ß-TCP/S53P4 Scaffolds Obtained by Gel Casting: Synthesis, Properties, and Biomedical Applications.

The objective of this study was to investigate the osteogenic and antimicrobial effect of bioactive glass S53P4 incorporated into ß-tricalcium phosphate (ß-TCP) scaffolds in vitro and the bone neoformation in vivo. ß-TCP and ß-TCP/S53P4 scaffolds were prepared by the gel casting method. Samples were morphologically and physically characterized through X-ray diffraction (XRD) and scanning electron microscope (SEM). In vitro tests were performed using MG63 cells. American Type Culture Collection reference strains were used to determine the scaffold's antimicrobial potential. Defects were created in the tibia of New Zealand rabbits and filled with experimental scaffolds. The incorporation of S53P4 bioglass promotes significant changes in the crystalline phases formed and in the morphology of the surface of the scaffolds. The ß-TCP/S53P4 scaffolds did not demonstrate an in vitro cytotoxic effect, presented similar alkaline phosphatase activity, and induced a significantly higher protein amount when compared to ß-TCP. The expression of Itg ß1 in the ß-TCP scaffold was higher than in the ß-TCP/S53P4, and there was higher expression of Col-1 in the ß-TCP/S53P4 group. Higher bone formation and antimicrobial activity were observed in the ß-TCP/S53P4 group. The results confirm the osteogenic capacity of ß-TCP ceramics and suggest that, after bioactive glass S53P4 incorporation, it can prevent microbial infections, demonstrating to be an excellent biomaterial for application in bone tissue engineering.

Effect of radiotherapy on the differentiation and osteogenic activity of mesenchymal stem cells on dental implants / Efeito da radioterapia na diferenciação e atividade osteogênica de células-tronco mesenquimais em implantes dentários

Objective: to evaluate the differentiation and gene expression of transcripts related to osteogenesis in a primary culture of Mesenchymal Stem Cells (MSCs) derived from rat femurs submitted to radiotherapy and the installation of pure titanium implants. Material and Methods: fifty-four rats received titanium implants in both femurs and were divided into three groups: Control: implant surgery (C); Implant + immediate irradiation (IrI), and Implant + late irradiation (IrL). Euthanasia occurred 3, 14, and 49 days after surgery. The bone marrow MSCs from the femurs were isolated and cultivated. The cell viability, total protein content, alkaline phosphatase (ALP) activity, and the formation of mineralization nodules and cellular genotoxicity were analyzed. The gene expression of Alkaline Phosphatase (phoA), Collagen 1 (COL1), Runt-related transcription factor 2 (RUNX2), Osterix (OSX), Osteopontin (OPN), Integrin ß1(ITGB1), Bone Sialoprotein (BSP), Osteonectin (SPARC), Osteocalcin (Bglap), Transforming Growth Factor ß-type (TGF-ß), Granulocyte Macrophage Colony Stimulating Factor (GM-CSF), Interleukin-6 (IL-6), Apolipoprotein E (APOE) and Prostaglandin E2 synthase (PGE2) were evaluated by qRT- PCR. Results: ionizing radiation suppresses the gene expression of essential transcripts for bone regeneration, as well as cellular viability, as observed in the IrI and IrL groups. Conclusion: although this can lead to the loss of osseointegration and failure of the implant, the MSCs showed more activity at 49 days than at 3 and 14 days. (AU)

Objetivo: avaliar a diferenciação e expressão gênica de transcritos relacionados à osteogênese em cultura primária de MSCs derivadas de fêmures de ratos submetidos à radioterapia e instalação de implantes de titânio puro. Material e Métodos: cinquenta e quatro ratos receberam implantes de titânio em ambos os fêmures e foram divididos em três grupos: Controle: cirurgia de implante (C); Implante + irradiação imediata (IrI) e Implante + irradiação tardia (IrL). A eutanásia ocorreu 3, 14 e 49 dias após a cirurgia. As MSCs de medula óssea dos fêmures foram isoladas e cultivadas. Foram analisadas a viabilidade celular, teor de proteína total, atividade da fosfatase alcalina (ALP), formação de nódulos de mineralização e genotoxicidade celular. A expressão gênica de Fosfatase Alcalina (phoA), Colágeno 1 (COL1), fator de transcrição relacionado a Runt 2 (RUNX2), Osterix (OSX), Osteopontina (OPN), Integrina ß1 (ITGB1), Sialoproteína Óssea (BSP), Osteonectina (SPARC), Osteocalcina (Bglap), Fator de Crescimento Transformador tipo ß (TGF-ß), Fator Estimulante de Colônia de Granulócitos-Macrófagos (GM-CSF), Interleucina-6 (IL-6), Apolipoproteína E (APOE) e Prostaglandina E2 sintase (PGE2) foram avaliados por qRT-PCR. Resultados: a radiação ionizante suprime a expressão gênica de transcritos essenciais para a regeneração óssea, bem como a viabilidade celular, como observado nos grupos IrI e IrL. Conclusão:embora isso possa levar à perda da osseointegração e falha do implante, as MSCs apresentaram maior atividade aos 49 dias do que aos 3 e 14 dias (AU)

Bioactivity of an Experimental Dental Implant with Anodized Surface.

