title: Growth factors and nonparenchymal cell conditioned media induce mitogenic responses in stable long-term adult rat hepatocyte cultures.
abstract: Most prior studies have characterized hepatocyte proliferative responses in culture systems that do not express a stable differentiated phenotype. We investigated the DNA synthetic response of long-term stable hepatocyte cultures to growth factor stimulation as well as conditioning with nonparenchymal cells (NPCs). Primary rat hepatocytes were cultured on a single layer of collagen (h/C) or Matrigel (h/M), or in a collagen sandwich (C/h/C) or collagen-Matrigel sandwich (M/h/C). Hepatocytes were cultured for 7 days to allow phenotypic stabilization before growth factor addition, except for h/C cultures, which are unstable, where growth factors were added 1 day after seeding. Culture medium was supplemented with a mixture of hepatocyte, epidermal, and vascular endothelial growth factors and interleukin-6, either directly or after conditioning with NPCs for 24 h. Growth factors alone induced hepatocyte DNA synthesis, as measured via [3H]thymidine uptake, in the h/C, C/h/C, and M/h/C configurations. h/M exhibited very low levels of DNA synthesis. In the C/h/C and M/h/C configurations, the greatest stimulation was obtained using NPC-conditioned growth factors. This response was sustained for several days and without decreasing albumin or urea synthesis. These results suggest that hepatocyte mitogens and NPC-derived factors can stimulate DNA synthesis in stable and differentiated hepatocyte cultures.
authors.affiliation: Center for Engineering in Medicine/Surgical Services, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.
authors.forename: Yoon H
authors.initials: YH
authors.lastname: Kang
authors.forename: François
authors.initials: F
authors.lastname: Berthiaume
authors.forename: Bharath D
authors.initials: BD
authors.lastname: Nath
authors.forename: Martin L
authors.initials: ML
authors.lastname: Yarmush
mesh_terms: D000418:Albumins / Q000378:metabolism; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002478:Cells, Cultured; D017077:Culture Media, Conditioned / Q000494:pharmacology*; D004247:DNA / Q000096:biosynthesis / Q000187:drug effects*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D006133:Growth Substances / Q000494:pharmacology*; D017228:Hepatocyte Growth Factor / Q000494:pharmacology; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015850:Interleukin-6 / Q000494:pharmacology; D008934:Mitogens / Q000494:pharmacology*; D008938:Mitosis / Q000187:drug effects* / Q000502:physiology; D051381:Rats; D011917:Rats, Inbred Lew; D014508:Urea / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
chemical_list: D000418:Albumins; D017077:Culture Media, Conditioned; D006133:Growth Substances; D015850:Interleukin-6; D008934:Mitogens; D042461:Vascular Endothelial Growth Factor A; D004815:Epidermal Growth Factor; D017228:Hepatocyte Growth Factor; D014508:Urea; D004247:DNA