Pore architecture effects on chondrogenic potential of patient-specific 3-dimensionally printed porous tissue bioscaffolds for auricular tissue engineering.

OBJECTIVE: This study aims to determine the effect of auricular scaffold microarchitecture on chondrogenic potential in an in vivo animal model. METHODS: DICOM computed tomography (CT) images of a human auricle were segmented to create an external anatomic envelope. Image-based design was used to generate 1) orthogonally interconnected spherical pores and 2) randomly interspersed pores, and each were repeated in three dimensions to fill the external auricular envelope. These auricular scaffolds were then 3D printed by laser sintering poly-l-caprolactone, seeded with primary porcine auricular chondrocytes in a hyaluronic acid/collagen hydrogel and cultured in a pro-chondrogenic medium. The auricular scaffolds were then implanted subcutaneously in rats and explanted after 4 weeks for analysis with Safranin O and Hematoxylin and Eosin staining. RESULTS: Auricular constructs with two micropore architectures were rapidly manufactured with high fidelity anatomic appearance. Subcutaneous implantation of the scaffolds resulted in excellent external appearance of both anterior and posterior auricular surfaces. Analysis on explantation showed that the defined, spherical micropore architecture yielded histologic evidence of more robust chondrogenic tissue formation as demonstrated by Safranin O and Hematoxylin and Eosin staining. CONCLUSIONS: Image-based computer-aided design and 3D printing offers an exciting new avenue for the tissue-engineered auricle. In early pilot work, creation of spherical micropores within the scaffold architecture appears to impart greater chondrogenicity of the bioscaffold. This advantage could be related to differences in permeability allowing greater cell migration and nutrient flow, differences in surface area allowing different cell aggregation, or a combination of both factors. The ability to design an anatomically correct scaffold that maintains its structural integrity while also promoting auricular cartilage growth represents an important step towards clinical applicability of this new technology.

Biomechanical evaluation of human and porcine auricular cartilage.

OBJECTIVES/HYPOTHESIS: The mechanical properties of normal auricular cartilage provide a benchmark against which to characterize changes in auricular structure/function due to genetic defects creating phenotypic abnormalities in collagen subtypes. Such properties also provide inputs/targets for auricular reconstruction scaffold design. Several studies report the biomechanical properties for septal, costal, and articular cartilage. However, analogous data for auricular cartilage are lacking. Therefore, our aim in this study was to characterize both whole-ear and auricular cartilage mechanics by mechanically testing specimens and fitting the results to nonlinear constitutive models. STUDY DESIGN: Mechanical testing of whole ears and auricular cartilage punch biopsies. METHODS: Whole human cadaveric ear and auricular cartilage punch biopsies from both porcine and human cartilage were subjected to whole-ear helix-down compression and quasistatic unconfined compression tests. Common hyperelastic constitutive laws (widely used to characterize soft tissue mechanics) were evaluated for their ability to represent the stress-strain behavior of auricular cartilage. RESULTS: Load displacement curves for whole ear testing exhibited compliant linear behavior until after significant displacement where nonlinear stiffening occurred. All five commonly used two-term hyperelastic soft tissue constitutive models successfully fit both human and porcine nonlinear elastic behavior (mean R(2) fit >0.95). CONCLUSIONS: Auricular cartilage exhibits nonlinear strain-stiffening elastic behavior that is similar to other soft tissues in the body. The whole ear exhibits compliant behavior with strain stiffening at high displacement. The constants from the hyperelastic model fits provide quantitative baselines for both human and porcine (a commonly used animal model for auricular tissue engineering) auricular mechanics. LEVEL OF EVIDENCE: NA

Computer aided-designed, 3-dimensionally printed porous tissue bioscaffolds for craniofacial soft tissue reconstruction.