BACKGROUND: Several studies proved that anodic oxidation improves osseointegration. This study aimed to optimize osseointegration through anodization in dental implants, obtaining anatase phase and controlled nanotopography. METHODS: The division of the groups with 60 titanium implants was: control (CG); sandblasted (SG); anodized (AG): anodized pulsed current (duty cycle 30%, 30 V, 0.2 A and 1000 Hz). Before surgery, surface characterization was performed using Atomic Force Microscopy (AFM), Scanning Electron Microscopy (SEM), X-ray Dispersive Energy Spectroscopy (EDS) and Raman Spectroscopy. For in vivo tests, 10 New Zealand white rabbits received an implant from each group. The sacrifice period was 2 and 6 weeks (n = 5) and the specimens were subjected to computed microtomography (µCT) and reverse torque test. RESULTS: AFM and SEM demonstrated a particular nanotopography on the surface in AG; the anatase phase was proved by Raman spectroscopy. In the µCT and in the reverse torque test, the AG group presented better results than the other groups. CONCLUSION: The chemical composition and structure of the TiO2 film were positively affected by the anodizing technique, intensifying the biological characteristics in osseointegration.

Efficacy of fluoride varnishes with different compositions on white spot lesions remineralization / Eficácia de vernizes fluoretados com diferentes composições na remineralização de manchas brancas

Objective: To evaluate the efficacy of different fluoride varnishes on white spot lesions (WSL) remineralization. Material and Methods: Polished bovine enamel specimens were obtained (n = 60) and had their initial surface Knoop microhardness (SMH) determined. WSL were created and the SMH was measured again. Then, specimens were allocated into six groups: C ­ Control (without varnish); BF ­ Bifluorid 12 (6% NaF + 6% CaF2); DP ­ Duraphat (5% NaF); PF ­ Profluorid (5% NaF); FP - Fluor Protector (0.2% NaF + 0.9% difluorsilane); CW - Clinpro White Varnish (5% NaF + 5% TCP). After varnishes application, specimens were immersed in artificial saliva for 24 h. Then, pH-cycling was performed for 8 days and SMH was measured. Data were analyzed by one-way ANOVA and Tukey's test. Results: Non-significant differences were observed among the groups at baseline (p = 0.187) and after WSL formation (p = 0.999). After treatments, significant differences were observed among the groups (p = 0.001). Mean % of alteration (SD) and results of Tukey test were: C- 92.40 (12.10)a; PF- 88.66 (10.66)a; FP- 85.90 (14.49)ab; BF- 67.85 (17.86)bc; CW- 66.60 (18.48)c; DP- 58.62 (8.69)c. Conclusion: Bifluorid 12, Clinpro White Varnish, and Duraphat showed higher efficacy than artificial saliva in promoting the remineralization of WSL, nevertheless, none of the treatments were able to recover sound enamel baseline microhardness (AU)

Objetivo: Avaliar a eficácia de diferentes vernizes fluoretados na remineralização de lesões de mancha branca (LMB). Material e métodos: Espécimes de esmalte bovino polido (n = 60) foram submetidos à análise de microdureza superficial Knoop (KMH) inicial. Foram então criadas LMB artificialmente e os espécimes foram alocados em seis grupos: C ­ Controle (sem aplicação de verniz); BF ­ Bifluorid 12 (6% NaF + 6% CaF2); DP ­ Duraphat (5% NaF); PF ­ Profluorid (5% NaF); FP - Fluor Protector (0.2% NaF + 0.9% difluorsilano); CW - Clinpro White Varnish (5% NaF + 5% TCP). Após a aplicação dos vernizes, os espécimes ficaram imersos em saliva artificial por 24h e uma ciclagem de pH foi realizada por 8 dias. Após a ciclagem, KMH final foi realizada. Os dados foram analisados por ANOVA e teste de Tukey (5%). Resultados: Não foi observada diferença significante para os grupos após a KHM inicial (p = 0.187) e após a formação de LMB (p = 0.999). Após os tratamentos, diferenças significativas foram observadas entre os grupos (p = 0.001). Valores de média de % de alteração superficial (desvio-padrão) e resultados do teste de Tukey foram: C- 92.40 (12.10)a; PF- 88.66 (10.66)a; FP- 85.90 (14.49)ab; BF- 67.85 (17.86)bc; CW- 66.60 (18.48)c; DP- 58.62 (8.69)c. Conclusão: Os vernizes Bifluorid 12, Clinpro White Varnish e Duraphat apresentaram maior eficácia na remineralização das LMB quando comparados à saliva artificial, entretanto, nenhum dos produtos testados foi capaz de recuperar os valores iniciais de microdureza. (AU)

