Desenvolvimento, caracterização e avaliação in vitro de cimento ósseo à base de nano hidroxiapatitas substituídas com magnésio, estrôncio e zinco / Development, characterization and in vitro evaluation of bone cement based on nano-hydroxyapatites substituted with magnesium, strontium and zinc

A incorporação de íons na estrutura da hidroxiapatita (HA) pode afetar sua estrutura, cristalinidade, solubilidade e citotoxicidade. Dentre os íons presentes na composição da apatita óssea, o magnésio (Mg2+), estrôncio (Sr2+) e zinco (Zn2+) são reconhecidos por promover a angiogênese e osteogênese. Portanto, as HAs substituídas podem apresentar melhor bioatividade, por fornecer íons com potencial de estimular a neoformação óssea nos locais enxertados. Nesse contexto, este estudo descreve a síntese, caracterização e comparação de uma série de nano-hidroxiapatitas (nHAs) substituídas e co-substituídas por Sr2+, Mg2+ ou Zn2+. Em seguida, foi desenvolvido um cimento ósseo à base das HAs com melhores resultados de citotoxicidade, associado ao DCPA, gelatina e quitosana. As nHAs foram caracterizadas físico-quimicamente usando diferentes técnicas. O método de co-precipitação foi eficaz para sintetizar HAs de dimensões nanométricas. Comparado a nHA pura, os difratogramas, espectros de FTIR e parâmetros de rede das nHAs substituídas e co-substituídas exibiram alterações, indicando que a incorporação de cátions resultou em distorções da rede da HA. Os testes de MTT demonstraram que as nHAs sintetizadas não foram citotóxicas, após contato direto com culturas de fibroblastos (L929) e pré-osteoblastos (MC3T3). Os resultados obtidos sugerem que as nHAs co-substituídas por Mg2+/Sr2+ e Zn2+/Sr2+ parecem induzir maior proliferação de células fibroblásticas e osteoblásticas, quando comparado a HA pura e substituída. Os cimentos ósseos desenvolvidos apresentaram capacidade de auto-endurecimento e resistência à lavagem. Além de possuírem alta molhabilidade e um perfil de liberação de íons Ca2+, Sr2+, Mg2+ e Zn2+, que está dentro das doses indicadas para estimular a proliferação de osteoblastos. Os cimentos exibiram excelente biocompatibilidade in vitro em culturas de células fibroblásticas, endoteliais e osteosblásticas. Os cimentos contendo nHAs co-substituídas por Mg2+/Sr2+ exibiram os melhores resultados de viabilidade celular. Após 24 horas de contato indireto com cultura de células fibroblásticas L929, o crescimento celular do grupo C2 foi maior que de todos os grupos em estudo (P < 0,01). Em cultura de células endoteliais EA.hy926, o percentual de células viáveis do grupo C3 foi significativamente maior que de todos os outros grupos, após 24 horas (p < 0,001). A citotoxicidade indireta em cultura de células pré-osteoblásticas MC3T3 revelou que após 48 horas, o grupo C3 apresentou maior viabilidade celular que todos os grupos em estudo (p < 0,01). O teste de formação de tubo sugere que todos os cimentos desenvolvidos possuem potencial angiogênico, sendo que os cimentos contendo nHAs co-substituídas por Zn2+/Sr2+ exibiram resultados significativamente superiores (p < 0,001). Apesar de ser necessário um maior número de testes de biocompatibilidade; a incorporação de íons na rede cristalina das nHAs, que são reconhecidos por afetar a angiogênese e a osteogênese, parece ter resultado no desenvolvimento de cimentos ósseos com potencial para promover a regeneração óssea.

