Optimal design and fabrication of scaffolds to mimic tissue properties and satisfy biological constraints

Bone tissue engineering scaffolds must shape regenerating tissue, provide temporary mechanical support and enhance tissue regeneration. These requirements result in conflicting design goals. For example, increased temporary mechanical function requires a dense scaffold while enhanced cell/gene delivery requires a porous scaffold. This paper demonstrates an image-based homogenization optimization approach that can design scaffold microstructure, scaffold material and regenerate tissue microstructure to meet conflicting design requirements. In addition, constraints to ensure adequate cell/gene delivery can be introduced using a minimum porosity threshold. Homogenization theory was used to compute relationships between scaffold microstructure and effective stiffness. The functional relationships were used in the MATLAB optimization toolbox to compute optimal pore dimensions and scaffold material such that the scaffold and regenerate tissue effective stiffness matched that of native bone stiffness. The scaffold design was converted into STL format for solid free-form fabrication. Scaffolds were designed that matched mandibular condyle trabecular bone properties. Results showed excellent agreement between native bone properties and designed scaffold properties (all R2 > 0.89). Finally, example scaffolds were built from hydroxyapatite using a SFF casting technique.     

Indirect solid free form fabrication of local and global porous,biomimetic and composite 3D polymer-ceramic scaffolds

Precise control over scaffold material, porosity, and internal pore architecture is essential for tissue engineering. By coupling solid free form (SFF) manufacturing with conventional sponge scaffold fabrication procedures, we have developed methods for casting scaffolds that contain designed and controlled locally porous and globally porous internal architectures. These methods are compatible with numerous bioresorbable and non-resorbable polymers, ceramics, and biologic materials. Phase separation, emulsion-solvent diffusion, and porogen leaching were used to create poly(L)lactide (PLA) scaffolds containing both computationally designed global pores (500, 600, or 800 microm wide channels) and solvent fashioned local pores (50-100 microm wide voids or 5-10 microm length plates). Globally porous PLA and polyglycolide/PLA discrete composites were made using melt processing. Biphasic scaffolds with mechanically interdigitated PLA and sintered hydroxyapatite regions were fabricated with 500 and 600 microm wide global pores. PLA scaffolds with complex internal architectures that mimicked human trabecular bone were produced. Our indirect fabrication using casting in SFF molds provided enhanced control over scaffold shape, material, porosity and pore architecture, including size, geometry, orientation, branching, and interconnectivity. These scaffolds that contain concurrent local and global pores, discrete material regions, and biomimetic internal architectures may prove valuable for multi-tissue and structural tissue interface engineering.     

Engineering craniofacial scaffolds

OBJECTIVE: To develop an integrated approach for engineering craniofacial scaffolds and to demonstrate that these engineered scaffolds would have mechanical properties in the range of craniofacial tissue and support bone regeneration for craniofacial reconstruction. EXPERIMENTAL VARIABLE: Scaffold architecture designed to achieve desired elasticity and permeability. Scaffold external shape designed to match craniofacial anatomy. OUTCOME MEASURE: Final fabricated biomaterial scaffolds. Compressive mechanical modulus and strength. Bone regeneration as measured by micro-CT scanning, mechanical testing and histology. SETTING: Departments of Biomedical Engineering, Oral/Maxillofacial Surgery, and Oral Medicine, Pathology and Oncology at the University of Michigan. RESULTS: Results showed that the design/fabrication approach could create scaffolds with designed porous architecture to match craniofacial anatomy. These scaffolds could be fabricated from a wide range of biomaterials, including titanium, degradable polymers, and degradable calcium phosphate ceramics. Mechanical tests showed that fabricated scaffolds had compressive modulus ranging 50 to 2900 MPa and compressive strength ranging from 2 to over 56 MPa, within the range of human craniofacial trabecular bone. In vivo testing of designed scaffolds showed that they could support bone regeneration via delivery of BMP-7 transduced human gingival fibroblasts in a mouse model. Designed hydroxyapatite scaffolds with pore diameters ranging from 400 to 1200 microns were implanted in minipig mandibular defects for 6 and 18 weeks. Results showed substantial bone ingrowth (between 40 and 50% at 6 weeks, between 70 and 80% at 18 weeks) for all scaffolds, with no significant difference based on pore diameter. CONCLUSION: Integrated image-based design and solid free-form fabrication can create scaffolds that attain desired elasticity and permeability while fitting any 3D craniofacial defect. The scaffolds could be manufactured from degradable polymers, calcium phosphate ceramics and titanium. The designed scaffolds supported significant bone regeneration for all pore sizes ranging from 300 to 1200 microns. These results suggest that designed scaffolds are clinically applicable for complex craniofacial reconstruction.     

Effect of surface modification on protein retention and cell proliferation under strain

When culturing cells on flexible surfaces, it is important to consider extracellular matrix treatments that will remain on the surface under mechanical strain. Here we investigate differences in laminin deposited on oxidized polydimethylsiloxane (PDMS) with plasma treatment (plasma-only) vs. plasma and aminopropyltrimethoxysilane treatment (silane-linked). We use specular X-ray reflectivity (SXR), transmission electron microscopy (TEM), and immunofluorescence to probe the quantity and uniformity of laminin. The surface coverage of laminin is approximately 45% for the plasma-only and 50% for the silane-linked treatment as determined by SXR. TEM and immunofluorescence reveal additional islands of laminin aggregates on the plasma-only PDMS compared with the relatively smooth and uniform silane-linked laminin surface. We also examine laminin retention under strain and vascular smooth muscle cell viability and proliferation under static and strain conditions. Equibiaxial stretching of the PDMS surfaces shows greatly improved retention of the silane-linked laminin over plasma-only. There are significantly more cells on the silane-linked surface after 4 days of equibiaxial strain.     

Tissue engineering osteochondral implants for temporomandibular joint repair

Tissue engineering has provided an alternative to traditional strategies to repair and regenerate temporomandibular joints (TMJ). A successful strategy to engineer osteochondral tissue, such as that found in the TMJ, will produce tissue that is both biologically and mechanically functional. Image-based design (IBD) and solid free-form (SFF) fabrication can be used to generate scaffolds that are load bearing and match patient and defect site geometry. The objective of this study was to demonstrate how scaffold design, materials, and biological factors can be used in an integrated approach to regenerate a multi-tissue interface. IBD and SFF were first used to create biomimetic scaffolds with appropriate bulk geometry and microarchitecture. Biphasic composite scaffolds were then manufactured with the same techniques and used to simultaneously generate bone and cartilage in discrete regions and provide for the development of a stable interface between cartilage and subchondral bone. Poly-l-lactic acid/hydroxyapatite composite scaffolds were differentially seeded with fibroblasts transduced with an adenovirus expressing bone morphogenetic protein-7 in the ceramic phase and fully differentiated chondrocytes in the polymeric phase, and were subcutaneously implanted into mice. Following implantation in the ectopic site, the biphasic scaffolds promoted the simultaneous growth of bone, cartilage, and a mineralized interface tissue. Within the ceramic phase, the pockets of tissue generated included blood vessels, marrow stroma, and adipose tissue. This combination of IBD and SFF-fabricated biphasic scaffolds with gene and cell therapy is a promising approach to regenerate osteochondral defects and, ultimately, the TMJ.