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  • Insulin storage and glucose homeostasis in mice null for the granule zinc transporter ZnT8 and studies of the type 2 diabetes-associated variants.

Insulin storage and glucose homeostasis in mice null for the granule zinc transporter ZnT8 and studies of the type 2 diabetes-associated variants.

Diabetes (2009-06-23)
Tamara J Nicolson, Elisa A Bellomo, Nadeeja Wijesekara, Merewyn K Loder, Jocelyn M Baldwin, Armen V Gyulkhandanyan, Vasilij Koshkin, Andrei I Tarasov, Raffaella Carzaniga, Katrin Kronenberger, Tarvinder K Taneja, Gabriela da Silva Xavier, Sarah Libert, Philippe Froguel, Raphael Scharfmann, Volodymir Stetsyuk, Philippe Ravassard, Helen Parker, Fiona M Gribble, Frank Reimann, Robert Sladek, Stephen J Hughes, Paul R V Johnson, Myriam Masseboeuf, Remy Burcelin, Stephen A Baldwin, Ming Liu, Roberto Lara-Lemus, Peter Arvan, Frans C Schuit, Michael B Wheeler, Fabrice Chimienti, Guy A Rutter
ABSTRACT

Zinc ions are essential for the formation of hexameric insulin and hormone crystallization. A nonsynonymous single nucleotide polymorphism rs13266634 in the SLC30A8 gene, encoding the secretory granule zinc transporter ZnT8, is associated with type 2 diabetes. We describe the effects of deleting the ZnT8 gene in mice and explore the action of the at-risk allele. Slc30a8 null mice were generated and backcrossed at least twice onto a C57BL/6J background. Glucose and insulin tolerance were measured by intraperitoneal injection or euglycemic clamp, respectively. Insulin secretion, electrophysiology, imaging, and the generation of adenoviruses encoding the low- (W325) or elevated- (R325) risk ZnT8 alleles were undertaken using standard protocols. ZnT8(-/-) mice displayed age-, sex-, and diet-dependent abnormalities in glucose tolerance, insulin secretion, and body weight. Islets isolated from null mice had reduced granule zinc content and showed age-dependent changes in granule morphology, with markedly fewer dense cores but more rod-like crystals. Glucose-stimulated insulin secretion, granule fusion, and insulin crystal dissolution, assessed by total internal reflection fluorescence microscopy, were unchanged or enhanced in ZnT8(-/-) islets. Insulin processing was normal. Molecular modeling revealed that residue-325 was located at the interface between ZnT8 monomers. Correspondingly, the R325 variant displayed lower apparent Zn(2+) transport activity than W325 ZnT8 by fluorescence-based assay. ZnT8 is required for normal insulin crystallization and insulin release in vivo but not, remarkably, in vitro. Defects in the former processes in carriers of the R allele may increase type 2 diabetes risks.

MATERIALS
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Product Description

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Formalin solution, neutral buffered, 10%, histological tissue fixative
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