULTS: Regarding substrates, the highest µTBS values in MPa were observed for CLTF enamel (26.0 ± 7.6), followed by S (22.0 ± 7.4) and CL (15.5 ± 4.9). A significant interaction was found between material and pH cycling (p = 0.0395). F (23.9 ± 7.6) showed higher µTBS values than H (18.3 ± 7.5) when submitted to pH cycling. The majority of samples presented mixed failure. CONCLUSIONS: Enamel substrate significantly affected µTBS, with the highest values for remineralized caries-like enamel lesions. Furthermore, µTBS values were dependent on both materials and pH cycling.

Enamel matrix derivative proteins for the treatment of proximal class II furcation involvements: a prospective 24-month randomized clinical trial.

OBJECTIVE: This study aimed to evaluate the response of proximal furcations treated with enamel matrix derivative proteins (EMD) in a 24-month follow-up. MATERIALS AND METHODS: Twelve patients presenting bilateral class II proximal furcation with vertical probing depth (PD) ≥5 mm and bleeding on probing were selected. The furcations were assigned to: a control group (n=12), open flap debridement (OFD)+EDTA and a test group (n=12) - OFD+EDTA+EMD. The gingival margin position, PD, relative vertical and horizontal clinical attachment level (RVCAL and RHCAL), vertical and horizontal bone level (VBL and HBL) and furcation closure were evaluated before treatment and after 6, 12 and 24 months. RESULTS: After follow-up, no statistical difference could be seen between groups. At 24 months, the test group showed 1.9 ± 1.6 mm PD reduction whereas the control group showed 1.0 ± 1.3 mm PD reduction. RHCAL gains of the control and the test group were 0.7 ± 1.3 and 1.4 ± 0.9 mm, respectively. However, at 24 months, the test group only presented five remaining class II furcations versus 10 furcations in the control group (p<0.05). CONCLUSION: It could be concluded that EMD therapy promoted a reduction in the number of proximal furcations presenting a diagnosis of class II after 24 months of treatment compared with OFD therapy.

Peri-implant reconstruction using autologous periosteum-derived cells and guided bone regeneration.

AIM: This investigation evaluated the bone healing in peri-implant defects treated with periosteum-derived cells (PCs) and guided bone regeneration (GBR). MATERIAL AND METHODS: PCs were harvested from six beagle dogs and characterized in vitro with regard to their osteogenic properties. The animals were subjected to teeth extraction in the mandible, and after 3 months of healing, implant sites were drilled, bone dehiscences were created and implants were placed. Dehiscences were randomly assigned to: PCs+GBR, GBR, PCs and non-treated defects. After 3 months, the implants/adjacent tissues were processed. Bone-to-implant contact (BIC) bone fill (BF) within implant threads, and bone area (BA) in a zone lateral to the implant were obtained. RESULTS: In vitro analyses confirmed the osteogenic potential of PCs. Histometrically, no statistically significant differences were observed among the PCs+GBR, GBR and PCs groups for both BF and BIC (p>0.05), whereas these groups showed statistically higher values, as compared with the non-treated group (p<0.05). With respect to BA, the PCs+GBR and GBR groups presented significantly higher means, as compared with the PCs and non-treated groups (p<0.05). CONCLUSION: Although successful outcomes have been promoted by using the combined approach, PCs in conjunction with membranes did not provide additional benefit during peri-implant bone regeneration, when compared with the therapeutic approaches used alone.

Treatment of dehiscence-type defects with collagen matrix and/or enamel matrix derivative: Histomorphometric study in minipigs.

