Influence of melatonin associated with the Bio-Gide® membrane on osteoblast activity: an in vitro Study.

Melatonin (MLT) is a hormone responsible for regulating several physiological processes. It has been shown that MLT can be an important mediator in bone formation and stimulation, promoting osteoblast differentiation. In clinical practice, in tissue regeneration procedures, it is necessary to use membranes or barriers, associated with biomaterials, or not. The aim of this in vitro study was to assess the effect of melatonin on the activity of osteoblastic cells, associated, or not, with a resorbable collagen membrane (Bio-Gideä). For this, mice-derived pre-osteoblastic cells MC3T3 obtained from the ATCC (American Type Culture Collection) were used. Cultured cells were subject to the following treatments: MLT with a concentration of 1mM, a Bio-Gideä membrane and a membrane associated with MLT (Bio-Gideä + MLT). Proliferation and cell viability assays and protein lysate (ELISA test) quantification for the BMP-2 protein were carried out, in periods of 72 hours, 7 days and 10 days. After analyzing the data (one-way ANOVA, alpha=5%) it was observed that when MLT was used in isolation, there was an increase in cell proliferation and viability in osteoblastic cells (p<0.05). But, when MLT was associated with resorbable membranes, there was an inverse behavior, both in terms of proliferation and viability (p<0.05). In the case of the ELISA test, no secretion of BMP-2 was detected in any of the analyzed groups. It is concluded that MLT has a stimulatory effect on osteoblasts, but, when associated with Bio-Gideä resorbable membranes, it does not show any viable action in osteoblastic cell stimulation.

A melatonina (MLT) é um hormônio responsável pela regulação de diversos processos fisiológicos no nosso organismo. Tem sido demonstrado que a melatonina possa ser um importante mediador na formação e estimulação óssea, promovendo a diferenciação dos osteoblastos. Clinicamente, para o procedimento de regeneração tecidual, faz-se necessário a utilização de membranas ou barreiras, associadas ou não a biomateriais. Assim, o objetivo deste estudo in vitro foi avaliar o efeito da melatonina na atividade de células osteoblásticas, associada ou não a uma membrana de colágeno reabsorvível (Bio-Gide®). Para isto foram utilizadas células pré-osteoblásticas MC3T3 do ATCC (American Type Culture Collection), de camundongos. As células em cultura foram submetidas aos seguintes tratamentos: MLT na concentração de 1mM, membrana Bio Gide® e membrana associada à MLT (Bio-Gide® + MLT). Foram realizados os ensaios de proliferação e viabilidade celular e quantificação do lisado proteico (teste ELISA), para a proteína BMP-2, nos períodos de 72 horas, 7 e 10 dias. Após a análise dos dados (ANOVA um critério, alfa=5%) pode-se observar que a MLT quando utilizada sozinha, resultou em um aumento na proliferação e viabilidade celular nas células osteoblásticas (p<0,05). Entretanto, quando a MLT foi associada à membrana reabsorvível foi observado um comportamento inverso, tanto na proliferação quanto na viabilidade (p<0,05). Para o teste ELISA realizado, não houve secreção detectável de BMP-2 para nenhum grupo analisado. Conclui-se que a melatonina possui uma ação estimuladora nos osteoblastos, mas quando associada à membrana reabsorvível Bio-Gide®, não demonstra uma ação viável na estimulação de células osteoblásticas.

Influence of melatonin associated with the Bio-Gide® membrane on osteoblast activity: an in vitro Study / Influência da melatonina associada à membrana bio gide® na atividade de osteoblastos: um estudo in vitro

ABSTRACT Melatonin (MLT) is a hormone responsible for regulating several physiological processes. It has been shown that MLT can be an important mediator in bone formation and stimulation, promoting osteoblast differentiation. In clinical practice, in tissue regeneration procedures, it is necessary to use membranes or barriers, associated with biomaterials, or not. The aim of this in vitro study was to assess the effect of melatonin on the activity of osteoblastic cells, associated, or not, with a resorbable collagen membrane (Bio-Gideä). For this, mice-derived pre-osteoblastic cells MC3T3 obtained from the ATCC (American Type Culture Collection) were used. Cultured cells were subject to the following treatments: MLT with a concentration of 1mM, a Bio-Gideä membrane and a membrane associated with MLT (Bio-Gideä + MLT). Proliferation and cell viability assays and protein lysate (ELISA test) quantification for the BMP-2 protein were carried out, in periods of 72 hours, 7 days and 10 days. After analyzing the data (one-way ANOVA, alpha=5%) it was observed that when MLT was used in isolation, there was an increase in cell proliferation and viability in osteoblastic cells (p<0.05). But, when MLT was associated with resorbable membranes, there was an inverse behavior, both in terms of proliferation and viability (p<0.05). In the case of the ELISA test, no secretion of BMP-2 was detected in any of the analyzed groups. It is concluded that MLT has a stimulatory effect on osteoblasts, but, when associated with Bio-Gideä resorbable membranes, it does not show any viable action in osteoblastic cell stimulation.

