RESUMO
AIM: To investigate the attachment and proliferation of dental pulp stem cells (DPSC) on dentine treated with various endodontic regeneration protocols. METHODOLOGY: Standardized dentine samples were irrigated with sodium hypochlorite (1.5% NaOCl) and ethylenediaminetetraacetic acid (17% EDTA) and randomized into four treatment groups and two control groups. The treatment groups were treated with a clinically used concentration of triple antibiotic paste (TAP), double antibiotic paste (DAP), calcium hydroxide (Ca(OH)2 ) or diluted TAP in a methylcellulose system (DTAP) for 1 week. Each sample in the treatment groups was then irrigated with EDTA. The two control groups were treated with EDTA or received no treatment. Dental pulp stem cells were seeded on each dentine specimen (10 000 cells). Lactate dehydrogenase activity assays were then performed to evaluate the attached DPSC after 1 day of incubation. Water-soluble tetrazolium assays were used to determine DPSC proliferation after three additional days of incubation. Friedman's test followed by least significant difference were used for statistical analyses (α = 0.05). RESULTS: Triple antibiotic paste and DTAP regeneration protocols, as well as EDTA-treated dentine, caused significant increases in DPSC attachment to dentine. Triple antibiotic paste, DAP and Ca(OH)2 regeneration protocols caused significant reductions in DPSC proliferation on dentine. However, the DTAP regeneration protocol did not have any significant negative effects on DPSC proliferation. CONCLUSIONS: The clinically used endodontic regeneration protocols that include the use of TAP, DAP or Ca(OH)2 medicament negatively affected DPSC proliferation on dentine. However, the use of DTAP medicament during regenerative endodontic treatment may not adversely affect the proliferation of DPSC.
Assuntos
Polpa Dentária/citologia , Dentina/efeitos dos fármacos , Antibacterianos/farmacologia , Hidróxido de Cálcio/farmacologia , Proliferação de Células , Ciprofloxacina/farmacologia , Ácido Edético/farmacologia , Humanos , Técnicas In Vitro , L-Lactato Desidrogenase/análise , Teste de Materiais , Metilcelulose/farmacologia , Metronidazol/farmacologia , Minociclina/farmacologia , Dente Molar , Distribuição Aleatória , Regeneração , Irrigantes do Canal Radicular/farmacologia , Hipoclorito de Sódio/farmacologia , Propriedades de SuperfícieRESUMO
This article presents details of fabrication, biological activity (i.e., anti-matrix metalloproteinase [anti-MMP] inhibition), cytocompatibility, and bonding characteristics to dentin of a unique doxycycline (DOX)-encapsulated halloysite nanotube (HNT)-modified adhesive. We tested the hypothesis that the release of DOX from the DOX-encapsulated nanotube-modified adhesive can effectively inhibit MMP activity. We incorporated nanotubes, encapsulated or not with DOX, into the adhesive resin of a commercially available bonding system (Scotchbond Multi-Purpose [SBMP]). The following groups were tested: unmodified SBMP (control), SBMP with nanotubes (HNT), and DOX-encapsulated nanotube-modified adhesive (HNT+DOX). Changes in degree of conversion (DC) and microtensile bond strength were evaluated. Cytotoxicity was examined on human dental pulp stem cells (hDPSCs). To prove the successful encapsulation of DOX within the adhesives-but, more important, to support the hypothesis that the HNT+DOX adhesive would release DOX at subantimicrobial levels-we tested the antimicrobial activity of synthesized adhesives and the DOX-containing eluates against Streptococcus mutans through agar diffusion assays. Anti-MMP properties were assessed via ß-casein cleavage assays. Increasing curing times (10, 20, 40 sec) led to increased DC values. There were no statistically significant differences (p > .05) in DC within each increasing curing time between the modified adhesives compared to SBMP. No statistically significant differences in microtensile bond strength were noted. None of the adhesives eluates were cytotoxic to the human dental pulp stem cells. A significant growth inhibition of S. mutans by direct contact illustrates successful encapsulation of DOX into the experimental adhesive. More important, DOX-containing eluates promoted inhibition of MMP-1 activity when compared to the control. Collectively, our findings provide a solid background for further testing of encapsulated MMP inhibitors into the synthesis of therapeutic adhesives that may enhance the longevity of hybrid layers and the overall clinical performance of adhesively bonded resin composite restorations.
