RESUMO
Low-intensity, high-frequency mechanical vibration (LHMV) has shown to increase bone formation. However, studies comparing the effectiveness of early- and late-treatments of LHMV to counteract bone loss have not been documented. This study was designed to compare the effects of early- and late-treatments of LHMV (at 30 Hz/0.6 g, 20 min per day/five days per week, for 12 weeks) on bone parameters in ovariectomized (Ovx) rats. Thirty days after ovariectomy, 40 adult rats were randomly divided into four groups: GI (early control group); GII treated with LHMV 3 weeks after Ovx (early treatment); GIII (late control group) and GIV treated with LHMV twelve weeks after Ovx (late treatment). Bone mineral density (BMD) was analyzed before Ovx and after treatments. Then, animals were killed, and the femurs were collected and their length and diaphysis diameter were measured; the distal femurs were taken and processed for histomorphometry and polarized light microscopy for collagen fibers analysis or subjected to immunohistochemistry of cleaved caspase-3 in osteocytes. Statistical analysis was done by ANOVA followed by the Bonferroni post hoc test (p < 0.05). BMD was similar among the groups before Ovx, but after treatments, it was significantly higher in GII and GIV compared with their control groups (p < 0.05). Femur length and cortical bone thickness were similar among the groups, but the diaphysis diameter of GII was higher compared with GI. Trabecular bone area was higher in the vibrated groups, but it was greater in GII (p < 0.05). Also, the vibrated groups showed the higher content collagen fibers and lower presence apoptotic osteocytes (positive caspase-3 immunoreactivity) when compared with the other groups (p < 0.05). These results suggest that both early- and late-treatments with LHMV counteract bone loss, being the early treatment more effective than the late treatment.
Assuntos
Densidade Óssea/fisiologia , Ovariectomia , Vibração/uso terapêutico , Animais , Apoptose , Caspase 3/análise , Colágeno/análise , Feminino , Fêmur , Humanos , Imuno-Histoquímica , Osteócitos/enzimologia , Osteogênese/fisiologia , Osteoporose Pós-Menopausa/prevenção & controle , Estimulação Física , Ratos , Fatores de TempoRESUMO
OBJECTIVE: The present study investigated the efficacy of a biphasic calcium phosphate as a bone grafting material for maxillary sinus augmentation in humans. MATERIALS AND METHODS: Half of the thirty patients selected for sinus augmentation were grafted with biphasic calcium phosphate, whereas the other half were grafted with autogenous bone chips harvested intraorally. After 9 months of healing, bone cores were retrieved from implant sites for histologic and histomorphometric evaluation. RESULTS: The areas augmented with autogenous bone chips showed newly formed bone with a pattern very similar to that of the native area. Histomorphometry demonstrated that the amount of newly formed bone in the autogenous bone group was significantly greater than in the biphasic calcium phosphate group (P < 0.05). In the biphasic calcium phosphate group, less bone formation was observed in the area further away from native bone interface than in the area closer to native bone interface (P < 0.05), whereas no significant differences were observed between both areas in the autogenous group. In both groups, the implant survival rate was 100% with a minimum 1-year follow-up. CONCLUSION: The data presented in this work confirm the osteoconductive properties of biphasic calcium phosphate, as well as its use in association with maxillary sinus floor augmentation procedures with successful outcomes.
