Efeito das condições de preparação e sinterização sobre a porosidade da hidroxiapatita

Este estudo foi realizado para verificar o efeito das condições de preparação e sinterização sobre a porosidade de amostras de hidroxiapatita, calculada como porcentagem da densidade teórica da hidroxiapatita. Foram verificados o efeito da carga utilizada para obter as amostras, o tamanho inicial das partículas de hidroxiapatita, a temperatura e o tempo de sinterização. Existiu uma correlação inversa entre carga e porosidade e entre temperatura de sinterização e porosidade, mas nenhuma correlação entre tempo de sinterização e porosidade. Amostras fabricadas com partículas de tamanho maior apresentaram maior porosidade. Foi concluído que para a fabricação de amostras de hidroxiapatita, a combinação entre a carga utilizada para a compactação das partículas e a temperatura de sinterização resulta na produção de amostras com uma ampla variação de porcentagens de porosidade.The aim of this study was to evaluate the effect of powder processing and sintering conditions on the resultant porosity of hydroxyapatite, which was calculated as percentage of the theoretical density of hydroxyapatite. The parameters evaluated were the effect of load during uniaxial pressing, the initial size of the particles of hydroxyapatite, and sintering temperature and time. There was an inverse correlation between load and porosity, and between sintering temperature and porosity, but there was no correlation between sintering time and porosity. An increase in the size of the particles resulted in greater porosity. It was concluded that a combination of loading conditions and sintering temperature can provide hydroxyapatite with a wide range of porosities

Surface topography of hydroxyapatite affects ROS17/2.8 cells response

A hidroxiapatita (HA) tem sido utilizada como revestimento de implantes e para substituição de tecido ósseo. O objetivo deste estudo foi avaliar o efeito da topografia de superfície da HA, resultante da presença de microporosidade, sobre a adesão, a morfologia e proliferação celulares, a medida de proteína total e a atividade de fosfatase alcalina. Discos de HA com diferentes porcentagens de microporosidade (< 5%, 15% e 30%) foram fabricados por uma combinação das técnicas de pressão uniaxial e sinterização. Células ROS17/2.8 foram cultivadas sobre os discos de HA. Para a adesão, as células foram cultivadas por duas horas. A morfologia foi avaliada após sete dias. A proliferação, medida de proteína total e atividade de ALP foram avaliadas após sete e quatorze dias. Os dados foram comparados por ANOVA e teste de Duncan quando apropriado. A adesão (p = 0,11) e a medida de proteína total (p = 0,31) não foram afetadas pela topografia de superfície. A proliferação após sete e quatorze dias (p = 0,0007 e p = 0,003, respectivamente), e a atividade de ALP (p = 0,0007) foram significantemente menores na superfície irregular (HA30). Esses resultados sugerem que eventos iniciais não são afetados pela topografia, enquanto superfícies com topografias mais regulares (microporosidade de 15% ou menos) favoreceram eventos intermediários e finais, como proliferação e atividade de ALP.Hydroxyapatite (HA) has been used in orthopedic, dental, and maxillofacial surgery as a bone substitute. The aim of this investigation was to study the effect of surface topography produced by the presence of microporosity on cell response, evaluating: cell attachment, cell morphology, cell proliferation, total protein content, and alkaline phosphatase (ALP) activity. HA discs with different percentages of microporosity (< 5%, 15%, and 30%) were confected by means of the combination of uniaxial powder pressing and different sintering conditions. ROS17/2.8 cells were cultured on HA discs. For the evaluation of attachment, cells were cultured for two hours. Cell morphology was evaluated after seven days. After seven and fourteen days, cell proliferation, total protein content, and ALP activity were measured. Data were compared by means of ANOVA and Duncans multiple range test, when appropriate. Cell attachment (p = 0.11) and total protein content (p = 0.31) were not affected by surface topography. Proliferation after 7 and 14 days (p = 0.0007 and p = 0.003, respectively), and ALP activity (p = 0.0007) were both significantly decreased by the most irregular surface (HA30). These results suggest that initial cell events were not affected by surface topography, while surfaces with more regular topography, as those present in HA with 15% or less of microporosity, favored intermediary and final events such as cell proliferation and ALP activity