Assuntos
Antibacterianos/química , Adesivos Dentinários/química , Doxiciclina/química , Nanotubos/química , Silicatos de Alumínio/síntese química , Silicatos de Alumínio/química , Silicatos de Alumínio/toxicidade , Antibacterianos/síntese química , Antibacterianos/toxicidade , Caseínas/efeitos dos fármacos , Técnicas de Cultura de Células , Argila , Colagem Dentária , Polpa Dentária/citologia , Polpa Dentária/efeitos dos fármacos , Dentina/efeitos dos fármacos , Dentina/ultraestrutura , Adesivos Dentinários/síntese química , Adesivos Dentinários/toxicidade , Doxiciclina/síntese química , Doxiciclina/toxicidade , Humanos , Teste de Materiais , Metaloproteinase 1 da Matriz/efeitos dos fármacos , Inibidores de Metaloproteinases de Matriz/química , Nanotubos/toxicidade , Polimerização , Cimentos de Resina/síntese química , Cimentos de Resina/química , Cimentos de Resina/toxicidade , Células-Tronco/efeitos dos fármacos , Streptococcus mutans/efeitos dos fármacos , Estresse Mecânico , Resistência à Tração , Fatores de TempoRESUMO
OBJECTIVE: Monomers such as triethylene glycol dimethacrylate (TEGDMA) can leach from dental composites. TEGDMA-induced apoptosis in human pulp has been reported. However, the apoptotic (pro or anti) proteins involved in this process remain unclear. Therefore, the purpose of this study was to determine which apoptotic proteins are enhanced or suppressed during TEGDMA-induced apoptosis. MATERIALS AND METHODS: Human pulp fibroblasts (HPFs) were incubated with different TEGDMA concentrations (0.125-1.0 mM) and cytotoxicity was determined. TEGDMA was shown to be cell cytotoxic at concentrations of 0.50 mM and higher. The highest concentration with no significant cytotoxicity was then incubated (0.25 mM TEGDMA) with the HPFs. Cell lysates were then prepared and the protein concentrations determined. Human Apoptosis Array kits were utilized to detect the relative levels of 43 apoptotic proteins. RESULTS: HPFs exposed to TEGDMA showed significant increases in multiple pro-apoptotic proteins such as Bid, Bim, Caspase 3, Caspase 8, and Cytochrome c at 24 hours. Some anti-apoptotic proteins were also altered. CONCLUSIONS: The results indicated that TEGDMA activates both the extrinsic and intrinsic apoptotic pathways.
Assuntos
Proteínas Reguladoras de Apoptose/biossíntese , Resinas Compostas/farmacologia , Polpa Dentária/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Polietilenoglicóis/farmacologia , Ácidos Polimetacrílicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/efeitos dos fármacos , Polpa Dentária/citologia , Relação Dose-Resposta a Droga , Humanos , L-Lactato Desidrogenase/metabolismo , Análise Serial de ProteínasRESUMO
OBJECTIVE: To investigate the effects of nicotine and cigarette smoke condensate (CSC) exposure on cytokine expression from human endothelial cells in order to identify one possible mechanism that smoking plays in the pathogenesis of both periodontal disease (PDD) and cardiovascular disease (CVD). METHODS: Human endothelial cells (HUVECs) were exposed to different concentrations of nicotine and CSC to examine the effects that they have on cell proliferation and cytotoxicity. Non-toxic levels were then used to examine cytokine expression using cytokine protein arrays. RESULTS: Exposure to nicotine caused significant down-regulation in the expression of IL-10 (P = 0.046), growth-regulated oncogene (GRO)α (P = 0.036), MCP-1 (P = 0.046), and GMCSF (P = 0.004) compared with the control untreated HUVECs. Exposure to CSC caused significant down-regulation in the expression of GRO (P = 0.04), GROα (P = 0.01), IL-6 (P = 0.03), and MCP-1 (P = 0.04) compared with the control untreated HUVECs. CONCLUSIONS: Exposure of HUVECs to nicotine or CSC affects the levels of cytokine expression including reduction in anti-inflammatory and chemoattractant cytokines. This may subsequently affect the host defensive mechanisms of the tissues. The action of toxic chemicals in tobacco smoke on endothelial cells is a potential pathogenic mechanism that may in part explain the association between tobacco, PDD, and CVD.
Assuntos
Citocinas/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Nicotiana , Nicotina/farmacologia , Fumaça , Técnicas de Cultura de Células , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Quimiocina CCL2/efeitos dos fármacos , Quimiocina CXCL1/efeitos dos fármacos , Regulação para Baixo , Células Endoteliais/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/efeitos dos fármacos , Humanos , Interleucina-10/análise , Interleucina-6/análise , Nicotina/toxicidade , Fumaça/análiseRESUMO
Cigarette smoke condensate (CSC) is the particulate matter of cigarette smoke. Porphyromonas gingivalis (P. gingivalis) is an opportunistic pathogen involved in periodontitis. It was hypothesized that the combination of CSC and P. gingivalis would increase the collagen-degrading ability of human gingival fibroblasts (HGFs). In this study, HGFs were exposed to CSC, P. gingivalis supernatant, and CSC plus P. gingivalis supernatant. The collagen-degrading ability and protein/mRNA levels of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) of HGFs were examined. The combined treatment increased collagen degradation, protein levels of active forms of MMP-1, MMP-2, MMP-3, and MMP-14 in conditioned media, and the low-molecular-weight fragment of MMP-14 in membrane extracts, as well as mRNA levels of MMP-1, MMP-2, and MMP-14. In conclusion, the combined effects of CSC and P. gingivalis increased HGF-mediated collagen degradation by destroying the balance between MMPs and TIMPs at multiple levels.
