In vitro testing of Nd:YAG laser processed calcium phosphate coatings.

Nd:YAG laser cladding is a new method for deposition of a calcium phosphate onto metallic surfaces of interest in implantology. The aim of this study was to compare the biologic response of MG-63 human osteoblast-like cells grown on Ti-6Al-4V substrates coated with a calcium phosphate layer applied using different methods: plasma spraying as reference material and Nd:YAG laser cladding as test material. Tissue culture polystyrene was used as negative control. The Nd:YAG laser clad material showed a behaviour similar to the reference material, plasma spray, respective to cell morphology (SEM observations), cell proliferation (AlamarBlue assay) and cytotoxicity of extracts (MTT assay). Proliferation, as measured by the AlamarBlue assay, showed little difference in the metabolic activity of the cells on the materials over an 18 day culture period. There were no significant differences in the cellular growth response on the test material when compared to the ones exhibited by the reference material. In the solvent extraction test all the extracts had some detrimental effect on cellular activity at 100% concentration, although cells incubated in the test material extract showed a proliferation rate similar to that of the reference material. To better understand the scope of these results it should be taken into account that the Nd:YAG clad coating has recently been developed. The fact that its in vitro performance is comparable to that produced by plasma spray, a material commercially available for more than ten years, indicates that this new laser based method could be of commercial interest in the near future.

Effect of the particle size on the micro and nanostructural features of a calcium phosphate cement: a kinetic analysis.

The aim of this work is to investigate the possibility of controlling the final micro and nanostructural features of a calcium phosphate cement by modifying the particle size of the starting powder, and to study the effect of this parameter on the kinetics of the setting reaction. The development of calcium phosphate materials with tailored structures at the micro and nanoscale levels could allow the modulation of some specific responses in biologic phenomena such as protein adsorption and cell adhesion, which strongly depend on the nano-sized roughness of the interface. It is shown that the higher specific surface, produced by the reduction of the particle size of the powder, strongly accelerates the hydrolysis of the alpha-TCP into calcium-deficient hydroxyapatite. The higher degree of supersaturation attained in the solution favours the nucleation of smaller crystals. Thus, by increasing the specific surface of the starting powder in a factor of 5, the size of the precipitated crystals is strongly reduced, and the specific surface of the set cement increases by a factor of 2. The reduction of the particle size produces a substantial decrease of the setting time and accelerates the hardening of the cement without significantly affecting the final strength attained. The mechanical strength achieved by the cement cannot be univocally related to the degree of reaction, without considering the microstructural features.

Calcium phosphate coatings obtained by Nd:YAG laser cladding: physicochemical and biologic properties.

The plasma spray (PS) technique is the most popular method commercially in use to produce calcium phosphate (CaP) coatings to promote fixation and osteointegration of the cementless prosthesis. Nevertheless, PS has some disadvantages, such as the poor coating-to-substrate adhesion, low mechanical strength, and brittleness of the coating. In order to overcome the drawbacks of plasma spraying, we introduce in this work a new method to apply a CaP coating on a Ti alloy using a well-known technique in the metallurgical field: laser surface cladding. The physicochemical characterization of the coatings has been carried out by means of X-ray diffraction (XRD), scanning electron microscopy (SEM), and energy dispersive X-ray analysis (EDX). The biologic properties of the coatings have been assessed in vitro with human osteoblast-like MG-63 cells. The overall results of this study affirm that the Nd:YAG laser cladding technique is a promising method in the biomedical field.

The Ca/P range of nanoapatitic calcium phosphate cements.

Nanoapatites are apatites consisting of nanometer size crystals. The commercial calcium phosphate cements set by the precipitation of nanoapatitic calcium phosphates in the range 1.5 < or = Ca/P < 1.8. In this study it is shown that a continuum of nanoapatites can precipitate in the range 0.8 < Ca/P< or = 1.5. In order to be formed these nanoapatites need to incorporate K+ ions. In addition they can incorporate some Na+ ions. Upon immersion in aqueous solutions these nanoapatites loose phosphate, K+ and Na+ so that in an open system they are transformed into calcium deficient hydroxyapatite Ca9(HPO4)(PO4)5OH within about 2 months.

Alloying of hydroxyapatite onto Ti6Al-4V by high power laser irradiation.

