Regulation of Insulin and Insulin-Like Growth Factor Responsiveness.

The primary objective of my thesis research was to investigate the regulation of hepatoma cell responsiveness to insulin and insulin-like growth factors (IGFs). IGFs stimulate many of the same biological actions as insulin, and although the IGFs bind weakly to the insulin receptor, two distinct types of IGF receptors have been defined. I have examined the insulin and IGF-II stimulation of tyrosine aminotransferase activity, glycogen synthase activity and amino acid transport in H-35 rat hepatoma cells to determine which receptor(s) mediate these metabolic responses. On the basis of both affinity cross-linking and competition binding studies, H-35 cells display insulin and type II IGF receptors, but not type I IGF receptors. IGF-II stimulates tyrosine aminotransferase activity, amino acid transport and glycogen synthase activity to the same magnitude as insulin; however, this stimulation occurs only at high concentrations of IGF-II, indicating action via the insulin receptor. Incubation of H-35 cells with insulin induces a state of unresponsiveness to the further actions of both IGF-II and insulin. In contrast, insulin-treated cells are fully responsive to 0.1 (mu)M dexamethasone, indicating that the loss of responsiveness is specific for insulin and IGFs. Incubation of H-35 cells with IGF-II also induces a state of unresponsiveness to the actions of both insulin and IGF-II. The concentration of insulin necessary for desensitization indicates that insulin-induced desensitization occurs via the insulin receptor; however, the high concentration of IGF-II necessary to induce desensitization is consistent with IGF-II action through the insulin receptor. Insulin- and IGF-II-induced desensitization is accompanied by little or no loss of insulin or IGF-II binding, indicating that desensitization is occurring primarily at a post-binding level. Using affinity cross-linking techniques, I have demonstrated the presence of type I and type II IGF receptors in Madin-Darby canine kidney (MDCK) cells, which lack insulin receptors and yet retain insulin-like actions. The IGF receptors were further characterized by competition binding studies and found to be similar to IGF receptors in other tissue types. MDCK cells may provide an opportunity to study IGF actions in the absence of an insulin receptor.Ph.D.GeneticsUniversity of Michiganhttp://deepblue.lib.umich.edu/bitstream/2027.42/160981/1/8612560.pd