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  • 1
    ISSN: 1432-0428
    Keywords: Clonal beta-cell line ; insulin secretion ; glucose transport ; glucose phosphorylation ; glucose utilization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary Glucose-stimulated insulin secretion, glucose transport, glucose phosphorylation and glucose utilization have been characterized in the insulinoma cell line MIN6, which is derived from a transgenic mouse expressing the large T-antigen of SV40 in pancreatic beta cells. Glucose-stimulated insulin secretion occurred progressively from 5 mmol/l glucose, reached the maximal level approximately seven-fold above the basal level at 25 mmol/l, and remained at this level up to 50 mmol/l. Glucose transport was very rapid with the half-maximal uptake of 3-O-methyl-d-glucose being reached within 15 s at 22 °C. Glucose phosphorylating activity in the cell homogenate was due mainly to glucokinase; the Vmax value of glucokinase activity was estimated to be 255±37 nmol·h−1·mg protein−1, constituting approximately 80% of total phosphorylating activity, whereas hexokinase activity constituted less than 20%. MIN6 cells exhibited mainly the high Km component of glucose utilization with a Vmax of 289±18 nmol·h−1·mg protein−1. Thus, glucose utilization quantitatively and qualitatively reflected glucose phosphorylation in MIN6 cells. In contrast, MIN7 cells, which exhibited only a small increase in insulin secretion in response to glucose, had 4.7-fold greater hexokinase activity than MIN6 cells with a comparable activity of glucokinase. These characteristics in MIN6 cells are very similar to those of isolated islets, indicating that this cell line is an appropriate model for studying the mechanism of glucose-stimulated insulin secretion in pancreatic beta cells.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-0428
    Keywords: Keywords Pancreatic beta-cell line ; mitochondrial calcium ; cytosolic calcium ; oscillation ; aequorin ; insulin secretagogue.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary We examined the relationship between cytosolic Ca2+ concentration ([Ca2+]c) and mitochondrial matrix Ca2+ concentration ([Ca2+]m) in the pancreatic beta-cell line, MIN6. [Ca2+]c was monitored in a single or a group (30 cells) of fura-2-loaded MIN6 cells, and [Ca2+]m was measured in a group (1 × 106 cells) of MIN6 cells stably transfected with aequorin targeted at the mitochondria. Exogenous ATP (0.25 mmol/l) produced a single transient increase in [Ca2+]c whereas 22 mmol/l KCl produced a sustained plateau increase. ATP and KCl evoked transient increases in [Ca2+]m but with distinct time courses of [Ca2+]m decline: the [Ca2+]m increase induced by ATP decreased more rapidly than that induced by KCl. Nitrendipine (3 μmol/l), a blocker of L-type Ca2+ channels, inhibited both [Ca2+]c and [Ca2+]m signals in response to KCl and tolbutamide, but not those to ATP. Peak levels of [Ca2+]m increase (around 2 μmol/l) exceeded those of [Ca2+]c increase (around 500 nmol/l). A rise in glucose concentration from 3 to 30 mmol/l induced oscillations of [Ca2+]c that overlay the sustained increases in [Ca2+]c in single cells. An oscillatory increase in [Ca2+]m was similarly observed in response to glucose. Addition of 10 mmol/l 2-ketoisocaproic acid at 20 mmol/l glucose further increased the plateau level of [Ca2+]c and the frequency of [Ca2+]c oscillations, which were correlated with a further increase in [Ca2+]m. In response to pulsatile exposure to KCl, [Ca2+]c and [Ca2+]m increased synchronously. These data suggest that an oscillatory increase in [Ca2+]m in beta cells, the signal which is thought to be necessary for continuous stimulation of mitochondrial metabolism, is produced synchronously with the [Ca2+]c oscillations. [Diabetologia (1998) 41: 279–286]
    Type of Medium: Electronic Resource
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