Assuntos
Fibroblastos/enzimologia , Gengiva/enzimologia , Nicotiana , Material Particulado , Porphyromonas gingivalis/fisiologia , Fumaça , Western Blotting , Membrana Celular/enzimologia , Proliferação de Células , Forma Celular , Sobrevivência Celular , Células Cultivadas , Colágeno/metabolismo , Meios de Cultivo Condicionados , Precursores Enzimáticos/análise , Gelatinases/análise , Humanos , Metaloproteinase 1 da Matriz/análise , Metaloproteinase 14 da Matriz/análise , Metaloproteinase 2 da Matriz/análise , Metaloproteinase 3 da Matriz/análise , Metaloproteinases da Matriz/análise , Peso Molecular , Material Particulado/efeitos adversos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fumaça/efeitos adversos , Inibidores Teciduais de Metaloproteinases/análiseRESUMO
Currently, the use of oral and systemic forms of bisphosphonates is increasing dramatically in a large group of patients either in the form of anti-osteoporosis medications or as a part of a chemotherapeutic regimen for several malignant diseases. As adult orthodontic treatment has become more widely accepted in most orthodontic practices, orthodontists must be aware of the risks, benefits, and effects of bisphosphonates use on the patient's general health status, as well as on their orthodontic treatment outcomes. This review aims to discuss the use of bisphosphonates, the complications associated with their use, and their impact on orthodontic treatment.
Assuntos
Conservadores da Densidade Óssea/efeitos adversos , Remodelação Óssea/efeitos dos fármacos , Difosfonatos/efeitos adversos , Doenças Maxilomandibulares/etiologia , Osteonecrose/etiologia , Técnicas de Movimentação Dentária , Processo Alveolar/efeitos dos fármacos , Animais , Conservadores da Densidade Óssea/química , Contraindicações , Análise do Estresse Dentário , Difosfonatos/química , Meia-Vida , Humanos , Procedimentos Cirúrgicos Bucais , Osteoclastos/efeitos dos fármacosRESUMO
BACKGROUND AND OBJECTIVE: This study investigated the expression of a key mediator that regulates differentiation of osteoclasts, receptor activator of nuclear factor kappaB ligand (RANKL), in rats with or without osteoporosis and periodontitis, to provide a better understanding of the association between these two diseases. MATERIAL AND METHODS: Forty adult Albino rats were divided into four groups: (1) control group; (2) experimentally induced periodontitis group; (3) experimentally induced osteoporosis group; and (4) experimentally induced osteoporosis and periodontitis group. At the end of the experimental period, blood samples were obtained and animals were sacrificed. Serum alkaline phosphatase (ALP) activity levels were measured. Histological evaluation and immunohistochemical detection of RANKL in the periodontal ligament and bone tissues were performed. RESULTS: There were significantly higher ALP levels in all of the experimental groups than in the control group. The pathology observed in the histological sections from group 4 was more severe than in either group 2 or group 3. The percentage of RANKL-immunoreactive cells in both the periodontal ligament and bone tissues in group 4 (16.8 +/- 5.1 and 11.2 +/- 5.2%, respectively) was significantly higher (p < 0.001) than in the other groups. In the periodontal ligament, the percentage of RANKL-immunoreactive cells in group 2 (10.1 +/- 1.9%) was significantly higher (p < 0.001) than in group 3 (5.3 +/- 2.7%) and the control group (4.12 +/- 1.5%). CONCLUSION: The increased bone loss observed in group 4 compared with either group 2 or group 3 supports the existence of an additive pathological effect of the two disease conditions. This is consistent with the increased RANKL expression observed in group 4.