Bone response to a Ca- and P-enriched titanium surface obtained by anodization

This study evaluated bone response to a Ca- and P- enriched titanium (Ti) surface treated by a multiphase anodic spark deposition coating (BSP-AK). Two mongrel dogs received bilateral implantation of 3 Ti cylinders (4.1 x 12 mm) in the humerus, being either BSP-AK treated or untreated (machined - control). At 8 weeks postimplantation, bone fragments containing the implants were harvested and processed for histologic and histomorphometric analyses. Bone formation was observed in cortical area and towards the medullary canal associated to approximately 1/3 of implant extension. In most cases, in the medullary area, collagen fiber bundles were detected adjacent and oriented parallel to Ti surfaces. Such connective tissue formation exhibited focal areas of mineralized matrix lined by active osteoblasts. The mean percentages of bone-to-implant contact were 2.3 (0.0-7.2 range) for BSP-AK and 0.4 (0.0-1.3 range) for control. Although the Mann-Whitney test did not detect statistically significant differences between groups, these results indicate a trend of BSP-AK treated surfaces to support contact osteogenesis in an experimental model that produces low bone-to-implant contact values.O objetivo desse estudo foi avaliar a resposta do tecido ósseo à superfície de titânio (Ti) enriquecida com Ca e P obtida por anodização (BSP-AK). Três cilindros de Ti (4,1 x 12 mm) BSP-AK ou usinado (controle) foram implantados bilateralmente nos úmeros de dois cães de raça indefinida. Oito semanas após a implantação, os fragmentos ósseos contendo os implantes foram removidos e processados para análises histológica e histomorfométrica. A formação óssea foi observada na região cortical e no canal medular até aproximadamente um terço da extensão do implante. Na maioria dos casos, feixes de fibras colágenas dispostos paralelamente à superfície do implante foram observados na região medular. Nessa região observaram-se também áreas focais de formação de matriz mineralizada e osteoblastos ativos. Os implantes do grupo BSP-AK apresentaram média de contato osso-implante 2,3%, com medidas variando de 0,0 a 7,2% e os do grupo controle tiveram média 0,4%, com medidas variando de 0,0 a 1,3%. Apesar do teste de Mann-Whitney não mostrar diferença estatisticamente significante entre os grupos, nossos resultados indicaram uma tendência para a ocorrência de osteogênese de contato na superfície BSP-AK em um modelo experimental que resulta em baixos valores de contato osso-implante.FAPES

Effect of focal adhesion kinase inhibition on osteoblastic cells grown on titanium with different topographies

Objective:The present study aimed to investigate the participation of focal adhesion kinases (FAK) in interactions between osteoblastic cells and titanium (Ti) surfaces with three different topographies, namely, untreated (US), microstructured (MS), and nanostructured (NS). Methodology; Osteoblasts harvested from the calvarial bones of 3-day-old rats were cultured on US, MS and NS discs in the presence of PF-573228 (FAK inhibitor) to evaluate osteoblastic differentiation. After 24 h, we evaluated osteoblast morphology and vinculin expression, and on day 10, the following parameters: gene expression of osteoblastic markers and integrin signaling components, FAK protein expression and alkaline phosphatase (ALP) activity. A smooth surface, porosities at the microscale level, and nanocavities were observed in US, MS, and NS, respectively. Results: FAK inhibition decreased the number of filopodia in cells grown on US and MS compared with that in NS. FAK inhibition decreased the gene expression of Alp, bone sialoprotein, osteocalcin, and ALP activity in cells grown on all evaluated surfaces. FAK inhibition did not affect the gene expression of Fak, integrin alpha 1 ( Itga1 ) and integrin beta 1 ( Itgb1 ) in cells grown on MS, increased the gene expression of Fak in cells grown on NS, and increased the gene expression of Itga1 and Itgb1 in cells grown on US and NS. Moreover, FAK protein expression decreased in cells cultured on US but increased in cells cultured on MS and NS after FAK inhibition; no difference in the expression of vinculin was observed among cells grown on all surfaces. Conclusions: Our data demonstrate the relevance of FAK in the interactions between osteoblastic cells and Ti surfaces regardless of surface topography. Nanotopography positively regulated FAK expression and integrin signaling pathway components during osteoblast differentiation. In this context, the development of Ti surfaces with the ability to upregulate FAK activity could positively impact the process of implant osseointegration

