Geometric Mismatch Promotes Anatomic Repair in Periorbital Bony Defects in Skeletally Mature Yucatan Minipigs.

Porous tissue-engineered 3D-printed scaffolds are a compelling alternative to autografts for the treatment of large periorbital bone defects. Matching the defect-specific geometry has long been considered an optimal strategy to restore pre-injury anatomy. However, studies in large animal models have revealed that biomaterial-induced bone formation largely occurs around the scaffold periphery. Such ectopic bone formation in the periorbital region can affect vision and cause disfigurement. To enhance anatomic reconstruction, geometric mismatches are introduced in the scaffolds used to treat full thickness zygomatic defects created bilaterally in adult Yucatan minipigs. 3D-printed, anatomically-mirrored scaffolds are used in combination with autologous stromal vascular fraction of cells (SVF) for treatment. An advanced image-registration workflow is developed to quantify the post-surgical geometric mismatch and correlate it with the spatial pattern of the regenerating bone. Osteoconductive bone growth on the dorsal and ventral aspect of the defect enhances scaffold integration with the native bone while medio-lateral bone growth leads to failure of the scaffolds to integrate. A strong positive correlation is found between geometric mismatch and orthotopic bone deposition at the defect site. The data suggest that strategic mismatch >20% could improve bone scaffold design to promote enhanced regeneration, osseointegration, and long-term scaffold survivability.

Point-of-care treatment of geometrically complex midfacial critical-sized bone defects with 3D-Printed scaffolds and autologous stromal vascular fraction.

Critical-sized midfacial bone defects present a unique clinical challenge due to their complex three-dimensional shapes and intimate associations with sensory organs. To address this challenge, a point-of-care treatment strategy for functional, long-term regeneration of 2 cm full-thickness segmental defects in the zygomatic arches of Yucatan minipigs is evaluated. A digital workflow is used to 3D-print anatomically precise, porous, biodegradable scaffolds from clinical-grade poly-ε-caprolactone and decellularized bone composites. The autologous stromal vascular fraction of cells (SVF) is isolated from adipose tissue extracts and infused into the scaffolds that are implanted into the zygomatic ostectomies. Bone regeneration is assessed up to 52 weeks post-operatively in acellular (AC) and SVF groups (BV/DV = 0.64 ± 0.10 and 0.65 ± 0.10 respectively). In both treated groups, bone grows from the adjacent tissues and restores the native anatomy. Significantly higher torque is required to fracture the bone-scaffold interface in the SVF (7.11 ± 2.31 N m) compared to AC groups (2.83 ± 0.23 N m). Three-dimensional microcomputed tomography analysis reveals two distinct regenerative patterns: osteoconduction along the periphery of scaffolds to form dense lamellar bone and small islands of woven bone deposits growing along the struts in the scaffold interior. Overall, this study validates the efficacy of using 3D-printed bioactive scaffolds with autologous SVF to restore geometrically complex midfacial bone defects of clinically relevant sizes while also highlighting remaining challenges to be addressed prior to clinical translation.

3D-printed oxygen-releasing scaffolds improve bone regeneration in mice.

Low oxygen (O2) diffusion into large tissue engineered scaffolds hinders the therapeutic efficacy of transplanted cells. To overcome this, we previously studied hollow, hyperbarically-loaded microtanks (µtanks) to serve as O2 reservoirs. To adapt these for bone regeneration, we fabricated biodegradable µtanks from polyvinyl alcohol and poly (lactic-co-glycolic acid) and embedded them to form 3D-printed, porous poly-ε-caprolactone (PCL)-µtank scaffolds. PCL-µtank scaffolds were loaded with pure O2 at 300-500 psi. When placed at atmospheric pressures, the scaffolds released O2 over a period of up to 8 h. We confirmed the inhibitory effects of hypoxia on the osteogenic differentiation of human adipose-derived stem cells (hASCs and we validated that µtank-mediated transient hyperoxia had no toxic impacts on hASCs, possibly due to upregulation of endogenous antioxidant regulator genes. We assessed bone regeneration in vivo by implanting O2-loaded, hASC-seeded, PCL-µtank scaffolds into murine calvarial defects (4 mm diameters × 0.6 mm height) and subcutaneously (4 mm diameter × 8 mm height). In both cases we observed increased deposition of extracellular matrix in the O2 delivery group along with greater osteopontin coverages and higher mineral deposition. This study provides evidence that even short-term O2 delivery from PCL-µtank scaffolds may enhance hASC-mediated bone tissue regeneration.

3D-Printing Technologies for Craniofacial Rehabilitation, Reconstruction, and Regeneration.

The treatment of craniofacial defects can present many challenges due to the variety of tissue-specific requirements and the complexity of anatomical structures in that region. 3D-printing technologies provide clinicians, engineers and scientists with the ability to create patient-specific solutions for craniofacial defects. Currently, there are three key strategies that utilize these technologies to restore both appearance and function to patients: rehabilitation, reconstruction and regeneration. In rehabilitation, 3D-printing can be used to create prostheses to replace or cover damaged tissues. Reconstruction, through plastic surgery, can also leverage 3D-printing technologies to create custom cutting guides, fixation devices, practice models and implanted medical devices to improve patient outcomes. Regeneration of tissue attempts to replace defects with biological materials. 3D-printing can be used to create either scaffolds or living, cellular constructs to signal tissue-forming cells to regenerate defect regions. By integrating these three approaches, 3D-printing technologies afford the opportunity to develop personalized treatment plans and design-driven manufacturing solutions to improve aesthetic and functional outcomes for patients with craniofacial defects.

Three-Dimensional Printing of Bone Extracellular Matrix for Craniofacial Regeneration.

Tissue-engineered approaches to regenerate bone in the craniomaxillofacial region utilize biomaterial scaffolds to provide structural and biological cues to stem cells to stimulate osteogenic differentiation. Bioactive scaffolds are typically comprised of natural components but often lack the manufacturability of synthetic materials. To circumvent this trade-off, we 3D printed materials comprised of decellularized bone (DCB) matrix particles combined with polycaprolactone (PCL) to create novel hybrid DCB:PCL scaffolds for bone regeneration. Hybrid scaffolds were readily printable at compositions of up to 70% bone by mass and displayed robust mechanical properties. Assessments of surface features revealed both collagenous and mineral components of bone were present. Qualitative and quantitative assessments showed increased surface roughness relative to that of pure PCL scaffolds. These findings correlated with enhanced cell adhesion on hybrid surfaces relative to that on pure surfaces. Human adipose-derived stem cells (hASCs) cultured in DCB:PCL scaffolds without soluble osteogenic cues exhibited significant upregulation of osteogenic genes in hybrid scaffolds relative to pure PCL scaffolds. In the presence of soluble phosphate, hybrid scaffolds resulted in increased calcification. The hASC-seeded scaffolds were implanted into critical-sized murine calvarial defects and yielded greater bone regeneration in DCB:PCL scaffolds compared to that in PCL-only at 1 and 3 months post-transplantation. Taken together, these results demonstrate that 3D printed DCB:PCL scaffolds might be effective for stimulating bone regeneration.