The influence of adrenergic blockade in rats with apical periodontitis under chronic stress conditions.

OBJECTIVE: To investigate the influence of chronic stress and adrenergic blockade in a rat model of apical periodontitis. METHODS: Thirty-two Wistar rats were submitted to an animal model of periapical lesion and randomly divided into 4 groups (n = 8): no stress (NS); stress + saline solution (SS); stress + ß-adrenergic blocker (Sß); stress + α-adrenergic blocker (Sα). The SS, Sß and Sα groups were submitted to an animal model of chronic stress for 28 days and received daily injections of saline solution, propranolol (ß adrenergic blocker) and phentolamine (α adrenergic blocker), respectively. After 28 days the animals were euthanized and the following analyses were carried out: a) serum corticosterone levels through Radioimmunoassay; b) measurement of serum levels of IL-1B, IL-6, IL-10 and IL-17 by enzyme-linked immunosorbent assay (ELISA); c) volume of periapical bone resorption by micro-computed tomography; d) histomorphometric analysis by staining with hematoxylin and eosin; e) expression of ß-AR, α-AR, receptor activator of nuclear factor kappa-B ligand (RANKL) and osteoprotegerin (OPG) by immunohistochemistry; f) tartrate-resistant acid phosphatase (TRAP) staining; g) ex-vivo cytokine release followed by the stimulation with LPS in superfusion system, by ELISA. RESULTS: SS group displayed significantly higher corticosterone levels than NS group (non-stressed). Higher IL-1ß serum level was observed in the NS group (p < .05); compared to all stressed groups. Other cytokines were present in similar amounts in the serum of all groups. All groups presented similar periapical lesions. All groups presented moderate inflammatory infiltrate, without statistically significant differences between them. No differences were observed regarding ß-AR, α-AR, Rank-L and OPG expression. The number of TRAP-positive cells was significantly decreased in the groups that received daily injections of adrenergic blockers. The IL-1ß release followed LPS stimulation was significantly suppressed when the superfusion media contained propranolol (p < .05). Perfusion containing phentolamine induced a greater release of IL-10. TGF-ß was significantly suppressed by phentolamine perfusion in the NS group (p < .05). CONCLUSIONS: Chronic stress can significantly change the inflammatory cytokines release. Rank-L/OPG system and periapical lesion volume were not affected following the current method applied. The administration of adrenergic blockers was not able to modulate the inflammatory response but presented effectivity in reducing the number of osteoclasts in the periapical region.

Effect of DLC Films with and without Silver Nanoparticles Deposited On Titanium Alloy.

Diamond-like carbon (DLC) film is a biocompatible hard coating material that can prevent the leaching of metal ions. This study evaluates the structural characteristics of DLC, with and without silver nanoparticles, deposited by plasma (PECVD) on titanium alloy (Ti-6Al-4V) and bone formation in contact with DLC films. Sixty Ti-6Al-4V samples were used divided in: uncoated, coated with DLC and coated with DLC-Ag. After structural characterization, samples were fixed bilaterally at the rabbit's mandible. After 15 and 90 days, samples were characterized again and bone formation in the area was analyzed by histomorphometry. Statistical analysis was performed by two-way ANOVA. Both the DLC and DLC-Ag films were firmly adhered and showed a high electrical resistance without significant changes in the Raman spectrum after in vivo integration. After 15 days, there were immature bone trabeculae in the interface and partially covering the surface. After 90 days, mature bone filled the interface and coved the surface. There was no statistically significant difference among the three groups in both periods. In conclusion, osseointegration with DLC, DLC-Ag and uncoated Ti-6Al-4V is similar. However, DLC and DLC-Ag coverings have the advantage of electrical insulation and can presumably control bacterial activity and ion leaching.

Silicon-based film on the yttria-stabilized tetragonal zirconia polycrystal: Surface and shear bond strength analysis.