Aging impairs osteoblast differentiation of mesenchymal stem cells grown on titanium by favoring adipogenesis

Aging negatively affects bone/titanium implant interactions. Our hypothesis is that the unbalance between osteogenesis and adipogenesis induced by aging may be involved in this phenomenon. Objective We investigated the osteoblast and adipocyte differentiation of mesenchymal stem cells (MSCs) from young and aged rats cultured on Ti. Material and Methods Bone marrow MSCs derived from 1-month and 21-month rats were cultured on Ti discs under osteogenic conditions for periods of up to 21 days and osteoblast and adipocyte markers were evaluated. Results Cell proliferation, alkaline phosphatase (ALP) activity, extracellular matrix mineralization and gene expression of RUNX2, osterix, ALP, bone sialoprotein, osteopontin, and osteocalcin were reduced in cultures of 21-month rats compared with 1-month rats grown on Ti. Gene expression of PPAR-γ , adipocyte protein 2, and resistin and lipid accumulation were increased in cultures of 21-month rats compared with 1-month rats grown on the same conditions. Conclusions These results indicate that the lower osteogenic potential of MSCs derived from aged rats compared with young rats goes along with the higher adipogenic potential in cultures grown on Ti surface. This unbalance between osteoblast and adipocyte differentiation should be considered in dental implant therapy to the elderly population

Effects of a novel calcium aluminate cement on the early events of the progression of osteogenic cell cultures

The present study evaluated the progression of osteogenic cell cultures exposed to a novel calcium aluminate cement (CAC+) in comparison with the gold standard mineral trioxide aggregate (MTA). Cells were enzimatically isolated from newborn rat calvarial bone, plated on glass coverslips containing either CAC+ or a control MTA samples in the center, and grown under standard osteogenic conditions. Over the 10-day culture period, roundening of sample edges was clearly noticed only for MTA group. Although both cements supported osteogenic cell adhesion, spreading, and proliferation, CAC+-exposed cultures showed significantly higher values in terms of total cell number at days 3 and 7, and total protein content and alkaline phosphatase activity at day 10. The present in vitro results indicate that the exposure to CAC+ supports a higher differentiation of osteogenic cells compared with the ones exposed to MTA. Further experimental studies should consider CAC+ as a potential alternative to MTA when the repair of mineralized tissues is one of the desired outcomes in endodontic therapy.O objetivo do presente estudo foi avaliar a progressão de cultura de células osteogênicas expostas a um novo cimento de aluminato de cálcio (CAC+) em comparação ao agregado de trióxido mineral (MTA). As células foram obtidas por digestão enzimática de calvária de ratos recém-nascidos, plaqueadas sobre lamínulas de vidro contendo em sua área central discos de CAC+ ou MTA e crescidas em condições osteogênicas por até 10 dias. Durante a cultura primária, observou-se o arredondamento das bordas das amostras de cimento apenas para MTA. Embora ambos os cimentos tenham permitido a adesão, o espraiamento e a proliferação celulares, as culturas crescidas em contato com CAC+ exibiram valores maiores de número total de células em 3 e 7 dias, e de conteúdo de proteína total e atividade de fosfatase alcalina em 10 dias. Os resultados indicam que a exposição ao CAC+ permite o desenvolvimento de uma proporção maior de células em estágios mais avançados da diferenciação osteoblástica, quando comparado ao MTA. Deve-se considerar em futuros estudos experimentais a utilização do CAC+ como um material alternativo ao MTA especialmente quando um dos objetivos do tratamento endodôntico é o de reparação dos tecidos mineralizados da região periapical.(FAPESP) São Paulo Research Foundation(CNPq) National Council of Scientific and Technological Developmen

