title: Glucose uptake and metabolism by cultured human skeletal muscle cells: rate-limiting steps.
abstract: To use primary cultures of human skeletal muscle cells to establish defects in glucose metabolism that underlie clinical insulin resistance, it is necessary to define the rate-determining steps in glucose metabolism and to improve the insulin response attained in previous studies. We modified experimental conditions to achieve an insulin effect on 3-O-methylglucose transport that was more than twofold over basal. Glucose phosphorylation by hexokinase limits glucose metabolism in these cells, because the apparent Michaelis-Menten constant of coupled glucose transport and phosphorylation is intermediate between that of transport and that of the hexokinase and because rates of 2-deoxyglucose uptake and phosphorylation are less than those of glucose. The latter reflects a preference of hexokinase for glucose over 2-deoxyglucose. Cellular NAD(P)H autofluorescence, measured using two-photon excitation microscopy, is both sensitive to insulin and indicative of additional distal control steps in glucose metabolism. Whereas the predominant effect of insulin in human skeletal muscle cells is to enhance glucose transport, phosphorylation, and steps beyond, it also determines the overall rate of glucose metabolism.
authors.affiliation: Department of Veterans Affairs Medical Center, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
authors.forename: L M
authors.initials: LM
authors.lastname: Perriott
authors.forename: T
authors.initials: T
authors.lastname: Kono
authors.forename: R R
authors.initials: RR
authors.lastname: Whitesell
authors.forename: S M
authors.initials: SM
authors.lastname: Knobel
authors.forename: D W
authors.initials: DW
authors.lastname: Piston
authors.forename: D K
authors.initials: DK
authors.lastname: Granner
authors.forename: A C
authors.initials: AC
authors.lastname: Powers
authors.forename: J M
authors.initials: JM
authors.lastname: May
mesh_terms: D019325:3-O-Methylglucose / Q000378:metabolism; D000328:Adult; D000368:Aged; D001693:Biological Transport, Active; D002454:Cell Differentiation; D002478:Cells, Cultured; D005260:Female; D005947:Glucose / Q000378:metabolism*; D006593:Hexokinase / Q000378:metabolism; D006801:Humans; D007004:Hypoglycemic Agents / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D008856:Microscopy, Fluorescence; D008875:Middle Aged; D018482:Muscle, Skeletal / Q000166:cytology / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D010766:Phosphorylation; D019289:Pyruvic Acid / Q000494:pharmacology; D013268:Stimulation, Chemical
chemical_list: D007004:Hypoglycemic Agents; D007328:Insulin; D019325:3-O-Methylglucose; D009249:NADP; D019289:Pyruvic Acid; D006593:Hexokinase; D005947:Glucose