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Fibrosis pp 223-233 | Cite as

Cell-Populated Collagen Lattice Models

  • Beate Eckes
  • Fang Wang
  • Laure Rittié
  • Gabriele Scherr
  • Paola Zigrino
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1627)

Abstract

Investigation of cell function is often hampered by the complexity of the tissue context. This problem is circumvented by isolating cells from tissues and analyzing their behavior in culture. Most cell types are cultured as monolayers on planar, rigid Petri dishes, an environment that does not reflect the spatial, three-dimensional cellular environment in vivo. Culture in three-dimensional collagen lattices has been devised to optimize in vitro culture conditions and to provide a more physiologic “in vivo-like” environment. Collagen lattices can easily be manipulated to suit diverse cell types and to provide variable mechanical forces. Cells can be imaged in such surroundings, and gene expression as well as protein production and activity can be monitored.

Key words

Fibroblast Three-dimensional culture Collagen-binding integrin MMP Morphology Mechanical force 

Notes

Acknowledgments

We thank all the members of the dermatology department in Cologne for their continued stimulating discussion; in particular we are grateful to Cornelia Mauch and Thomas Krieg for their constructive criticism and support. Work in the Eckes and Zigrino labs is supported by the German Research Foundation (Deutsche Forschungsgemeinschaft) through SFB 829.