Surface topography of hydroxyapatite affects ROS17/2.8 cells response

Hydroxyapatite (HA) has been used in orthopedic, dental, and maxillofacial surgery as a bone substitute. The aim of this investigation was to study the effect of surface topography produced by the presence of microporosity on cell response, evaluating: cell attachment, cell morphology, cell proliferation, total protein content, and alkaline phosphatase (ALP) activity. HA discs with different percentages of microporosity (< 5%, 15%, and 30%) were confected by means of the combination of uniaxial powder pressing and different sintering conditions. ROS17/2.8 cells were cultured on HA discs. For the evaluation of attachment, cells were cultured for two hours. Cell morphology was evaluated after seven days. After seven and fourteen days, cell proliferation, total protein content, and ALP activity were measured. Data were compared by means of ANOVA and Duncan’s multiple range test, when appropriate. Cell attachment (p = 0.11) and total protein content (p = 0.31) were not affected by surface topography. Proliferation after 7 and 14 days (p = 0.0007 and p = 0.003, respectively), and ALP activity (p = 0.0007) were both significantly decreased by the most irregular surface (HA30). These results suggest that initial cell events were not affected by surface topography, while surfaces with more regular topography, as those present in HA with 15% or less of microporosity, favored intermediary and final events such as cell proliferation and ALP activity

Characterization and in vitro cytocompatibility of an acid-etched titanium surface

The aims of this study were to characterize the microstructure of a commercially pure titanium (cpTi) surface etched with HCl/H2SO4 (AE-cpTi) and to investigate its in vitro cytocompatibility compared to turned cpTi (T-cpTi). T-cpTi showed a grooved surface and AE-cpTi revealed a surface characterized by the presence of micropits. Surface parameters indicated that the AE-cpTi surface is more isotropic and present a greater area compared to T-cpTi. The oxide film thickness was similar between both surfaces; however, AE-cpTi presented more Ti and O and less C. Osteoblastic cell proliferation, alkaline phosphatase activity, and bone-like nodule formation were greater on T-cpTi than on AE-cpTi. These results show that acid etching treatment produced a surface with different topographical and chemical features compared to the turned one, and such surface modification affected negatively the in vitro cytocompatibility of cpTi as demonstrated by decreasing culture growth and expression of osteoblastic phenotype.O objetivo deste estudo foi caracterizar a microestrutura de uma superfície de titânio comercialmente puro (cpTi) condicionada com HCl/H2SO4 (acid etched) (AE-cpTi) e investigar sua citocompatibilidade in vitro, comparada à do cpTi usinado (turned) (T-cpTi). O T-cpTi apresentou uma superfície com sulcos e o AE-cpTi exibiu uma superfície caracterizada pela presença de micro-vales. Os parâmetros de superfície indicaram que a superfície AE-cpTi é mais isotrópica e apresenta uma área maior quando comparada à superfície T-cpTi. A espessura da camada de óxido foi similar para as duas superfícies; no entanto, a AE-cpTi apresentou maiores quantidades de Ti e O e menor, de C. A proliferação de células osteoblásticas, a atividade de fosfatase alcalina e a formação de matriz mineralizada foram maiores na superfície T-cpTi que na AE-cpTi. Esses resultados mostram que o condicionamento ácido produziu uma superfície com características topográficas e químicas diferentes quando comparadas às da superfície usinada. Além disso, observou-se que essas modificações de superfície afetaram de forma negativa a citocompatibilidade in vitro do cpTi como demonstrado pela inibição da proliferação celular e da expressão do fenótipo osteoblástico

Bone tissue response to plasma-nitrided titanium implant surfaces

A current goal of dental implant research is the development of titanium (Ti) surfaces to improve osseointegration. Plasma nitriding treatments generate surfaces that favor osteoblast differentiation, a key event to the process of osteogenesis. Based on this, it is possible to hypothesize that plasma-nitrided Ti implants may positively impact osseointegration. Objective The aim of this study was to evaluate the in vivo bone response to Ti surfaces modified by plasma-nitriding treatments. Material and Methods Surface treatments consisted of 20% N2 and 80% H2, 450°C and 1.5 mbar during 1 h for planar and 3 h for hollow cathode. Untreated surface was used as control. Ten implants of each surface were placed into rabbit tibiae and 6 weeks post-implantation they were harvested for histological and histomorphometric analyses. Results Bone formation was observed in contact with all implants without statistically significant differences among the evaluated surfaces in terms of bone-to-implant contact, bone area between threads, and bone area within the mirror area. Conclusion Our results indicate that plasma nitriding treatments generate Ti implants that induce similar bone response to the untreated ones. Thus, as these treatments improve the physico-chemical properties of Ti without affecting its biocompatibility, they could be combined with modifications that favor bone formation in order to develop new implant surfaces