Hierarchical Nanofibrous Microspheres with Controlled Growth Factor Delivery for Bone Regeneration

The integration of controlled growth factor delivery and biomimetic architecture into a microsphere is a challenging but attractive strategy for developing new injectable biomaterials. In this work, a unique hierarchical nanosphere-encapsulated-in-microsphere scaffolding system is developed. First, heparin-conjugated gelatin (HG) is synthesized, which provides binding domains for bone morphogenetic protein 2 (BMP2) to stabilize this growth factor, protect it from denaturation and proteolytic degradation, and subsequently prolong its sustained release. Next, a unique approach is developed which includes a water-in-oil-in-oil double emulsion process and a thermally induced phase separation to encapsulate BMP2-binding HG nanospheres into nanofibrous microspheres. The nanofibrous microsphere is self-assembled from synthetic nanofibers, and has superior surface area, high porosity, low density, and is an excellent carrier to support cell adhesion and tissue in-growth. BMP2 in the hierarchical microsphere is released in a multiple-controlled manner by the binding with heparin and encapsulation of the nanosphere and microsphere. An in vivo calvarial defect model confirms that this microsphere is an excellent osteoinductive scaffold for enhanced bone regeneration. By choosing different growth factors, this hierarchical microsphere system can easily be applied to other types of tissue regeneration. The work expands the ability to develop new injectable biomaterials for advanced regenerative therapies.