RESUMO A melatonina (MLT) é um hormônio responsável pela regulação de diversos processos fisiológicos no nosso organismo. Tem sido demonstrado que a melatonina possa ser um importante mediador na formação e estimulação óssea, promovendo a diferenciação dos osteoblastos. Clinicamente, para o procedimento de regeneração tecidual, faz-se necessário a utilização de membranas ou barreiras, associadas ou não a biomateriais. Assim, o objetivo deste estudo in vitro foi avaliar o efeito da melatonina na atividade de células osteoblásticas, associada ou não a uma membrana de colágeno reabsorvível (Bio-Gide®). Para isto foram utilizadas células pré-osteoblásticas MC3T3 do ATCC (American Type Culture Collection), de camundongos. As células em cultura foram submetidas aos seguintes tratamentos: MLT na concentração de 1mM, membrana Bio Gide® e membrana associada à MLT (Bio-Gide® + MLT). Foram realizados os ensaios de proliferação e viabilidade celular e quantificação do lisado proteico (teste ELISA), para a proteína BMP-2, nos períodos de 72 horas, 7 e 10 dias. Após a análise dos dados (ANOVA um critério, alfa=5%) pode-se observar que a MLT quando utilizada sozinha, resultou em um aumento na proliferação e viabilidade celular nas células osteoblásticas (p<0,05). Entretanto, quando a MLT foi associada à membrana reabsorvível foi observado um comportamento inverso, tanto na proliferação quanto na viabilidade (p<0,05). Para o teste ELISA realizado, não houve secreção detectável de BMP-2 para nenhum grupo analisado. Conclui-se que a melatonina possui uma ação estimuladora nos osteoblastos, mas quando associada à membrana reabsorvível Bio-Gide®, não demonstra uma ação viável na estimulação de células osteoblásticas.

Influence of abutment angulation on loss of prosthetic abutment torque under mechanical cycling.

STATEMENT OF PROBLEM: Internal conical connections provide mechanical stability for the prosthetic abutment and implant connection. However, some clinical situations require the use of angled prosthetic abutments that may increase stress on supportive implants by difference force vectors under cyclic loading. PURPOSE: The purpose of this in vitro study was to measure the screw loosening values of prosthetic abutments with internal conical connections (indexed and nonindexed) having different angles under mechanical cycling. MATERIAL AND METHODS: Thirty-six implants (4.0×13 mm, Titamax) with internal conical connections and their respective universal prosthetic abutments (n=36, 3.5×3.3 mm) were divided into indexed and nonindexed groups (n=18) with abutment inclinations of 0 (straight), 17, and 30 degrees. An insertion torque of 15 Ncm was applied according to the manufacturer's specifications. The specimens underwent fatigue testing of 500 000 cycles at a frequency of 2 Hz with a dynamic compressive load of 120 N at an angle of 30 degrees. The detorque values were measured by using a digital torque meter and tabulated for statistical analyses. RESULTS: The specimens with indexed abutments had mean ±standard deviation detorque values of 6.72 ±2.29 Ncm under mechanical cycling, whereas those with nonindexed abutments had values of 8.98 ±1.84 Ncm. In the indexed group, the lowest detorque value was observed for abutments at 30 degrees compared with the straight group (P<.05). As for nonindexed abutments, similar detorque values were observed after increasing the abutment inclination (P>.05). CONCLUSIONS: A decrease in detorque values in the indexed abutments related to their inclination was found under mechanical cycling, whereas the prosthetic abutments with 30 degrees of angulation had the lowest values. No decrease was found in the nonindexed abutments.

Melatonin has a stimulatory effect on osteoblasts by upregulating col-i and opn expression/secretion.

Melatonin (MLT) is a potential signaling molecule in the homeostasis of bone metabolism and may be an important mediator of bone formation and stimulation. The aim of this in vitro study was to evaluate the effect of MLT on the viability, mRNA/protein expression and mineralization of pre-osteoblastic cells. The concentrations 5, 2.5, 1, 0.1 and 0.01 mM MLT were tested on pre-osteoblastic cells (MC3T3) compared to control (no MLT), evaluating proliferation and cell viability (C50), gene expression (RT-PCR) and secretion (ELISA) of COL-I and OPN at 24h, 48h and 72h, and the formation of mineral nodules (alizarin red and fast red) after 10 days of treatment. MLT at 5 and 2.5 mM proved to be cytotoxic (C50), so only 0.01, 0.1 and 1 mM were used for the subsequent analyses. OPN mRNA expression increased with MLT at 0.1 mM - 1 mM, which was followed by increased secretion of OPN both at 24h and 72h compared to the remaining groups (p <0.05). COL-I mRNA and COL-1 secretion followed the same pattern as OPN at 0.1 mM MLT at 72h of treatment (p <0.05). Regarding mineralization, all MLT doses (except 1mM) caused an increase (p <0.05) in the formation of mineral nodules compared to the control. Melatonin at 0.01mM - 1mM had a stimulatory effect on osteoblasts by upregulating COL-I and OPN expression/ secretion and mineralization, thereby fostering osteogenesis.