The incorporation of ions into the HA lattice can affect its structure, crystallinity, solubility and cytotoxicity. From the ions present in the composition of bone apatite, Mg2+, Sr2+ and Zn2+ are recognized for promoting angiogenesis and osteogenesis. The substituted HAs can be present better bioactivity for supplying ions with potential to stimulate bone neoformation in grafted sites. This study described the synthesis, characterization and comparison of a range of substituted and co-substituted nHAs contained Sr2+, Mg2+ or Zn2+. Then, it developed bone cement based on HAs with better cytotoxicity results, associated with DCPA, gelatin and chitosan. The nHAs were physicochemically characterized using different techniques. The co-precipitation method was effective for synthesizing HAs with nanometric dimensions. Compared to pure nHA, the diffractograms, FTIR spectra and lattice parameters of the substituted and co-substituted nHAs showed changes, indicating that the incorporation of cations resulted in distortions of the HA lattice. MTT tests demonstrated that the all synthesized nHAs were not cytotoxic after direct contact with fibroblasts (L929) and pre-osteoblasts (MC3T3) cultures. MTT results suggest that Mg2+/Sr2+ and Zn2+/Sr2+ co-substituted nHAs seem to induce more proliferation of fibroblastic and osteoblastic than pure and Mg2+, Sr2+ and Zn2+ substituted nHAs. Bone cements developed showed self-hardening and washout resistance. Also, they Exhibited high wettability and ion release profile with non-toxic concentrations of Ca2+, Sr2+, Mg2+ and Zn2+, range within indicated doses to stimulate the proliferation of osteoblasts. The cement exhibited excellent in vitro cytocompatibility in fibroblastic, endothelial and osteoblastic cell cultures. Cement containing Mg2+/Sr2+ co-substituted nHAs showed better results of the cell viability. After 24 hours of indirect contact with L929 fibroblast culture, the cell growth in the C2 group was highest than all study groups (P <0.01). In EA.hy926 endothelial culture, the cell viability of the C3 group was significantly highest than all other groups after 24 hours (p <0.001). The indirect cytotoxicity in MC3T3 pre-osteoblastic culture revealed that after 48 hours, the C3 group showed the greatest cell viability than all the study groups (p <0.01). The tube formation assay suggests that all cement have angiogenic potential, being that the cements containing Zn2+ / Sr2+ co-substituted nHAs exhibited significantly better results (p < 0,001). Despite being necessary to perform a more significant number of biocompatibility tests, the incorporation of ions into the nHA lattice, which are recognized for affects angiogenesis and osteogenesis, may have resulted in the development of bone cements with the potential to promoting bone regeneration.

Estudio comparativo de la bioactividad de dos materiales biocerámicos / Comparative Study of the Bioactivity of Two Bioceramic Materials

RESUMEN Los materiales a base de silicato de calcio han demostrado ser bioactivos debido a su capacidad para producir apatita carbonatada biológicamente compatible. El objetivo de este estudio fue analizar la bioactividad de Biodentine™ y MTA Repair HP® en contacto con discos de dentina humana, que se obturaron y dividieron aleatoriamente para formar cuatro grupos: grupo 1 Biodentine™, grupo 2 MTA Repair HP®, grupo control positivo MTA Angelus® y grupo control negativo IRM®, los cuales se incubaron en solución PBS durante 10 días, para posterior análisis por medio de MEB-EDS y Espectroscopía Raman. Los tres materiales a base de silicato de calcio analizados en este estudio demostraron ser bioactivos pues al entrar en contacto con una solución a base de fosfato desencadenaron la precipitación inicial de fosfato de calcio amorfo, que actúa como precursor durante la formación de apatita carbonatada.

ABSTRACT Calcium silicate-based materials have been shown to be bioactive due to their ability to produce biologically compatible carbonated apatite. The objective of this study was to analyze the bioactivity of Biodentine ™ and MTA Repair HP® in contact with human dentine discs, which were sealed and divided randomly to form four groups: group 1 Biodentine™, group 2 MTA Repair HP®, positive control group MTA Angelus® and negative control group IRM®, which were incubated in PBS solution for 10 days, for a subsequent analysis by means of MEB-EDS and Raman spectroscopy. The three calcium-based materials analyzed in this study proved to be bioactive because upon contact with a phosphate-based solution they were triggered at the onset of amorphous calcium phosphate, as the precursor during the formation of carbonated apatite.

Fluoride release and uptake in enhanced bioactivity glass ionomer cement ("glass carbomer™") compared with conventional and resin-modified glass ionomer cements

Abstract Objectives To study the fluoride uptake and release properties of glass carbomer dental cements and compare them with those of conventional and resin-modified glass ionomers. Materials and Methods Three materials were used, as follows: glass carbomer (Glass Fill), conventional glass ionomer (Chemfil Rock) and resin-modified glass ionomer (Fuji II LC). For all materials, specimens (sets of six) were matured at room temperature for time intervals of 10 minutes, 1 hour and 6 weeks, then exposed to either deionized water or sodium fluoride solution (1000 ppm in fluoride) for 24 hours. Following this, all specimens were placed in deionized water for additional 24 hours and fluoride release was measured. Results Storage in water led to increase in mass in all cases due to water uptake, with uptake varying with maturing time and material type. Storage in aqueous NaF led to variable results. Glass carbomer showed mass losses at all maturing times, whereas the conventional glass ionomer gained mass for some maturing times, and the resin-modified glass ionomer gained mass for all maturing times. All materials released fluoride into deionized water, with glass carbomer showing the highest release. For both types of glass ionomer, uptake of fluoride led to enhanced fluoride release into deionized water. In contrast, uptake by glass carbomer did not lead to increased fluoride release, although it was substantially higher than the uptake by both types of glass ionomer. Conclusions Glass carbomer resembles glass ionomer cements in its fluoride uptake behavior but differs when considering that its fluoride uptake does not lead to increased fluoride release.