Assuntos
Substitutos Ósseos/uso terapêutico , Hidroxiapatitas/uso terapêutico , Levantamento do Assoalho do Seio Maxilar/métodos , Adolescente , Adulto , Idoso , Autoenxertos , Biópsia/métodos , Transplante Ósseo/métodos , Colágeno , Implantação Dentária Endóssea/métodos , Seguimentos , Humanos , Processamento de Imagem Assistida por Computador/métodos , Arcada Parcialmente Edêntula/reabilitação , Arcada Parcialmente Edêntula/cirurgia , Maxila/patologia , Maxila/cirurgia , Seio Maxilar/patologia , Membranas Artificiais , Pessoa de Meia-Idade , Osteogênese/fisiologia , Estudos Prospectivos , Resultado do Tratamento , Cicatrização/fisiologia , Adulto JovemRESUMO
The low level laser therapy (LLLT) has been used as an option to accelerate the regeneration of bone tissue. In this study, both femurs of male Wistar rats (30 animals) were injured with a drill and the effect of LLLT using a laser diode (100 mW at 660 nm) in the bone matrix on the left paw measured. LLLT effect on the healing bone tissue matrix was evaluated by a combination of immunohistochemical histomorphometry, confocal immunofluorescence microscopy and isolation and characterization of glycosaminoglycans. Histomorphometric analysis showed that LLLT increased bone matrix and showing more organized. Alcian Blue and PAS staining seems to suggest differential glycosaminoglycans and glycoproteins. The data showed increased expression of chondroitin sulfate and hyaluronic acid, after reduction as the LLLT and mature bone, resembling the expression of osteonectin and biglycan. The difference in expression of siblings (DMP-1, OPN and BSP) is in accordance with the repair accelerated bone formation after the application of LLLT as compared with control. The expression of osteonectin and osteocalcin supports their role in bone mineralization protein, indicating that LLLT accelerates this process. The overall data show that LLLT bone changes dynamic array, shortening the time period involved in the bone repair.
Assuntos
Matriz Óssea/efeitos da radiação , Regeneração Óssea/efeitos da radiação , Fêmur/efeitos da radiação , Terapia com Luz de Baixa Intensidade , Azul Alciano , Animais , Matriz Óssea/lesões , Sulfatos de Condroitina/biossíntese , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Fêmur/lesões , Expressão Gênica/efeitos da radiação , Ácido Hialurônico/biossíntese , Imuno-Histoquímica , Sialoproteína de Ligação à Integrina/genética , Sialoproteína de Ligação à Integrina/metabolismo , Lasers , Masculino , Microscopia de Fluorescência , Osteocalcina/genética , Osteocalcina/metabolismo , Osteonectina/genética , Osteonectina/metabolismo , Reação do Ácido Periódico de Schiff , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Ratos , Ratos WistarRESUMO
This study investigated structural and functional features of apoptotic alveolar bone osteoclasts in estrogen-treated rats. For this purpose, 15 female rats 22 days old were divided into three groups: Estrogen (EG), Sham (SG) and Control (CG). The rats of EG received daily intramuscular injection of estrogen for 7 days. The SG received only the oil vehicle. Maxillary fragments containing alveolar bone were removed and processed for light and transmission electron microscopy. Area (OcA) and number of nuclei (OcN) and bone resorption surface per TRAP-positive osteoclasts (BS/OC) were obtained. Vimentin, caspase-3 and MMP-9 immunoreactions, TUNEL/TRAP and MMP-9/TUNEL combined reactions were performed. In EG, the OcA, OcN and BS/Oc were reduced. Moreover, osteoclasts showed cytoplasm immunolabelled by caspase-3 and a different pattern of vimentin expression in comparison with CG and SG. MMP-9 expression was not affected by estrogen and the TUNEL-positive osteoclasts were MMP-9-immunolabelled. In EG, ultrastructural images showed that apoptotic osteoclasts did not exhibit ruffled borders or clear zones and were shedding mononucleated portions. TRAP-positive structures containing irregular and dense chromatin were partially surrounded by fibroblast-like cells. In conclusion, the reduction in the BS/Oc may be due to reduction in OcA and OcN; these effects seem to be related to vimentin disarrangement rather than to an interference of estrogen with osteoclast MMP-9 expression. Osteoclast apoptosis involves caspase-3 activity and vimentin degradation; these cells release portions containing one apoptotic nucleus and, subsequently, undergo fragmentation, giving rise to apoptotic bodies.