Assuntos
Osteoporose/metabolismo , Periodontite/metabolismo , Ligante RANK/análise , Fosfatase Alcalina/sangue , Perda do Osso Alveolar/metabolismo , Perda do Osso Alveolar/patologia , Processo Alveolar/metabolismo , Processo Alveolar/patologia , Animais , Biomarcadores/sangue , Medula Óssea/patologia , Contagem de Células , Feminino , Regulação da Expressão Gênica , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica , Leucócitos Mononucleares/patologia , Odontoblastos/patologia , Osteoblastos/patologia , Osteoclastos/patologia , Osteócitos/patologia , Osteoporose/patologia , Ligamento Periodontal/metabolismo , Ligamento Periodontal/patologia , Periodontite/patologia , RatosRESUMO
BACKGROUND AND OBJECTIVE: Cigarette smoke condensate, the particulate matter of cigarette smoke, is composed of thousands of chemicals, including nicotine. Cigarette smoking is a risk factor for periodontal disease. This study investigated the influence of cigarette smoke condensate on the collagen-degrading ability of human gingival fibroblasts and its mechanism. MATERIAL AND METHODS: Human gingival fibroblasts were exposed for 72 h to various concentrations of total particulate matter cigarette smoke condensate. Cell proliferation and cytotoxicity were evaluated using water-soluble tetrazolium-1 and lactate dehydrogenase, respectively. The collagen-degrading ability of human gingival fibroblasts was evaluated in collagen-coated six-well plates. Conditioned media and membrane extracts were collected for zymography and western blot analyses of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). RESULTS: Cell proliferation decreased and cytotoxicity increased in human gingival fibroblasts with increasing concentrations of cigarette smoke condensate. Cell proliferation decreased by more than 50% (p < 0.05) when the concentrations of total particulate matter cigarette smoke condensate were above 200 microg/mL, and cytotoxicity increased to more than 30% (p < 0.05) when the concentrations of total particulate matter cigarette smoke condensate were above 400 microg/mL. Cigarette smoke condensate increased the collagen-degrading ability of human gingival fibroblasts, especially at a concentration of 100 microg/mL (1.5-fold increase, p < 0.05) compared with the control. Cigarette smoke condensate increased the production of proMMP-1, proMMP-2, MMP-14 and TIMP-1, and decreased the production of TIMP-2, in conditioned media. Furthermore, compared with the control group, cigarette smoke condensate increased the production of MMP-2, MMP-14 and TIMP-2 in membrane extracts, especially at concentrations of 50-100 microg/mL. CONCLUSION: Cigarette smoke condensate affects human gingival fibroblast proliferation and is toxic at total particulate matter cigarette smoke condensate concentrations of >or= 400 microg/mL. Cigarette smoke condensate can increase the collagen-degrading ability of human gingival fibroblasts by altering the production and localization of MMPs and TIMPs.
Assuntos
Colágeno/metabolismo , Fibroblastos/patologia , Gengiva/patologia , Nicotiana/efeitos adversos , Fumaça/efeitos adversos , Adesão Celular , Morte Celular , Proliferação de Células , Forma Celular , Sobrevivência Celular , Células Cultivadas , Meios de Cultivo Condicionados , Precursores Enzimáticos/análise , Fibroblastos/enzimologia , Gelatinases/análise , Gengiva/enzimologia , Humanos , Indicadores e Reagentes , L-Lactato Desidrogenase/análise , Metaloproteinase 1 da Matriz/análise , Metaloproteinase 14 da Matriz/análise , Metaloproteinase 2 da Matriz/análise , Metaloproteinases da Matriz/análise , Mitocôndrias/enzimologia , Oxirredutases/análise , Sais de Tetrazólio , Fatores de Tempo , Inibidor Tecidual de Metaloproteinase-1/análise , Inibidor Tecidual de Metaloproteinase-2/análise , Inibidores Teciduais de Metaloproteinases/análiseRESUMO
BACKGROUND AND OBJECTIVE: The objective of this study was to determine the effects that nicotine and the combination of nicotine and Porphyromonas gingivalis supernatant have on human gingival fibroblast-mediated collagen degradation. MATERIAL AND METHODS: Human gingival fibroblasts were cultured with 25-500 microg/ml of nicotine in collagen-coated six-well plates. On days 1-5, the conditioned media was collected for zymography and western blot analyses of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). The cells were then removed and the collagen cleavage visualized by Coomassie blue staining. To examine the combined effect, 250 microg/ml of nicotine and 10% v/v culture supernatant of P. gingivalis ATCC 33277 were added to the human gingival fibroblasts. The mRNA levels of multiple MMPs and TIMPs were monitored. RESULTS: Nicotine increased the human gingival fibroblast-mediated collagen cleavage. The MMP-14 and MMP-2 produced by the nicotine-treated human gingival fibroblasts more readily underwent zymogen activation. Nicotine treatment resulted in TIMP-2 redistribution to the cell surface. The mRNAs of multiple MMPs and TIMPs were unaltered by nicotine. An additive collagen cleavage effect was observed when the human gingival fibroblasts were treated with both nicotine and P. gingivalis. CONCLUSION: Nicotine increased human gingival fibroblast-mediated collagen degradation, in part through the activation of membrane-associated MMPs. Nicotine and P. gingivalis had an additive effect on human gingival fibroblast-mediated collagen degradation.