Enhanced bioactive properties of BiodentineTM modified with bioactive glass nanoparticles

Abstract Objective To prepare nanocomposite cements based on the incorporation of bioactive glass nanoparticles (nBGs) into BiodentineTM (BD, Septodent, Saint-Maur-des-Fosses Cedex, France) and to assess their bioactive properties. Material and Methods nBGs were synthesised by the sol-gel method. BD nanocomposites (nBG/BD) were prepared with 1 and 2% nBGs by weight; unmodified BD and GC Fuji IX (GIC, GC Corporation, Tokyo, Japan) were used as references. The in vitro ability of the materials to induce apatite formation was assessed in SBF by X-ray diffraction (XRD), attenuated total reflectance with Fourier transform infrared spectroscopy (ATR-FTIR), and scanning electron microscopy (SEM) with energy dispersive X-ray (EDX) analysis. BD and nBG/BD were also applied to dentine discs for seven days; the morphology and elemental composition of the dentine-cement interface were analysed using SEM-EDX. Results One and two percent nBG/BD composites accelerated apatite formation on the disc surface after short-term immersion in SBF. Apatite was detected on the nBG/BD nanocomposites after three days, compared with seven days for unmodified BD. No apatite formation was detected on the GIC surface. nBG/BD formed a wider interfacial area with dentine than BD, showing blockage of dentine tubules and Si incorporation, suggesting intratubular precipitation. Conclusions The incorporation of nBGs into BD improves its in vitro bioactivity, accelerating the formation of a crystalline apatite layer on its surface after immersion in SBF. Compared with unmodified BD, nBG/BD showed a wider interfacial area with greater Si incorporation and intratubular precipitation of deposits when immersed in SBF.

Enamel crystals of mice susceptible or resistant to dental fluorosis: an AFM study

Objective: This study aimed to assess the overall apatite crystals profile in the enamel matrix of mice susceptible (A/J strain) or resistant (129P3/J strain) to dental fluorosis through analyses by atomic force microscopy (AFM). Material and Methods: Samples from the enamel matrix in the early stages of secretion and maturation were obtained from the incisors of mice from both strains. All detectable traces of matrix protein were removed from the samples by a sequential extraction procedure. The purified crystals (n=13 per strain) were analyzed qualitatively in the AFM. Surface roughness profile (Ra) was measured. Results: The mean (±SD) Ra of the crystals of A/J strain (0.58±0.15 nm) was lower than the one found for the 129P3/J strain (0.66±0.21 nm) but the difference did not reach statistical significance (t=1.187, p=0.247). Crystals of the 129P3/J strain (70.42±6.79 nm) were found to be significantly narrower (t=4.013, p=0.0013) than the same parameter measured for the A/J strain (90.42±15.86 nm). Conclusion: enamel crystals of the 129P3/J strain are narrower, which is indicative of slower crystal growth and could interfere in the occurrence of dental fluorosis. .

Concentraçäo de flúor no esmalte humano, após profilaxia com jato de bicarbonato ou taça de borracha, seguidos da aplicaçäo tópica de flúor em gel / Concentration of fluoride in human dental enamel, after the prophylaxis with bicarbonate jet or rubber cup, following topical application of fluoride gel

Estudou-se o efeito do tipo de profilaxia na reatividade do flúor tópico, com o esmalte dental humano. As concentraçöes de fluoreto de cálcio (CaF2) e apatita fluoretada (FA) foram determinadas no esmalte, após a profilaxia com jato de bicabornato (PROFI IIr) ou taça de borracha, seguidas de aplicaçäo tópica de fúor em gel. Os resultados obtidos mostraram que formou-se mais CaF2 sobre o esmalte, devido a profilaxia com jato de bicarbonato do que com taça de borracha, embora näo tenha sido observada uma diferença estatisticamente significativa. Com relaçäo à concentraçäo de FA, observou-se nas regiöes mais internas do esmalte uma maior concentraçäo em funçäo da profilaxia com jato de bicarbonato, porém sem significado estatístico

Reatividade dos dentifrícios fluoretados comercializados no Brasil com o esmalte dental humano / The reactivity of fluorated dentifrices sold in Brazil to the human dental enamel

A reatividade dos dentifrícios fluoretados com o esmalte dental humano foi avaliada «in vitro¼. As concentrações de flúor total, fluoreto de cálcio e apatita fluoretada, foram determinadas no esmalte após a reação com as soluções dos dentifrícios. A capacidade de formar estes produtos variou amplamente quando os dentifrícios foram comparados. Estes resultados foram discutidos em termos do potencial caristático dos mesmos