UNSPECIFIED (2005) Uncoupling of nutrient metabolism from insulin secretion by overexpression of cytosolic phospholipase A(2). DIABETES, 54 (1). pp. 116-124. ISSN 0012-1797

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Abstract

We have generated MIN6 beta-cells that stably overexpress cytosolic phospholipase A(2) (cPLA(2)) and show a ninefold increase in cPLA2 activity. Overexpression of cPLA2 did not affect the capacity of MINE cells to show elevations in intracellular Ca2+ concentration ([Ca2+](i)) in response to tolbutamide and KCl, and these depolarizing stimuli produced insulin secretion profiles in cPLA(2)-overexpressing cells similar to those they produced in passage-matched nontransfected MINE cells. However, cPLA2-overexpressing MIN6 cells did not respond to elevations in extracellular glucose with increases in ATP, [Ca2+](i), or insulin secretion. Nontransfected MIN6 cells showed a rapid and sustained increase in NAD(P)H autofluorescence in response to 25 mmol/l glucose, and this was reduced by similar to95% in MIN6 cells overexpressing cPLA2. This effect was mimicked in nontransfected MIN6 cells by p-(trifluoromethoxy) phenylylhydrazone, a mitochondrial uncoupler. Quantitative RT-PCR indicated that mRNA for uncoupling protein-2 (UCP-2) was increased in the cPLA2-overexpressing MIN6 cells, and this could be prevented by exposure to 100 mumol/l methyl arachidonyl fluorophosphate, a cPLA2 inhibitor. Glucose caused a decrease in rhodamine 123 fluorescence in control cells, but not in those overexpressing cPLA2, consistent with the transfected cells being unable to maintain mitochondrial proton gradients as a consequence of UCP-2 upregulation. Our data indicate that overexpression of cPLA2 results in severe impairment of the calcium and secretory responses of beta-cells to glucose through upregulation of UCP-2 and uncoupling of mitochondrial metabolism from ATP generation.

Item Type:	Journal Article
Subjects:	R Medicine > RC Internal medicine
Journal or Publication Title:	DIABETES
Publisher:	AMER DIABETES ASSOC
ISSN:	0012-1797
Date:	January 2005
Volume:	54
Number:	1
Number of Pages:	9
Page Range:	pp. 116-124
Publication Status:	Published
URI:	http://wrap.warwick.ac.uk/id/eprint/7493

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