FUNDAMENTAL METABOLIC DIFFERENCES BETWEEN HEPATOCYTES AND ISLET BETA-CELLS REVEALED BY GLUCOKINASE OVEREXPRESSION

Adenovirus-mediated overexpression of the glucose phosphorylating enzy me glucokinase causes large changes in glycolytic flux and glucose sto rage in isolated rat hepatocytes, but not in pancreatic islets. We hav e used the well-differentiated insulinoma cell line INS-1 to investiga te the basis for these apparent cell-type specific differences. We fin d that 2- or 5-[H-3]glucose usage is increased at low (less than or eq ual to 5 mM) but not high glucose concentrations in INS-1 cells treate d with a recombinant adenovirus containing the glucokinase cDNA (AdCMV -GKI), while glucose usage is increased at both low and high glucose c oncentrations in similarly treated hepatocytes. Utilization of 2-[H-3] glucose in INS-1 cells is suppressed in glucokinase overexpressing INS -1 cells in a rapid, glucose concentration-dependent, and reversible f ashion, while such regulation is largely absent in hepatocytes. Levels of hexose phosphates (glucose-6-phosphate, fructose-6-phosphate, and fructose-1,6-bisphosphate) were profoundly and rapidly elevated follow ing the switch to high glucose in either AdCMV-GKI-treated INS-1 cells or hepatocytes relative to controls. In contrast, triose phosphate le vels (glyceraldehyde-3-phosphate + dihydroxyacetone phosphate) were mu ch higher in AdCMV-GKI-treated INS-1 cells than in similarly treated h epatocytes, suggesting limited flux throught the glyceraldehyde-3-phos phate dehydrogenase (G3PDH) step in the former cells. Hepatocytes were found to contain approximately 62 times more lactate dehydrogenase (L DH) activity than INS-1 cells, and this was reflected in a 3-fold incr ease in lactate production in AdCMV-GKI-treated hepatocytes relative t o similarly treated INS-1 cells. Since the amounts of G3PDH activity i n INS-1 and hepatocyte extracts are similar, we suggest that flux thro ugh this step in INS-1 cells is limited by failure to regenerate NAD i n the LDH reaction and that a fundamental difference between hepatocyt es and islet beta-cells is the limited capacity of the latter to metab olize glycolytic intermediates beyond the G3PDH step.