Functionalisation of Ti6Al4V and hydroxyapatite surfaces with combined peptides based on KKLPDA and EEEEEEEE peptides

Surface modifications are usually performed on titanium alloys to improve osteo-integration and surface bioactivity. Modifications such as alkaline and acid etching, or coating with bioactive materials such as hydroxyapatite, have previously been demonstrated. The aim of this work is to develop a peptide with combined titanium oxide and hydroxyapatite binders in order to achieve a biomimetic hydroxyapatite coating on titanium surfaces. The technology would also be applicable for the functionalisation of titanium and hydroxyapatite surfaces for selective protein adsorption, conjugation of antimicrobial peptides, and adsorption of specialised drugs for drug delivery. In this work, functionalisation of Ti6Al4V and hydroxyapatite surfaces was achieved using combined titanium-hydroxyapatite (Ti-Hap) peptides based on titanium binder (RKLPDA) and hydroxyapatite binder (EEEEEEEE) peptides. Homogeneous peptide coatings on Ti6Al4V surfaces were obtained after surface chemical treatments with a 30 wt % aqueous solution of H2O2 for 24 and 48 hours. The treated titanium surfaces presented an average roughness of Sa=197 nm (24 h) and Sa=128 nm (48 h); an untreated mirror polished sample exhibited an Sa of 13 nm. The advancing water contact angle of the titanium oxide layer after 1 hour of exposure to 30 wt % aqueous solution of H2O2 was around 65°, decreasing gradually with time until it reached 35° after a 48 hour exposure, suggesting that the surface hydrophilicity increased over etching time. The presence of a lysine (L) amino acid in the sequence of the titanium binder resulted in fluorescence intensity roughly 16 % higher compared with the arginine (R) amino acid analogue and therefore the lysine containing titanium binder was used in this work. The Ti-Hap peptide KKLPDAEEEEEEEE (Ti-Hap1) was not adsorbed by the treated Ti6Al4V surfaces and therefore was modified. The modifications involved the inclusion of a glycine spacer between the binding terminals (Ti-Hap2) and the addition of a second titanium binder (KKLPDA) (Ti-Hap3 and Ti-Hap4). The Ti-Hap peptide aptamer which exhibited the strongest intensity after the titanium dip coating was KKLPDAKKLPDAEEEEEEEE (Ti-Hap4). On the other hand, hydroxyapatite surfaces, exhibiting an average roughness of Sa=1.42 µm, showed a higher fluorescence for all peptides compared with titanium surfaces

Revêtements d'hydroxyapatite réalisés par projection plasma : vers de nouvelles fonctionnalités

Dans les stratégies de remplacement/reconstruction des os et des articulations, il est souvent fait appel à des matériaux revêtus. Le matériau de structure permet d'assurer une bonne tenue mécanique et le revêtement doit faciliter et pérenniser l'ancrage dans le site osseux. Dans ce contexte, la technique de projection plasma est largement utilisée pour réaliser industriellement des revêtements d'hydroxyapatite. Nous proposons ici de faire une revue sur les avancées en termes de procédé (projection plasma dc ou rf, de suspensions/solutions, basse énergie, …) couplé à des compositions d'hydroxyapatite modifiées

Formation of nanosized strontium substituted hydroxyapatites

Incorporation of specific elements into calcium phosphates offers the combination of a bioactive material and a therapeutic effect. This is important for improving the integration of implants as well as treating medical conditions. Strontium is a suitable candidate and displays the ability to stimulate bone growth and reducing bone resorption. This study investigated the formation of strontium carbonated hydroxyapatite nanoparticles from an amorphous phase. Crystallization of carbonated hydroxyapatite occurred at 585 oC, but samples with an intended 25% and 75% replacement of calcium with strontium crystallized at 624 oC. Heat treatment at the crystallization temperature revealed that strontium free apatite does not crystallize in 5 minutes, but an increasing strontium concentration leads to a higher rate of crystallization. X-ray diffraction patterns suggest that it may be difficult to include strontium, but higher strontium concentrations are possibly included with ease in the lattice. This work has produced a nanosized apatite accompanied by an amorphous phase after a short heat-treatment time. This offers a range of features that collectively show great promise for significantly enhancing the release of strontium for improved bone therapeutic effects

Hydroxyapatite coating on titanium by a low energy plasma spraying mini-gun

Plasma-sprayed hydroxyapatite (HA) coatings are used on metallic implants to improve osseointegration and bone growth. The purpose of this work was to determine the microstructure and composition of HA coatings obtained with a newly developed low energy plasma spray mini-gun employing an HA feedstock powder with smaller granulometry than that commonly used. The microstructure and the phase composition of the coatings obtained by varying the number of mini-gun runs were examined using scanning electron microscopy, X-ray diffraction and Fourier transform infrared and micro-Raman spectroscopy. In all cases, the results indicate the presence of an amorphous phase and oxyapatite in the coatings due to hydroxyl group removal. No other foreign crystalline phases were detected. The absence of foreign phases was attributed to the fast cooling rate of the small particles used in the experiments and the low amount of energy employed with the mini-gun. Decomposition in the υ1PO4 region of the Raman spectra allowed a semi-quantitative evaluation of the phase contents as a function of the number of runs. Micro-Raman spectroscopy appears to be a powerful technique providing comprehensive and localised information concerning calcium phosphate phases in coatings

Biomimetic apatite sintered at very low temperature by spark plasma sintering: Physico-chemistry and microstructure aspects

