Abstract
A bioengineered model of 3D brain-like tissue was developed using silk-collagen protein scaffolds seeded with primary cortical neurons. The scaffold design provides compartmentalized control for spatial separation of neuronal cell bodies and neural projections, which resembles the layered structure of the brain (cerebral cortex). Neurons seeded in a donut-shaped porous silk sponge grow robust neuronal projections within a collagen-filled central region, generating 3D neural networks with structural and functional connectivity. The silk scaffold preserves the mechanical stability of the engineered tissues, allowing for ease of handling, long-term culture in vitro and anchoring of the central collagen gel to avoid shrinkage, and enabling neural network maturation. This protocol describes the preparation and manipulation of silk-collagen constructs, including the isolation and seeding of primary rat cortical neurons. This 3D technique is useful for mechanical injury studies and as a drug screening tool, and it could serve as a foundation for brain-related disease models. The protocol of construct assembly takes 2 d, and the resulting tissues can be maintained in culture for several weeks.
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Acknowledgements
We thank S. Moss from Tufts University for providing embryonic rat brain tissues. This work was funded by a US National Institutes of Health (NIH) P41 Tissue Engineering Resource Center Grant (EB002520) and by the German Research Foundation (DFG; CH 1400/2-1, Postdoctoral Fellowship for K.C.).
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K.C. performed experimental work, data analysis and wrote the paper; M.D.T.-S., D.L.K. and F.G.O. conceived the idea; M.D.T.-S. developed the design and protocol; and D.L.K and F.G.O. supervised the project. All authors commented on the results and the manuscript.
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Chwalek, K., Tang-Schomer, M., Omenetto, F. et al. In vitro bioengineered model of cortical brain tissue. Nat Protoc 10, 1362–1373 (2015). https://doi.org/10.1038/nprot.2015.091
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DOI: https://doi.org/10.1038/nprot.2015.091
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