A melatonina (MLT) é uma molécula potencial de sinalização na homeostase do metabolismo ósseo e pode ser um importante mediador da formação e estimulação óssea. O objetivo deste estudo in vitro foi avaliar o efeito da MLT na viabilidade, na expressão do mRNA da proteína e mineralização de células préosteoblásticas. As concentrações de MLT 5, 2,5, 1, 0,1 e 0,01 mM foram testadas em células pré-osteoblásticas da linhagem MC3T3 em comparação ao controle (sem MLT), avaliando a proliferação e a viabilidade celular (C50), expressão gênica (rtPCR) e secreção (Elisa) de Colágeno tipo 1 (COL-I) e osteopontina (OPN) às 24, 48 e 72 horas, além da formação de nódulos minerais por meio do teste vermelho de Alizarina fast red após 10 dias de tratamento. MLT a 5 e 2,5 mM provou ser tóxico (C50). Portanto, as concentrações de 0,01, 0,1 e 1 mM foram utilizadas para as análises subsequentes. A expressão do mRNA da OPN aumentou com MLT a 0,1 mM-1mM, seguida pela secreção aumentada de OPN às 24 e 72 horas em comparação aos demais grupos (p<0,05). O mRNA de COL-I e a secreção de COL-I seguiram o mesmo padrão do OPN a 0,1 mM de MLT em 72 horas de tratamento (p<0,05). Em relação à mineralização, todas as doses de MLT (exceto 1mM) causaram aumento (p<0,05) na formação de nódulos minerais em comparação ao controle. A MLT na concentração entre 0,01mM a 1 mM teve um efeito estimulador sobre os osteoblastos, ao regular positivamente a expressão e secreção de COL-I e OPN, além da mineralização, favorecendo a osteogênese.

Melatonin has a stimulatory effect on osteoblasts by upregulating COL-I and OPN expression/secretion / Melatonina tem efeito estimulador em osteoblastos, pela regulação positiva da expressão/secreção de COL-I e OPN

ABSTRACT Melatonin (MLT) is a potential signaling molecule in the homeostasis of bone metabolism and may be an important mediator of bone formation and stimulation. The aim of this in vitro study was to evaluate the effect of MLT on the viability, mRNA/protein expression and mineralization of pre-osteoblastic cells. The concentrations 5, 2.5, 1, 0.1 and 0.01 mM MLT were tested on pre-osteoblastic cells (MC3T3) compared to control (no MLT), evaluating proliferation and cell viability (C50), gene expression (RT-PCR) and secretion (ELISA) of COL-I and OPN at 24h, 48h and 72h, and the formation of mineral nodules (alizarin red and fast red) after 10 days of treatment. MLT at 5 and 2.5 mM proved to be cytotoxic (C50), so only 0.01, 0.1 and 1 mM were used for the subsequent analyses. OPN mRNA expression increased with MLT at 0.1 mM - 1 mM, which was followed by increased secretion of OPN both at 24h and 72h compared to the remaining groups (p <0.05). COL-I mRNA and COL-1 secretion followed the same pattern as OPN at 0.1 mM MLT at 72h of treatment (p <0.05). Regarding mineralization, all MLT doses (except 1mM) caused an increase (p <0.05) in the formation of mineral nodules compared to the control. Melatonin at 0.01mM - 1mM had a stimulatory effect on osteoblasts by upregulating COL-I and OPN expression/ secretion and mineralization, thereby fostering osteogenesis.

RESUMO A melatonina (MLT) é uma molécula potencial de sinalização na homeostase do metabolismo ósseo e pode ser um importante mediador da formação e estimulação óssea. O objetivo deste estudo in vitro foi avaliar o efeito da MLT na viabilidade, na expressão do mRNA da proteína e mineralização de células préosteoblásticas. As concentrações de MLT 5, 2,5, 1, 0,1 e 0,01 mM foram testadas em células pré-osteoblásticas da linhagem MC3T3 em comparação ao controle (sem MLT), avaliando a proliferação e a viabilidade celular (C50), expressão gênica (rtPCR) e secreção (Elisa) de Colágeno tipo 1 (COL-I) e osteopontina (OPN) às 24, 48 e 72 horas, além da formação de nódulos minerais por meio do teste vermelho de Alizarina fast red após 10 dias de tratamento. MLT a 5 e 2,5 mM provou ser tóxico (C50). Portanto, as concentrações de 0,01, 0,1 e 1 mM foram utilizadas para as análises subsequentes. A expressão do mRNA da OPN aumentou com MLT a 0,1 mM-1mM, seguida pela secreção aumentada de OPN às 24 e 72 horas em comparação aos demais grupos (p<0,05). O mRNA de COL-I e a secreção de COL-I seguiram o mesmo padrão do OPN a 0,1 mM de MLT em 72 horas de tratamento (p<0,05). Em relação à mineralização, todas as doses de MLT (exceto 1mM) causaram aumento (p<0,05) na formação de nódulos minerais em comparação ao controle. A MLT na concentração entre 0,01mM a 1 mM teve um efeito estimulador sobre os osteoblastos, ao regular positivamente a expressão e secreção de COL-I e OPN, além da mineralização, favorecendo a osteogênese.

Clinical and tomographic comparison of dental implants placed by guided virtual surgery versus conventional technique: A split-mouth randomized clinical trial.

AIM: Our objective was to compare guided virtual surgery to conventional surgery in terms of angular deviation of single dental implants placed in the posterior mandible. MATERIALS AND METHODS: Patients with bilateral homologous single teeth missing in the posterior mandible were eligible for this split-mouth randomized clinical trial. Cone beam computed tomography (CBCT) was performed for virtual planning of implant position and manufacturing of the stereolithographic guides. One week after the surgery, a second CBCT scan was superimposed to the initial planning. Primary endpoint was the angular deviation between virtual and clinical implant position. Secondary endpoints were linear deviations and patient-reported outcomes collected with a questionnaire. RESULTS: Data from 12 patients were available for analysis. Angular deviation was significantly lower using stereolithographic guides as compared to conventional guides (2.2 ± 1.1° vs. 3.5 ± 1.6°, p = .042). Linear deviations were similar for both techniques in the coronal (2.34 ± 1.01 vs. 1.93 ± 0.95 mm) and apical (2.53 ± 1.11 vs. 2.19 ± 1.00 mm) dimensions (p Ëƒ .05). The selection of the surgical technique had no significant impact on the patient-reported outcomes. CONCLUSION: Our data suggest that the angular discrepancy between the virtual and the clinical implant position is slightly lower when using stereolithographic guides as compared to conventional guides.