BACKGROUND: This study aimed to evaluate, histomorphometrically, the use of collagen matrix (CM) and/or enamel matrix derivative (EMD) for the treatment of dehiscence-type recession defects in minipigs. METHODS: Eight healthy, male, young BR-1 minipigs, with no periodontal disease were treated. Bilateral dehiscence-type defects were surgically created on the buccal of the mandibular premolars (PI and PII). After 30 days, the defects were randomly assigned to four groups: coronally advanced flap (CAF); CAF + CM; CAF + EMD; and CAF + CM + EMD (split-mouth design). The evaluated parameters (mm): total defect length; new cementum (NC); new bone (NB); gingival margin position; total epithelium length; epithelium on the root; connective tissue adaptation; and soft tissue thickness (STT). RESULTS: The EMD-treated groups showed a superior length of NC [4.13 ± 1.22 (CAF + EMD); 3.95 ± 1.11 (CAF + CM + EMD); 2.94 ± 0.77 (CAF + CM); 2.72 ± 0.81 (CAF), P = 0.02] and NB [3.21 ± 0.68 (CAF + CM + EMD); 3.01 ± 0.56 (CAF + EMD); 2.15 ± 0.47 (CAF + CM); 2.29 ± 0.82 (CAF), P = 0.005]. The CAF and CAF + CM groups showed a superior epithelial length when compared to EMD-treated groups after 3 months. A superior STT was observed for CAF + CM + EMD group (1.5 ± 0.33) when compared with the other groups [1.09 ± 0.26 (CAF + EMD); 1.04 ± 0.34 (CAF + CM); and 1.14 ± 0.29 (CAF), P = 0.03]. CONCLUSION(S): The results of the present study indicate that EMD application, irrespective of the combination with CM, may improve the periodontal regeneration of dehiscence-type defects in this animal model.

Synthesis of multifunctional chlorhexidine-doped thin films for titanium-based implant materials.

Our goal was to create bio-functional chlorhexidine (CHX)-doped thin films on commercially pure titanium (cpTi) discs using the glow discharge plasma approach. Different plasma deposition times (50, 35 and 20 min) were used to create bio-functional surfaces based on silicon films with CHX that were compared to the control groups [no CHX and bulk cpTi surface (machined)]. Physico-chemical and biological characterizations included: 1. Morphology, roughness, elemental chemical composition, film thickness, contact angle and surface free energy; 2. CHX-release rate; 3. Antibacterial effect on Streptococcus sanguinis biofilms at 24, 48 and 72 h; 4. Cytotoxicity and metabolic activity using fibroblasts cell culture (NIH-F3T3 cells) at 1, 2, 3 and 4 days; 5. Protein expression by NIH-F3T3 cells at 1, 2, 3 and 4 days; and 6. Co-culture assay of fibroblasts cells and S. sanguinis to assess live and dead cells on the confocal laser scanning microscopy, mitochondrial activity (XTT), membrane leakage (LDH release), and metabolic activity (WST-1 assay) at 1, 2 and 3 days of co-incubation. Data analysis showed that silicon films, with or without CHX coated cpTi discs, increased surface wettability and free energy (p < 0.05) without affecting surface roughness. CHX release was maintained over a 22-day period and resulted in a significant inhibition of biofilm growth (p < 0.05) at 48 and 72 h of biofilm formation for 50 min and 20 min of plasma deposition time groups, respectively. In general, CHX treatment did not significantly affect NIH-F3T3 cell viability (p > 0.05), whereas cell metabolism (MTT assay) was affected by CHX, with the 35 min of plasma deposition time group displaying the lowest values as compared to bulk cpTi (p < 0.05). Moreover, data analysis showed that films, with or without CHX, significantly affected the expression profile of inflammatory cytokines, including IL-4, IL-6, IL-17, IFN-y and TNF-α by NIH-F3T3 cells (p < 0.05). Co-culture demonstrated that CHX-doped film did not affect the metabolic activity, cytotoxicity and viability of fibroblasts cells (p > 0.05). Altogether, the findings of the current study support the conclusion that silicon films added with CHX can be successfully created on titanium discs and have the potential to affect bacterial growth and inflammatory markers without affecting cell viability/proliferation rates.

TiO2 nanotubes improve physico-mechanical properties of glass ionomer cement.

