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1.
Int J Nanomedicine ; 14: 3471-3490, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31190805

RESUMO

Background: Zinc-doped hydroxyapatite has been proposed as a graft biomaterial for bone regeneration. However, the effect of zinc on osteoconductivity is still controversial, since the release and resorption of calcium, phosphorus, and zinc in graft-implanted defects have rarely been studied. Methods: Microspheres containing alginate and either non-doped carbonated hydroxyapatite (cHA) or nanocrystalline 3.2 wt% zinc-doped cHA (Zn-cHA) were implanted in critical-sized calvarial defects in Wistar rats for 1, 3, and 6 months. Histological and histomorphometric analyses were performed to evaluate the volume density of newly formed bone, residual biomaterial, and connective tissue formation. Biomaterial degradation was characterized by transmission electron microscopy (TEM) and synchrotron radiation-based X-ray microfluorescence (SR-µXRF), which enabled the elemental mapping of calcium, phosphorus, and zinc on the microsphere-implanted defects at 6 months post-implantation. Results: The bone repair was limited to regions close to the preexistent bone, whereas connective tissue occupied the major part of the defect. Moreover, no significant difference in the amount of new bone formed was found between the two microsphere groups. TEM analysis revealed the degradation of the outer microsphere surface with detachment of the nanoparticle aggregates. According to SR-µXRF, both types of microspheres released high amounts of calcium, phosphorus, and zinc, distributed throughout the defective region. The cHA microsphere surface strongly adsorbed the zinc from organic constituents of the biological fluid, and phosphorus was resorbed more quickly than calcium. In the Zn-cHA group, zinc and calcium had similar release profiles, indicating a stoichiometric dissolution of these elements and non-preferential zinc resorption. Conclusions: The nanometric size of cHA and Zn-cHA was a decisive factor in accelerating the in vivo availability of calcium and zinc. The high calcium and zinc accumulation in the defect, which was not cleared by the biological medium, played a critical role in inhibiting osteoconduction and thus impairing bone repair.


Assuntos
Alginatos/química , Regeneração Óssea , Cálcio/metabolismo , Durapatita/química , Microesferas , Nanopartículas/química , Zinco/química , Zinco/metabolismo , Animais , Materiais Biocompatíveis/química , Disponibilidade Biológica , Regeneração Óssea/efeitos dos fármacos , Carbonatos/química , Morte Celular , Linhagem Celular , Sobrevivência Celular , Feminino , Camundongos , Nanopartículas/ultraestrutura , Ratos Wistar , Crânio/fisiologia , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios X
2.
J Mech Behav Biomed Mater ; 80: 143-154, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29414470

RESUMO

After insertion into bone, dental implants may be subjected to tribocorrosive conditions resulting in the release of metallic ions and solid wear debris, which can induce to peri-implant inflammatory reactions accompanied by bone loss, and ultimately implant loosening. Despite the promising ability of TiO2 nanotubes (NTs) to improve osseointegration and avoid infection-related failures, the understanding of their degradation under the simultaneous action of wear and corrosion (tribocorrosion) is still very limited. This study aims, for the first time, to study the tribocorrosion behavior of bio-functionalized TiO2 NTs submitted to two-cycle sliding actions, and compare it with conventional TiO2 NTs. TiO2 NTs grown by anodization were doped with bioactive elements, namely calcium (Ca), phosphorous (P), and zinc (Zn), through reverse polarization anodization treatments. Characterization techniques such as scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDS) and scanning transmission electron microscopy (STEM), were used to characterize the films. Tribocorrosion tests were carried out in artificial saliva (AS) by applying two cycles of reciprocating sliding actions. The open circuit potential (OCP) was monitored before, during, and after both cycles of sliding, during which the coefficient of friction (COF) was calculated. The resulting wear scars were analyzed by SEM and EDS, and wear volume measurements were performed by 2D profilometry. Finally, the mechanical features of TiO2 NTs were accessed by nanoindentation. The results show that bio-functionalized TiO2 NTs display an enhanced tribocorrosion performance, ascribed to the growth of a nano-thick oxide film at Ti/TiO2 NTs interface, which significantly increased their adhesion strength to the substrate and consequently their hardness. Furthermore, it was discovered that during tribo-electrochemical solicitations, the formation of a P-rich tribofilm takes place, which grants both electrochemical protection and resistance to mechanical wear. This study provides fundamental and new insights for the development of multifunctional TiO2 NTs with long-term biomechanical stability and improved clinical outcomes.