Assuntos
Fibroblastos/efeitos dos fármacos , Estimulantes Ganglionares/efeitos adversos , Gengiva/efeitos dos fármacos , Nicotina/efeitos adversos , Porphyromonas gingivalis/metabolismo , Western Blotting/métodos , Células Cultivadas/efeitos dos fármacos , Colágeno/efeitos dos fármacos , Fibroblastos/enzimologia , Gengiva/citologia , Humanos , Metaloproteinases da Matriz/efeitos dos fármacos , Metaloproteinases da Matriz/metabolismo , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Inibidores Teciduais de Metaloproteinases/efeitos dos fármacos , Inibidores Teciduais de Metaloproteinases/metabolismoRESUMO
BACKGROUND AND OBJECTIVE: The purpose of this study was to characterize the heterogeneity of the collagen-degrading ability of different human gingival fibroblast cell lines treated with Porphyromonas gingivalis supernatant. MATERIAL AND METHODS: Seven human gingival fibroblast cell lines were analyzed for their ability to cleave Type I collagen in the presence and absence of culture supernatant from P. gingivalis ATCC 33277 (10% v/v). The matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) produced by these human gingival fibroblasts were monitored at the protein level by zymography and/or western blot analyses, as well as at the mRNA level by reverse transcription-polymerase chain reaction. RESULTS: The collagen-degrading ability of the human gingival fibroblasts increased in four cell lines (aggressive) and was only slightly altered in the other three cell lines (nonaggressive) in the presence of P. gingivalis supernatant. MMP-1, MMP-2, and MMP-3 more readily underwent activation while the TIMP-1 level was decreased in the conditioned media from a P. gingivalis-treated human gingival fibroblast aggressive cell line. None of these was altered in a nonaggressive cell line. The mRNA levels of the MMPs and TIMPs were only slightly different between these two cell lines. CONCLUSION: Heterogeneity exists in human gingival fibroblasts in regard to their collagenolytic activity in the presence of P. gingivalis.
Assuntos
Colágeno/metabolismo , Fibroblastos/metabolismo , Gengiva/metabolismo , Porphyromonas gingivalis/metabolismo , Linhagem Celular , Colágeno Tipo I/metabolismo , Meios de Cultivo Condicionados , Heterogeneidade Genética , Gengiva/citologia , Humanos , Metaloproteinase 1 da Matriz/metabolismo , Metaloproteinase 14 da Matriz/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Inibidor Tecidual de Metaloproteinase-2/metabolismoRESUMO
Degenerative temporomandibular joint (TMJ) disorders are characterized by the excessive turnover of collagen. In addition to the matrix metalloproteinase (MMP) and cathepsin mediated collagen degradation pathways, a serine proteinase dependent pathway has recently been identified in TMJ fibroblasts. This study focused on further characterizing this serine proteinase pathway utilizing a media-mediated collagen degradation assay and zymography. The conditioned media from cell-mediated collagen degradation assays were incubated with Type I collagen at pH 7.5 with or without a MMP inhibitor (1,10-phenanthroline), serine proteinase inhibitors (alpha1-antitrypsin and soybean trypsin inhibitor, STI), or cysteine proteinase inhibitors. The data showed that 1,10-phenanthroline and STI reduced the collagen cleavage by 12.33% and 47.78%, respectively. The cysteine proteinase inhibitors had no effect. The combination of alpha1-antitrypsin and 1,10-phenanthroline inhibited the cleavage by 79.22%, while STI and 1,10-phenanthroline together blocked the cleavage by 85.44%. Zymography identified a proteinase at approximate 22.5 kDa, which was more effectively blocked by serine proteinase inhibitors than by MMP or cysteine proteinase inhibitors. Reverse transcript-PCR and real-time PCR results demonstrated that TMJ cells did not express trypsinogen-2 mRNA, a collagen cleaving serine proteinase. This study demonstrated that TMJ fibroblasts can predominantly utilize a serine proteinase to mediate collagen degradation, which is not trypsinogen-2.
Assuntos
Colágeno Tipo I/metabolismo , Fibroblastos/metabolismo , Serina Endopeptidases/metabolismo , Articulação Temporomandibular/metabolismo , Linhagem Celular Tumoral , Meios de Cultivo Condicionados , Gelatina/metabolismo , Humanos , Fenantrolinas/farmacologia , Fluoreto de Fenilmetilsulfonil/farmacologia , Membrana Sinovial/citologia , Membrana Sinovial/metabolismo , Tripsina/análise , Tripsinogênio/análise , alfa 1-Antitripsina/farmacologiaRESUMO
Matrix metalloproteinases (MMPs) and their inhibitors have long been believed to play a major role in the collagen loss seen in destructive temporomandibular joint (TMJ) disorders. This project was originally designed to examine the expression of MMPs and the tissue inhibitors of MMPs (TIMPs) by diseased human TMJ synovial fibroblasts and to determine their ability to degrade Type I collagen. Reverse transcriptase-PCR indicated that these TMJ synovial fibroblasts expressed mRNA for multiple MMPs and TIMPs. The collagen degradation assay showed that these TMJ synovial fibroblasts at passage 3 to 8 were capable of digesting the collagen underneath them on collagen-coated plates. This degradation was inhibited by GM6001, a synthetic MMP inhibitor. During passage 8 to 13, these TMJ fibroblasts were able to digest all the collagen in the wells. This degradation was completely inhibited by combining GM6001 and soybean trypsin inhibitor (STI), a serine proteinase inhibitor. The collagen cleavage activity of collected conditioned media was dramatically inhibited by STI but not by 1,10-phenanthroline, an MMP inhibitor. The data suggest that these TMJ cells utilize a MMP-dependent pathway and a novel MMP-independent pathway to digest Type I collagen.
