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Fundamental Metabolic Differences between Hepatocytes and Islet β-cells Revealed by Glucokinase Overexpression

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Gifford Laboratories for Diabetes Research and Departments of Biochemistry and Internal Medicine, University of Texas Southwestern Medical Center, Dallas, Texas 75235
Cite this: Biochemistry 1998, 37, 13, 4543–4552
Publication Date (Web):March 13, 1998
https://doi.org/10.1021/bi9726133
Copyright © 1998 American Chemical Society

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    Abstract

    Adenovirus-mediated overexpression of the glucose phosphorylating enzyme glucokinase causes large changes in glycolytic flux and glucose storage in isolated rat hepatocytes, but not in pancreatic islets. We have used the well-differentiated insulinoma cell line INS-1 to investigate the basis for these apparent cell-type specific differences. We find that 2- or 5-[3H]glucose usage is increased at low (≤5 mM) but not high glucose concentrations in INS-1 cells treated with a recombinant adenovirus containing the glucokinase cDNA (AdCMV-GKI), while glucose usage is increased at both low and high glucose concentrations in similarly treated hepatocytes. Utilization of 2-[3H]glucose in INS-1 cells is suppressed in glucokinase overexpressing INS-1 cells in a rapid, glucose concentration-dependent, and reversible fashion, 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 following the switch to high glucose in either AdCMV-GKI-treated INS-1 cells or hepatocytes relative to controls. In contrast, triose phosphate levels (glyceraldehyde-3-phosphate + dihydroxyacetone phosphate) were much higher in AdCMV-GKI-treated INS-1 cells than in similarly treated hepatocytes, suggesting limited flux throught the glyceraldehyde-3-phosphate dehydrogenase (G3PDH) step in the former cells. Hepatocytes were found to contain approximately 62 times more lactate dehydrogenase (LDH) activity than INS-1 cells, and this was reflected in a 3-fold increase in lactate production in AdCMV-GKI-treated hepatocytes relative to similarly treated INS-1 cells. Since the amounts of G3PDH activity in INS-1 and hepatocyte extracts are similar, we suggest that flux through this step in INS-1 cells is limited by failure to regenerate NAD in the LDH reaction and that a fundamental difference between hepatocytes and islet β-cells is the limited capacity of the latter to metabolize glycolytic intermediates beyond the G3PDH step.

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     This was supported by a National Institutes of Health/Juvenile Diabetes Foundation International Interdisciplinary Research Program and National Institutes of Health Grant RO1 DK 46492 (to C.B.N.).

    *

     Author to whom correspondence should be addressed at Gifford Laboratories for Diabetes Research, Room Y8.212, University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75235. Telephone:  (214) 648-2930. Fax:  (214) 648-9191. E-mail:  [email protected].

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    18. Gary D Lopaschuk. Glycolysis Regulation. 2001https://doi.org/10.1038/npg.els.0000617
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