AIM: To analyze the effect of a silicon (Si)-based film deposited on yttria-stabilized tetragonal zirconia polycrystal (Y-TZP) on the topography and bond strength of resin cement. METHODS: Specimens of zirconia were obtained and randomly divided into 4 groups, according to surface treatment: polished group (PG) zirconia; sandblasted group (SG) zirconia with aluminum oxide (100 µm); after polished, zirconia was coated with Si-based film group (SiFG); and after sandblasted, zirconia was coated with Si-based film group (SiFSG). The Si-based films were obtained through plasma-enhanced chemical vapor deposition. Surface roughness and contact angle analysis were performed. Resin cement cylinders were built up on the treated surface of blocks, after applying Monobond-S. The specimens were submitted to thermocycling aging and shear bond strength testing. The Kruskal-Wallis and Mann-Whitney U-tests were performed. RESULTS: There were significant differences between the surface treatments for each roughness parameter measured. Si-based film increased roughness and decreased the contact angle. Si-based film groups also demonstrated significantly lower bond strength values. CONCLUSION: Si-based film produced using plasma deposition provided lower bond strength to resin cement compared with conventional treatment; however, the film deposition reduced the contact angle and improved roughness, favorable properties in the long way to prepare an optimum material.

Diamond-like carbon films over reconstructive TMJ prosthetic materials: Effects in the cytotoxicity, chemical and mechanical properties.

Increasingly more young patients have been submitted to reconstruction of the Temporomandibular Joint (TMJ), so, the prostheses must to present more functional longevity. OBJECTIVE: To evaluate the effect of diamond-like carbon film (DLC) over titanium alloy (Ti6Al4V) and polyethylene (UHWPE) samples, their mechanical and chemical properties and cellular cytotoxicity. METHODS: Titanium and UHWPE specimens, with 2.5 cm in diameter and 2 mm thickness were coated through plasma enhanced chemical vapor deposition (PECVD) with DLC or DLC doped with silver (DLC-Ag). Scanning electron microscopy (SEM) morphological analysis, Energy-dispersive spectroscopy (EDS) chemical analysis, scratching test, mechanical fatigue test, surface roughness analysis, and cellular cytotoxicity were performed. Data were statistically analyzed using one-way ANOVA (p < 0.05) or two-way ANOVA and multiple comparison Tukey test. RESULTS: In the SEM analysis, morphological differences were observed on substrates after DLC deposition. The film chemically modified the substrate surfaces, according to the EDS analysis. The initial critical load failure occurred at 6.1 N for DLC and 9.7 N for the DLC-Ag film. The DLC film deposition over the polyethylene promoted a decrease in the polymer's damaged area after mechanical fatigue cycling. The cytotoxicity analysis demonstrated less biocompatibility in experimental groups, when compared to control, however, increased biocompatibility was observed, at 10 days, in all groups. CONCLUSION: The diamond-like carbon coating enhanced the chemical and mechanical properties from substrates, however modified biological interaction course of the titanium alloy (Ti6Al4V) and polyethylene (UHWPE) samples. Parameters for film deposition remain to be improved in order to obtain best biocompatibility.

Correlation Between Volume of Root Canal, Cultivable Bacteria, Bacterial Complexes and Endotoxins in Primary Infection.

This clinical study was conducted to correlate the levels of endotoxins and culturable bacteria found in primary endodontic infection (PEI) with the volume of root canal determined by using Cone Beam Computed Tomography (CBCT); and to evaluate the bacterial diversity correlating with clinical features. Twenty patients with PEI were selected and clinical features were recorded. The volume (mm3) of root canal was determined by CBCT analysis. Root canal samples were analyzed by using kinetic LAL-assay test to determine the levels of endotoxins and anaerobic technique to determine the bacterial count (CFU/mL). DNA was extracted from all samples to determine bacterial diversity and quantified by using Checkerboard-DNA-DNA- Hybridization. Culturable bacteria and endotoxins were detected in 100% of the root canal samples. Linear regression analysis revealed a correlation between root canal volume and presence of anaerobic bacteria (p<0.05). Positive correlations were found between bacteria species and presence of different clinical features (p<0.05). After grouping the bacteria species into bacterial complexes, positive associations were found between green, orange and red complexes with presence of sinus tract (p<0.05). This clinical study revealed that larger root canals hold higher levels of culturable bacteria in PEI. Thus, the interaction of different virulent bacteria species in complexes seems to play an important role in the development of clinical features.

