In Vitro Cellular & Developmental Biology

, Volume 25, Issue 2, pp 174–182

Analysis of plasma protein and lipoprotein synthesis in long-term primary cultures of baboon hepatocytes maintained in serum-free medium

  • Robert E. Lanford
  • Kenneth D. Carey
  • Larry E. Estlack
  • G. Con Smith
  • Rick V. Hay
Regular Papers

DOI: 10.1007/BF02626175

Cite this article as:
Lanford, R.E., Carey, K.D., Estlack, L.E. et al. In Vitro Cell Dev Biol (1989) 25: 174. doi:10.1007/BF02626175

Summary

The analysis of lipoprotein synthesis and secretion in primary hepatocytes has been restricted by the short-term viability and low proliferative response of hepatocytes in vitro. During this investigation a serum-free medium formulation was developed that supports long-term maintenance (>70 d) and active proliferation of primary baboon hepatocytes. Examination of proliferating cells by electron microscopy revealed a distinctive hepatocyte ultrastructure including intercellular bile canaliculi and numerous surface microvilli. High levels of secreted apolipoproteins A-I and E were detected in the tissue culture medium by gel electrophoresis and immunoblot analysis. Immunoprecipitation of proteins from [35S]-methionine labeled tissue culture medium revealed the synthesis and secretion of numerous plasma proteins. Metabolic labeling of cells with [35S]-methionine followed by single-spin density gradient flotation of the media demonstrated that apolipoproteins were being secreted in the form of lipoprotein particles with buoyant densities corresponding to the very low density lipoprotein and low density lipoprotein range, and to the high density lipoprotein range. The labeled apolipoproteins included Bh, E, and A-I. This system for primary hepatocyte culture should prove very useful in future investigations on the regulation of lipoprotein production by hepatocytes.

Key words

hepatocyteslipoprotein synthesisserum-free medium

Copyright information

© Tissue Culture Association, Inc 1989

Authors and Affiliations

  • Robert E. Lanford
    • 1
  • Kenneth D. Carey
    • 1
  • Larry E. Estlack
    • 1
  • G. Con Smith
    • 1
  • Rick V. Hay
    • 2
  1. 1.Department of Virology and ImmunologySouthwest Foundation for Biomedical ResearchSan Antonio
  2. 2.Department of PathologyThe University of ChicagoChicago