Extracel-X™

Extracel-X™ Hydrogel Kit

Extracel-X™ is recommended for growing tumor xenografts since it has been shown to support growth of a wide variety of tumor cancer cell types (both orthotopically and subcutaneously) in mice1.

Extracel-X™ is a synthetic extracellular matrix (ECM) used to deliver and cultivate cancer cells in vivo and for tissue-engineering applications (with and without cells). The Extracel-X™ solution can be injected and crosslinked in situ. Unlike animal-derived extracellular matrices (ECM), Extracel-X™ is chemically defined and nonimmunogenic. Extracel-X™ is based on three biocompatible components: thiol-modified hyaluronan (a major constituent of native ECM), thiol-modified gelatin (denatured collagen), and a thiol-reactive crosslinker (polyethylene glycol diacrylate, PEGDA).

Gelation

Reconstituted Extracel-X™ components remain liquid at 15 to 37°C. The hydrogel is formed when the crosslinking agent Extralink™ (PEGDA) is added to Glycosil™ (thiol-modified hyaluronan) and Gelin-S™ (thiol-modified gelatin). Gelation occurs from twenty to more than 120 minutes after all three components are mixed, depending on user requirements. No steps depend on low temperature or low pH.

Volume and Composition

The Extracel-X™ Hydrogel Kits come in two sizes:

  • 12.5 ml total hydrogel with the components in three vials (for large-volume applications)
    • 5.0 ml of Glycosil™, 5.0 ml of Gelin-S™, 2.5 ml of Extralink™
  • 7.5 ml total hydrogel with three sets of vials that make 2.5 ml each (for small-volume applications)
    • 3x 1.0 ml of Glycosil™, 3x 1.0 ml of Gelin-S™, 3x 0.5 ml of Extralink™

Applications

Tumor Xenografts

Extracel-X™ hydrogels have been used to grow subcutaneous and orthotopic tumors produced from breast, colon, and ovarian cancer cells; tumors were developed by seeding the hydrogel with organotypic cells prior to gelation and injecting them subcutaneously into mammary fat pads, subserosally into colons, and intracapsularly into ovaries of nude mice1. Extracel-X™ has also delivered red fluorescent protein-tagged metastatic pancreatic cancer cells directly into the pancreas of nude mice, thus permitting real-time intravital imaging of the primary tumor as well as metastases2. The Extracel-X™ hydrogels themselves can be fluorescently labeled for tracking in vivo.

The orthotopic delivery of cancer cells in Extracel-X™ hydrogels showed the following advantages when compared with orthotopic injection of cells in serum-free medium at four weeks following injection1:

  • increased incidence of cancer formation and reduced variability in tumor size
  • enhanced growth of organ-specific cancers with good tumor-tissue integration
  • improved vascularization and reduced necrosis in tumors
  • reduced cancer seeding on adjacent tissues or organs
  • better general animal health

3-D Cell Culture

Extracel-X™ is designed for both in vivo and in vitro applications, which allows direct comparison between lab and animal experiments. Cells can be encapsulated during crosslinking3, where they attach and grow within the hydrogel matrix, or they can be plated on top of the hydrogel for pseudo 3-D growth4. Cells are recovered from the hydrogel either by enzyme digestion for cells encapsulated in the hydrogel4,5 or by trypsinization for cells grown on the surface.

The following cancer cell lines2,6 have been cultured in Extracel-X™:

  • MDA-MB-468 (breast)
  • MCF-7 (breast)
  • MDA-MB-231 (breast)
  • SK-Br-3 (breast)
  • OVCAR-3 (ovarian)
  • SK-OV-3 (ovarian)
  • HCT-116 (colon)
  • Caco-2 (colon)
  • MiaPaCa-2 (pancreatic)

Tissue Engineering

The versatility of Extracel-X™ allows it to be used as a hydrogel7, sponge (lyophilized hydrogel)8, or film (dried hydrogel)9. It can be injected or implanted into animals for tissue-engineering applications10. In vivo degradation occurs in four to eight weeks, depending on its form and the amount of Gelin-S™ present11.

The combination of biocompatibility and flexibility in Extracel-X™ facilitates a wide range of tissue-engineering applications. For example, Extracel-X™ has been used to repair cartilage7, bone8, and vocal folds9 and cast into tubes to create blood vessels12.

To watch a video showing how easy it is to use Extracel-X™ for tumor xenografts click here.

References

  1. Y. Liu, X. Z. Shu, and G. D. Prestwich, “Tumor Engineering: Orthotopic Cancer Models in Mice Using Cell-Loaded, Injectable, Crosslinked Hyaluronan-Derived Hydrogels,” in press, Tissue Engineering (2007).
  2. Unpublished data from C. Scaife, et al, University of Utah.
  3. 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).
  4. X. Z. Shu, S. Ahmad, Y. Liu, and G. D. Prestwich, “Synthesis and Evaluation of Injectable, In Situ Crosslinkable Synthetic Extracellular Matrices (sECMs) for Tissue Engineering,” J. Biomed Mater. Res. A, 79A(4), 901-912 (2006).
  5. X. Z. Shu, Y. Liu, F. Palumbo, G. D. Prestwich, “Disulfide-Crosslinked Hyaluronan-Gelatin Hydrogel Films: A Covalent Mimic of the Extracellular Matrix for In Vitro Cell Growth,” Biomaterials, 24, 3825-3834 (2003).
  6. Unpublished data from G. D. Prestwich, et al, University of Utah.
  7. Y. Liu, X. Z. Shu, G. D. Prestwich, “Osteochondral defect repair with autologous bone marrow derived MSC cells in an injectable in situ crosslinked synthetic extracellular matrix,” Tissue Engineering, Tissue Eng., 12(12), 3405-3416 (2006).
  8. Y. Liu, S. Ahmad, X. Z. Shu, R. K. Sanders, S.A. Kopesec, G. D. Prestwich, “Accelerated repair of cortical bone defects using a synthetic extracellular matrix to deliver human demineralized bone matrix,” J Orthop Res, 24(7), 1454-1462 (2006).
  9. S. Duflo, S. L. Thibeault, W. Li, X. Z. Shu, G. D. Prestwich, “Vocal fold tissue repair in vivo using a synthetic extracellular matrix,” Tissue Engineering, 12(8), 2171-2180 (2006).
  10. X. Z. Shu, Y. Liu, F. Palumbo, Y. Luo, and G. D. Prestwich, “In Situ Crosslinkable Glycosaminoglycan Hydrogels for Tissue Engineering,” Biomaterials, 25, 1339-1348 (2004).
  11. X. Z. Shu, S. Ahmad, Y. Liu, and G. D. Prestwich, “Synthesis and Evaluation of Injectable, In Situ Crosslinkable Synthetic Extracellular Matrices (sECMs) for Tissue Engineering,” J. Biomed Mater. Res. A, 79A(4), 901-912 (2006).
  12. V. Mironov, V. Kasyanov, X. Z. Shu, C. Eisenberg, L. Eisenberg, S. Gonda, T. Trusk, R. R. Markwald, G. D. Prestwich, “Fabrication of tubular tissue constructs by centrifugal casting of cells suspended in an in situ crosslinkable hyaluronan-gelatin hydrogel,” Biomaterials, 26, 7628-7625 (2005).