H2S in periodontal immuneinflammatory response and bone loss: a study in rats.

Halitosis is highly prevalent in periodontitis and attributed mainly to the presence of volatile sulfur compounds (VSC), where hydrogen sulfide (H2S) is the chief culprit in the characteristic malodor of periodontitis and thus may play an active role in its pathogenesis. The aim of this study was to evaluate the effect of H2S in the acute, intermediate and chronic immuneinflammatory host response and alveolar bone loss in vivo by using an animal model of induced periodontal disease. Thirtysix rats were divided into 2 groups: test group (n = 18), rats exposed to H2S (NaHS H2S donor molecule) and control group (n = 18), rats treated with saline only (Ctrl). All animals had one of their lower second molars ligated to induce periodontal disease (PD). The sound contralateral molar was used as control (H). Each group was subdivided into 3 (n = 6), according to followup time (3h, 5 days and 14 days). The gingival tissue was used for mRNA expression analysis (IL1, IL6, RANKL, OPG and SOFAT) by realtime PCR and the mandibles were analyzed morphometrically. Data analysis showed that the ligature promoted alveolar bone loss, observed mainly at 14 days, both in the group exposed to H2S and in the Ctrl group. H2S administration did not result in additional bone loss. Gene expression showed a significant increase in IL1, IL6, RANKL and SOFAT only in the CtrlPD group (p<0.05). A significant downregulation in OPG expression was observed over time in the CtrlPD group (p<0.05). In conclusion, H2S had no effect on alveolar bone loss in the absence of a ligature. In the presence of a ligature, however, exposure to H2S had an immunoregulatory effect on the expression of proinflammatory and proresorptive cytokines.

A halitose é altamente prevalente na periodontite e é atribuída principalmente à presença de compostos sulforosos voláteis (CSV), sendo o sulfeto de hidrogênio (H2S) o principal gás relacionado ao mau odor e que pode estar envolvido na patogênese da doença periodontal. O objetivo deste estudo foi avaliar o efeito agudo, intermediário e crônico do H2S na resposta imunoinflamatória e na perda óssea alveolar em ratos, com e sem doença periodontal induzida. Trinta e seis ratos foram divididos em 2 grupos: teste (n = 18), ratos expostos ao H2S (NaHS molécula doadora de H2S) e grupo controle (n = 18), ratos tratados apenas com solução salina (Ctrl). Todos os animais tiveram um dos seus segundos molares inferiores submetidos à colocação de uma ligadura para o desenvolvimento da doença periodontal (DP), em comparação com o dente contralateral saudável (H). Cada grupo foi subdividido em 3 (n = 6), de acordo com o tempo de eutanásia (3h, 5 dias e 14 dias). Os tecidos gengivais foram utilizados para a análise da expressão gênica (IL1, IL6, RANKL, OPG e SOFAT) por PCR em tempo real e as mandíbulas foram analisadas morfometricamente. Análise dos dados demonstrou que a ligadura promoveu perda óssea alveolar, observada principalmente aos 14 dias, tanto no grupo exposto ao H2S quanto no grupo Ctrl. A administração de H2S não resultou em perda óssea adicional. A expressão gênica demonstrou aumento significativo de IL1, IL6, RANKL e SOFAT apenas no grupo CtrlPD (p <0,05). Uma significativa regulação negativa na expressão de OPG foi observada ao longo do tempo no grupo CtrlPD (p <0,05). Podese concluir que o H2S não teve efeito adicional na perda óssea alveolar, na ausência de ligadura. Entretanto, na presença de ligadura, a exposição ao H2S teve um efeito imunorregulatório na expressão de citocinas próinflamatórias e próreabsortivas.

Rehabilitation of the atrophic mandible with short implants in different positions: A finite elements study.

OBJECTIVE: The aim of this study was to analyze whether the use of inclined short implants without lower transcortical involvement (test model - SI), thus preserving the mandibular lower cortical bone, could optimize stress distribution. MATERIALS AND METHODS: Six identical atrophic mandible models were created featuring 8mm of height at the symphysis. Two study factors were evaluated: implant length and angulation. Implant length was represented either by short implants (7mm) with preservation of the mandibular lower cortical bone or standard implants (9mm) with a bicortical approach and 3 possible implant positioning configurations: 4 distally-inclined implants at 45° (experimental model), all-on-four, 4 vertical implants. All tridimensional (3D) models were analyzed using the Finite Element Method (FEM) and the Ansys Workbench software. RESULTS: The maximum stress on the bone at the cervical region of the implants in the experimental model was 132MPa and transcortical involvement with implant inclination yielded higher values (171MPa). Regarding von Mises stress on the retaining screw of the prosthesis, 61MPa was recorded for the experimental model while upright implants had the highest values (223MPa). At the acrylic base, 4MPa was recorded for the experimental model whereas models with upright implants showed the highest stress values (11MPa). CONCLUSION: Rehabilitation of severely resorbed mandibles with 4 short implants placed distally at 45°, without lower transcortical involvement, were biomechanically more favorable, generating lower stress peaks, than the models with short implants on an all-on-four, or on an upright configuration, with or without lower transcortical involvement.

