Analysis of ciliogenesis in primary culture mouse tracheal epithelial cells

Eszter K. Vladar, Steven L. Brody

Research output: Chapter in Book/Report/Conference proceedingChapter

26 Scopus citations

Abstract

Cell biological and molecular characterization of structural and functional ciliary components and regulators of mammalian motile ciliogenesis is made possible by the development of a robust and biologically faithful mouse tracheal epithelial cell (MTEC) culture system and complementary research techniques. Here, we describe the air-liquid interface culture of mouse airway epithelial progenitor cells that undergo motile ciliogenesis de novo. Multiciliated cells differentiate rapidly, and distinct stages of the ciliogenesis pathway can be identified and characterized with centriolar and ciliary immunofluorescence markers. Immunolabeled structures correlate with morphological features previously identified by electron microscopy, facilitating light microscopy analysis. MTEC cultures can be successfully transduced by lentiviral RNAi or epitope-tagged cDNA constructs to perturb gene expression. Also, motile ciliogenesis can be manipulated by drug treatment. Distinct cell populations can be isolated by cell sorting to facilitate comparison among the multiciliated and other cell types in the in vitro differentiated epithelium. The MTEC system uniquely offers the study of ciliogenesis in cells from genetically modified mouse strains.

Original languageEnglish
Title of host publicationCilia, Part B
PublisherAcademic Press Inc.
Pages285-309
Number of pages25
ISBN (Print)9780123979445
DOIs
StatePublished - 2013

Publication series

NameMethods in Enzymology
Volume525
ISSN (Print)0076-6879
ISSN (Electronic)1557-7988

Keywords

  • Air-liquid interface
  • Ciliogenesis
  • Electron microscopy
  • Fluorescence-activated cell sorting
  • Immunofluorescence
  • Lentiviral gene transfer
  • Motile cilia
  • Mouse tracheal epithelial cell (MTEC)
  • Primary culture

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  • Cite this

    Vladar, E. K., & Brody, S. L. (2013). Analysis of ciliogenesis in primary culture mouse tracheal epithelial cells. In Cilia, Part B (pp. 285-309). (Methods in Enzymology; Vol. 525). Academic Press Inc.. https://doi.org/10.1016/B978-0-12-397944-5.00014-6