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A B S T R A C TCeramic/polymer-based biocomposites have emerged as potential biomaterials to fill, replace, repair or regenerate injured or diseased bone, due to their outstanding features in terms of biocompatibility, bioactivity, injectability, and biodegradability. However, these properties can be dependent on the amount of ceramic component present in the polymer-based composite. Therefore, in the present study, the influence of nanohydroxyapatite content (30 to 70 wt%) on alginate-based hydrogels was studied in order to evaluate the best formulation for maximizing bone tissue regeneration. The composite system was characterized in terms of physic-chemical properties and biological response, with in vitro cytocompatibility assessment with human osteoblastic cells and ex vivo functional evaluation in embryonic chick segmental bone defects. The main morphological characteristics of the alginate network were not affected by the addition of nanohydroxyapatite. However, physic-chemical features, like water-swelling rate, stability at extreme pH values, apatite formation, and Ca 2+ release were nanoHA dose-dependent. Within in vitro cytocompatibility assays it was observed that hydrogels with nanoHA 30% content enhanced osteoblastic cells proliferation and expression of osteogenic transcription factors, while those with higher concentrations (50 and 70%) decreased the osteogenic cell response. Ex vivo data underlined the in vitro findings, revealing an enhanced collagenous deposition, trabecular bone formation and matrix mineralization with Alg-nanoHA30 composition, while compositions with higher nanoHA content induced a diminished bone tissue response.The outcomes of this study indicate that nanohydroxyapatite concentration plays a major role in physicchemical properties and biological response of the composite system and the optimization of the components ratio must be met to maximize bone tissue regeneration.

The aim of this work was to develop a bioactive bone substitute with an effective antibacterial ability based on a cerium (Ce) doped glass-reinforced hydroxyapatite (GR-HA) composite. Developed composites were physicochemically characterized, using x-ray diffraction (XRD) analysis, SEM, energy dispersive x-ray spectroscopy (EDS), and flexural bending strength (FBS) tests. X-ray photoelectron spectroscopy (XPS) analysis was performed to analyze the oxidation state of Ce in the prepared doped glass. The antimicrobial activity of the composites was evaluated against Staphylococcus aureus, Staphylococcus epidermidis and Pseudomonas aeruginosa; whether the cytocompatibility profile was assayed with human osteoblastic-like cells (Mg-63 cell line). The results revealed that the Ce inclusion in the GR-HA matrix induced the antimicrobial ability of the composite. In addition, Ce-doped materials reported an adequate biological behavior following seeding of osteoblastic populations, by inducing cell adhesion and proliferation. Developed materials were also found to enhance the expression of osteoblastic-related genes. Overall, the developed GR-HA_Ce composite is a prospective candidate to be used within the clinical scenario with a successful performance due to the effective antibacterial properties and capability of enhancing the osteoblastic cell response.

The high number of biomaterial associated infections demands new strategies to prevent this problem. In this study the suitability of nanohydroxyapatite (nanoHA)-based surfaces containing two Brazilian extracts of propolis (green and red ones) to prevent bacterial growth and biofilm formation, as well as its non-cytotoxic nature, was investigated. Optical density, colony forming units and MTT reduction assay were used to assess the materials' antibacterial activity against planktonic and sessile growth of Staphylococcus aureus. NanoHA matrix was able to absorb both types of propolis and the obtained results revealed the antibacterial effectiveness of the novel materials expressed as the reduction of bacterial growth and biofilm formation ability. Additionally, cell culture tests showed the growth of fibroblasts with high metabolic activity and without membrane damage. Therefore, these nanoHA-based surfaces containing natural products deriving from bees may be a promising bioactive biomaterial to be further studied with the aim of application to orthopaedic or dental devices.

This study concerns the preparation, physical, and in vitro characterization of two different types of hydroxyapatite (HA) microspheres, which are intended to be used as drug-delivery systems and bone-regeneration matrices. Hydroxyapatite nanoparticles (HA-1 and HA-2) were prepared using the chemical precipitation synthesis with H 3 PO 4 , Ca(OH) 2 , and a surfactant, SDS (sodium dodecyl sulfate), as starting reagents. The HA powders were dispersed in a sodium alginate solution, and spherical particles were obtained by droplet extrusion coupled with ionotropic gelation in the presence of Ca 2þ . These were subsequently sintered to produce HA-1 and HA-2 microspheres with a uniform size and interconnected microporosity. Both powders and microspheres were characterized using FTIR and X-ray diffraction. Moreover, SEM and mercury intrusion porosimetry were used to analyze the microspheres, and TEM was used to analyze the powders. Results showed that pure HA and mixtures of HA/b-TCP in the nanometer range and needlelike shape were obtained for HA-1 and HA-2 powders, respectively. Neutral Red, scanning electron microscopy and confocal microscopy were used to evaluate the behavior of osteoblastic-like MG-63 cells cultured on HA microspheres surfaces for 7 days. Results showed that good adhesion and proliferation of osteoblasts on the HA microspheres surface. Cells built bridges between adjacent microspheres, forming microspheres-cells clusters in both types of materials.

An innovative delivery system based on bacteriophages-loaded alginate-nanohydroxyapatite hydrogel was developed as a multifunctional approach for local tissue regeneration and infection prevention and control. Bacteriophages were efficiently encapsulated, without jeopardizing phage viability and functionality, nor affecting hydrogel morphology and chemical composition. Bacteriophage delivery occurred by swelling-disintegration-degradation process of the alginate structure and was influenced by environmental pH. Good tissue response was observed following the implantation of bacteriophages-loaded hydrogels, sustaining their biosafety profile. Bacteriophagesloaded hydrogels did not affect osteoblastic cells' proliferation and morphology. A strong osteogenic and mineralization response was promoted through the implantation of hydrogels system with nanohydroxyapatite. Lastly, bacteriophages-loaded hydrogel showed excellent antimicrobial activity inhibiting the attachment and colonization of multidrug-resistant E. faecalis surrounding and within femoral tissues. This new local delivery approach could be a promising approach to prevent and control bacterial contamination during implantation and bone integration.