Plastic Surgery Research Council
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PSRC 60th Annual Meeting
Program and Abstracts

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Co-Implantation of Auricular Chondrocytes and Mesenchymal Stem Cells for the Generation of Auricular Elastic Neocartilage: Strategy Towards the Translation of Tissue Engineered Ears
Ope A. Asanbe, MD1, Benjamin P. Cohen, BS2, Peipei Zhang, PhD1, Wilmina N. Landford, BA1, Adam M. Jacoby, BA1, Lawrence J. Bonassar, PhD2, Jason A. Spector, MD, FACS1.
1Weill Cornell Medical College, New York, NY, USA, 2Cornell University, Ithaca, NY, USA.

PURPOSE: We previously fabricated high fidelity patient-specific ears using injection molding and bovine chondrocytes. A major obstacle to the clinical translation of this approach is the large number (approximately 250 million) of auricular chondrocytes (ACs) required for the fabrication of a full sized ear scaffold. Unfortunately, in patients with microtia, only a few million ACs can be isolated from patient donor sites (the microtic remnant and the contralateral conchal bowl), necessitating strategies for the augmentation of the limited chondrocyte supply. As chondrocyte expansion causes de-differentiation and loss of chondrogenic capacity, we hypothesize that mesenchymal stem cells (MSCs), which can be obtained reliably in large numbers, expanded significantly and are known to release chondrogenic trophic factors, co-implanted with ACs will form elastic cartilage while requiring significantly fewer ACs.
METHODS: Bovine ACs (BACs) and bovine MSCs (bMSCs) were encapsulated into 10 mg/mL type I collagen hydrogels in ratios of 100:0, 50:50 and 0:100 BACs:bMSCs with a constant cell density of 25 million cells/mL hydrogel. Two mm thick collagen sheet gels were fabricated, and 8mm diameter disc constructs were extracted using a biopsy punch. The discs were then implanted subcutaneously in the dorsa of nude mice. Constructs were explanted at 1 and 3 month time points and thereafter analyzed for gross morphology, histology and biochemical composition.
RESULTS: Gross inspection of the explants showed that the BACs:bMSCs combination constructs maintained their size and exhibited cartilage-like elasticity after 1 and 3 months of implantation. Comparatively, 100% BACs and 100% bMSCs scaffolds were significantly contracted and resorbed after 3 months. H&E staining demonstrated BACs:bMSCs scaffolds developed auricular cartilage features, including cellular lacunae and perichondrial layering. Verhoeff staining revealed BACs:bMSCs combination constructs deposited more dense elastin fibers than did 100% BACs scaffolds. Furthermore, biochemical analysis confirmed that mixed cell constructs featured significantly more proteoglycan content than either the 100% BACs or 100% bMSCs groups. Additionally, significantly more proteoglycan content was noted after 3 months compared to 1 month in the BACs:bMSCs combination explants.
DISCUSSION: 1:1 BACs:bMSCs co-implanted constructs maintained their size and shape after 1 and 3 months of implantation, while constructs featuring only BACs or bMSCs contracted and resorbed. Mixed cell constructs developed auricular cartilage features and deposited critical cartilage molecular components more readily than BACs or bMSCs alone. The in vivo maturation of mixed cell constructs into elastic cartilage demonstrates the potential for MSCs to supplement the limited pool of ACs for tissue engineered auricular reconstruction techniques. Using human cells, methods similar to those elaborated here are currently being investigated, bringing us significantly closer to the fabrication of patient-specific, high fidelity human auricles for the treatment of microtia.


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