Plastic Surgery Research Council

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Scaffolding the Scaffold: Mitigating Loss of Volume and Topography of Engineered Auricular Cartilage Using 3D Printed Contour Matching Cages
Alexandra Lin, BA1, Jaime Bernstein, BS1, Benjamin Cohen, BS2, Justin Buro, BA1, Karel-Bart Celie, BA1, Yoshiko Toyoda, BA1, Andrew Miller, BS1, Alice Harper, BA1, Lawrence J. Bonassar, PhD2, John P. Morgan, PhD1, Jason A. Spector, MD FACS1.
1Laboratory of Bioregenerative Medicine and Surgery, New York, NY, USA, 2Meinig School of Biomedical Engineering, Ithaca, NY, USA.

PURPOSE: Autologous reconstruction of the ear, whether for microtia or acquired deformity, is a complex procedure with substantial donor site morbidity and suboptimal aesthetic outcomes. An engineered auricular scaffold would obviate donor morbidity and provide improved aesthetic outcomes. A major obstacle to clinical translation of tissue-engineered auricles is the significant contraction and loss of topography that occurs during maturation of the soft collagen/chondrocyte matrix into elastic cartilage. Previously, we demonstrated that a 3D-printed biodegradable cage significantly mitigated contraction of simple disc-shaped collagen hydrogels seeded with human auricular chondrocytes (HAuCs) in vivo without impeding the development of elastic cartilage. Herein we fabricate cages to invest chondrocyte-collagen hydrogels with more intricate “anatomic” topographic features.
METHODS: Custom external cages were designed with a geometric element representative of the helical rim using SolidWorks (Dassault Systèmes, Vélizy-Villacoublay, France), then 3D-printed using polylactic acid (PLA) on a 5th generation MakerBot printer (MakerBot, New York, NY). Using auricular cartilage from discarded otoplasty specimens, HAuCs were harvested and expanded to passage 2. The chondrocytes were encapsulated into type I collagen hydrogels at a density of 25million cells/mL with high fidelity contour matching to the cages. The hydrogels, either protected or unprotected by the PLA cages, were implanted into nude rats and explanted after 3 months.
RESULTS: After 3 months in vivo, all constructs developed a glossy white cartilaginous appearance, similar to native auricular cartilage. Histologic analysis demonstrated development of an organized perichondrium composed of collagen, a rich proteoglycan matrix, cellular lacunae, and a dense elastin fibrin network by safranin-O and Verhoeff's stain. Biochemical analysis confirmed similar amounts of proteoglycan and hydroxyproline content in the constructs when compared to native auricular cartilage. Cage-protected constructs contracted significantly less than unprotected constructs on base area comparison (14.33% vs. 56%, p=0.0023), retained volume (213.4mm3 vs. 117.2mm3 compared to original volume of 280mm3 and corresponding to 76.2% vs. 41.9% retention, p=0.0290), and maintenance of the topographic “helical rim” feature compared to unprotected constructs. Constructs were imaged via computed tomography with an Inveon Pre-clinical MicroPET/CT/SPECT (CTI/Siemens, Knoxville, TN), then digitally reconstructed with Imaris (Bitplane, Belfast, UK). Preservation of the “helical rim” feature was evaluated subjectively by gross examination and objectively by measuring the angle between the rim and base of the constructs, a measurement that demonstrated a significant difference between protected and unprotected constructs, respectively (151.8° vs. 197.7°, p=0.0445), and that indicated protected constructs better maintain the initial angle (110°) between rim and base.
CONCLUSIONS: We have shown that custom contour matched 3D-printed biocompatible/biodegradable external cages significantly mitigate contraction and maintain the complex topography of HAuC constructs. Furthermore, cages do not impede formation of mature elastic cartilage. This technique can be used to create custom cages that contour to any form, enabling the fabrication of engineered autologous cartilage tailored to individual patient anatomy, without the contraction and loss of topography that has thus far impeded translation to the clinic.


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