title: Characterization of hepatocyte-derived mitogenic activity on hepatic stellate cells.
abstract: OBJECTIVE
  Hepatic stellate cells (HSC) are located in close proximity to hepatocytes in Disse's space. Hepatocyte derived factors have earlier been implicated in the paracrine regulation of HSC proliferation. The aim of the present study was to further characterize this mitogenic activity of the parenchymal cell conditioned medium (PCcM).
  
  
  METHODS
  Primary rat HSC were cultured for 4 days. DNA synthesis was measured by 3H-thymidine incorporation. TGFbeta1 immunoreactivity was quantified by ELISA. PCcM was obtained from hepatocytes cultured in medium without serum or hormones for two days.
  
  
  RESULTS
  Incubation of 4-day-old HSC on plastic surface with PCcM for 2 days increased DNA synthesis, while no effect was seen in HSC cultured on Matrigel. Heat-, acid-, and protease-treatment of PCcM abolished its stimulatory effect. Size fractionations with spin columns indicated that the stimulatory effect was contained in the fractions of a molecular size between 30 and 100 kD. The addition of LY 294002, a phosphatidylinositol 3-kinase (PI3-K) inhibitor, dose-dependently inhibited the PCcM induced increase in DNA synthesis to about 9% of the control values. The specific MAP kinase (MAPK) inhibitor, PD 98059 only suppressed the PCcM induced DNA synthesis to 35% of control cultures at the highest dose (10 microM). DNA content in the cultures was not affected by either blocker. HSC seemed to produce immunoreactive TGFbeta1. However, addition of latency-associated peptide (LAP), a potent TGFbeta1 blocker, stimulated DNA synthesis to a much less extent than PCcM.
  
  
  CONCLUSIONS
  The factor(s) that stimulate DNA synthesis in HSC from hepatocytes are most likely protein(s) with a molecular size between 30-100 kD. These factor(s) rely more on PI3-K than on MAPK for their mitogenic effect and are probably not acting via TGFbeta1 inhibition.
authors.affiliation: Research Center for Endocrinology and Metabolism, Sahlgrenska University Hospital, Göteborg, Sweden.
authors.forename: S
authors.initials: S
authors.lastname: Skrtic
authors.forename: K
authors.initials: K
authors.lastname: Wallenius
authors.forename: A M
authors.initials: AM
authors.lastname: Gressner
authors.forename: J O
authors.initials: JO
authors.lastname: Jansson
mesh_terms: D000818:Animals; D002417:Cattle; D002469:Cell Separation; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D017077:Culture Media, Conditioned / Q000378:metabolism / Q000494:pharmacology; D004247:DNA / Q000096:biosynthesis; D004261:DNA Replication / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005312:Fetal Blood / Q000502:physiology; D005419:Flavonoids / Q000494:pharmacology; D006358:Hot Temperature; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D008099:Liver / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008934:Mitogens / Q000737:chemistry; D008938:Mitosis / Q000187:drug effects; D008970:Molecular Weight; D009025:Morpholines / Q000494:pharmacology; D010446:Peptide Fragments; D011498:Protein Precursors; D011506:Proteins / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta / Q000378:metabolism / Q000494:pharmacology; D053773:Transforming Growth Factor beta1
chemical_list: D002867:Chromones; D017077:Culture Media, Conditioned; D004791:Enzyme Inhibitors; D005419:Flavonoids; D008934:Mitogens; D009025:Morpholines; D010446:Peptide Fragments; D011498:Protein Precursors; D011506:Proteins; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D007334:Insulin-Like Growth Factor I; D004247:DNA; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one