Avaliação da eficácia das escovas convencionais e multifilamentadas: Um estudo randomizado cego controlado / Effectiveness of conventional and multifi lamentary toothbrushes: a randomized, blinded, controlled trial

Objetivo: comparar o efeito de escovas multifilamentadas com as escovas dentais convencionais, na formação do biofilme dental bacteriano na área dentogengival, em indivíduos saudáveis. Material e métodos: para a realização deste estudo de delineamento prospectivo, cruzado, cego e randomizado, foram selecionados 16 voluntários periodontalmente saudáveis, os quais inicialmente foram submetidos a uma adequação bucal. Após sete dias de adequação, os indivíduos foram aleatoriamente divididos em quatro grupos: A) escova multifilamentos nacional (Sanifill Infinite); B) escova multifilamentos importada (Curaprox); C) escova convencional 1 (Bitufo Class macia); e D) escova convencional 2 (Oral B Indicator ), utilizando o mesmo dentifrício para os quatro grupos. Os voluntários foram instruídos a usarem somente o método de higiene referente ao grupo a que foram designados, por um período de 14 dias, com intervalos (washout) de sete dias entre os períodos experimentais. Durante o washout, todos os indivíduos fizeram uso de escovas, dentifrícios e fio dental padronizados. Os seguintes parâmetros clínicos foram avaliados nos tempos 0 e 14 dias: índice de placa visível e corada (IPV e IPC) e índice de sangramento gengival (ISG). Resultados: após análise dos dados, não foram observadas diferenças estatísticas (p > 0,05), nem intragrupo e nem intergrupo, para todos os parâmetros analisados. Conclusão: escovas convencionais e multifilamentadas foram igualmente eficazes no controle do biofilme dental bacteriano, na área dentogengival.

Objective: to compare the effect of multifilament toothbrushes and the conventional ones relating it to the formation of dental bacterial biofilm in the dentogingival region in healthy individuals. Material and methods: to conduct this study in a prospective, crossed, blind and randomized outlining way, sixteen periodontal healthy volunteers were selected and initially submitted to an oral adjustment. After seven days of adjustment, the individuals were randomly divided into four groups: A) the national multifilament toothbrush (Sanifi ll Infi nite); B) the imported multifilament toothbrush (Curaprox); C) the conventional toothbrush 1 (Bitufo Class Macia); and D) the conventional toothbrush 2 (Oral B Indicator); the same toothpaste was utilized by the four groups. The volunteers were instructed to the usage of only one method of oral hygiene which is related to the group they were designed for a period of fourteen days, with intervals (washout) of seven days between the experimental periods. During the washout, all the individuals made use of the toothbrushes, toothpastes and standardized dental floss. The following clinical parameters were evaluated at 0 day and 14 days: visible plaque index and disclosed plaque index (VPI and DPI) and gingival bleeding index (GBI). Results: no statistically significant differences were observed (p > 0,05), neither with the intragroup nor the intergroup in all the parameters analyzed. Conclusion: conventional toothbrushes and the multifilamented ones were equally effective in controling dental bacterial biofilm in the dentogingival region.

In Vitro Microbiological Analysis of Bacterial Seal in Hybrid Zirconia Abutment Tapered Connection.

PURPOSE: The aim of this study was to evaluate the bacterial seal at the implant-hybrid zirconia abutment interface and Morse taper-type connections through in vitro microbiological analysis. MATERIALS AND METHODS: Sixteen implants and their respective abutments were divided into 3 groups: test (10 sets), positive control (3 sets), and negative control (3 sets). In the test group, 10 implants were contaminated with Escherichia coli using a sterile inoculating loop to the inner portion of the implants, followed by torque application to the abutment (30 N·cm). The positive controls were also contaminated, but no torque was applied to the abutment screw. The negative control consisted of uncontaminated sets. All specimens were immersed in test tubes containing 5 mL brain heart infusion (BHI) broth, maintained in a microbiological incubator for 14 days at 37°C under aerobic conditions, and monitored every 24 hours for evidence of bacterial growth. RESULTS: During the 14 days of incubation, no significant increase in the number of cloudy culture media was observed in the test group (P = 0.448). No significant difference in broth turbidity ratio was observed (P > 0.05). CONCLUSION: Hybrid zirconia abutments can create an effective seal at the tapered abutment-implant interface with a 30-N·cm installation torque.

Microbiological Seal of Two Types of Tapered Implant Connections

Abstract Tapered implant connections have gained wide popularity for being more resistant to fatigue and for promoting a better seal against bacterial infiltration than conventional connections. The aim of this study was to evaluate the bacterial seal at the implant-abutment interface using two Morse taper implant models, by in vitro microbiological analysis. Eleven non-indexed and 11 indexed abutments were selected and connected to their respective implants with a 20 N torque, according to manufacturer's recommendation. Microbiological analysis was carried out using colonies of Escherichia coli transported directly from a culture dish to the prosthetic component. For control, one non-contaminated abutment-implant set from each group (negative control) and one contaminated implant with no abutment (positive control) were used. The specimens were immersed in BHI broth and maintained in an incubator at 37 °C for 14 days to assess the development of bacterial contamination. The results revealed that 36.4% (n=4) of the indexed components and 90.9% (n=10) of the non-indexed components allowed bacterial leakage, with significant difference between groups (p=0.0237). In conclusion, both tapered components failed to provide adequate sealing to bacterial leakage, although the indexed type components showed a superior seal compared with non-indexed components.

