Bone tissue engineering is a promising strategy to treat the huge number of bone fractures caused by progressive population ageing and diseases i.e., osteoporosis. The bioactive and biomimetic materials design modulating cell behaviour can support healthy bone tissue regeneration. In this frame, type I collagen and hydroxyapatite (HA) have been often combined to produce biomimetic scaffolds. In addition, mesoporous bioactive glasses (MBGs) are known for their ability to promote the deposition of HA nanocrystals and their potential to incorporate and release therapeutic ions. Furthermore, the use of 3D printing technologies enables the effective design of scaffolds reproducing the natural bone architecture.

This study aims to design biomimetic and bioactive 3D printed scaffolds that mimic healthy bone tissue natural features in terms of chemical composition, topography and biochemical cues. Optimised collagenous hybrid systems will be processed by means of extrusion 3D printing technologies to obtain high resolution bone-like structures. Protocols of human co-cultures of osteoblasts and osteoclasts will be developed and used to test the 3D scaffolds.

Type I collagen has been combined with rod-like nano-HA and strontium containing MBGs (micro- and nano-sized particles) in order to obtain hybrid systems resembling the composition of native bone tissue. A comprehensive rheological study has been performed to investigate the potential use of the hybrid systems as biomaterial inks. Mesh-like structures have been obtained by means of extrusion-based technologies exploiting the freeform reversible embedding of suspended hydrogels (FRESH) approach. Different crosslinking methods have been tested to improve final constructs mechanical properties. Both crosslinked and non-crosslinked biomaterials were cultured with human osteoblasts and osteoclasts to assay the hybrid matrix biocompatibility as well as its influence on cell behaviour.

Homogeneous hybrid systems have been successfully developed and characterised, proving their suitability as biomaterial inks for 3D printing technologies. Mesh-like structures have been extruded in a thermo-reversible gelatine slurry, exploiting the sol-gel transition of the systems under physiological conditions. Covalent bonds between collagen molecules have been promoted by genipin treatment, leading to a significant increase in matrix strength and stability. The collagen methacrylation and the further UV-crosslinking are under investigation as alternative promising method to reinforce the 3D structure during the printing process. Biological tests showed the potential of the developed systems especially for genipin treated samples, with a significant adhesion of primary cells.

Collagenous hybrid systems proved their suitability for bioactive 3D printed structures design for bone tissue engineering. The multiple stimuli provided by the scaffold composition and structure will be investigated on both direct and indirect human osteoblasts and osteoclasts co-culture, according to the developed protocols.

Summary Statement

This experimental study showed that platelet rich fibrin matrix can improve muscle regeneration and long-term vascularization without local adverse effects.