In the biomedical field, the synthetic hydroxyapatite [Ca(10)(PO)(4)(OH)(2)], with similarity to the inorganic component of bone but brittle, has been considered as the appropriate coating on stronger implant materials, such as metallic implants, for presenting a surface which is conductive to bone formation. Many industrial and laboratory techniques were developed to apply hydroxyapatite onto metallic substrates, such as electrophoretic deposition, ion sputtering, hot isostatic pressing, pulsed laser deposition and the only widely used method commercially available: plasma spraying. This work presents a new approach on how to bind calcium phosphate (CaP) to the Ti alloy with a well-known technique in the metallurgical field: laser surface alloying, in order to overcome the drawbacks of plasma spraying. The analysis of the results obtained and the description of the phenomena that take place in the coating process will complete this explorative study.

Transforming growth factor-beta1 incorporation in a calcium phosphate bone cement: material properties and release characteristics.

The bone regenerative properties of calcium phosphate cements (CPCs) may be improved by the addition of growth factors, such as recombinant human transforming growth factor-beta1 (rhTGF-beta1). Previously, we showed that rhTGF-beta1 in CPC stimulated the differentiation of preosteoblastic cells from adult rat long bones. The intermixing of rhTGF-beta1 in CPC, which was subsequently applied to rat calvarial defects, enhanced bone growth around the cement and increased the degradation of the cement. However, it is unknown whether the addition of rhTGF-beta1 changes the material properties of CPC and what the characteristics of the release of rhTGF-beta1 from CPC are. Therefore, we determined in this study the release of rhTGF-beta1, in vitro, from the cement pellets as implanted in the rat calvariae. The possible intervening effects of rhTGF-beta1 intermixing on the clinical compliance of CPC were studied through an assessment of its compressive strength and setting time, as well as its crystallinity, calcium-to-phosphorus ratio, porosity, and microscopic structure. We prepared CPC by mixing calcium phosphate powder (58% alpha-tricalcium phosphate, 25% anhydrous dicalcium phosphate, 8.5% calcium carbonate, and 8.5% hydroxyapatite) with a liquid (3 g/mL). The liquid for standard CPC consisted of water with 4% disodium hydrogen phosphate, whereas the liquid for modified CPC was mixed with an equal amount of 4 mM hydrochloride with 0.2% bovine serum albumin. The hydrochloride liquid contained rhTGF-beta1 in different concentrations for the release experiments. Most of the rhTGF-beta1 incorporated in the cement pellets was released within the first 48 h. For all concentrations of intermixed rhTGF-beta1 (100 ng to 2.5 mg/g of CPC), approximately 0.5% was released in the first 4 h, increasing to 1.0% after 48 h. Further release was only about 0.1% from 2 days to 8 weeks. CPC modification slightly increased the initial setting time at 20 degrees C from 2.6 to 5 min but had no effect on the final setting time of CPC at 20 degrees C or the initial and final setting times at 37 degrees C. The compressive strength was increased from 18 MPa in the standard CPC to 28 MPa in the modified CPC only 4 h after mixing. The compressive strength diminished in the modified CPC between 24 h and 8 weeks from 55 to 25 MPa. No other significant change was found with the CPC modification for rhTGF-beta1. X-ray diffraction revealed that standard and modified CPCs changed similarly from the original components, alpha-tricalcium phosphate and anhydrous dicalcium phosphate, into an apatite cement. The calcium-to-phosphorus ratio, as determined with an electron microprobe, did not differ for standard CPC and modified CPC. Standard and modified CPCs became dense and homogeneous structures after 24 h, but the modified CPC contained more crystal plaques than the standard CPC, as observed with scanning electron microscopy (SEM). SEM and back- scattered electron images revealed that after 8 weeks the cements showed equally and uniformly dense structures with microscopic pores (<1 microm). Both CPCs showed fewer crystal plaques at 8 weeks than at 24 h. This study shows that CPC is not severely changed by its modification for rhTGF-beta1. The prolonged setting time of modified cement may affect the clinical handling but is still within acceptable limits. The compressive strength for both standard and modified cements was within the range of thin trabecular bone; therefore, both CPCs can withstand equal mechanical loading. The faster diminishing compressive strength of modified cement from 24 h to 8 weeks likely results in early breakdown and so might be favorable for bone regeneration. Together with the beneficial effects on bone regeneration from the addition of rhTGF-beta1 to CPC, as shown in our previous studies, we conclude that the envisaged applications for CPC in bone defects are upgraded by the intermixing of rhTGF-beta1. Therefore, the combination of CPC and rhTGF-beta1 forms a promising synthetic bone graft.