Resumo Conexões de implantes cônicos cresceram em popularidade por serem mais resistentes à fadiga e por promover uma melhor vedação contra infiltração bacteriana do que as conexões convencionais. O objetivo deste estudo foi avaliar o selamento bacteriano na interface implante-pilar utilizando dois modelos de implantes cone Morse, por meio de análise microbiológica in vitro. Onze pilares não indexados e 11 pilares indexados foram selecionados e conectados aos seus respectivos implantes com um torque de 20 N, de acordo com a recomendação do fabricante. A análise microbiológica foi realizada utilizando colônias de Escherichia coli retirados diretamente a partir de uma placa de cultura para o componente protético. Para os grupos de controle, foi utilizado um pilar-implante não contaminado de cada grupo (controle negativo) e um implante contaminado sem pilar (controle positivo). Os espécimes foram imersos em caldo BHI e mantidos numa incubadora a 37 °C durante 14 dias, para monitorar o desenvolvimento de contaminação bacteriana. Os resultados revelaram que 36,4% (n=4) dos componentes indexados e 90,9% (n=10) dos componentes não indexados obtiveram infiltração bacteriana, com diferença significativa entre os grupos (p=0,0237). Como conclusão, os dois componentes cônicos não conseguiram proporcionar uma vedação adequada contra infiltração bacteriana, embora os componentes do tipo indexados mostrassem uma vedação superior, quando comparados com componentes não indexados.

Microbiological Seal of Two Types of Tapered Implant Connections.

Tapered implant connections have gained wide popularity for being more resistant to fatigue and for promoting a better seal against bacterial infiltration than conventional connections. The aim of this study was to evaluate the bacterial seal at the implant-abutment interface using two Morse taper implant models, by in vitro microbiological analysis. Eleven non-indexed and 11 indexed abutments were selected and connected to their respective implants with a 20 N torque, according to manufacturer's recommendation. Microbiological analysis was carried out using colonies of Escherichia coli transported directly from a culture dish to the prosthetic component. For control, one non-contaminated abutment-implant set from each group (negative control) and one contaminated implant with no abutment (positive control) were used. The specimens were immersed in BHI broth and maintained in an incubator at 37 °C for 14 days to assess the development of bacterial contamination. The results revealed that 36.4% (n=4) of the indexed components and 90.9% (n=10) of the non-indexed components allowed bacterial leakage, with significant difference between groups (p=0.0237). In conclusion, both tapered components failed to provide adequate sealing to bacterial leakage, although the indexed type components showed a superior seal compared with non-indexed components.

Expression of SOFAT by T- and B-lineage cells may contribute to bone loss.

A novel T cell-secreted cytokine, termed secreted osteoclastogenic factor of activated T cells (SOFAT) that induces osteoclastic bone resorption in a RANKL-independent manner, has been described. Our group have previously reported that SOFAT is highly expressed in gingival tissues of patients with chronic periodontitis suggesting a putative role in the bone loss associated with periodontal disease. The aim of the present study was to identify other potential cellular sources of SOFAT in the bone resorptive lesions of patients with periodontal disease. Gingival tissues were biopsied from systemically healthy subjects without periodontal disease (n=5) and patients with chronic periodontitis (n=5), and the presence of SOFAT was analyzed by immunohistochemistry and immunofluorescence staining. The present data demonstrated marked SOFAT staining in diseased periodontal tissues that was predominantly associated with the lymphocytic infiltration of gingival tissues. Notably, in addition to CD3+ T cells, B­lineage cells including plasma cells also exhibited strong staining for SOFAT. As SOFAT has not previously been reported in B­lineage cells, splenic T cells and B cells were further purified from BALB/c mice and activated using CD3/CD28 and lipopolysaccharide, respectively. SOFAT was quantified by reverse transcription­quantitative polymerase chain reaction and was shown to be significantly expressed (P<0.05) in both activated T cells and B cells compared with unstimulated cells. These data support a putative role of SOFAT in the bone loss associated with chronic periodontal disease. In addition, to the best of our knowledge, this study demonstrates for the first time that in addition to T cells, B-lineage cells may also be a significant source of SOFAT in inflammatory states.

Evaluation of the Biological Behavior of Mucograft® in Human Gingival Fibroblasts: An In Vitro Study

Mucograft(r) is a resorbing porcine matrix composed of type I and type III collagen, used for soft tissue augmentation in guided tissue bony regeneration procedures. This in vitro study aimed to evaluate the biological behavior of Mucograft(r) in human gingival fibroblasts, as well as the ability of the matrix to induce production of extracellular matrix. Six resorbing Mucograft(r) matrices (MCG) were cut into 3 x 2 mm rectangles and 5 x 5 mm squares and were placed in 96- and 24-well plates, respectively. The control group (CTRL) consisted of cells plated on polystyrene without the MCG. After one, two, three and seven days, cell proliferation and viability were assessed using the Trypan exclusion method and MTT test, respectively. Type III collagen (COL 3A1) and vimentin (VIM) expression were also evaluated at 10 and 14 days, using Western blotting. Statistical analysis, using ANOVA with post hoc Bonferroni test, revealed that human gingival fibroblasts from MCG showed similar results (p>0.05) for proliferation and viability as the cells cultured on CTRL. After 14 days, a significant decrease in COL 3A1 expression (p<0.05) was observed when cultured with the MCG. VIM expression showed no significant difference at any time period (p>0.05). Although no increase in extracellular matrix secretion was observed in this in vitro study, Mucograft(r) presented cellular compatibility, being an option for a scaffold whenever it is required.

