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Pharmaceutical Research

, Volume 15, Issue 10, pp 1533–1539 | Cite as

Partial Maintenance of Taurocholate Uptake by Adult Rat Hepatocytes Cultured in a Collagen Sandwich Configuration

  • Xingrong Liu
  • Kim L. R. Brouwer
  • Liang-Sheng L. Gan
  • Kenneth R. Brouwer
  • Bruno Stieger
  • Peter J. Meier
  • Kenneth L. Audus
  • Edward L. LeCluyse
Article

Abstract

Purpose. This study was designed to characterize taurocholate uptake properties in primary cultures of rat hepatocytes maintained under different matrix conditions.

Methods. Hepatocytes isolated from male Wistar rats (230−280 g) were cultured on a simple collagen film, on a substratum of gelled collagen or between two layers of gelled collagen (sandwich configuration). Hepatocyte morphology, taurocholate uptake properties, and expression of the sinusoidal transport protein, Na+/taurocholate-cotransporting polypeptide (Ntcp) were examined in these cultures at day 0 and day 5.

Results. By day 5, monolayer integrity had deteriorated in simple collagen cultures. In contrast, cell morphology was preserved in hepatocytes maintained in a sandwich configuration. At day 5, taurocholate accumulation at 5 min in hepatocytes cultured on a simple collagen film, on a substratum of gelled collagen, and in a sandwich configuration was ∼13%, 20% and 35% of day-0 levels, respectively, and occurred predominately by a Na+-dependent mechanism. The initial taurocholate uptake rate vs. concentration (1-200 μM) profile was best described by a combined Michaelis-Menten and first-order function. In all cases, the estimated apparent Km values were comparable for day-0 and day-5 hepatocytes (32−41 μM). In contrast, the Vmax values of hepatocytes cultured on a simple collagen film, on gelled collagen and in a sandwich configuration were ∼5, 6 and 14% of the values at day 0, respectively; values for the first-order rate constant were 5-, 3- and 2-fold lower, respectively. Immunoblot analysis indicated that at day 5 Ntcp expression in hepatocytes cultured in a sandwich configuration was greater than in hepatocytes cultured on a simple collagen film.

Conclusions. A collagen sandwich configuration reestablishes normal morphology and partially restores bile acid uptake properties in primary cultures of rat hepatocytes.

taurocholate transport Na+/taurocholate-cotransporting polypeptide (Ntcp) hepatocyte culture collagen-sandwich configuration 

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Copyright information

© Plenum Publishing Corporation 1998

Authors and Affiliations

  • Xingrong Liu
    • 1
  • Kim L. R. Brouwer
    • 1
  • Liang-Sheng L. Gan
    • 2
  • Kenneth R. Brouwer
    • 2
  • Bruno Stieger
    • 3
  • Peter J. Meier
    • 3
  • Kenneth L. Audus
    • 4
  • Edward L. LeCluyse
    • 1
  1. 1.Division of Drug Delivery and Disposition, School of PharmacyUniversity of North Carolina at Chapel HillChapel Hill
  2. 2.Division of Bioanalysis and Drug MetabolismGlaxo Wellcome, Inc.
  3. 3.Division of Clinical Pharmacology, Department of MedicineUniversity HospitalZurichSwitzerland
  4. 4.Department of Pharmaceutical ChemistryUniversity of KansasLawrence

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