Abstract
The physiologic environment of neuronal and glial cells is the three-dimensional (3D) tissue. We and others have demonstrated that more physiologically relevant cell responses occur in 3D culture environments as opposed to flat stiff culture substrates such as tissue culture plates and coverslips. Hydrogels provide significant advantages towards providing cells a biocompatible, all-encompassing culture environment, but pose challenges towards adapting standard biochemical and molecular biology assay techniques. Herein, we provide methods for encapsulating and culturing cells inside 3D type I collagen gels. We also provide several basic methods to assess cell viability, fix and stain cells while encapsulated in the gels, and digest the gels to retrieve live cells for subculturing or other experiments (e.g., flow cytometry).
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References
Matsusaki M, Case CP, Akashi M (2014) Three-dimensional cell culture technique and pathophysiology. Adv Drug Deliv Rev 74C:95–103
Baker BM, Chen CS (2012) Deconstructing the third dimension: how 3D culture microenvironments alter cellular cues. J Cell Sci 125:3015–3024
Tibbitt MW, Anseth KS (2009) Hydrogels as extracellular matrix mimics for 3D cell culture. Biotechnol Bioeng 103:655–663
Banker G, Goslin K (1998) Culturing nerve cells. MIT Press, Cambridge, MA
McCarthy KD, de Vellis J (1980) Preparation of separate astroglial and oligodendroglial cell cultures from rat cerebral tissue. J Cell Biol 85:890–902
Ribeiro A, Balasubramanian S, Hughes D et al (2013) Beta1-Integrin cytoskeletal signaling regulates sensory neuron response to matrix dimensionality. Neuroscience 248C:67–78
Ribeiro A, Vargo S, Powell EM et al (2012) Substrate three-dimensionality induces elemental morphological transformation of sensory neurons on a physiologic timescale. Tissue Eng A 18:93–102
Zustiak SP, Leach JB (2010) Hydrolytically degradable poly(ethylene glycol) hydrogel scaffolds with tunable degradation and mechanical properties. Biomacromolecules 11:1348–1357
Zustiak SP, Pubill S, Ribeiro A et al (2013) Hydrolytically degradable poly(ethylene glycol) hydrogel scaffolds as a cell delivery vehicle: characterization of PC12 cell response. Biotechnol Prog 29:1255–1264
Kadler KE, Holmes DF, Trotter JA et al (1996) Collagen fibril formation. Biochem J 316(Pt 1):1–11
Miron-Mendoza M, Seemann J, Grinnell F (2010) The differential regulation of cell motile activity through matrix stiffness and porosity in three dimensional collagen matrices. Biomaterials 31:6425–6435
Willits RK, Skornia SL (2004) Effect of collagen gel stiffness on neurite extension. J Biomater Sci Polym Ed 15:1521–1531
Sanz-Ramos P, Mora G, Ripalda P et al (2012) Identification of signalling pathways triggered by changes in the mechanical environment in rat chondrocytes. Osteoarthr Cartilage 20:931–939
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Balasubramanian, S., Powell, E.M., Leach, J.B. (2015). Culturing Neurons, Glia, and Progenitor Cells in Three-Dimensional Hydrogels. In: Leach, J., Powell, E. (eds) Extracellular Matrix. Neuromethods, vol 93. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2083-9_9
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DOI: https://doi.org/10.1007/978-1-4939-2083-9_9
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Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2082-2
Online ISBN: 978-1-4939-2083-9
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