Resumo A Mucograft(r) é uma matriz reabsorvível, de origem suína, composta de colágenos do tipo I e III, utilizada para aumento de tecido mole em regeneração óssea guiada. Este estudo in vitro teve como objetivo avaliar o comportamento biológico da Mucograft(r), em fibroblastos gengivais humanos, bem como a indução da síntese de matriz extracelular. Seis matrizes reabsorvíveis de Mucograft(r) (MCG) foram cortadas em retângulos e quadrados medindo 3 x 2 mm e 5 x 5 mm e alocadas em placas de 96 e 24 poços, respectivamente. O grupo controle (CTRL) consistiu no plaqueamento celular em poliestireno, sem MCG. Após um, dois, três e sete dias, a proliferação e a viabilidade celular foram avaliadas utilizando o corante vital azul de Trypan e o teste MTT, respectivamente. Além disso, a expressão de colágeno tipo III (COL 3A1) e vimentina (VIM) foi avaliada após 10 e 14 dias, por meio de Western-blotting. Após análise estatística (Anova e pós teste de Bonferroni), pode-se observar que os fibroblastos gengivais humanos, cultivados sobre MCG, apresentaram proliferação e viabilidade semelhantes em comparação às células que foram cultivadas apenas no poliestireno (CTRL). Após 14 dias, notou-se uma diminuição significativa da expressão de COL 3A1 (p<0,05) quando as células foram cultivadas sobre a MCG. A expressão da VIM não mostrou diferença significativa em nenhum dos períodos estudados (p>0,05). No presente estudo in vitro pode-se concluir que apesar de não ter sido observado aumento da síntese de matriz extracelular, a Mucograft(r) apresentou compatibilidade celular, sendo uma opção de biomaterial em casos que o arcabouço é necessário.

Evaluation of the Biological Behavior of Mucograft® in Human Gingival Fibroblasts: An In Vitro Study.

Mucograft(r) is a resorbing porcine matrix composed of type I and type III collagen, used for soft tissue augmentation in guided tissue bony regeneration procedures. This in vitro study aimed to evaluate the biological behavior of Mucograft(r) in human gingival fibroblasts, as well as the ability of the matrix to induce production of extracellular matrix. Six resorbing Mucograft(r) matrices (MCG) were cut into 3 x 2 mm rectangles and 5 x 5 mm squares and were placed in 96- and 24-well plates, respectively. The control group (CTRL) consisted of cells plated on polystyrene without the MCG. After one, two, three and seven days, cell proliferation and viability were assessed using the Trypan exclusion method and MTT test, respectively. Type III collagen (COL 3A1) and vimentin (VIM) expression were also evaluated at 10 and 14 days, using Western blotting. Statistical analysis, using ANOVA with post hoc Bonferroni test, revealed that human gingival fibroblasts from MCG showed similar results (p>0.05) for proliferation and viability as the cells cultured on CTRL. After 14 days, a significant decrease in COL 3A1 expression (p<0.05) was observed when cultured with the MCG. VIM expression showed no significant difference at any time period (p>0.05). Although no increase in extracellular matrix secretion was observed in this in vitro study, Mucograft(r) presented cellular compatibility, being an option for a scaffold whenever it is required.

Impact of supragingival therapy on subgingival microbial profile in smokers versus non-smokers with severe chronic periodontitis.

BACKGROUND: The aim of this study was to assess subgingival microbiological changes in smokers versus non-smokers presenting severe chronic periodontitis after supragingival periodontal therapy (ST). METHODS: Non-smokers (n=10) and smokers (n=10) presenting at least nine teeth with probing pocket depth (PPD) (≥5 mm), bleeding on probing (BoP), and no history of periodontal treatment in the last 6 months were selected. Clinical parameters assessed were plaque index (PI), BoP, PPD, relative gingival margin position (rGMP) and relative clinical attachment level (rCAL). Subgingival biofilm was collected before and 21 days after ST. DNA was extracted and the 16S rRNA gene was amplified with the universal primer pair, 27F and 1492R. Amplified genes were cloned, sequenced, and identified by comparison with known 16S rRNA sequences. Statistical analysis was performed by Student's t and Chi-Square tests (α=5%). RESULTS: Clinically, ST promoted a significant reduction in PI and PPD, and gain of rCAL for both groups, with no significant intergroup difference. Microbiologically, at baseline, data analysis demonstrated that smokers harbored a higher proportion of Porphyromonas endodontalis, Bacteroidetes sp., Fusobacterium sp. and Tannerella forsythia and a lower number of cultivated phylotypes (p<0.05). Furthermore, non-smokers featured significant reductions in key phylotypes associated with periodontitis, whereas smokers presented more modest changes. CONCLUSION: Within the limits of the present study, ST promoted comparable clinical improvements in smokers and non-smokers with severe chronic periodontitis. However, in smokers, ST only slightly affected the subgingival biofilm biodiversity, as compared with non-smokers.

Evidence that metyrapone in the presence of inflammation modulates cytokine mRNA expression.