Bilateral mandibular buccal bifurcation cyst: a case report emphasizing the role of imaging examination in the diagnosis

A mandibular buccal bifurcation cyst is an inflammatory cyst that usually occurs on the buccal aspect of the permanent mandibular first molar of children. This lesion is diagnosed by an association of radiographic, clinical, and histological features. We report a bilateral case of mandibular buccal bifurcation cyst and discuss the main findings of this entity. A 7-year-old girl presented pain and delayed dental eruption in the posterior mandibular region. A cone beam computed tomography was performed and revealed hypodense lesions involving the crown and root of the mandibular first molars, with expansion of the buccal cortical and lingual tilting of the molar roots. A biopsy was carried out, and the common features of an inflammatory odontogenic cyst were histologically observed. The final diagnosis was bilateral mandibular buccal bifurcation cyst. Clinicians need to be aware of this diagnostic possibility in cases of mandibular cysts in children­especially when bilateral­to perform the correct treatment, which should not involve the extraction of the affected tooth.

CaP Coating and Low-Level Laser Therapy to Stimulate Early Bone Formation and Improve Fixation of Rough Threaded Implants.

PURPOSE: This study aimed to compare in vivo osteogenesis on rough threaded dental implants with and without calcium phosphate (CaP) coating deposition, alone or in association with low-level laser therapy (LLLT) by gallium aluminum arsenide. MATERIAL AND METHODS: Four groups were studied: G1: implant; G2: implant + CaP coating; G3: implant + LLLT; and G4: implant + CaP coating + LLLT. LLLT was applied for 7 days at the surgical site before and after placing the implant. Topographic characterization was performed before surgery using scanning electron microscopy and energy dispersion spectrophotometry. Bone-implant contact (BIC) was measured after 1, 2, and 6 weeks and reverse torque after 6 weeks. In short periods, G2, G3, and G4 showed significantly greater BIC than G1 (P < 0.05), but no difference in BIC was observed at 6 weeks. However, the values for the removal torque test at 6 weeks were higher in G2 and G4 (P < 0.05). CONCLUSION: Both CaP coating alone and using LLLT induce cellular stimulation and improve BIC in short-term healing, resulting in higher implant fixation, and should be considered in clinical practice due to their low cost and high effectiveness.

Colonization of oropharynx and lower respiratory tract in critical patients: Risk of ventilator-associated pneumonia.

OBJECTIVE: To investigate the microbial diversity existing in oral cavity and respiratory tract samples (from mini-bronchoalveolar lavage (BAL), endotracheal aspirate, and orotracheal tube) of patients on mechanical ventilation by using the checkerboard DNA-DNA hybridisation. Also, the study aimed to evaluate whether the microbial profile in the oral cavity is found in respiratory tract samples, at different periods of mechanical ventilation time (12h, 48h, 96h) in attempt to identification of relationship between VAP (ventilator-associated pneumonia) and bacterial species studied. The last objective was to analyses correlation between blood cultures and VAP. DESIGN: The samples were collected from ten patients in intensive care unit with medical prescription of orotracheal intubation and mechanical ventilation. Clinical data were tabulated and blood cultures were performed according medical indication. For checkerboard samples collection, chosen sites were the dorsal side of the tongue and gingival sulcus at 12h, 48h, 96h, BAL, at 12h, endotracheal aspirate at 48 and 96h, and orotracheal tube at extubation time, when feasible. RESULTS: It was possible to identify the presence of bacterial species in mouth and in the BAL/endotracheal aspirate. The data demonstrated an increase in the quantity of bacterial associated with prolonged use of mechanical ventilation (48 and 96h). CONCLUSIONS: Bacterial species may migrate rapidly from mouth and upper airways during orotracheal intubation which contributes to the pathogenesis of VAP. There were associations between VAP and Enterococcus faecalis, Fusobacterium periodonticum, Gemella morbillorum, Neisseria mucosa, Propionibacterium acnes, Prevotella melaninogenica, Streptococcus oralis, Streptococcus sanguinis, Treponema denticola, Treponema socransckii, and Veillonella parvula.

Expression of BMP II by human osteoblasts cultivated on dense or porous titanium / Expressão da BMP II por osteoblastos humanos cultivados sobre titânio denso ou poroso

