Mineralized, hierarchical, bioinspired materials for tissue engineering

The project aims at using a multidisciplinary approach to develop new, organic-inorganic, hydrogel based, composite materials for application in bone tissue engineering and other biomedical research. In particular, we aim reproducing hierarchical structures often found in natural composites (like bone). In these, precise arrangement of different component is controlled at a large range of length scales, in this way allowing for optimization of material properties. By combination of engineering, microfabrication, molecular self-assembly and mineral deposition controlled by molecular interactions we hope to control the structure of materials at the length scales from mm to nm. A successful outcome of this project will provide comprehensive insight into biomaterial fabrication using cell-friendly methods, and mechanistic insight into interactions between cells and structurally well organized (anisotropic, hierarchical) materials. Due to a highly complex architecture across wide length scales (nm-cm), the hierarchical structure of bone is not easily replicated in the laboratory. One strategy to overcome this obstacle when engineering synthetic bone tissue scaffolds is to provide a simpler material which inherent or implanted cells can remodel into natural bone. We have therefore investigated different forms of calcium phosphate, especially focusing on these which are more soluble and can be remodelled under physiological conditions. We have developed methods to prepare alginate mineralizaed with DCPD (dicalcium phosphate dihydrate), as well as investigated how crystallization of this CaP phase is effected by the alginate hydrogel. The method is based on controlling nucleation of desired crystal phase using well defined seed crystals, as due to non-homogeneous and time dependent driving forces for mineral phase formation and due to interactions between biopolymer network and crystal growth, controlled mineralisation of hydrogel-based materials is challenging. Biocompatibility of the material as well as transformation under physiological conditions have also been investigated. In addition we have developed a new model system in which hydrogel mineralization can be studied in great details. This system allows for the mineralization process to be studied in correlative manner using a variety of experimental techniques. It allowed us to in details describe mineral deposition process, showing conclusively that within a hydrogel matrix and under used experimental condition, amorphous calcium phosphate is formed first, and this phase quickly transform into more stable crystalline polymorphs. At the same time we have focused on the use of microfluidics to prepare alginate composites with precisely controlled geometry and composition. Significant progress have been made in this area, including development and adaptation of existing methods to make mineralizaed hydrogel beads and fibres, encapsulation of cells and fabrication of composite materials. We have discovered a new method to form hydrogels of ionotropic polymers using competitive displacement of chelated ions, thus making specific ions available to induce interactions between polymer chains and form a hydrogel. This strategy enables control of ion release kinetics within an aqueous polymer solution and thus control over gelation kinetics across a wide range of pH. Previously, we have shown that mineralized alginate can support osteoblast cultures and the ability to form synthetic bone like constructs. We build on this experience to develop a co-culture of osteoblasts and osteoclasts to create a synthetic bone-like 3D platform using mineralised alginate microbeads. As natural human bone undergoes continuous remodelling by a co-ordinated process involving three main cell types: bone formation is mediated by osteoblasts and osteocytes and bone resorption by osteoclasts, with all the cell types affecting each other. Imbalances in this process may cause pathological loss of bone mass as seen in conditions of delayed bone healing after fracture, osteoporosis and metabolic bone diseases. As this imbalance cannot be considered in mono-cellular systems, development of co-culture systems is of a major importance.

Proposed project aims at using a multidisciplinary material science and fabrication approach to address important challenges related to hierarchical, hydrogel-based biomaterials, with specific focus on materials for bone tissue engineering. We propose a novel approach to make complex, organic-inorganic composites with hierarchical structure and inherent anisotropy. The main challenge in making materials which mimic complex tissue, is in control of the structure on different length scales, from nm to mm. We propose to achieved this by a combination of engineering, micro-fabrication, molecular self-assembly and mineral deposition controlled by molecular interactions. A successful outcome of this project will provide comprehensive insight into fabrication of hierarchical composite biomaterials using cell-friendly methods, and mechanistic insight into interactions between cells and structurally well organized (anisotropic, hierarchical) materials.