Abstract
In order to elucidate the role of polyamines in the replication and insulin production of insulin-secreting cells, we have investigated the impact of partial polyamine depletion on the proliferation, metabolism, insulin synthesis and ultrastructure of clonal rat insulinoma cells (RINm5F). For this purpose RINm5F cells were exposed for 4 days to the specific ornithine decarboxylase (ODC) inhibitor difluoromethylornithine (DFMO). This resulted in a profound decrease in ODC activity and cytoplasmic polyamine contents. The polyamine content of cell nuclei was, however, not altered by DFMO. Addition of small amounts of putrescine during culture elevated the intracellular content of this diamine and suppressed ODC activity. The decrease in polyamine contents was accompanied by a pronounced inhibition of the cellular proliferative activity. The rates of glucose utilization, oxygen uptake and activity of the pentose cycle were decreased in DFMO-treated cells, whereas the glucose oxidation rate, oxidation/utilization ratio, ATP content and ATP/ADP ratio were increased. Insulin mRNA content and synthesis of proinsulin, insulin and total protein were not altered by DFMO. In contrast, there was a sizeable increase in the cellular insulin content, despite a lowered total protein content. Electron-microscopic analysis revealed an accumulation of insulin-secretion granules in the DFMO-treated cells. In addition, short-term insulin release was increased after DFMO exposure, but was not rendered glucose-sensitive. It is concluded that polyamines are necessary for the maintenance of rapid insulinoma-cell replication and that DFMO-treated RINm5F cells acquire an enhanced substrate oxidation and increased content of insulin and ATP.