Objective: Modifications of titanium have been described as an important tool improving bone repair and boneimplant contact. The aim of this research was to quantified the expression of the morphogenetic bone protein II (BMP II) produced by human cells with osteoblast differentiation, after cultured over dense or porous samples of pure titanium grade II. Material and Methods: The experimental groups were: control group, dense titanium, porosity of 33.79% and porosity of 41,79% (n=36). The samples were produced by powder metallurgy technique. Mesenquimal steam cells isolated from alveolar bone of healthy donors were stimulated to differentiate, assuming an osteoblastic phenotype, by supplemented medium and plated over the samples. After 7 and 14 days, the RNA was collected to perform reverse transcriptase polymerase chain reaction (RT-PCR) in real time. Data was analysed by t-Student and ANOVA tests. The porosity, the pore morphology and interconnection were evaluated by Scanning Electron Microscopy (SEM). Results: Total porosity (obtained after apply dimensions and density formulas) and surface porosity (SEM) presented significant differencesamong the groups. For the group of total porosity of 33.79%, the superficial porosity was 32.5% (± 7.74%) and for the group of 41.79%, the superficial porosity was 37.4% (± 7.95%), significantly lower. The expression of BMP II was similar in all groups. Conclusion: The present study demonstrated that powder metallurgy has a reduced ability to standardize the porosity in the samples and that the porosity does not interfere in the cellular response of BMP II production, an important inducer of osteoblastic differentiation. (AU)

Objetivo: As modificações do titânio são descritas como importantes ferramentas na melhora do reparo ósseo no contato osso implante. O objetivo deste estudo foi quantificar a expressão da proteína óssea morfogênica II (BMP II) por células humanas com diferenciação osteoblastica, quando cultivadas sobre amostras de titânio puro grau II, denso ou poroso. Material e Métodos: Os grupos experimentais foram: controle, titânio denso, titânio de maior porosidade e titânio de menor porosidade, sendo que, as amostras foram confeccionadas pela técnica da metalurgia do pó. As células isoladas de doadores saudáveis foram plaqueadas sobre as amostras. Após 7 e 14 dias, o RNA foi extraído das células. A qualidade e integridade do RNA foram analisadas qualitativamente por eletroforese e quantitativamente por espectrofotômetro. O cDNA foi confeccionado e a foi utilizada técnica de reação em cadeia da polimerase (PCR) em tempo real. Os dados foram utilizados para quantificação relativa, e o gene constitutivo foi a BetaActina. A morfologia e a interligação dos poros foram comprovadas por Microscopia Eletrônica de Varredura (MEV). Resultados: A porosidade superficial (MEV) teve diferença significativa em relação a porosidade obtida analisando-se volume e massa das amostras. Para o grupo 3,79%, a superficial foi de 32,5% (±7,74%) e para o grupo 41,79% a porosidade superficial foi de 37,4% (±7,95%), significativamente menor. A expressão da BMP II foi semelhante em todos os grupos. Conclusão: Concluiu-se a metalurgia do pó tem reduzida capacidade de padronização da porosidade das amostras por ela confeccionas e que a porosidade não interfere na resposta celular de produção da BMP II, importante indutor de diferenciação osteoblastica.(AU)

Analysis of indicators of osteogenesis, cytotoxicity and genotoxicity of an experimental ß-TCP compared to other bone substitutes / Análise dos indicadores da osteogênese, citotoxicidade e genotoxicidade de um ß-TCP experimental comparado a outros substitutos ósseos

The aim of this study was to evaluate the indicators of osteogenesis, cytotoxicity and genotoxicity of an experimental beta tri-calcium phosphate (experimental ß-TCP) compared with two other bone substitutes: bovine hydroxyapatite (HA) (Bio-Oss® - Geistlich) and beta tri-calcium phosphate (ß-TCP - Bionnovation). The cell viability and genotoxicity were measured by MTT and MNT assay, respectively. The indicators of osteogenesis were analyzed by alkaline phosphatase activity, total protein content, and calcium deposition. The MTT and MNT assay showed that none of the tested materials was cytotoxic nor genotoxic. Concerning the indicators of osteogenesis, it was observed that cells in contact with all the materials were able to induce the osteogenesis and this process was influenced by the period of the cell culture in contact with bone substitutes. Based on the results of this study, it was concluded that this experimental ß-TCP appears to be a promising material as a bone substitute.

O objetivo deste estudo foi avaliar os indicadores da osteogênese, citotoxicidade e genotoxicidade de um beta-tricálcio fosfato (ß-TCP experimental) comparado com dois outros substitutos ósseos : Hidroxiapatita Bovina (HA) (Bio-Oss® - Geistlich) e beta-tricálcio fosfato (ß-TCP - Bionnovation). A viabilidade celular e genotoxicidade foram mensuradas pelos ensaios MTT e MNT, respectivamente. Os indicadores da osteogênese foram analisados pela atividade de fosfatase alcalina (ALP), conteúdo de proteína total, e deposição de cálcio. Os ensaios MTT e MNT mostraram que nenhum dos materiais testados foi citotóxico ou genotóxico. Em relação aos indicadores da osteogênese, foi observado que as células em contato com todos os materiais foram capazes de induzir a osteogênese, e que esse processo foi influenciado pelo período da cultura celular em contato com os substitutos ósseos. Baseado nos resultados desse estudo, conclui-se que este ß-TCP experimental parece ser um material promissor para ser utilizado como substituto ósseo.

