3-D Cell Culture: Primary Cells

Primary Cells

The Extracel™ Hydrogel Kits (Extracel™ and Extracel™-HP) emulate a simplified version of the extracellular matrix (ECM). Extracel is composed of Glycosil™ (thiol-modified hyaluronan), Gelin-S™ (thiol-modified denatured collagen), and Extralink™ (thiol-reactive crosslinking agent). Thiol-modified heparin has been added to Extracel to produce Extracel-HP.

Gelin-S serves as a general attachment signal for cell growth. However, some primary cells require specific ECM proteins to expand and retain function (e.g. adult, human, polyploid hepatocytes require collagen I; breast epithelial cells require laminin). In these situations, ECM proteins can be non-covalently added into the hydrogels prior to crosslinking to mimic a specific ECM environment.

The rigidity of the Extracel-based hydrogel can be altered either by varying the concentrations of the hydrogel solutions or by changing the amount of Extralink used to crosslink the hydrogel. Cells can be encapsulated during crosslinking, where they attach and grow within the hydrogel matrix, or they can be plated on top of the hydrogel for pseudo three-dimensional growth. All Extracel Hydrogel Kits are tested for bacterial growth and endotoxins.

The following primary cells have been cultured in Extracel-based hydrogels:

  • human primary hepatocytes1,2
  • rat primary hepatocytes1,2
  • human tracheal scar fibroblasts3,4
  • human dermal fibroblasts5,6
  • human fetal osteoblasts7
  • chick dorsal root ganglia8

References

  1. G. D. Prestwich, Y. Liu, M. Serban, B. Yu, X. Z. Shu, and A. Scott, “3-D Culture in Synthetic Extracellular Matrices: New Tissue Models for Drug Toxicology and Cancer Drug Discovery,” invited, Adv. Enz. Res., in press (2007).
  2. Unpublished data from G. D. Prestwich, et al, University of Utah.
  3. Y. Liu, Z. X. Shu, S. D. Gray, G. D. Prestwich, “Disulfide-crosslinked Hyaluronan-Gelatin Sponge: Growth of Fibrous Tissue In Vivo,” J Biomed Mat Res, 68A, 142-149 (2004).
  4. X. Z. Shu, Y. Liu, F. Palumbo, Y. Luo, G. D. Prestwich, “In Situ Crosslinkable Hyaluronan Hydrogels for Tissue Engineering,” Biomaterials, 25, 1339-1348 (2004).
  5. G. D. Prestwich, X. Z. Shu, Y. Liu, S. Cai, J. F. Walsh, C. W. Hughes, K. R. Kirker, R. R. Orlandi, A. H. Park, S. L. Thibeault, M. E. Smith, “Injectable Synthetic Extracellular Matrices for Tissue Engineering and Repair,” Adv. Exp. Med. Biol., 585, 125-133 (2006).
  6. K. Ghosh, Z. Pan, E. Guan, S. Ge, Y. Liu, T. Nakamura, X. Ren, M. Rafailovich, R. Clark, “Cell Adaptation to a Physiologically Relevant ECM Mimic with Different Viscoelastic Properties,” Biomaterials 28, 671-679 (2007).
  7. Unpublished data from Dietmar Hutmacher, et al, University of Singapore.
  8. E. M. Horn, M. Beaumont, X. Z. Shu, A. Harvey, G. D. Prestwich, K. M. Horn, A. R. Gibson, M. C. Preul, A. Panitch, “Influence of cross-linker hyaluronic acids on neurite outgrowth and recovery from spinal cord injury,” J of Neurosurg Spine, 6(2), 133-40 (2007).