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SLC30A3 responds to glucose- and zinc variations in beta-cells and is critical for insulin production and in vivo glucose-metabolism during beta-cell stress.

Smidt K, Jessen N, Petersen AB, Larsen A, Magnusson N, Jeppesen JB, Stoltenberg M, Culvenor JG, Tsatsanis A, Brock B, Schmitz O, Wogensen L, Bush AI, Rungby J - PLoS ONE (2009)

Bottom Line: In INS-1E cells 2 mM glucose down-regulated ZnT-3 and up-regulated ZnT-5 expression relative to 5 mM. 16 mM glucose increased ZnT-3 and decreased ZnT-8 expression.ZnT-3, which is pivotal in the development of cellular changes as also seen in type 2 diabetes (e.g. amyloidosis in Alzheimer's disease) but not previously described in beta-cells, is present in this cell type, up-regulated by glucose in a concentration dependent manner and up-regulated by zinc depletion which by contrast decreased ZnT-3 protein levels.Knock-down of the ZnT-3 gene lowers insulin secretion in vitro and affects in vivo glucose metabolism after streptozotocin treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Aarhus, Arhus, Denmark.

ABSTRACT

Background: Ion transporters of the Slc30A- (ZnT-) family regulate zinc fluxes into sub-cellular compartments. beta-cells depend on zinc for both insulin crystallization and regulation of cell mass.

Methodology/principal findings: This study examined: the effect of glucose and zinc chelation on ZnT gene and protein levels and apoptosis in beta-cells and pancreatic islets, the effects of ZnT-3 knock-down on insulin secretion in a beta-cell line and ZnT-3 knock-out on glucose metabolism in mice during streptozotocin-induced beta-cell stress. In INS-1E cells 2 mM glucose down-regulated ZnT-3 and up-regulated ZnT-5 expression relative to 5 mM. 16 mM glucose increased ZnT-3 and decreased ZnT-8 expression. Zinc chelation by DEDTC lowered INS-1E insulin content and insulin expression. Furthermore, zinc depletion increased ZnT-3- and decreased ZnT-8 gene expression whereas the amount of ZnT-3 protein in the cells was decreased. Zinc depletion and high glucose induced apoptosis and necrosis in INS-1E cells. The most responsive zinc transporter, ZnT-3, was investigated further; by immunohistochemistry and western blotting ZnT-3 was demonstrated in INS-1E cells. 44% knock-down of ZnT-3 by siRNA transfection in INS-1E cells decreased insulin expression and secretion. Streptozotocin-treated mice had higher glucose levels after ZnT-3 knock-out, particularly in overt diabetic animals.

Conclusion/significance: Zinc transporting proteins in beta-cells respond to variations in glucose and zinc levels. ZnT-3, which is pivotal in the development of cellular changes as also seen in type 2 diabetes (e.g. amyloidosis in Alzheimer's disease) but not previously described in beta-cells, is present in this cell type, up-regulated by glucose in a concentration dependent manner and up-regulated by zinc depletion which by contrast decreased ZnT-3 protein levels. Knock-down of the ZnT-3 gene lowers insulin secretion in vitro and affects in vivo glucose metabolism after streptozotocin treatment.

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Relative gene expression of ZnTs in INS-1E cells and mouse islets after 24 hours of glucose stimulation.A) INS-1E cells: Stimulations were performed at 2, 5 or 16 mM glucose. Normalised to UBC-7, CycA and β-actin. Data are mean and SEM (*p<0.05). N = 9. B) Mouse islets: Stimulations were performed at 2, 5 or 16 mM glucose. Normalised to UBC-7, CycA and HPRT. Data are mean and SEM (*p<0.05). N = 6.
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pone-0005684-g001: Relative gene expression of ZnTs in INS-1E cells and mouse islets after 24 hours of glucose stimulation.A) INS-1E cells: Stimulations were performed at 2, 5 or 16 mM glucose. Normalised to UBC-7, CycA and β-actin. Data are mean and SEM (*p<0.05). N = 9. B) Mouse islets: Stimulations were performed at 2, 5 or 16 mM glucose. Normalised to UBC-7, CycA and HPRT. Data are mean and SEM (*p<0.05). N = 6.

Mentions: We cultured INS-1E cells for 24 hours at 2, 5 or 16 mM glucose in order to study the gene expression of selected ZnTs. ZnT-1 and ZnT-3-8 were expressed at all glucose concentrations. ZnT-5 expression was significantly up-regulated at the lowest glucose-concentration (p<0.01) while ZnT-3 was significantly down-regulated (p<0.05). At the highest glucose concentration, ZnT-8 was significantly down-regulated (p<0.05) whereas ZnT-3 was significantly up-regulated (p<0.01) (Fig. 1A). All comparisons are relative to 5 mM glucose. Islets from mice cultured for 24 hours at 2, 5 or 16 mM glucose had significantly higher ZnT-3 expression at 16 mM glucose compared to 2 mM glucose (p<0.05). ZnT-8 expression was not significantly different at the various glucose concentrations (Fig. 1B).


