Nano-hydroxyapatite (nano-HAp) materials show an analogous chemical composition to the biogenic mineral components of calcified tissues and depending on their topography they may mimic the specific arrangement of the crystals in bone. In this work, we have evaluated the potential of four synthesized nano-HAp superstructures for the in vitro conditions of bone-repair. Experiments are underway to investigate the effects of the material microstructure, surface roughness and hydrophilicity on their osseo-integration, osteo-conduction and osteo-induction abilities. Materials were tested in the presence of both, rat primary osteoblasts and rabbit mesenchymal stem cells. The following aspects are discussed: (i) cytotoxicity and material degradation; (ii) rat osteoblast spreading, proliferation and differentiation; and (iii) rabbit mesenchymal stem cell adhesion on nano-HAp and nano-HAp/collagen type I coatings. We effectively prepared a material based on biomimetic HAp nano-rods displaying the appropriate surface topography, hydrophilicity and degradation properties to induce the in vitro desired cellular responses for bone bonding and healing. Cells seeded on the selected material readily attached, proliferated and differentiated, as confirmed by cell viability, mitochondrial metabolic activity, alkaline phosphatase (ALP) activity and cytoskeletal integrity analysis by immunofluorescence localization of alpha-smooth muscle actin (α-SMA) protein. These results highlight the influence of material's surface characteristics to determine their tissue regeneration potential and their future use in engineering osteogenic scaffolds for orthopedic implants.

Ionic substitution can affect essential physicochemical properties leading to a specific biological behavior upon implantation. Therefore, it has been proposed as a tool to increase the biological efficiency of calcium phosphate based materials. In the following study, we have evaluated the contribution of an important cation in nature, Mg²⁺, into the structure of previously studied biocompatible and biodegradable hydroxyapatite (HA) nanorods and its subsequent effect on its chemical, morphology, and bone mimetic articulation. Mg²⁺-substituted HA samples were synthesized by an aqueous wet-chemical precipitation method, followed by an hydrothermal treatment involving a Mg²⁺ precursor that partially replace Ca²⁺ ions into HA crystal lattice; Mg²⁺ concentrations were modulated to obtain a nominal composition similar to that exists in calcified tissues. Hydrothermally synthesized Mg²⁺-substituted HA nanoparticles were characterized by X-ray powder diffraction, FT-NIR and EDX spectroscopies, field emission scanning and high resolution transmission electron microscopies (FE-SEM, H-TEM). Molecular modeling combining ab initio methods and power diffraction data were also performed. Results showed that Mg²⁺-substitution promoted the formation of calcium deficient HA (cdHA) where Mg²⁺ replacement is energetically favored at Ca(1) position in a limited and specific amount directing the additional Mg²⁺ toward the surface of the crystal. The control of Mg²⁺ incorporation into HA nanorods gave rise to a tailored crystallinity degree, cell parameters, morphology, surface hydration, solubility, and degradation properties in a dose-replacement dependent manner. The obtained materials show qualities that conjugated together to drive an optimal in vitro cellular viability, spreading, and proliferation confirming their biocompatibility. In addition, an improved adhesion of osteoblast was evidenced after Mg²⁺–Ca²⁺ substitution.

Membranes for guided bone regeneration should have a mechanical structure and a chemical composition suitable for mimicking biological structures. In this work, we pursue the development of periosteum-inspired bilayered membranes obtained by crosslinking alginate with different amounts of nanohydroxyapatite. Alginate-nanohydroxyapatite interaction was studied by rheology and infrared spectroscopy measurements. The membranes were characterized regarding their tensile strength, degradation and surface morphology. Finally, cell cultures were performed on each side of the membranes. The ionic bonding between alginate polysaccharide networks and nanohydroxyapatite was proven, and had a clear effect in the strength and microstructure of the hydrogels. Distinct surface characteristics were achieved on each side of the membranes, resulting in a highly porous fibrous side and a mineral-rich side with higher roughness and lower porosity. Moreover, the effect of amount of nanohydroxyapatite was reflected in a decrease of the membranes' plasticity and an increment of degradation rate. Finally, it was proved that osteoblast-like cells proliferated and differentiated on the mineral-rich side, specially when a higher amount of nanohydroxyapatite was used, whereas fibroblasts-like cells were able to proliferate on the fibrous side. These periosteum-inspired membranes are promising biomaterials for guided tissue regeneration applications.