Titanium-35niobium alloy as a potential material for biomedical implants: In vitro study.

Research on new titanium alloys and different surface topographies aims to improve osseointegration. The objective of this study is to analyze the behavior of osteogenic cells cultivated on porous and dense samples of titanium-niobium alloys, and to compare them with the behavior of such type of cells on commercial pure titanium. Samples prepared using powder metallurgy were characterized using scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), X-ray diffraction (XRD), and metallographic and profilometer analyses. Osteogenic cells from newborn rat calvaria were plated over different groups: dense or porous samples composed of Ti or Ti-35niobium (Nb). Cell adhesion, cell proliferation, MTT assay, cell morphology, protein total content, alkaline phosphatase activity, and mineralization nodules were assessed. Results from XRD and EDS analysis confirmed the presence of Ti and Nb in the test alloy. Metallographic analysis revealed interconnected pores, with pore size ranging from 138 to 150µm. The profilometer analysis detected the greatest rugosity within the dense alloy samples. In vitro tests revealed similar biocompatibility between Ti-35Nb and Ti; furthermore, it was possible to verify that the association of porous surface topography and the Ti-35Nb alloy positively influenced mineralized matrix formation. We propose that the Ti-35Nb alloy with porous topography constitutes a biocompatible material with great potential for use in biomedical implants.

Osteoblast response to porous titanium and biomimetic surface: In vitro analysis.

OBJECTIVE: This study analyzed the behavior of human osteoblasts cultured on porous titanium specimens, with and without biomimetic treatment, compared to dense titanium. DESIGN: The experiment had seven groups: Group 1: cells cultured on polystyrene of culture plate wells; Group 2: cells cultured on dense titanium specimen; Group 3: specimen with 33.79% of pores; Group 4: 41.79% of pores; Groups 5, 6 and 7: specimens similar to groups 2, 3 and 4, yet with biomimetic treatment. Real time-polymerase chain reaction with reverse transcription of the following genes was performed: prostaglandin E2 synthase, integrin ß1, osterix, Runx2, Interleukin 6, macrophage colony stimulating factor, apolipoprotein E and others. The study achieved data on cell adhesion, growth and viability, total protein content, alkaline phosphatase activity and quantity of mineralized nodule formations. Data were statistically evaluated. RESULTS: Adherent cells and alkaline phosphatase activity were similar in titanium specimens, regardless of the groups. Biomimetic treatment reduced the total protein activity and the viability of tested cells. Most tested genes had statistically similar expression in all groups. CONCLUSION: The tested porosities did not cause alterations in osteoblast behavior and the biomimetic treatment impaired the biocompatibility of titanium causing cytotoxicity.

Immunolocalization of markers for bone formation during guided bone regeneration in osteopenic rats

Objective The aim of this paper was to evaluate the repair of onlay autogenous bone grafts covered or not covered by an expanded polytetrafluoroethylene (e-PTFE) membrane using immunohistochemistry in rats with induced estrogen deficiency. Material and Methods Eighty female rats were randomly divided into two groups: ovariectomized (OVX) and with a simulation of the surgical procedure (SHAM). Each of these groups was again divided into groups with either placement of an autogenous bone graft alone (BG) or an autogenous bone graft associated with an e-PTFE membrane (BGM). Animals were euthanized on days 0, 7, 21, 45, and 60. The specimens were subjected to immunohistochemistry for bone sialoprotein (BSP), osteonectin (ONC), and osteocalcin (OCC). Results All groups (OVX+BG, OVX+BMG, SHAM+BG, and SHAM+BMG) showed greater bone formation, observed between 7 and 21 days, when BSP and ONC staining were more intense. At the 45-day, the bone graft showed direct bonding to the recipient bed in all specimens. The ONC and OCC showed more expressed in granulation tissue, in the membrane groups, independently of estrogen deficiency. Conclusions The expression of bone forming markers was not negatively influenced by estrogen deficiency. However, the markers could be influenced by the presence of the e-PTFE membrane. .

The RANK/ RANKL/ OPG interaction in the repair of autogenous bone grafts in female rats with estrogen deficiency.