Assuntos
Processo Alveolar/efeitos dos fármacos , Reabsorção Óssea/prevenção & controle , Estrogênios/farmacologia , Osteoclastos/efeitos dos fármacos , Processo Alveolar/citologia , Processo Alveolar/metabolismo , Animais , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Núcleo Celular/efeitos dos fármacos , Feminino , Injeções Intramusculares , Metaloproteinase 9 da Matriz/metabolismo , Microscopia Eletrônica de Transmissão , Osteoclastos/metabolismo , Osteoclastos/ultraestrutura , Ratos , Ratos Sprague-Dawley , Vimentina/metabolismoRESUMO
Osteoclasts, the multinucleated bone-resorbing cells, arise through fusion of precursors from the myeloid lineage. However, not all osteoclasts are alike; osteoclasts at different bone sites appear to differ in numerous respects. We investigated whether bone marrow cells obtained from jaw and long bone differed in their osteoclastogenic potential. Bone marrow cells from murine mandible and tibiae were isolated and cultured for 4 and 6 days on plastic or 6 and 10 days on dentin. Osteoclastogenesis was assessed by counting the number of TRAP(+) multinucleated cells. Bone marrow cell composition was analyzed by FACS. The expression of osteoclast- and osteoclastogenesis-related genes was studied by qPCR. TRAP activity and resorptive activity of osteoclasts were measured by absorbance and morphometric analyses, respectively. At day 4 more osteoclasts were formed in long bone cultures than in jaw cultures. At day 6 the difference in number was no longer observed. The jaw cultures, however, contained more large osteoclasts on plastic and on dentin. Long bone marrow contained more osteoclast precursors, in particular the myeloid blasts, and qPCR revealed that the RANKL:OPG ratio was higher in long bone cultures. TRAP expression was higher for the long bone cultures on dentin. Although jaw osteoclasts were larger than long bone osteoclasts, no differences were found between their resorptive activities. In conclusion, bone marrow cells from different skeletal locations (jaw and long bone) have different dynamics of osteoclastogenesis. We propose that this is primarily due to differences in the cellular composition of the bone site-specific marrow.
Assuntos
Células da Medula Óssea/fisiologia , Osso e Ossos/citologia , Diferenciação Celular , Arcada Osseodentária/citologia , Osteoclastos/fisiologia , Animais , Medula Óssea/metabolismo , Medula Óssea/fisiologia , Células da Medula Óssea/metabolismo , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/metabolismo , Contagem de Células , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Células Cultivadas , Feminino , Regulação da Expressão Gênica , Arcada Osseodentária/diagnóstico por imagem , Arcada Osseodentária/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Células Mieloides/citologia , Células Mieloides/fisiologia , Osteoclastos/metabolismo , Osteoprotegerina/genética , Osteoprotegerina/metabolismo , Ligante RANK/genética , Ligante RANK/metabolismo , Microtomografia por Raio-XRESUMO
Paradoxical growth (PG) has been described for echinocandins and is characterized by cell growth at drug concentrations above the MIC. In this study, two isolates each of Candida albicans, C. tropicalis, C. orthopsilosis, and C. parapsilosis, all of which displaying PG in response to caspofungin, were subjected to MIC, minimal fungicidal concentration (MFC), and time-kill curve assays to evaluate the levels of PG. Cell wall components and ultrastructural modifications of the PG cells were also investigated. The results showed that when cell growth and survival were evaluated by MFC or time-kill curve assays, high concentrations of caspofungin did not show fungicidal activity against PG cells. Furthermore, for C. parapsilosis and C. orthopsilosis, time-kill curves were more discriminatory than MFCs in detecting the PG effect. The four different Candida species studied demonstrated similar alterations in cell wall components and ultrastructure associated with PG. In PG cells, ß-1,3-glucan content decreased from 2.7- to 7.8-fold, whereas chitin content increased from 4.0- to 6.6-fold. An electron microscopy study of the PG cells revealed morphological alterations, clumping of cells, enlarged cells, the absence of filamentation, abnormal septa, and accumulation of chitin in the cell wall. Also, PG cells basically exhibited a single dark high-density layer in the cell wall, indicating the loss of the ß-1,3-glucan layer. Our results present novel details about the ultrastructural alterations that occur in C. albicans, C. parapsilosis, C. orthopsilosis, and C. tropicalis during PG and show that chitin is the major component of the cell walls of PG cells. Stimulation of chitin synthesis may represent a rescue mechanism against caspofungin activity.
Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Candida/ultraestrutura , Parede Celular/efeitos dos fármacos , Parede Celular/ultraestrutura , Equinocandinas/farmacologia , Caspofungina , Lipopeptídeos , Testes de Sensibilidade Microbiana , Microscopia de Força Atômica , Microscopia Eletrônica de Transmissão , Microscopia de FluorescênciaRESUMO
We investigated the effects of gamma-radiation on cells isolated from the longitudinal smooth muscle layer of the guinea pig ileum, a relatively radioresistant tissue. Single doses (up to 50 Gy) reduced the amount of sarcoplasmatic reticulum and condensed the myofibrils, as shown by electron microscopy 3 days post-irradiation. After that, contractility of smooth muscle strips was reduced. Ca(2+) handling was altered after irradiation, as shown in fura-2 loaded cells, with elevated basal intracellular Ca(2+), reduced amount of intrareticular Ca(2+), and reduced capacitive Ca(2+) entry. Radiation also induced apoptosis, judged from flow cytometry of cells loaded with proprium iodide. Electron microscopy showed that radiation caused condensation of chromatin in dense masses around the nuclear envelope, the presence of apoptotic bodies, fragmentation of the nucleus, detachment of cells from their neighbors, and reductions in cell volume. Radiation also caused activation of caspase 12. Apoptosis was reduced by the administration of the caspase inhibitor Z-Val-Ala-Asp-fluoromethyl-ketone methyl ester (Z-VAD-FMK) during the 3 day period after irradiation, and by the chelator of intracellular Ca(2+), 1,2-bis(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA), from 1 h before until 2 h after irradiation. BAPTA also reduced the effects of radiation on contractility, basal intracellular Ca(2+), amount of intrareticular Ca(2+), capacitative Ca(2+) entry, and apoptosis. In conclusion, the effects of gamma radiation on contractility, Ca(2+) handling, and apoptosis appear due to a toxic action of intracellular Ca(2+). Ca(2+)-induced damage to the sarcoplasmatic reticulum seems a key event in impaired Ca(2+) handling and apoptosis induced by gamma-radiation.
Assuntos
Apoptose/efeitos da radiação , Raios gama , Íleo/efeitos da radiação , Miócitos de Músculo Liso/efeitos da radiação , Retículo Sarcoplasmático/efeitos da radiação , Animais , Cálcio/metabolismo , Caspases/fisiologia , Células Cultivadas , Ácido Egtázico/análogos & derivados , Ácido Egtázico/farmacologia , Cobaias , Íleo/fisiologia , Contração Muscular/efeitos da radiação , Miócitos de Músculo Liso/fisiologia , Retículo Sarcoplasmático/fisiologiaRESUMO
O estudo avaliou clinica e histologicamente a utilização de substituto ósseo (Bonefill - xenoenxerto bovino inorgânico) como material de preenchimento ósseo em cirurgias de levantamento de seio maxilar em humanos. Cinco pacientes cuidadosamente selecionados foram submetidos a cirurgias de aumento ósseo sinusal bilateral, sendo um lado enxertado com osso bovino inorgânico e o outro com enxerto ósseo autógeno removido de área doadora intrabucal. Após nove meses de reparação óssea sem complicações, realizaram-se as cirurgias para colocação de implantes osseointegrados nas áreas enxertadas de cada paciente, momento em que foram colhidas amostras do tecido neoformado para avaliação histológica. Clinicamente, o osso bovino inorgânico originou um tecido de consistência amolecida, diferente do tecido ósseo normal. Histologicamente, o osso bovino inorgânico foi incapaz de propiciar neoformação óssea previsível entre suas partículas mineralizadas, as quais encontravam-se completamente envoltas por um tecido conjuntivo denso fibroso. Concluiu-se, portanto que, clínica e histologicamente, o osso bovino inorgânico não foi eficaz como material de enxerto ósseo em humanos.