3.
J Biomed Mater Res A ; 106(2): 428-439, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29044948

RESUMO

Internalization of hydroxyapatite nanoparticles in SAOS-2 osteoblasts for 2 and 24 h was investigated in vitro using 5 and 50 µg/mL nanoparticles in culture medium. No cytotoxic effects were observed in a PrestoBlue viability assay. Focused ion beam-scanning electron microscopy and transmission electron microscopy were used to study nanoparticle trafficking inside cells and to characterize the physicochemical properties of the remodeled nanoparticles. Nanoparticles were actively internalized by cells and maintained in intracellular membrane-bound compartments. Dissolution of hydroxyapatite nanoparticles was observed inside phagolysosome in all samples. After 24 h of internalization in cell culture assays, reprecipitation of calcium phosphate minerals was observed in membrane-bound compartments in 5 and 50 µg/mL samples. Compared to the original nanoparticles, the reprecipitated calcium phosphate phase presented a different morphology, structure, and chemical composition. Two sample preparation methods were used and confirmed that reprecipitation of the calcium phosphate crystallites occurred in the intracellular environment and not during electron microscopy sample preparation. Reprecipitation of calcium phosphate prevented the release of large amounts of calcium and phosphate ions inside the cells. This phenomenon may be linked to physiological processes in the cell that control the concentration and trafficking of intracellular calcium ions, which are highly controlled by cells. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 428-439, 2018.


Assuntos
Durapatita/química , Nanopartículas/química , Osteoblastos/citologia , Linhagem Celular , Sobrevivência Celular , Humanos , Nanopartículas/ultraestrutura , Espectrometria por Raios X
4.
Sci Rep ; 7(1): 14843, 2017 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-29093500

RESUMO

ABSTARCT: This work explores a new class of vortex/magnetite/iron oxide nanoparticles designed for magnetic hyperthermia applications. These nanoparticles, named Vortex Iron oxide Particles (VIPs), are an alternative to the traditional Superparamagnetic Iron Oxide Nanoparticles (SPIONs), since VIPs present superior heating power while fulfilling the main requirements for biomedical applications (low cytotoxicity and nonremanent state). In addition, the present work demonstrates that the synthesized VIPs also promote an internalization and aggregation of the particles inside the cell, resulting in a highly localized hyperthermia in the presence of an alternating magnetic field. Thereby, we demonstrate a new and efficient magnetic hyperthermia strategy in which a small, but well localized, concentration of VIPs can promote an intracellular hyperthermia process.

5.
Micron ; 80: 122-34, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26546967

RESUMO

The interest in effects of strontium (Sr) on bone has greatly increased in the last decade due to the development of the promising drug strontium ranelate. This drug is used for treating osteoporosis, a major bone disease affecting hundreds of millions of people worldwide, especially postmenopausal women. The novelty of strontium ranelate compared to other treatments for osteoporosis is its unique effect on bone: it simultaneously promotes bone formation by osteoblasts and inhibits bone resorption by osteoclasts. Besides affecting bone cells, treatment with strontium ranelate also has a direct effect on the mineralized bone matrix. Due to the chemical similarities between Sr and Ca, a topic that has long been of particular interest is the incorporation of Sr into bones replacing Ca from the mineral phase, which is composed by carbonated hydroxyapatite nanocrystals. Several groups have analyzed the mineral produced during treatment; however, most analysis were done with relatively large samples containing numerous nanocrystals, resulting thus on data that represents an average of many crystalline domains. The nanoscale analysis of the bone apatite crystals containing Sr has only been described in a few studies. In this study, we review the current knowledge on the effects of Sr on bone mineral and discuss the methodological approaches that have been used in the field. In particular, we focus on the great potential that advanced microscopy and microanalytical techniques may have on the detailed analysis of the nanostructure and composition of bone apatite nanocrystals produced during treatment with strontium ranelate.


Assuntos
Osso e Ossos/química , Osso e Ossos/metabolismo , Minerais/análise , Estrôncio/metabolismo , Animais , Feminino , Humanos , Masculino , Microscopia/métodos , Osteoporose/tratamento farmacológico , Análise Espectral/métodos , Estrôncio/uso terapêutico
6.
J Mater Sci Mater Med ; 26(4): 166, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25791461