Assuntos
Colágeno/metabolismo , Metaloproteinases da Matriz/fisiologia , Transtornos da Articulação Temporomandibular/metabolismo , Articulação Temporomandibular/metabolismo , Células Cultivadas , Meios de Cultivo Condicionados , Fibroblastos/metabolismo , Humanos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Membrana Sinovial/citologia , Membrana Sinovial/metabolismoRESUMO
Diverse reports have described how various types of adhesive systems cause disastrous pulp necrosis, chronic severe inflammation or failure to stimulate any pulp reactions. This article reports on the effects of five common adhesive systems and how they compare in terms of pulp injury as measured by odontoblast survival or dentin regeneration and reactionary dentin formation. One hundred and thirty Class V pulp, non-exposed cavities were prepared in non-human primate teeth and were restored with five different adhesive systems. After a period of time between 3 and 172 days, the teeth were extracted, fixed, processed and examined histomorphometrically. Bacterial microleakage was detected with McKays stain and inflammation was categorized according to the International Organization for Standardization (ISO) criteria. The number of odontoblasts and the area of reactionary dentin were measured. Pulp reactions of all adhesive systems were generally minimal, although some systems permitted bacterial microleakage in 33% of restorations, and some other systems were associated with pulp inflammation in 22% of restorations. These observations suggest that adhesive systems provide acceptable biocompatibility, however, there is strong potential for improvement.
Assuntos
Resinas Compostas/farmacologia , Polpa Dentária/efeitos dos fármacos , Adesivos Dentinários/farmacologia , Análise de Variância , Animais , Materiais Biocompatíveis/farmacologia , Sobrevivência Celular , Distribuição de Qui-Quadrado , Preparo da Cavidade Dentária , Infiltração Dentária/microbiologia , Polpa Dentária/patologia , Restauração Dentária Permanente , Dentina/efeitos dos fármacos , Dentina/patologia , Dentina Secundária/efeitos dos fármacos , Dentina Secundária/patologia , Macaca mulatta , Metacrilatos/farmacologia , Odontoblastos/efeitos dos fármacos , Odontoblastos/patologia , Pulpite/induzido quimicamente , Pulpite/patologia , Cimentos de Resina/farmacologiaRESUMO
OBJECTIVES: To investigate the changes in morphology and activity of pulp odontoblasts in response to cavity restoration variables and patient factors. METHODS: Class V non exposed cavities were prepared in the intact 1st or 2nd premolar teeth of 27 patients, aged between 9 and 17 years-old. Following tooth extraction, the area of reactionary dentine and the area of the odontoblasts were measured using computerised histomorphometry. RESULTS: The cytoplasm to nucleus ratio of the odontoblasts was found to increase beneath cut dentinal tubules, following the secretion of reactionary dentine. However, none of the patient or preparation variables were found to be correlated with changes in the odontoblast cytoplasm to nucleus ratio. CONCLUSIONS: Morphological changes in human odontoblasts is directly related to their capacity to repair dentine injuries and provide pulp protection. Changes in odontoblast morphology reflect secretory activity.
Assuntos
Preparo da Cavidade Dentária/classificação , Dentina Secundária/fisiologia , Odontoblastos/citologia , Adolescente , Dente Pré-Molar , Hidróxido de Cálcio/uso terapêutico , Núcleo Celular/ultraestrutura , Criança , Citoplasma/ultraestrutura , Amálgama Dentário/química , Forramento da Cavidade Dentária , Polpa Dentária/patologia , Restauração Dentária Permanente/métodos , Dentina/ultraestrutura , Feminino , Humanos , Masculino , Metilmetacrilatos/química , Odontoblastos/metabolismo , Pulpite/patologia , Fatores de Tempo , Cimento de Óxido de Zinco e Eugenol/químicaRESUMO
AIM: To evaluate pulp responses as a function of remaining dentine thickness (RDT) of 98 class V cavity preparations in 49 teeth of 31 patients aged 10-16 years. METHODOLOGY: Shallow cavities were restored with amalgam, deeper cavities or pulp exposures were restored with amalgam lined with calcium hydroxide or with zinc oxide eugenol. Teeth were extracted after 3-89 days for orthodontic reasons. Following processing for light microscope analysis, the number of odontoblasts, pulp inflammation, and repair was recorded. RESULTS: In comparison with independent odontoblasts, the numbers of odontoblasts were reduced by 13.6% beneath a RDT of 2.5-0.5 mm, 33.7% beneath a RDT of 0.5-0.01 mm and 99.0% beneath pulp-exposed cavities. Reparative dentine was observed following pulp exposure and reactionary dentine was observed with a mean RDT of 0.77 mm (2.5-0.01 mm). Reactionary dentine secretion was influenced by RDT and restorative materials. Pulp inflammation was not influenced by RDT in the present study. CONCLUSIONS: Cavity RDT mediates a powerful influence on underlying pulp tissue vitality but it has little effect on reactionary dentine secretion and inflammatory activity. Gross tissue injury explains the poor pulp capping prognosis following exposure and underlies the need to avoid this type of injury. Following restoration, a RDT of 0.5 mm or greater is necessary to avoid evidence of pulp injury.