References

  1. 1.
    Bell E, Ivarsson B, Merrill C (1979) Production of a tissue-like structure by contraction of collagen lattices by human fibroblasts of different proliferative potential in vitro. Proc Natl Acad Sci U S A 76(3):1274–1278. doi: 10.1073/pnas.76.3.1274
  2. 2.
    Eckes B, Krieg T, Nusgens BV et al (1995) In vitro reconstituted skin as a tool for biology, pharmacology and therapy: a review. Wound Repair Regen 3(3):248–257. doi: 10.1046/j.1524-475X.1995.30304.x CrossRefPubMedGoogle Scholar
  3. 3.
    Kurschat P, Zigrino P, Nischt R et al (1999) Tissue inhibitor of matrix metalloproteinase-2 regulates matrix metalloproteinase-2 activation by modulation of membrane-type 1 matrix metalloproteinase activity in high and low invasive melanoma cell lines. J Biol Chem 274(30):21056–21062. doi: 10.1074/jbc.274.30.21056 CrossRefPubMedGoogle Scholar
  4. 4.
    Zigrino P, Mauch C, Fox JW et al (2005) Adam-9 expression and regulation in human skin melanoma and melanoma cell lines. Int J Cancer 116(6):853–859. doi: 10.1002/ijc.21087 CrossRefPubMedGoogle Scholar
  5. 5.
    Grinnell F, Petroll WM (2010) Cell motility and mechanics in three-dimensional collagen matrices. Annu Rev Cell Dev Biol 26:335–361. doi: 10.1146/annurev.cellbio.042308.113318 CrossRefPubMedGoogle Scholar
  6. 6.
    Eckes B, Dogic D, Colucci-Guyon E et al (1998) Impaired mechanical stability, migration and contractile capacity in vimentin-deficient fibroblasts. J Cell Sci 111(Pt 13):1897–1907PubMedGoogle Scholar
  7. 7.
    Grinnell F (1994) Fibroblasts, myofibroblasts, and wound contraction. J Cell Biol 124(4):401–404. doi: 10.1083/jcb.124.4.401 CrossRefPubMedGoogle Scholar
  8. 8.
    Smola H, Thiekotter G, Fusenig NE (1993) Mutual induction of growth factor gene expression by epidermal-dermal cell interaction. J Cell Biol 122(2):417–429. doi: 10.1083/jcb.122.2.417 CrossRefPubMedGoogle Scholar
  9. 9.
    Barczyk M, Carracedo S, Gullberg D (2010) Integrins. Cell Tissue Res 339(1):269–280. doi: 10.1007/s00441-009-0834-6 CrossRefPubMedGoogle Scholar
  10. 10.
    Zhang ZG, Bothe I, Hirche F et al (2006) Interactions of primary fibroblasts and keratinocytes with extracellular matrix proteins: contribution of alpha2beta1 integrin. J Cell Sci 119(Pt 9):1886–1895. doi: 10.1242/jcs.02921 CrossRefPubMedGoogle Scholar
  11. 11.
    Zweers MC, Davidson JM, Pozzi A et al (2007) Integrin alpha2beta1 is required for regulation of murine wound angiogenesis but is dispensable for reepithelialization. J Invest Dermatol 127(2):467–478. doi: 10.1038/sj.jid.5700546 CrossRefPubMedGoogle Scholar
  12. 12.
    Popova SN, Barczyk M, Tiger CF et al (2007) Alpha11 beta1 integrin-dependent regulation of periodontal ligament function in the erupting mouse incisor. Mol Cell Biol 27(12):4306–4316. doi: 10.1128/MCB.00041-07 CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Rosenfeldt H, Grinnell F (2000) Fibroblast quiescence and the disruption of ERK signaling in mechanically unloaded collagen matrices. J Biol Chem 275(5):3088–3092. doi: 10.1074/jbc.275.5.3088 CrossRefPubMedGoogle Scholar
  14. 14.
    Mauch C, Hatamochi A, Scharffetter K et al (1988) Regulation of collagen synthesis in fibroblasts within a three-dimensional collagen gel. Exp Cell Res 178(2):493–503. doi: 10.1016/0014-4827(88)90417-X CrossRefPubMedGoogle Scholar
  15. 15.
    Mauch C, Adelmann-Grill B, Hatamochi A et al (1989) Collagenase gene expression in fibroblasts is regulated by a three-dimensional contact with collagen. FEBS Lett 250(2):301–305. doi: 10.1016/0014-5793(89)80743-4 CrossRefPubMedGoogle Scholar
  16. 16.
    Zigrino P, Drescher C, Mauch C (2001) Collagen-induced proMMP-2 activation by MT1-MMP in human dermal fibroblasts and the possible role of alpha2beta1 integrins. Eur J Cell Biol 80(1):68–77. doi: 10.1078/0171-9335-00134 CrossRefPubMedGoogle Scholar
  17. 17.
    Grinnell F (2003) Fibroblast biology in three-dimensional collagen matrices. Trends Cell Biol 13(5):264–269. doi: 10.1016/S0962-8924(03)00057-6 CrossRefPubMedGoogle Scholar
  18. 18.
    Chiquet M, Matthisson M, Koch M et al (1996) Regulation of extracellular matrix synthesis by mechanical stress. Biochem Cell Biol 74(6):737–744. doi: 10.1006/excr.1998.4363 CrossRefPubMedGoogle Scholar
  19. 19.
    Kessler D, Dethlefsen S, Haase I et al (2001) Fibroblasts in mechanically stressed collagen lattices assume a “synthetic” phenotype. J Biol Chem 276(39):36575–36585. doi: 10.1074/jbc.M101602200 CrossRefPubMedGoogle Scholar
  20. 20.
    Trachslin J, Koch M, Chiquet M (1999) Rapid and reversible regulation of collagen XII expression by changes in tensile stress. Exp Cell Res 247(2):320–328. doi: 10.1006/excr.1998.4363 CrossRefPubMedGoogle Scholar
  21. 21.
    Tomasek JJ, Gabbiani G, Hinz B et al (2002) Myofibroblasts and mechano-regulation of connective tissue remodelling. Nat Rev Mol Cell Biol 3(5):349–363. doi: 10.1038/nrm809 CrossRefPubMedGoogle Scholar
  22. 22.
    Murad S, Grove D, Lindberg KA et al (1981) Regulation of collagen synthesis by ascorbic acid. Proc Natl Acad Sci U S A 78(5):2879–2882CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Rittié L, Fisher GJ (2005) Isolation and culture of skin fibroblasts. Methods Mol Med 117:83–98. doi: 10.1385/1-59259-940-0:083 PubMedGoogle Scholar
  24. 24.
    Kuroda Y, Wakao S, Kitada M et al (2013) Isolation, culture and evaluation of multilineage-differentiating stress-enduring (muse) cells. Nat Protoc 8(7):1391–1415. doi: 10.1038/nprot.2013.076 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Beate Eckes
    • 1
  • Fang Wang
    • 1
  • Laure Rittié
    • 2
    • 3
  • Gabriele Scherr
    • 1
  • Paola Zigrino
    • 1
  1. 1.Department of DermatologyUniversity of CologneCologneGermany
  2. 2.Department of DermatologyUniversity of Michigan Medical SchoolAnn ArborUSA
  3. 3.Dermatology Therapeutic AreaGlaxoSmithKlineCollegevilleUSA

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