OBJECTIVE: To determine the potential of an integrated, image-based computer-aided design (CAD) and 3-dimensional (3D) printing approach to engineer scaffolds for head and neck cartilaginous reconstruction for auricular and nasal reconstruction. STUDY DESIGN: Proof of concept revealing novel methods for bioscaffold production with in vitro and in vivo animal data. SETTING: Multidisciplinary effort encompassing 2 academic institutions. SUBJECTS AND METHODS: Digital Imaging and Communications in Medicine (DICOM) computed tomography scans were segmented and utilized in image-based CAD to create porous, anatomic structures. Bioresorbable polycaprolactone scaffolds with spherical and random porous architecture were produced using a laser-based 3D printing process. Subcutaneous in vivo implantation of auricular and nasal scaffolds was performed in a porcine model. Auricular scaffolds were seeded with chondrogenic growth factors in a hyaluronic acid/collagen hydrogel and cultured in vitro over 2 months' duration. RESULTS: Auricular and nasal constructs with several types of microporous architecture were rapidly manufactured with high fidelity to human patient anatomy. Subcutaneous in vivo implantation of auricular and nasal scaffolds resulted in an excellent appearance and complete soft tissue ingrowth. Histological analysis of in vitro scaffolds demonstrated native-appearing cartilaginous growth that respected the boundaries of the scaffold. CONCLUSION: Integrated, image-based CAD and 3D printing processes generated patient-specific nasal and auricular scaffolds that supported cartilage regeneration.

Mechanical characterization and non-linear elastic modeling of poly(glycerol sebacate) for soft tissue engineering.

Scaffold tissue engineering strategies for repairing and replacing soft tissue aim to improve reconstructive and corrective surgical techniques whose limitations include suboptimal mechanical properties, fibrous capsule formation and volume loss due to graft resorption. An effective tissue engineering strategy requires a scaffolding material with low elastic modulus that behaves similarly to soft tissue, which has been characterized as a nonlinear elastic material. The material must also have the ability to be manufactured into specifically designed architectures. Poly(glycerol sebacate) (PGS) is a thermoset elastomer that meets these criteria. We hypothesize that the mechanical properties of PGS can be modulated through curing condition and architecture to produce materials with a range of stiffnesses. To evaluate this hypothesis, we manufactured PGS constructs cured under various conditions and having one of two architectures (solid or porous). Specimens were then tensile tested according to ASTM standards and the data were modeled using a nonlinear elastic Neo-Hookean model. Architecture and testing conditions, including elongation rate and wet versus dry conditions, affected the mechanical properties. Increasing curing time and temperature led to increased tangent modulus and decreased maximum strain for solid constructs. Porous constructs had lower nonlinear elastic properties, as did constructs of both architectures tested under simulated physiological conditions (wetted at 37 °C). Both solid and porous PGS specimens could be modeled well with the Neo-Hookean model. Future studies include comparing PGS properties to other biological tissue types and designing and characterizing PGS scaffolds for regenerating these tissues.

Effect of polycaprolactone scaffold permeability on bone regeneration in vivo.

Successful bone tissue engineering depends on the scaffold's ability to allow nutrient diffusion to and waste removal from the regeneration site, as well as provide an appropriate mechanical environment. Since bone is highly vascularized, scaffolds that provide greater mass transport may support increased bone regeneration. Permeability encompasses the salient features of three-dimensional porous scaffold architecture effects on scaffold mass transport. We hypothesized that higher permeability scaffolds will enhance bone regeneration for a given cell seeding density. We manufactured poly-ɛ-caprolactone scaffolds, designed to have the same internal pore design and either a low permeability (0.688×10(-7)m(4)/N-s) or a high permeability (3.991×10(-7)m(4)/N-s), respectively. Scaffolds were seeded with bone morphogenic protein-7-transduced human gingival fibroblasts and implanted subcutaneously in immune-compromised mice for 4 and 8 weeks. Micro-CT evaluation showed better bone penetration into high permeability scaffolds, with blood vessel infiltration visible at 4 weeks. Compression testing showed that scaffold design had more influence on elastic modulus than time point did and that bone tissue infiltration increased the mechanical properties of the high permeability scaffolds at 8 weeks. These results suggest that for polycaprolactone, a more permeable scaffold with regular architecture is best for in vivo bone regeneration. This finding is an important step toward the end goal of optimizing a scaffold for bone tissue engineering.