Assuntos
Preparo da Cavidade Dentária/efeitos adversos , Dentina Secundária/metabolismo , Dentina/anatomia & histologia , Pulpite/etiologia , Adolescente , Análise de Variância , Hidróxido de Cálcio/efeitos adversos , Contagem de Células , Criança , Restauração Dentária Permanente/efeitos adversos , Dentina Secundária/crescimento & desenvolvimento , Humanos , Odontoblastos/metabolismo , Análise de Regressão , Cimento de Óxido de Zinco e Eugenol/efeitos adversosRESUMO
AIM: The purpose of this study was to collect quantitative information about the numbers and dentine bridge secretory activity of odontoblast-like cells following dental pulp exposure. METHODOLOGY: The numbers and secretory activity of odontoblast-like cells were measured histomorphometrically between 7 days and 2 years in 161 pulp-exposed nonhuman primate teeth. The area of dentine bridges and the dimensions of cavity preparations were measured. The density of odontoblast-like cells and subjacent reorganizing tissue cells were measured beneath dentine bridge formation. The presence of operative dentine debris and tunnel defects in bridges was noted. Pulp inflammation was categorized according to ISO standards. Bacteria were detected using McKay's stain. RESULTS: The area of dentine bridges was mediated by the density and secretory activity of odontoblast-like cells over time. The cell density of subjacent reorganizing tissue was found to be strongly associated with that of odontoblast-like cells. Bacterial microleakage was found to impede dentine bridge secretion by odontoblast-like cells. CONCLUSIONS: Pulp reparative activity occurs naturally beneath capping materials in the absence of bacterial microleakage. The outcome of pulp-capping treatments could be beneficially influenced by concentrating attention on limiting the width of pulp exposure, minimizing pulp injury by limiting the creation of operative debris and placing materials which prevent bacterial microleakage.
Assuntos
Exposição da Polpa Dentária/fisiopatologia , Dentina Secundária/metabolismo , Odontoblastos/metabolismo , Análise de Variância , Animais , Contagem de Células , Infiltração Dentária/prevenção & controle , Capeamento da Polpa Dentária , Exposição da Polpa Dentária/terapia , Dentina Secundária/crescimento & desenvolvimento , Macaca mulatta , CicatrizaçãoRESUMO
OBJECTIVES: The purpose of this study is to compare and contrast differences of pulp responses between non-exposed and exposed cavity preparations in terms of inflammation, frequency of bacterial microleakage, odontoblast and odontoblastoid cell numbers, and tertiary dentine formation. METHODS: Class V non-exposed cavities (n=161) and exposed cavities (n=161 teeth) were prepared in non-human primate teeth. Cavities were restored with calcium hydroxide [Ca(OH)(2)], resin modified glass ionomer, or resin composite. Following extraction (7-730 days), bacteria were detected with McKays stain and pulp reactions were categorized according to ISO guidelines. Teeth were analyzed histomorphometrically and statistically using analysis of variance tests. RESULTS: Exposed cavities in comparison with non-exposed cavities were found to have more severe inflammation (p=0.0001), greater quantities of tertiary dentine (p=0.0001), and an increased frequency of bacterial microleakage (p=0.0034). The density of odontoblastoid cells beneath pulp exposed tertiary dentine was found to be 47.8% of odontoblast cell density beneath non-exposed dentine (p=0.0001). CONCLUSIONS: The restoration of exposed cavity preparations is associated with more traumatic pulp injury and repair responses. Consequently, efforts should be made to minimize iatrogenic dentine removal during cavity preparation and the creation of pulp exposures whenever possible.
