IGF-1 production by primary cultured hepatocytes from rats and sheep
Methods were established for the serum-free primary culture of rat and sheep hepatocytes. These were used to study the IGF-1 production by the cells. IGF-1 in media was separated by acid gel filtration or HPLC prior to RIA. IGF-1 production by both rat and sheep hepatocytes were time-dependent over 30 hours of culture. Production rates by rat cells in modified Eagle's medium, sheep cells modified Eagle's medium and Waymouth's medium were 13.1, 4.4 and 6.4 pmol/mg cell DNA/10 hours respectively. IGF-1 production by rat cells was very sensitive to altering concentrations of amino acids, glucose and both in the medium. IGF-1 production by sheep cells was also controlled by nutrients in modified Eagle's medium, but was not as sensitive as in rat cells. Growth hormone (GH), unlike insulin and T3, had no effect on hepatic IGF-1 production in rat cells. In contrast, omitting GH from the medium for sheep cells decreased IGF-1 production by 20-60% in sheep cells. Insulin also controlled hepatic IGF-1 production in sheep. The results show the species difference in the hepatice IGF-1 production and control between rats and sheep. Data in this thesis are thought to provide the first published evidence of a direct nutritional control over IGF-1 production in rat hepatocytes and to give the first direct evidence that sheep hepatocytes produce IGF-1 and this is directly controlled by GH.