Plastic Surgery Research Council
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PSRC 60th Annual Meeting

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Beyond Cotton Candy: Fabrication of Capillary Networks within Biocompatible Tissue- Engineered Constructs from Kerria Lacca Resin (Shellac)
Adam Jacoby, BA, Rachel C. Hooper, MD, Jeremiah Joyce, BA, Remco Bleecker, BA, Jason A. Spector, MD, FACS.
Weill Cornell Medical College, New York, NY, USA.

PURPOSE: Creation of synthetic tissues with microvascular networks, which adequately mimics the size and density of capillaries found in vivo, remains one of the foremost challenges in the field of tissue engineering. In our previous work, we utilized a sacrificial microfiber technique whereby 1.5 mm Pluronic F127 microfibers were embedded within a collagen matrix, leaving a patent internal channel, which was subsequently seeded with endothelial and smooth muscle cells forming a neointima and neomedia respectively. Here we describe two approaches to synthesize a tissue-engineered construct with macro-inlet and outlet vessels, bridged by a dense network of microvessels, which recapitulates the hierarchical organization of an arteriole, venule, and capillary bed found in vivo.
METHODS: 1.5 mm longitudinal Pluronic F127 macrofibers were created using pre-formed polydimethylsiloxane (PDMS) molds. Dense three dimensional matrices were fabricated using one of two techniques: 1) manual extrusion of 100-500 µm Pluronic F127 “mesh” or “checkerboard” 2) melt-spinning 10-400 µm Shellac (SSB 55 Astra FL) microfiber “cotton candy” or “fluff,” both of which are FDA approved materials. Microfibers were melt-fused with Pluronic F127 macrofibers to complete the circuit. Pre-vascular networks were embedded in type I collagen, sacrificed and intraluminally seeded with 5x10^6 cells/mL human aortic smooth muscle cells (HASMC) followed by 5x10^6 cells/mL HUVEC 24 hours after. Constructs were prepared for 7 day static culture and a subset underwent dynamic culture via gravity driven perfusion. Architecture and cell viability were confirmed via histology and imaging.
RESULTS: Pluronic and Shellac/Pluronic three-dimensional constructs were successfully embedded and sacrificed in type I collagen, leaving behind patent microchannels ranging in size from 10 to 500 µm. Patency was confirmed via perfusion with colored buffer solution, whole rat blood, and gadolinium injection during microMRI. Hematoxylin and eosin staining after 7 days of static culture confirmed patency and the presence of a dense tangle of interconnecting microchannels with adherent cells along the luminal surfaces of both micro and macro channels. Additionally, following dynamic pressure-gradient driven perfusion, cells remained adherent along the luminal surfaces of the macro- and microchannels.
CONCLUSION: We have modified our previously described sacrificial approach for in vivo application and have developed two novel techniques to create dense, three-dimensional microvascular networks within tissue-engineered constructs using Pluronic F127 and Shellac sacrificial microfibers. Both techniques leave micro/macro channels that support adhesion and growth of endothelial cells crucial to providing thrombosis-free flow through the network. These results represent significant progress towards the fabrication of constructs with a hierarchical vascular network analogous to that seen in vivo, a necessity for the production of human-scale engineered tissues.


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