OBJECTIVES: The aim of this study was to determine the physico-mechanical properties of a high viscosity glass ionomer cement (GIC) reinforced with TiO2 nanotubes (TiO2-nt). METHODS: TiO2-nt was incorporated into the GIC powder components (Ketac Molar EasyMix™) in concentrations of 0% (control group), 3%, 5%, 7% by weight. Compressive strength (n = 10/group), three point bending for flexural strength (n = 18/group), microshear bond strength to dentin and failure mode (n = 20/group), and surface roughness and weight loss before and after brushing simulation (30,000 cycles) (n = 8/group) were evaluated. Data were submitted to Shapiro-Wilk, ANOVA, Tukey and Chi-square tests (α ≤ 0.05). RESULTS: Addition of 5% of TiO2-nt into GIC presented the highest values for compressive strength and differed from the control, 3% and 7% groups (p = 0.023). There were no significant differences in flexural strength (p = 0.107) and surface roughness before and after the dental brushing (p = 0.287) among the groups. GIC added with 5% TiO2-nt showed the lowest weight loss values (p = 0.01), whereas the control, 3% or 5% TiO2-nt groups presented similar microshear bond strength values (p ≥ 0.05). The 5% TiO2-nt group featured higher microshear bond strength than the 7% TiO2-nt group (p = 0.034). Cohesive in material was the most representative failure mode for all groups. SIGNIFICANCE: The incorporation of TiO2-nt did not affect GIC's adhesiveness to dentin, but improved its compressive strength at 5%. Furthermore, TiO2-nt decreased the percentage of weight loss after GIC's surface wear.

Inflammatory and bone-related genes are modulated by aging in human periodontal ligament cells.

Periodontal ligament cells (PDLC) play a major role in periodontal tissues homeostasis and destruction. Most age-associated diseases seem to be closely related to an underlying chronic inflammatory state. Thus, the present study aimed at evaluating in PDLC the effect of aging on the basal levels of inflammatory and bone-related genes. Primary PDLC cultures were obtained from subjects aged 15-20 years (control- n=5), and subjects aged more than 60 years (test- n=5). Proliferation, cell viability and total secreted protein assays were performed, and mRNA levels were quantitatively assessed for interleukin (IL)-1beta, IL-4, IL-6 and IL-8, and for receptor activator of nuclear factor-kappaB ligand (RANKL) and osteoprotegerin (OPG) by real time PCR. Data analysis demonstrated that aging negatively influenced cell proliferation, whereas cell viability and total secreted protein were not affected (p>0.05). Gene expression analysis showed that mRNA levels for RANKL and IL-8 were not affected by aging (p>0.05) whereas, mRNA levels for IL-4 was significantly lower in aged cells (p<0.05) and OPG, IL-1beta and IL-6 mRNA levels were higher (p<0.05). Data analysis suggests that aging decreased the ability of PDLC to proliferate and modulated the expression of important inflammatory and bone-related genes in periodontal ligament cells, favoring a proinflammatory and an antiresorptive profile.

Cervical restoration and the amount of soft tissue coverage achieved by coronally advanced flap: a 2-year follow-up randomized-controlled clinical trial.

BACKGROUND: The aim of this study was to evaluate the 2-year follow-up success of the treatment of gingival recession associated with non-carious cervical lesions by a coronally advanced flap (CAF) alone or in combination with a resin-modified glass ionomer restoration (CAF+R). MATERIAL AND METHODS: Sixteen patients with bilateral Miller Class I buccal gingival recessions, associated with non-carious cervical lesions, were selected. The defects received either CAF or CAF+R. Bleeding on probing (BOP), probing depth (PD), relative gingival recession (RGR), clinical attachment level (CAL) and cervical lesion height (CLH) coverage were measured at the baseline and 6, 12 and 24 months after the treatment. RESULTS: Both groups showed statistically significant gains in CAL and soft tissue coverage. The differences between groups were not statistically significant in BOP, PD, RGR and CAL, after 2 years. The percentages of CLH covered were 51.57 +/- 17.2% for CAF+R and 53.87 +/- 12.6% for CAF (p>0.05). The estimated root coverage was 80.37 +/- 25.44% for CAF+R and 83.46 +/- 20.79% for CAF (p>0.05). CONCLUSION: Within the limits of the present study, it can be concluded that both procedures provide acceptable soft tissue coverage after 2 years, with no significant differences between the two approaches.