SLC30A3 responds to glucose- and zinc variations in beta-cells and is critical for insulin production and in vivo glucose-metabolism during beta-cell stress.

Smidt K, Jessen N, Petersen AB, Larsen A, Magnusson N, Jeppesen JB, Stoltenberg M, Culvenor JG, Tsatsanis A, Brock B, Schmitz O, Wogensen L, Bush AI, Rungby J - PLoS ONE (2009)

Relative gene expression of ZnTs in INS-1E cells and mouse islets after 24 hours of glucose stimulation.A) INS-1E cells: Stimulations were performed at 2, 5 or 16 mM glucose. Normalised to UBC-7, CycA and β-actin. Data are mean and SEM (*p<0.05). N = 9. B) Mouse islets: Stimulations were performed at 2, 5 or 16 mM glucose. Normalised to UBC-7, CycA and HPRT. Data are mean and SEM (*p<0.05). N = 6.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2683566&req=5

pone-0005684-g001: Relative gene expression of ZnTs in INS-1E cells and mouse islets after 24 hours of glucose stimulation.A) INS-1E cells: Stimulations were performed at 2, 5 or 16 mM glucose. Normalised to UBC-7, CycA and β-actin. Data are mean and SEM (*p<0.05). N = 9. B) Mouse islets: Stimulations were performed at 2, 5 or 16 mM glucose. Normalised to UBC-7, CycA and HPRT. Data are mean and SEM (*p<0.05). N = 6.
Mentions: We cultured INS-1E cells for 24 hours at 2, 5 or 16 mM glucose in order to study the gene expression of selected ZnTs. ZnT-1 and ZnT-3-8 were expressed at all glucose concentrations. ZnT-5 expression was significantly up-regulated at the lowest glucose-concentration (p<0.01) while ZnT-3 was significantly down-regulated (p<0.05). At the highest glucose concentration, ZnT-8 was significantly down-regulated (p<0.05) whereas ZnT-3 was significantly up-regulated (p<0.01) (Fig. 1A). All comparisons are relative to 5 mM glucose. Islets from mice cultured for 24 hours at 2, 5 or 16 mM glucose had significantly higher ZnT-3 expression at 16 mM glucose compared to 2 mM glucose (p<0.05). ZnT-8 expression was not significantly different at the various glucose concentrations (Fig. 1B).

Bottom Line: In INS-1E cells 2 mM glucose down-regulated ZnT-3 and up-regulated ZnT-5 expression relative to 5 mM. 16 mM glucose increased ZnT-3 and decreased ZnT-8 expression.ZnT-3, which is pivotal in the development of cellular changes as also seen in type 2 diabetes (e.g. amyloidosis in Alzheimer's disease) but not previously described in beta-cells, is present in this cell type, up-regulated by glucose in a concentration dependent manner and up-regulated by zinc depletion which by contrast decreased ZnT-3 protein levels.Knock-down of the ZnT-3 gene lowers insulin secretion in vitro and affects in vivo glucose metabolism after streptozotocin treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Aarhus, Arhus, Denmark.

ABSTRACT

Background: Ion transporters of the Slc30A- (ZnT-) family regulate zinc fluxes into sub-cellular compartments. beta-cells depend on zinc for both insulin crystallization and regulation of cell mass.

Methodology/principal findings: This study examined: the effect of glucose and zinc chelation on ZnT gene and protein levels and apoptosis in beta-cells and pancreatic islets, the effects of ZnT-3 knock-down on insulin secretion in a beta-cell line and ZnT-3 knock-out on glucose metabolism in mice during streptozotocin-induced beta-cell stress. In INS-1E cells 2 mM glucose down-regulated ZnT-3 and up-regulated ZnT-5 expression relative to 5 mM. 16 mM glucose increased ZnT-3 and decreased ZnT-8 expression. Zinc chelation by DEDTC lowered INS-1E insulin content and insulin expression. Furthermore, zinc depletion increased ZnT-3- and decreased ZnT-8 gene expression whereas the amount of ZnT-3 protein in the cells was decreased. Zinc depletion and high glucose induced apoptosis and necrosis in INS-1E cells. The most responsive zinc transporter, ZnT-3, was investigated further; by immunohistochemistry and western blotting ZnT-3 was demonstrated in INS-1E cells. 44% knock-down of ZnT-3 by siRNA transfection in INS-1E cells decreased insulin expression and secretion. Streptozotocin-treated mice had higher glucose levels after ZnT-3 knock-out, particularly in overt diabetic animals.

Conclusion/significance: Zinc transporting proteins in beta-cells respond to variations in glucose and zinc levels. ZnT-3, which is pivotal in the development of cellular changes as also seen in type 2 diabetes (e.g. amyloidosis in Alzheimer's disease) but not previously described in beta-cells, is present in this cell type, up-regulated by glucose in a concentration dependent manner and up-regulated by zinc depletion which by contrast decreased ZnT-3 protein levels. Knock-down of the ZnT-3 gene lowers insulin secretion in vitro and affects in vivo glucose metabolism after streptozotocin treatment.

Show MeSH
Related in: MedlinePlus