3D-Printed Scaffold with Halloysite Nanotubes Laden As a Sequential Drug Delivery System Regulates Vascularized Bone Tissue Healing

Bone repair is a complex, multi-stage process, involving angiogenesis and osteogenesis, under the control of sequential stimulation from multiple factors such as FGFs, VEGF, and BMP2. Promoting the bone repair processes at different stages can greatly shorten the period and improve the quality of repair. How to deliver angiogenic and osteogenic cues in sequence still remains a challenge. Therefore, this research proposed a 3D-printed scaffold with a sequential delivery platform loaded with nanotubes and microspheres to realize the coupling regeneration of blood vessels and bones. Deferoxamine was loaded onto halloysite nanotubes by electrostatic interaction to promote the pre-vascularization of the defect area, which can provide a better blood supply for the subsequent regeneration of bone tissue. BMP2 was encapsulated into the microspheres to achieve continuous long-term osteogenic induction. The PLGA/TCP solution mixed with microspheres and halloysite nanotubes was shaped into a scaffold by 3D low-temperature deposition printing to ensure drug inactivation did not occur. The drug delivery system inside the scaffold released pro-angiogenic drugs within the first week of repair and maintained an effective level of bone regeneration-promoting growth factor for an extended period, consistent with the cascade of the natural bone repair process. In vitro results revealed that the scaffold could enhance the proliferation capacity and osteogenic process of bone marrow mesenchymal stem cells (BMSCs), as well as the proliferation and adherent junctions of human umbilical endothelial cells (HUVECs). Furthermore, subcutaneous ectopic experiments demonstrated better angiogenesis and osteoinductive effects in vivo, which is characterized by collagen deposition, the number of capillaries and calcium nodules. Current study highlights the great potential of scaffold with a sequential delivery platform for clinical application.