OBJECTIVE: Metyrapone (MT) has been used clinically to decrease glucocorticoid levels in human and animal studies. However, the potential effects of MT in the presence of inflammation are poorly understood. Thus, the aim of this study was to evaluate the effects of the administration of MT on the mRNA levels of pro-inflammatory cytokines in the presence of inflammation induced by the well-established model of ligature-induced periodontitis in rats. MATERIAL AND METHODS: Sixty animals were randomly assigned into three experimental groups of 20 rats each: G1-control; G2-periodontal disease (PD) induced by cotton ligature; G3-PD associated with 3 daily doses of MT (50mg/kg/3×3h). After 30 days, all animals were killed by decapitation. Blood samples were taken and the concentrations of corticosterone and catecholamines measured. Marginal tissues around ligated and non-ligated teeth were harvested and gene expression was assessed by quantitative polymerase chain reaction technique (qPCR). Moreover, the area of interradicular bone loss (ABL) was histometrically determined. RESULTS: Data analysis showed that: (i) ligature placement resulted in a significant ABL, as compared to non-ligated sites of G1 group; (ii) mRNA levels of all the pro-inflammatory factors assessed (INF-γ, TNF-α, IL-1ß and IL-6) were increased in the PD group (G2) (p<0.05) when compared to G1; (iii) there were no significant differences in corticosterone and catecholamine plasmatic levels between the three groups; (iv) MT administration, in the presence of inflammation, induces an increased ABL and significantly increased mRNA levels of all pro-inflammatory cytokines analyzed (p<0.05). CONCLUSION: Within the limits of this study, it can be concluded that MT in the presence of inflammation may modulate expression of pro-inflammatory cytokines, regardless of its effect on plasma corticosterone levels.

Chronic stress may modulate periodontal disease: a study in rats.

BACKGROUND: The present study aimed to evaluate whether chronic stress (CS) affects ligature-induced periodontal disease and to investigate the impact of CS on the mRNA levels of interleukin (IL)-1beta, -1 receptor antagonist, -6, and -10, interferon-gamma (IFN-gamma), receptor activator of nuclear factor-kappa B ligand (RANKL), and osteoprotegerin in the gingival tissues of rats. METHODS: Sixty male Wistar rats were assigned randomly to three groups: G1 (control; non-ligated sites), G2 (periodontal disease), and G3 (periodontal disease associated with restraint stress for 12 hours/day for the entire study). After 30 days, all animals were sacrificed by decapitation. Blood samples were taken, and the concentrations of corticosterone and catecholamines were measured as biomarkers of CS. Marginal tissues around ligated and non-ligated teeth were harvested, and gene expression was assessed by quantitative polymerase chain reaction. Moreover, the area of bone loss (ABL) was determined histometrically. RESULTS: Data analysis showed that CS increased serum levels of stress biomarkers (P <0.05), ligature placement resulted in a significant ABL compared to non-ligated sites, CS significantly increased the amount of ABL in inflamed sites (P <0.001), and CS significantly increased mRNA levels of proinflammatory (IL-1beta and -6 and IFN-gamma) and anti-inflammatory (IL-10) cytokines and proresorptive factor (RANKL) in ligated sites (P <0.05). CONCLUSION: CS significantly increased bone loss resulting from ligature-induced periodontitis by a local increase in proinflammatory and proresorptive factors.

A systematic review of stress and psychological factors as possible risk factors for periodontal disease.

BACKGROUND: Clinical observations and epidemiologic studies suggest that some negative life events and psychological factors may contribute to an increased susceptibility to periodontal disease. The aim of the present study was to systematically review the evidence from case-control studies, cross-sectional studies, and prospective clinical trials reporting on the influence of stress and psychological factors on periodontal disease. The focused question addressed in this systematic review was whether the scientific evidence is enough to consider stress and psychological factors as risk factors for periodontal disease. METHODS: A literature search was conducted using two databases (MEDLINE and the Cochrane Oral Health Group specialist trials register) in addition to searching reference lists of original and review articles. The search strategy used was the combination of the terms: "stress," "periodontal disease," and "psychosocial disorders." Studies were selected if they were published in dental journals between January 1, 1990 and April 1, 2006; only human studies and studies with adults and middle-aged subjects were included. Suitable variables included control for the potential effect of confounding factors, adequate criteria to define periodontal disease, adequate criteria for establishing stress, and methodologic quality. Only English-language articles were considered, and unpublished data were not sought. Two reviewers independently extracted information regarding quality and study characteristics in duplicate. The studies were assessed regarding their methodologic characteristics, statistical analysis, characteristics of the periodontal outcome measures, and psychological measurements. RESULTS: Of the 58 articles identified in the search, 10 were excluded because they were reviews and 34 did not comply with the selection criteria. Fourteen articles (seven case-control studies, six cross-sectional studies, and one prospective clinical trial) were included in the analysis; their quality and main study characteristics were assessed according to the criteria preestablished in the protocol of the study. With regard to the results of the studies, 57.1% found a positive outcome between psychosocial factors/stress and periodontal disease, 28.5% observed a positive outcome for some characteristics and a negative outcome for others, and 14.2% found a negative outcome. CONCLUSIONS: Within the limitations of this systematic review, the majority of studies showed a positive relationship between stress/psychological factors and periodontal disease. However, in the future, well-designed and more representative studies should be considered to confirm these factors as a risk for periodontal disease.