title: Insulin secretion, insulin content and glucose phosphorylation in RINm5F insulinoma cells after transfection with human GLUT2 glucose-transporter cDNA.
abstract: The insulin-secretory response to glucose is defective in the RINm5F insulin-producing tumour cell line. Stable transfection with human low-affinity GLUT2 glucose-transporter cDNA revealed a significant improvement in stimulus-secretion coupling in these insulinoma cells. 3-O-Methylglucose uptake increased 10-fold in the concentration range 10-20 mM, whereas non-transfected control cells were unresponsive. Northern-blot analysis revealed a 7-fold increase in expression of the insulin gene in the GLUT2-transfected RINm5F cell clone T1. In contrast, glucokinase and GLUT1 glucose-transporter mRNA gene expression were not affected by transfection with GLUT2 glucose-transporter cDNA. The insulin content of transfected RINm5F cells was 7-fold higher after tissue culture at high glucose concentrations than in non-transfected controls. GLUT2-transfected RINm5F cells also regained insulin-secretory responsiveness toward high glucose concentrations. Tissue culture for 72 h in 20 mM glucose induced glucokinase activity in the GLUT2-transfected RINm5F clone T1, raising the glucokinase/hexokinase phosphorylation ratio from 0.2 to 0.6. The experiments demonstrate that an increased glucose uptake via a low-affinity glucose transporter and an increased metabolic flux rate are important factors in the induction of insulin-gene expression and glucokinase activity and thus improved glucose-induced biosynthesis and secretion of insulin in RINm5F insulinoma cells.
authors.affiliation: Institute of Pharmacology and Toxicology, University of Göttingen, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Tiedge
authors.forename: M
authors.initials: M
authors.lastname: Höhne
authors.forename: S
authors.initials: S
authors.lastname: Lenzen
mesh_terms: D019325:3-O-Methylglucose; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002460:Cell Line; D018076:DNA, Complementary / Q000378:metabolism*; D005941:Glucokinase / Q000378:metabolism; D005947:Glucose / Q000378:metabolism*; D051273:Glucose Transporter Type 2; D006593:Hexokinase / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000378:metabolism / Q000502:physiology*; D000078790:Insulin Secretion; D007340:Insulinoma; D007700:Kinetics; D008757:Methylglucosides / Q000378:metabolism*; D009004:Monosaccharide Transport Proteins / Q000096:biosynthesis / Q000378:metabolism*; D010190:Pancreatic Neoplasms; D012333:RNA, Messenger / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D018076:DNA, Complementary; D051273:Glucose Transporter Type 2; D007328:Insulin; D008757:Methylglucosides; D009004:Monosaccharide Transport Proteins; D012333:RNA, Messenger; D019325:3-O-Methylglucose; D006593:Hexokinase; D005941:Glucokinase; D005947:Glucose