Recently, bone tissue engineering emerged as a viable therapeutic alternative, comprising bone implants and new personalized scaffolds to be used in bone replacement and regeneration. In this study, biocompatible scaffolds were produced by freeze-drying, using different formulations (chitosan, chitosan/gelatin, chitosan/β-TCP and chitosan/gelatin/β-TCP) to be used as temporary templates during bone tissue regeneration. Sample characterization was performed through attenuated total reflectance-Fourier transform infrared spectroscopy, X-ray diffraction and energy dispersive spectroscopy analysis. Mechanical characterization and porosity analysis were performed through uniaxial compression test and liquid displacement method, respectively. In vitro studies were also done to evaluate the biomineralization activity and the cytotoxic profile of the scaffolds. Scanning electron and confocal microscopy analysis were used to study cell adhesion and proliferation at the scaffold surface and within their structure. Moreover, the antibacterial activity of the scaffolds was also evaluated through the agar diffusion method. Overall, the results obtained revealed that the produced scaffolds are bioactive and biocompatible, allow cell internalization and show antimicrobial activity against Staphylococcus aureus. Such, make these 3D structures as potential candidates for being used on the bone tissue regeneration, since they promote cell adhesion and proliferation and also prevent biofilm development at their surfaces, which is usually the main cause of implant failure.

Bone is the main component of the human skeleton. This tissue is responsible for several functions at the structural and biological level. It is a tissue that is highly vascularized and dynamic, with excellent mechanical properties and also has the capacity for selfregeneration. However, several factors as aging, disorders at metabolic level and fractures, can lead to a loss of integrity of this tissue. In this context, it is necessary to develop bone substitutes that mimic the structure of extracellular matrix and induce regeneration of the damaged tissue. Currently, the most commonly clinical treatments used are based in the application of bone grafts obtained from the patient or from another. However, these therapeutic approaches present several limitations, namely immunologic responses and infectious risks. Tissue Engineering, an area that aims to develop biological substitutes that are able to enhance, restore or replace the damaged tissue, has emerged as an area of research that is looking for new solutions to improve bone regeneration. In this field of research, several types of biomaterials have been developed for this purpose. Among these, three dimensional structures, also known as scaffolds are the most promising because these structures possess the essential properties needed for the bone regeneration, such as biocompatibility, biodegradability and porosity. Besides, scaffolds provide a structure that mimic extracellular matrix which is fundamental to promote cell adhesion and proliferation until tissue regeneration occurs. In this context, novel biomaterials based in chitosan, gelatin and beta-tricalcium phosphate were developed in this work, in order to be applied in a near future in bone regeneration. Several physical-chemical and mechanical studies were performed, to study the influence of the incorporation of gelatin and beta tricalcium phosphate in the structure of chitosan scaffolds. Besides, biological studies were performed in vitro with human osteoblasts to evaluate the materials cytotoxic profile. The results obtained demonstrate that the scaffolds developed provide a suitable support for cell adhesion and proliferation. Moreover, the scaffolds do not present toxicity, highlighting its application in regenerative medicine.

The growing need to treat bone-related diseases in an elderly population compels the development of novel bone substitutes to improve patient quality of life. In this context, the advent of affordable and effective rapid prototyping equipment, such as the Fab@home plotter, has contributed to the development of novel scaffolds for bone tissue engineering. In this study, we report for the first time the use of a Fab@home plotter for the production of 3D scaffolds composed by beta-tricalcium phosphate (β-TCP)/alginate hybrid materials. β-TCP/alginate mixtures were used in a proportion of 50/50% (w/w), 30/70% (w/w) and 20/80% (w/w). The printing parameters were optimized to a nozzle diameter of 20 Gauge for the production of rigid scaffolds with pre-defined architectures. We observed that, despite using similar printing parameters, both the precision and resolution of the scaffolds were significantly affected by the blend's viscosity. In particular, we demonstrate that the higher viscosity of 50/50 scaffolds (150.0 ± 3.91 mPa s) provides a higher precision in the extrusion process. The physicochemical and biological characterization of the samples demonstrated that the 50/50 scaffolds possessed a resistance to compression comparable to that of native trabecular bone. Moreover, this particular formulation also exhibited a Young's modulus that was higher than that of trabecular bone. Scanning electron microscopy and fluorescence microscopy analysis revealed that osteoblasts were able to adhere, proliferate and also penetrate into the scaffold's architecture. Altogether, our findings suggest that the Fab@home printer can be employed in the manufacture of reproducible scaffolds, using a formulation 50/50 alginate-β-TCP that has suitable properties to be applied as bone substitutes in the future.

The regeneration of large bone defects remains a challenging scenario from a therapeutic point of view. In fact, the currently available bone substitutes are often limited by poor tissue integration and severe host inflammatory responses, which eventually lead to surgical removal. In an attempt to address these issues, herein we evaluated the importance of alginate incorporation in the production of improved and tunable β-tricalcium phosphate (β-TCP) and hydroxyapatite (HA) three-dimensional (3D) porous scaffolds to be used as temporary templates for bone regeneration. Different bioceramic combinations were tested in order to investigate optimal scaffold architectures. Additionally, 3D β-TCP/HA vacuum-coated with alginate, presented improved compressive strength, fracture toughness and Young's modulus, to values similar to those of native bone. The hybrid 3D polymeric–bioceramic scaffolds also supported osteoblast adhesion, maturation and proliferation, as demonstrated by fluorescence microscopy. To the best of our knowledge this is the first time that a 3D scaffold produced with this combination of biomaterials is described. Altogether, our results emphasize that this hybrid scaffold presents promising characteristics for its future application in bone regeneration.