Assuntos
Preparo da Cavidade Dentária/métodos , Exposição da Polpa Dentária/fisiopatologia , Polpa Dentária/fisiopatologia , Restauração Dentária Permanente , Análise de Variância , Animais , Bactérias/isolamento & purificação , Hidróxido de Cálcio/química , Contagem de Células , Distribuição de Qui-Quadrado , Corantes , Resinas Compostas/química , Forramento da Cavidade Dentária , Preparo da Cavidade Dentária/efeitos adversos , Infiltração Dentária/microbiologia , Polpa Dentária/microbiologia , Polpa Dentária/patologia , Capeamento da Polpa Dentária , Exposição da Polpa Dentária/etiologia , Exposição da Polpa Dentária/terapia , Dentina/microbiologia , Dentina/patologia , Dentina Secundária/patologia , Corantes Fluorescentes , Cimentos de Ionômeros de Vidro/química , Macaca mulatta , Análise Multivariada , Neutrófilos/patologia , Odontoblastos/patologia , Pulpite/etiologia , Pulpite/patologia , Cimentos de Resina/químicaRESUMO
OBJECTIVES: Following tooth pulp exposure, pulpal repair is accomplished by dentine bridge secretion by odontoblast-like cells. However, little information is available about the hierarchy of variables, which influence odontoblast-like cell numbers. The purpose of this study was to examine correlations between pulp capping events and odontoblast-like cell numbers. METHODS: Two hundred and fifty standardised pulp exposed cavities were prepared in non-human primate teeth according to ISO usage guidelines. Exposed pulps were capped with Calcium hydroxide [Ca(OH)(2)], and multi-step and self-etching primer composite resins. Teeth were collected from 3 to 60-days to observe pulp reactions. Statistical analysis was evaluated using analysis of variance. RESULTS: The hierarchy of variables correlated to odontoblast-like cells were the dentine bridge area (P = 0.0001), time since pulp exposure (P = 0.0001), odontoblast numbers opposite the exposure site (P = 0.0002), and pulp capping materials (P = 0.0313). Other pulp capping variables were found to be less likely to be correlated with odontoblast-like cell numbers. CONCLUSIONS: The area of dentine bridge formation is directly related to the numbers of odontoblast-like cells, cell activity is time dependent, and the cell numbers are much lower than original odontoblast cells. The time-lag between the appearance of odontoblast-like cells at the site of pulp exposure, and the limited numbers of these cells, explain why pulpal repair is difficult to achieve successfully following pulp exposure.
Assuntos
Capeamento da Polpa Dentária/métodos , Polpa Dentária/citologia , Dentina Secundária/efeitos dos fármacos , Dentina Secundária/metabolismo , Odontoblastos/metabolismo , Animais , Hidróxido de Cálcio/uso terapêutico , Polpa Dentária/efeitos dos fármacos , Capeamento da Polpa Dentária/classificação , Exposição da Polpa Dentária/terapia , Dentina Secundária/citologia , Adesivos Dentinários/uso terapêutico , Macaca , Metacrilatos/uso terapêutico , Minerais/uso terapêutico , Odontoblastos/citologia , Odontoblastos/efeitos dos fármacos , Fatores de TempoRESUMO
Oral squamous cell carcinomas are highly invasive lesions that destroy adjacent tissues and invade bone and muscle, which is most likely the result of matrix metalloproteinase (MMP) activity. We examined three cell lines derived from squamous cell carcinoma of the tongue for their intrinsic capacities to degrade interstitial collagen with the goal of identifying the matrix-degrading enzymes. SCC-25 and SCC-15 cells degrade reconstituted fibrillar type I collagen in the absence of exogenous growth factors or cytokines when seeded as a colony on dried films. Degradation is confined to the subjacent matrix, is enhanced 2-3-fold by phorbol ester, and is strictly MMP-dependent, as it is blocked by BB-94 and tissue inhibitor of metalloproteinases-2 but not by inhibitors of serine and cysteine proteinases. Both cell lines express active (M(r) 57,000) membrane type I-MMP (MT1-MMP) on their surfaces, as detected by surface biotinylation and immunoprecipitation. Concomitantly, both cell lines activate endogenous MMP-2 when cultured on type I collagen films, as assessed by zymography. Phorbol ester treatment enhances collagen-induced MMP-2 activation, which is accompanied by the appearance of a surface-labeled M(r) 43,000 form of MT1-MMP. Treatment of cells with a synthetic furin inhibitor, which inhibits processing of the MT1-MMP zymogen, blocks collagen degradation. In contrast, CAL 27 cells do not degrade collagen under either basal or phorbol 12-myristate 13-acetate-stimulated conditions. Although proMT1-MMP (M(r) 63,000/65,000) is detectable in these cells by immunoblot analysis, they express greatly reduced levels of active MT1-MMP on their surfaces relative to SCC-25 and SCC-15 cells. Correspondingly, CAL 27 cells cultured on collagen express neither latent nor active gelatinases. Immunoblots of lysates and conditioned media revealed the constitutive expression of proMMP-1 and proMMP-13 in all three cell lines. We conclude that in the absence of exogenous growth factors or accessory stromal cells, degradation of interstitial collagen by oral squamous cell carcinoma cells requires a threshold level of active MT1-MMP on cell surfaces.