Nanocrystalline apatites analogous to bone mineral are very promising materials for the preparation of highly bioactive ceramics due to their unique intrinsic physico-chemical characteristics. Their surface reactivity is indeed linked to the presence of a metastable hydrated layer on the surface of the nanocrystals. Yet the sintering of such apatites by conventional techniques, at high temperature, strongly alters their physico-chemical characteristics and biological properties, which points out the need for "softer" sintering processes limiting such alterations. In the present work a non-conventional technique, spark plasma sintering, was used to consolidate such nanocrystalline apatites at non-conventional, very low temperatures (T° < 300 °C) so as to preserve the surface hydrated layer present on the nanocrystals. The bioceramics obtained were then thoroughly characterized by way of complementary techniques. In particular, microstructural, nanostructural and other major physico-chemical features were investigated and commented on. This work adds to the current international concern aiming at improving the capacities of present bioceramics, in view of elaborating a new generation of resorbable and highly bioactive ceramics for bone tissue engineering

Hydroxyapatite 3D-printed scaffolds with Gyroid-Triply periodic minimal surface porous structure:Fabrication and an in vivo pilot study in sheep

Bone repair is a major challenge in regenerative medicine, e.g. for large defects. There is a need for bioactive, highly percolating bone substitutes favoring bone ingrowth and tissue healing. Here, a modern 3D printing approach (VAT photopolymerization) was exploited to fabricate hydroxyapatite (HA) scaffolds with a Gyroid-“Triply periodic minimal surface” (TPMS) porous structure (65% porosity, 90.5% HA densification) inspired from trabecular bone. Percolation and absorption capacities were analyzed in gaseous and liquid conditions. Mechanical properties relevant to guided bone regeneration in non-load bearing sites, as for maxillofacial contour reconstruction, were evidenced from 3-point bending tests and macrospherical indentation. Scaffolds were implanted in a clinically-relevant large animal model (sheep femur), over 6 months, enabling thorough analyses at short (4 weeks) and long (26 weeks) time points. In vivo performances were systematically compared to the bovine bone-derived Bio-OssⓇ standard. The local tissue response was examined thoroughly by semi-quantitative histopathology. Results demonstrated the absence of toxicity. Bone healing was assessed by bone dynamics analysis through epifluorescence using various fluorochromes and quantitative histomorphometry. Performant bone regeneration was evidenced with similar overall performances to the control, although the Gyroid biomaterial slightly outperformed Bio-OssⓇ at early healing time in terms of osteointegration and appositional mineralization. This work is considered a pilot study on the in vivo evaluation of TPMS-based 3D porous scaffolds in a large animal model, for an extended period of time, and in comparison to a clinical standard. Our results confirm the relevance of such scaffolds for bone regeneration in view of clinical practice. Statement of significance: Bone repair, e.g. for large bone defects or patients with defective vascularization is still a major challenge. Highly percolating TPMS porous structures have recently emerged, but no in vivo data were reported on a large animal model of clinical relevance and comparing to an international standard. Here, we fabricated TPMS scaffolds of HA, determined their chemical, percolation and mechanical features, and ran an in-depth pilot study in the sheep with a systematic comparison to the Bio-OssⓇ reference. Our results clearly show the high bone-forming capability of such scaffolds, with outcomes even better than Bio-OssⓇ at short implantation time. This preclinical work provides quantitative data validating the relevance of such TMPS porous scaffolds for bone regeneration in view of clinical evaluation.</p

Comparative study of conversion of coral with ammonium dihydrogen phosphate and orthophosphoric acid to produce calcium phosphates

© 2014, Australasian Ceramic Society. All rights reserved. Biogenic materials like corals, which are readily available, could be used to produce bioceramic materials and address significant advantages due to their unique structures and chemical compositions that contain Mg and Sr. Many conversion processes has been in the past proposed. In this work, a comparison study between the conversion of coral with orthophosphoric acid and ammonium dihydrogen phosphate was conducted. The resultant structures and compositions were studied using XRD, ICP-MS, SEM and FTIR. The results show that with phosphoric acid the coral was converted into mainly monetite (92%). The ammonium dihydrogen phosphate converted approximately 76% of the coral to hydroxyapatite through solid state reactions. The two routes proved to be effective in producing bioceramic materials from corals under moderate conditions of temperature with a basic condition favouring the yield of hydroxyapatite

Synthesis of fluor-hydroxyapatite powder for plasma sprayed biomedical coatings: Characterization and improvement of the powder properties

Fluor-hydroxyapatite (FHA) powder was synthesized by double decomposition in a view to produce bioactive and thermally stable coatings by plasma spray process. This work aims at studying the influence of chemical composition, microstructure and surface energy on the flowability of the FHA particles which is known as a determinant property during plasma spraying for the quality of the product in terms of yield and homogeneity of composition. The as-synthesized FHA powder was sieved in order to obtain two ranges of particles size: 50– 80μm and 80–100μm. The phase composition and structure and physical characteristics of FHA particles have been determined by complementary analytical techniques (X-ray diffraction, Fourier transform infrared spectroscopy, scanning electron spectroscopy and laser granulometry). Then, the FHA flowability was determined as recommended by the European Pharmacopeia. The results showed that the 80–100μm FHA powder has a longer flow time compared to a reference hydroxyapatite powder whereas the 50–80μm FHA particles did not flow. We propose a simple treatment of the FHA particles using ethanol as solventwhich showed to significantly improve the flowability of FHA powders