Skip to main content

A Quantifiable In Vitro Model to Assess Effects of PAI-1 Gene Targeting on Epithelial Cell Motility

  • Protocol
Book cover Wound Healing

Part of the book series: Methods in Molecular Medicine™ ((MIMM,volume 78))

  • 2092 Accesses

Abstract

Gene “knockout” studies and analysis of physiologically relevant in vitro models have clarified basic mechanisms in the tissue response to injury (1-5). Fundamental to this process is the genetic reprogramming required for conversion of normally sedentary cells to an actively motile, invasive phenotype (5,6). It is difficult, however, to identify important events among the diverse changes associated with the initiation and maintenance of migration or the acquisition of “plasticity” in a specific cell type in the intact animal. Thus, the development of in vitro approaches that closely mimic defined stages in epidermal wound repair facilitates discovery of the critical elements and provides an opportunity to probe the underlying molecular mechanisms.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Garlick, J. A. and Taichman, L. B. (1994) Fate of human keratinocytes during reepithelialization in an organotypic culture model. Lab. Invest. 70, 916–924.

    PubMed  CAS  Google Scholar 

  2. Romer, J., Lund, L. R., Kriksen, J., Pyke, C., Kristensen, P., and Dano, K. (1996) Impaired wound healing in mice with a disrupted plasminogen gene. Nat. Med. 2, 287–292.

    Article  PubMed  CAS  Google Scholar 

  3. Providence, K. M., Kutz, S. M., Staiano-Coico, L., and Higgins, P. J. (2000) PAI-1 gene expression is regionally induced in wounded epithelial cell monolayers and required for injury repair. J. Cell. Physiol. 182, 269–280.

    Article  PubMed  CAS  Google Scholar 

  4. Coulombe, P. A. (1997) Towards a molecular definition of keratinocyte activation after acute injury to stratified epithelia. Biochem. Biophys. Res. Commun. 236, 231–238.

    Article  PubMed  CAS  Google Scholar 

  5. Sato, Y. and Rifkin, D. B. (1988) Autocrine activities of basic fibroblast growth factor: regulation of endothelial cell movement, plasminogen activator synthesis, and DNA synthesis. J. Cell Biol. 107, 1199–1205.

    Article  PubMed  CAS  Google Scholar 

  6. Ando, Y. and Jensen, P.J. (1996) Protein kinase C mediates up-regulation of urokinase and its receptor in migrating keratinocytes of wounded cultures, but urokinase is not required for movement across a substratum in vitro. J. Cell. Physiol. 167, 500–511.

    Article  PubMed  CAS  Google Scholar 

  7. Reidy, M., Irwin, C., and Lindner, V. (1995) Migration of arterial wall cells. Circ. Res. 78, 405–414.

    Article  Google Scholar 

  8. Zahm, J. M., Kaplan, H., Herard, A. L., Doriot, F., Pierrot, D., Somelette, P., and Puchelle, E. (1997) Cell migration and proliferation during the in vitro wound repair of the respiratory epithelium. Cell Motil. Cytoskel. 37, 33–43.

    Article  CAS  Google Scholar 

  9. Andreasen, P. A., Kjoller, L., Christensen, L., and Duffy, M. J. (1997) The urokinase-type plasminogen activator system in cancer metastasis: a review. Int. J. Cancer 72, 1–22.

    Article  PubMed  CAS  Google Scholar 

  10. Blasi, F. (1997) uPA, uPAR, PAI-1: key intersection of proteolytic, adhesive and chemotactic highways? Immunol. Today 18, 415–417.

    Article  PubMed  CAS  Google Scholar 

  11. Loskutoff, D. J., Curriden, S. A., Hu, G., and Deng, G. (1999) Regulation of cell adhesion by PAI-1. APMIS 107, 54–61.

    Article  PubMed  CAS  Google Scholar 

  12. Stefansson, S. and Lawrence, D. A. (1996) The serpin PAI-1 inhibits cell migration by blocking integrin αvβ5 binding to vitronectin. Nature 383, 441–443.

    Article  PubMed  CAS  Google Scholar 

  13. Bajou, K., Noel, A., Gerard, R. D., Masson, V., Brunner, N., Holst-Hansen, C., Skobe, M., Fusenig, N. E., Carmeliet, P., Collen, D., and Foidart, J. M. (1998) Absence of host plasminogen activator inhibitor 1 prevents cancer invasion and vascularization. Nat. Med. 4, 923–938.

    Article  PubMed  CAS  Google Scholar 

  14. Romer, J., Lund, L. R., Eriksen, J., Ralfkiaer, E., Zeheb, R., Gelehrter, T. D., Dano, K., and Kristensen, P. (1991) Differential expression of urokinase-type plasminogen activator and its type-1 inhibitor during healing of mouse skin wounds. J. Invest. Dermatol. 97, 803–811.

    Article  PubMed  CAS  Google Scholar 

  15. Boehm, J. R., Kutz, S. M., Sage, E. H., Staiano-Coico, L., and Higgins, P. J. (1999) Growth state-dependent regulation of plasminogen activator inhibitor type-1 gene expression during epithelial cell stimulation by serum and transforming growth factor-β1. J. Cell. Physiol. 181, 96–106.

    Article  PubMed  CAS  Google Scholar 

  16. Higgins, P. J., Ryan, M. P., and Jelley, D. M. (1997) p52PAI-1 gene expression in butyrate-induced flat revertants of v-ras-transformed rat kidney cells: mechanism of induction and involvement in the morphologic response. Biochem. J. 321, 431–437.

    PubMed  CAS  Google Scholar 

  17. Li, F., Goncalves, J., Faughnan, K., Steiner, M. G., Pagan-Charry, I., Esposito, D., Chin, B., Providence, K. M., Higgins, P. J., and Staiano-Coico, L. (2000) Targeted inhibition of wound-induced PAI-1 expression alters migration and differentiation in human epidermal keratinocytes. Exp. Cell Res. 258, 245–253.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2003 Humana Press Inc., Totowa, NJ

About this protocol

Cite this protocol

Providence, K.M., Staiano-Coico, L., Higgins, P.J. (2003). A Quantifiable In Vitro Model to Assess Effects of PAI-1 Gene Targeting on Epithelial Cell Motility. In: DiPietro, L.A., Burns, A.L. (eds) Wound Healing. Methods in Molecular Medicine™, vol 78. Humana Press, Totowa, NJ. https://doi.org/10.1385/1-59259-332-1:293

Download citation

  • DOI: https://doi.org/10.1385/1-59259-332-1:293

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-0-89603-999-5

  • Online ISBN: 978-1-59259-332-3

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics