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Permeability of Differentiated Human Urothelium In Vitro

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Permeability Barrier

Part of the book series: Methods in Molecular Biology ((MIMB,volume 763))

Abstract

The urothelium plays a critical role in the bladder as a permeability barrier to urine. Whereas it was once considered a simple physical barrier, it is increasingly evident that urothelium has a regulatory role in maintaining the barrier both through self-repair and by mediating the transport of ions and small molecules across the transcellular and paracellular interfaces. The development of cell culture systems that replicate the morphological and differentiated features of human urothelium provides a versatile in vitro tool for exploring molecular and functional relationships in normal bladder physiology and for examining inherent changes in the urothelia of patients with dysfunctional bladder syndromes. In addition, it provides a useful platform to study the effect of pharmacological treatment on urothelial barrier function. In this review, we describe the development of differentiated urothelial cell constructs from in vitro-propagated normal human urothelial cells, and the application of methods to assess barrier function using transepithelial electrical resistance, water, urea, and dextran transport as objective and quantifiable parameters.

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References

  1. Hu, P., Meyers, S., Liang, F. X., Deng, F. M., Kachar, B., Zeidel, M. L., and Sun, T. T. (2002) Role of membrane proteins in permeability barrier function: uroplakin ablation elevates urothelial permeability. Am J Physiol Renal Physiol 283, F1200–7.

    PubMed  CAS  Google Scholar 

  2. Olsburgh, J., Harnden, P., Weeks, R., Smith, B., Joyce, A., Hall, G., Poulsom, R., Selby, P., and Southgate, J. (2003) Uroplakin gene expression in normal human tissues and locally advanced bladder cancer. J Pathol 199, 41–9.

    Article  PubMed  Google Scholar 

  3. Wu, X. R., Manabe, M., Yu, J., and Sun, T. T. (1990) Large scale purification and immunolocalization of bovine uroplakins I, II, and III. Molecular markers of urothelial differentiation. J Biol Chem 265, 19170–9.

    Google Scholar 

  4. Varley, C. L., Garthwaite, M. A., Cross, W., Hinley, J., Trejdosiewicz, L. K., and Southgate, J. (2006) PPARgamma-regulated tight junction development during human urothelial cytodifferentiation. J Cell Physiol 208, 407–17.

    Article  PubMed  CAS  Google Scholar 

  5. Reznikoff, C. A., Johnson, M. D., Norback, D. H., and Bryan, G. T. (1983) Growth and characterization of normal human urothelium in vitro. In Vitro 19, 326–43.

    Article  PubMed  CAS  Google Scholar 

  6. Liebert, M., Wedemeyer, G., Chang, J. H., Stein, J. A., McKeever, P. E., Carey, T. E., Flint, A., Steplewski, Z., Buchsbaum, D. J., Wahl, R. L., and et al. (1990) Comparison of antigen expression on normal urothelial cells in tissue section and tissue culture. J Urol 144, 1288–92.

    PubMed  CAS  Google Scholar 

  7. Hutton, K. A., Trejdosiewicz, L. K., Thomas, D. F., and Southgate, J. (1993) Urothelial tissue culture for bladder reconstruction: an experimental study. J Urol 150, 721–5.

    PubMed  CAS  Google Scholar 

  8. Southgate, J., Hutton, K. A., Thomas, D. F., and Trejdosiewicz, L. K. (1994) Normal human urothelial cells in vitro: proliferation and induction of stratification. Lab Invest 71, 583–94.

    PubMed  CAS  Google Scholar 

  9. Cross, W.R., and Southgate, J. (2004) Biomi­metic Urothelium. Patent WO/2004/011630.

    Google Scholar 

  10. Cross, W. R., Eardley, I., Leese, H. J., and Southgate, J. (2005) A biomimetic tissue from cultured normal human urothelial cells: analysis of physiological function. Am J Physiol Renal Physiol 289, F459–68.