RESUMO

Hydroxyapatite (HA) has been investigated as a delivery system for antimicrobial and antibacterial agents to simultaneously stimulate bone regeneration and prevent infection. Despite evidence supporting the bactericidal efficiency of these HA carriers, few studies have focused on the effect of this association on bone regeneration. In this work, we evaluated the physico-chemical properties of hydroxyapatite microspheres loaded with chlorhexidine (CHX) at two different concentrations, 0.9 and 9.1 µgCHX/cm2 HA, and characterized their effects on in vitro osteoblast viability and bone regeneration. Ultraviolet-visible spectroscopy, scanning and transmission electron microscopy associated with energy-dispersive X-ray spectroscopy and electron energy loss spectroscopy were used to characterize the association of CHX and HA nanoparticles. The high CHX loading dose induced formation of organic CHX plate-like aggregates on the HA surface, whereas a Langmuir film was formed at the low CHX surface concentration. Quantitative evaluation of murine osteoblast viability parameters, including adhesion, mitochondrial activity and membrane integrity of cells exposed to HA/CHX extracts, revealed a cytotoxic effect for both loading concentrations. Histomorphological analysis upon implantation into the dorsal connective tissues and calvaria of rats for 7 and 42 days showed that the high CHX concentration induced the infiltration of inflammatory cells, resulting in retarded bone growth. Despite a strong decrease in in vitro cell viability, the low CHX loading dose did not impair the biocompatibility and osteoconductivity of HA during bone repair. These results indicate that high antimicrobial doses may activate a strong local inflammatory response and disrupt the long-term osteoconductive properties of CHX-HA delivery systems.


Assuntos
Fenômenos Fisiológicos Bacterianos/efeitos dos fármacos , Substitutos Ósseos/administração & dosagem , Clorexidina/administração & dosagem , Implantes de Medicamento/administração & dosagem , Osteoblastos/fisiologia , Osteogênese/fisiologia , Células 3T3 , Animais , Antibacterianos/administração & dosagem , Antibacterianos/química , Substitutos Ósseos/síntese química , Cápsulas/administração & dosagem , Cápsulas/síntese química , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Clorexidina/química , Terapia Combinada , Difusão , Implantes de Medicamento/química , Durapatita/administração & dosagem , Durapatita/química , Masculino , Camundongos , Osteoblastos/citologia , Osteogênese/efeitos dos fármacos , Ratos , Ratos Wistar
7.
Acta Biomater ; 10(9): 3875-84, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24487057

RESUMO

We investigated the ultrastructure and crystallographic orientation of spicules from the calcareous sponge Paraleucilla magna (subclass Calcaronea) by transmission and scanning electron microscopy using two different methods of sample preparation: ultramicrotomy and focused ion beam (FIB). It was found that the unpaired actine from the spicules was oriented in the [211] zone axis. The plane that contains the unpaired actine and divides symmetrically the paired actines is the (-120). This plane is a mirror plane of the hexagonal lattice system. All the spicule types analyzed presented the same crystallographic orientation. Electron nanodiffraction maps from 4µm×4µm regions prepared by FIB showed disorientation of <2° between diffraction patterns obtained from neighbor regions, indicating the presence of a unique, highly aligned calcite crystalline phase. Among the eight FIB sections obtained, four presented high pore density. In one section perpendicular to the actine axis pores were observed only in the center of the spicule aligned in a circular pattern and surrounded by a faint circular contour with a larger radius. The presence of amorphous carbon representative of organic molecules detected by electron energy loss spectroscopy was correlated neither with porosity nor with specific lattice planes.


Assuntos
Carbonato de Cálcio/química , Poríferos/anatomia & histologia , Poríferos/química , Animais , Carbono/química , Cristalografia , Poríferos/ultraestrutura , Porosidade , Análise Espectral
8.
Micron ; 56: 29-36, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24207060

RESUMO

Strontium ranelate has been used to prevent bone loss and stimulate bone regeneration. Although strontium may integrate into the bone crystal lattice, the chemical and structural modifications of the bone when strontium interacts with the mineral phase are not completely understood. The objective of this study was to evaluate apatite from the mandibles of rats treated with strontium ranelate in the drinking water and compare its characteristics with those from untreated rats and synthetic apatites with and without strontium. Electron energy loss near edge structures from phosphorus, carbon, calcium and strontium were obtained by electron energy loss spectroscopy in a transmission electron microscope. The strontium signal was detected in the biological and synthetic samples containing strontium. The relative quantification of carbon by analyzing the CK edge at an energy loss of ΔE = 284 eV showed an increase in the number of carbonate groups in the bone mineral of treated rats. A synthetic strontium-containing sample used as control did not exhibit a carbon signal. This study showed physicochemical modifications in the bone mineral at the nanoscale caused by the systemic administration of strontium ranelate.


Assuntos
Conservadores da Densidade Óssea/farmacologia , Densidade Óssea/efeitos dos fármacos , Mandíbula/fisiologia , Tiofenos/farmacologia , Animais , Apatitas/análise , Cálcio/química , Carbonatos/análise , Masculino , Mandíbula/efeitos dos fármacos , Microscopia Eletrônica de Transmissão , Compostos Organometálicos/farmacologia , Ratos , Ratos Wistar , Espectroscopia de Perda de Energia de Elétrons , Estrôncio/metabolismo
9.
ACS Appl Mater Interfaces ; 5(19): 9435-45, 2013 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-24059686

RESUMO

Right angle radio frequency magnetron sputtering technique (RAMS) was redesigned to favor the production of high-quality hydroxyapatite (HA) thin coatings for biomedical applications. Stoichiometric HA films with controlled crystallinity, thickness varying from 254 to 540 nm, crystallite mean size of 73 nm, and RMS roughness of 1.7 ± 0.9 nm, were obtained at room temperature by tuning the thermodynamic properties of the plasma sheath energy. The plasma energies were adjusted by using a suitable high magnetic field confinement of 143 mT (1430 G) and a substrate floating potential of 2 V at the substrate-to-magnetron distance of Z = 10 mm and by varying the sputtering geometry, substrate-to-magnetron distance from Z = 5 mm to Z = 18 mm, forwarded RF power and reactive gas pressure. Measurements that were taken with a Langmuir probe showed that the adjusted RAMS geometry generated a plasma with an adequate effective temperature of Teff ≈ 11.8 eV and electron density of 2.0 × 10(15) m(-3) to nucleate nanoclusters and to further crystallize the nanodomains of stoichiometric HA. The deposition mechanism in the RAMS geometry was described by the formation of building units of amorphous calcium phosphate clusters (ACP), the conversion into HA nanodomains and the crystallization of the grain domains with a preferential orientation along the HA [002] direction.

10.
Mater Sci Eng C Mater Biol Appl ; 33(5): 2981-6, 2013 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-23623122

RESUMO

Single phase, stoichiometrically pure, hollow nanotubes of hydroxyapatite have been synthesized and single-particle analysis has been performed to successfully prove the sole formation of Ca10(PO4)6(OH)2 phase. The facile synthesis involves a sol-gel process under neutral conditions in the presence of a sacrifical anodic alumina template. The structures formed are hollow nanotubes that have been characterized by XRD, SEM, TEM, SAED, EELS, EDS and BET measurements. The diameter of the resulting tubes is in the range of 140-350 nm, length is on the order of a few microns and the wall thickness of the tubes was found to be ca. 30 nm. Moreover these tubes had a large BET surface area of 115 m(2)/g and were found to be biocompatible. They displayed inertness in the presence of NIH 3T3 mouse fibroblast cells as dictated by an MTT assay.


Assuntos
Materiais Biocompatíveis , Durapatita/química , Nanotubos , Animais , Camundongos , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Células NIH 3T3 , Difração de Pó
11.
Bone ; 50(1): 301-10, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22057083

RESUMO

We report the ultrastructure of regenerated bone surrounding two types of biomaterials: hydroxyapatite-alginate composite and sintered hydroxyapatite. Critical defects in the calvaria of Wistar rats were filled with micrometer-sized spherical biomaterials and analyzed after 90 and 120 days of implantation by high-resolution transmission electron microscopy and Fourier transform infrared attenuated total reflectance microscopy, respectively. Infrared spectroscopy showed that hydroxyapatite of both biomaterials became more disordered after implantation in the rat calvaria, indicating that the biological environment induced modifications in biomaterials structure. We observed that the regenerated bone surrounding both biomaterials had a lamellar structure with type I collagen fibers alternating in adjacent lamella with angles of approximately 90°. In each lamella, plate-like apatite crystals were aligned in the c-axis direction, although a rotation around the c-axis could be present. Bone plate-like crystal dimensions were similar in regenerated bone around biomaterials and pre-existing bone in the rat calvaria. No epitaxial growth was observed around any of the biomaterials. A distinct mineralized layer was observed between new bone and hydroxyapatite-alginate biomaterial. This region presented a particular ultrastructure with crystallites smaller than those of the bulk of the biomaterial, and was possibly formed during the synthesis of alginate-containing composite or in the biological environment after implantation. Round nanoparticles were observed in regions of newly formed bone. The findings of this work contribute to a better understanding of the role of hydroxyapatite based biomaterials in bone regeneration processes at the nanoscale.


Assuntos
Alginatos/química , Materiais Biocompatíveis/química , Matriz Óssea/química , Matriz Óssea/ultraestrutura , Regeneração Óssea/fisiologia , Hidroxiapatitas/química , Animais , Calcificação Fisiológica , Masculino , Teste de Materiais , Microscopia Eletrônica de Transmissão , Ratos , Ratos Wistar , Crânio/química , Crânio/patologia , Crânio/fisiologia , Crânio/ultraestrutura , Espectroscopia de Infravermelho com Transformada de Fourier
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