title: Switch to anaerobic glucose metabolism with NADH accumulation in the beta-cell model of mitochondrial diabetes. Characteristics of betaHC9 cells deficient in mitochondrial DNA transcription.
abstract: To elucidate the mechanism underlying diabetes caused by mitochondrial gene mutations, we created a model by applying 0.4 microg/ml ethidium bromide (EtBr) to the murine pancreatic beta cell line betaHC9; in this model, transcription of mitochondrial DNA, but not that of nuclear DNA, was suppressed in association with impairment of glucose-stimulated insulin release (Hayakawa, T., Noda, M., Yasuda, K., Yorifuji, H., Taniguchi, S., Miwa, I., Sakura, H., Terauchi, Y., Hayashi, J.-I., Sharp, G. W. G., Kanazawa, Y., Akanuma, Y., Yazaki, Y., and Kadowaki, T. (1998) J. Biol. Chem. 273, 20300-20307). To elucidate fully the metabolism-secretion coupling in these cells, we measured glucose oxidation, utilization, and lactate production. We also evaluated NADH autofluorescence in betaHC9 cells using two-photon excitation laser microscopy. In addition, we recorded the membrane potential and determined the ATP and ADP contents of the cells. The results indicated 22.2 mm glucose oxidation to be severely decreased by EtBr treatment compared with control cells (by 63% on day 4 and by 78% on day 6; both p < 0.01). By contrast, glucose utilization was only marginally decreased. Lactate production under 22.2 mm glucose was increased by 2.9- and 3.5-fold by EtBr treatment on days 4 and 6, respectively (both p < 0.01). Cellular NADH at 2.8 mm glucose was increased by 35 and 43% by EtBr on days 4 and 6 (both p < 0.01). These data suggest that reduced expression of the mitochondrial electron transport system causes NADH accumulation in beta cells, thereby halting the tricarboxylic acid cycle on one hand, and on the other hand facilitating anaerobic glucose metabolism. Glucose-induced insulin secretion was lost rapidly along with the EtBr treatment with concomitant losses of membrane potential depolarization and the [Ca(2+)](i) increase, whereas glibenclamide-induced changes persisted. This is the first report to demonstrate the connection between metabolic alteration of electron transport system and that of tricarboxylic acid cycle and its impact on insulin secretion.
authors.affiliation: Department of Metabolic Diseases, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Japan.
authors.forename: Mitsuhiko
authors.initials: M
authors.lastname: Noda
authors.forename: Shigeo
authors.initials: S
authors.lastname: Yamashita
authors.forename: Noriko
authors.initials: N
authors.lastname: Takahashi
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Eto
authors.forename: Lin-Ming
authors.initials: LM
authors.lastname: Shen
authors.forename: Kazuo
authors.initials: K
authors.lastname: Izumi
authors.forename: Samira
authors.initials: S
authors.lastname: Daniel
authors.forename: Yoshiharu
authors.initials: Y
authors.lastname: Tsubamoto
authors.forename: Tomomi
authors.initials: T
authors.lastname: Nemoto
authors.forename: Masamitsu
authors.initials: M
authors.lastname: Iino
authors.forename: Haruo
authors.initials: H
authors.lastname: Kasai
authors.forename: Geoffrey W G
authors.initials: GW
authors.lastname: Sharp
authors.forename: Takashi
authors.initials: T
authors.lastname: Kadowaki
mesh_terms: D000693:Anaerobiosis; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002452:Cell Count; D002478:Cells, Cultured; D004272:DNA, Mitochondrial / Q000235:genetics*; D003920:Diabetes Mellitus / Q000378:metabolism*; D004996:Ethidium / Q000494:pharmacology; D005947:Glucose / Q000378:metabolism*; D005905:Glyburide / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000378:metabolism*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D019344:Lactic Acid / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D009243:NAD / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D004272:DNA, Mitochondrial; D007328:Insulin; D009243:NAD; D019344:Lactic Acid; D007770:L-Lactate Dehydrogenase; D004996:Ethidium; D005947:Glucose; D005905:Glyburide; D002118:Calcium