    Article  PubMed  CAS  Google Scholar 

  11. Turner, A. M., Subramaniam, R., Thomas, D. F., and Southgate, J. (2008) Generation of a functional, differentiated porcine urothelial tissue in vitro. Eur Urol 54, 1423–32.

    Article  PubMed  Google Scholar 

  12. Rubenwolf PC, Georgopoulos NT, Clements LA, Feather S, Holland P, Thomas DF, Southgate J. (2009) Expression and localisation of aquaporin water channels in human urothelium in situ and in vitro. Eur Urol. 56, 1013–1024.

    Article  PubMed  CAS  Google Scholar 

  13. Fraser, M., Thomas, D. F., Pitt, E., Harnden, P., Trejdosiewicz, L. K., and Southgate, J. (2004) A surgical model of composite cystoplasty with cultured urothelial cells: a controlled study of gross outcome and urothelial phenotype. BJU Int 93, 609–16.

    Article  PubMed  CAS  Google Scholar 

  14. Southgate, J., Masters, J. R., and Trejdosiewicz, L. K. (2002) in “Culture of Epithelial Cells.” (Freshney, R. I., and Freshney, M. G., Eds.), pp. 381–400, J Wiley and Sons, Inc., New York.

    Google Scholar 

  15. Lobban, E. D., Smith, B. A., Hall, G. D., Harnden, P., Roberts, P., Selby, P. J., Trejdosiewicz, L. K., and Southgate, J. (1998) Uroplakin gene expression by normal and neoplastic human urothelium. Am J Path 153, 1957–67.

    Article  PubMed  CAS  Google Scholar 

  16. Varley, C., Hill, G., Pellegrin, S., Shaw, N. J., Selby, P. J., Trejdosiewicz, L. K., and Southgate, J. (2005) Autocrine regulation of human urothelial cell proliferation and migration during regenerative responses in vitro. Exp Cell Res 306, 216–29.

    Article  PubMed  CAS  Google Scholar 

  17. Fromter, E., and Diamond, J. (1972) Route of passive ion permeation in epithelia. Nat New Biol 235, 9–13.

    Article  PubMed  CAS  Google Scholar 

  18. Negrete, H. O., Lavelle, J. P., Berg, J., Lewis, S. A., and Zeidel, M. L. (1996) Permeability properties of the intact mammalian bladder epithelium. Am J Physiol 271, F886–94.

    PubMed  CAS  Google Scholar 

  19. Lewis, S. A., and Diamond, J. M. (1976) Na+ transport by rabbit urinary bladder, a tight epithelium. J Membr Biol 28, 1–40.

    Article  PubMed  CAS  Google Scholar 

  20. Hicks, R. M. (1975) The mammalian urinary bladder: an accommodating organ. Biol Rev Camb Philos Soc 50, 215–46.

    Article  PubMed  CAS  Google Scholar 

  21. Spector, D. A., Wade, J. B., Dillow, R., Steplock, D. A., and Weinman, E. J. (2002) Expression, localization, and regulation of aquaporin-1 to -3 in rat urothelia. Am J Physiol Renal Physiol 282, F1034–42.

    PubMed  CAS  Google Scholar 

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Acknowledgements

We are grateful to Mr. William Cross, PhD, Consultant Urologist and Mr Nicholas Smith, PhD, Specialist Registrar in Urology at St. James’s University Hospital, Leeds, for their input into the TER and dextran permeability sections. Lisa Clements and Ros Unwin are thanked for careful reading of the manuscript.

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Correspondence to Jennifer Southgate .

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Rubenwolf, P., Southgate, J. (2011). Permeability of Differentiated Human Urothelium In Vitro. In: Turksen, K. (eds) Permeability Barrier. Methods in Molecular Biology, vol 763. Humana Press. https://doi.org/10.1007/978-1-61779-191-8_14

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  • DOI: https://doi.org/10.1007/978-1-61779-191-8_14

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-190-1

  • Online ISBN: 978-1-61779-191-8

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