Characterization of apical and basolateral plasma membrane domains derived from cultured rat cholangiocytes

Pamela Tietz, Susan Levine, Ralph Holman, Chris Fretham, Nicholas F La Russo

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Cholangiocytes, the epithelial cells that line intrahepatic bile ducts, are composed of plasma membranes with discrete apical (lumenal) and basolateral domains that contain different channels, transporters, and receptors. In recent work, we developed a long-term, primary culture system of normal rat cholangiocytes (NRC). Our aims here were to prepare and characterize apical and basolateral plasma membrane vesicles from NRC. Using serial isopycnic centrifugation on sucrose gradients, we generated separate apical and basolateral plasma membrane vesicles. We characterized these vesicles by transmission electron microscopy, specific marker enzyme assays, and immunoblotting; we also determined the percentage of sealed vesicles and their intravesicular volume and sidedness using biochemical approaches. Our results showed that vesicles derived from the apical and basolateral plasma membrane domains of NRC were highly purified, predominately sealed, right- side-out vesicles that differed in size and lipid and protein composition. This experimental model represents a novel tool that will be useful for additional functional studies on the channels, transporters, and receptors differentially distributed in the plasma membrane of biliary epithelia.

Original languageEnglish (US)
Pages (from-to)192-199
Number of pages8
JournalAnalytical Biochemistry
Volume254
Issue number2
DOIs
StatePublished - Dec 15 1997

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Cell membranes
Rats
Cell Membrane
Isopycnic Centrifugation
Intrahepatic Bile Ducts
Centrifugation
Enzyme Assays
Transmission Electron Microscopy
Immunoblotting
Ducts
Sucrose
Assays
Theoretical Models
Epithelium
Epithelial Cells
Transmission electron microscopy
Lipids
Cell Line
Enzymes
Chemical analysis

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Characterization of apical and basolateral plasma membrane domains derived from cultured rat cholangiocytes. / Tietz, Pamela; Levine, Susan; Holman, Ralph; Fretham, Chris; La Russo, Nicholas F.

In: Analytical Biochemistry, Vol. 254, No. 2, 15.12.1997, p. 192-199.

Research output: Contribution to journalArticle

Tietz, Pamela ; Levine, Susan ; Holman, Ralph ; Fretham, Chris ; La Russo, Nicholas F. / Characterization of apical and basolateral plasma membrane domains derived from cultured rat cholangiocytes. In: Analytical Biochemistry. 1997 ; Vol. 254, No. 2. pp. 192-199.
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