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Formation of multicellular epithelial structures.

Keith Mostov1, Paul Brakeman, Anirban Datta

  • 1Department of Anatomy, University of California, San Francisco 94143-2140, USA.

Novartis Foundation Symposium
|December 17, 2005
PubMed
Summary
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Researchers found that altering Rac1 in kidney cells grown in 3D gels inverts epithelial polarity. This suggests polarity establishment and orientation can be separated, with laminin assembly playing a key role.

Area of Science:

  • Cell Biology
  • Renal Physiology
  • Epithelial Biology

Background:

  • Epithelial cells, crucial in kidney function, exhibit polarization.
  • External cues typically dictate epithelial cell polarity orientation in experimental models.
  • Three-dimensional (3D) culture systems offer alternative models to study intrinsic polarity mechanisms.

Purpose of the Study:

  • To investigate the role of small GTPase Rac1 in regulating epithelial polarity orientation.
  • To determine if polarity establishment and orientation can be experimentally dissociated.
  • To elucidate the role of the basement membrane, specifically laminin, in polarity orientation.

Main Methods:

  • Utilized Madin-Darby canine kidney (MDCK) cells cultured in 3D collagen gels to form cysts.

Related Experiment Videos

  • Expressed a dominant-negative (DN) form of Rac1 to disrupt its function.
  • Assessed epithelial polarity by observing apical surface orientation within the cysts.
  • Analyzed basement membrane assembly and laminin distribution.
  • Investigated the effect of exogenous laminin on polarity defects.

    • Main Results:

    • DN Rac1 expression inverted epithelial polarity in 3D cysts, with apical surfaces facing outwards.
    • This inversion demonstrated a separation between polarity establishment and orientation.
    • DN Rac1 disrupted laminin assembly into the basement membrane network.
    • Supplementation with exogenous laminin rescued the polarity inversion phenotype.

    Conclusions:

    • Rac1 is critical for maintaining the correct orientation of epithelial polarity in 3D cultures.
    • The basement membrane, particularly laminin organization, is essential for proper polarity orientation.
    • 3D culture systems provide a valuable tool to decouple polarity establishment from orientation cues.