The aim of this study was to evaluate the resorption process during the repair of autogenous bone grafts with or without coverage by an expanded polytetrafluoroethylene (e-PTFE) membrane in female rats with estrogen deficiency using the immunohistochemical technique. Eighty female rats were randomly divided into two groups (OVX and SHAM). The 40 female rats in the OVX group were subjected to ovariectomy, and the 40 female rats in the SHAM group were subjected to simulated ovariectomy. The two groups were further divided in subgroup E, which was subjected to surgery for placement of autogenous bone graft (ABG), and subgroup ME, in which the ABG was covered with an e-PTFE membrane. The animals were killed at 0, 7, 21, 45 and 60 days. The specimens were analyzed using immunohistochemistry for the bone resorption markers RANK, RANK-L and Osteoprotegerin (OPG). A higher remodeling rate was observed at 7 and 21 days after the autogenous bone grafts, when the markers were more intensely expressed. At the final time point, the specimens presented similar characteristics to those observed at the initial time point. The expression of immunohistochemical markers was not altered by the estrogen deficiency. The presence of the e-PTFE membrane delayed the bone resorption process, influencing the immunohistochemical expression of markers.

Immunolocalization of markers for bone formation during guided bone regeneration in osteopenic rats.

OBJECTIVE: The aim of this paper was to evaluate the repair of onlay autogenous bone grafts covered or not covered by an expanded polytetrafluoroethylene (e-PTFE) membrane using immunohistochemistry in rats with induced estrogen deficiency. MATERIAL AND METHODS: Eighty female rats were randomly divided into two groups: ovariectomized (OVX) and with a simulation of the surgical procedure (SHAM). Each of these groups was again divided into groups with either placement of an autogenous bone graft alone (BG) or an autogenous bone graft associated with an e-PTFE membrane (BGM). Animals were euthanized on days 0, 7, 21, 45, and 60. The specimens were subjected to immunohistochemistry for bone sialoprotein (BSP), osteonectin (ONC), and osteocalcin (OCC). RESULTS: All groups (OVX+BG, OVX+BMG, SHAM+BG, and SHAM+BMG) showed greater bone formation, observed between 7 and 21 days, when BSP and ONC staining were more intense. At the 45-day, the bone graft showed direct bonding to the recipient bed in all specimens. The ONC and OCC showed more expressed in granulation tissue, in the membrane groups, independently of estrogen deficiency. CONCLUSIONS: The expression of bone forming markers was not negatively influenced by estrogen deficiency. However, the markers could be influenced by the presence of the e-PTFE membrane.

Pleomorphic adenoma versus basal cell adenoma: an immunohistochemical analysis with B-catenin / Adenoma pleomorfo versus adenoma de células basais: Análise imunohistoquímica com B-catenina

Objective: The objective of this study was to investigate the distribution of β-catenin in pleomorphic adenomas and the basal cell adenomas to clarify its possible role in the etiopathogenesis of these two lesions. Material and Methods: The expression of β catenin (BD Transduction Laboratories) was analyzed by immunohistochemistry in formalinfixed, paraffin-embedded specimens by the avidinbiotin- peroxidase complex method in 10 pleomorphic adenomas and 2 basal cell adenomas. The specimens were analyzed taking into account intensity, distribution and association with myoepithelial cells. Results: The results showed that all cases of pleomorphic adenomas exhibited membranous and cytoplasmic immunostaining and the 2 cases of basal cell adenomas displayed nuclear staining. Higher β-catenin index rates were seen mainly in ductal structures of pleomorphic adenomas and in the nuclei of myoepithelial stromal and myoepithelial cells of solid clusters in basal cell adenomas. Conclusion: In conclusion, this immunohistochemical study may suggests the different degree of differentiation of the myoepithelial cells in these two tumors

Objetivos: O objetivo deste estudo foi investigar a distribuição da β-catenina em adenomas pleomorfos e os adenomas de células basais para esclarecer o seu possível papel na etiopatogenia dessas duas lesões. Material e Métodos: A expressão de β-catenina (BD-Transdução Laboratories) foi analisada por imunohistoquímica em espécimes parafinados fixados em formalina pelo método avidina-biotinaperoxidase em 10 adenomas pleomórficos e 2 adenomas de células basais. Os espécimes foram analisados levando-se em conta a intensidade, quantidade de células marcadas e distribuição em estruturas morfológicas. Resultados: Os resultados mostraram que todos os casos de adenomas pleomorfos exibiram imuno marcação membranosa e citoplasmática e os 2 casos de adenomas de células basais exibiram coloração nuclear. Marcação mais intensa de β-catenina foi vista principalmente em estruturas ductais de adenomas pleomorfos e nos núcleos de células mioepiteliais de estruturas tubulares e trabeculares em adenomas de células basais. Conclusão: Concluindo, este estudo imunohistoquímico sugere distintos graus de diferenciação das células mioepiteliais em cada um destes dois tumores