Adenoviral overexpression of the glutamylcysteine ligase catalytic subunit protects pancreatic islets against oxidative stress.
The catalytic subunit of glutamylcysteine ligase (GCLC) primarily regulates de novo synthesis of glutathione (GSH) in mammalian cells and is central to the antioxidant capacity of the cell. However, GCLC expression in pancreatic islets has not been previously examined. We designed experiments to ascertain whether GCLC is normally expressed in islets and whether it is up-regulated by interleukin-1 beta (IL-1 beta). GCLC expression levels were intermediate compared with other metabolic tissues (kidney, liver, muscle, fat, and lung). IL-1 beta up-regulated GCLC expression (10 ng/ml IL-1 beta, 3.76 +/- 0.86; 100 ng/ml IL-1 beta, 4.22 +/- 0.68-fold control) via the p38 form of mitogen-activated protein kinase and NF kappa B and also increased reactive oxygen species levels (10 ng/ml IL-1 beta, 5.41 +/- 1.8-fold control). This was accompanied by an increase in intraislet GSH/GSSG ratio (control, 7.1 +/- 0.1; 10 ng/ml IL-1 beta, 8.0 +/- 0.5; 100 ng/ml IL-1 beta, 8.2 +/- 0.5-fold control; p < 0.05). To determine whether overexpression of GCLC increases the antioxidant capacity of the islet and prevents the adverse effects of IL-1 beta on glucose-induced insulin secretion, islets were infected with an adenovirus encoding GCLC. IL-1 beta significantly decreased glucose-stimulated insulin secretion (control, 123.8 +/- 17.7; IL-1 beta, 40.2 +/- 3.9 microunits/ml insulin/islet). GCLC overexpression increased intraislet GSH levels and partially prevented the decrease in glucose-stimulated insulin secretion caused by IL-1 beta. These data provide the first report of GCLC expression in the islet and demonstrate that adenoviral overexpression of GCLC increases intracellular GSH levels and protects the beta cell from the adverse effects of IL-1 beta.
Pacific Northwest Research Institute, Seattle, Washington 98122, USA.
Phuong Oanh T
PO
Tran
Sarah M
SM
Parker
Eric
E
LeRoy
Christopher C
CC
Franklin
Terrance J
TJ
Kavanagh
Tao
T
Zhang
Huarong
H
Zhou
Portia
P
Vliet
Elizabeth
E
Oseid
Jamie S
JS
Harmon
R Paul
RP
Robertson
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D005434:Flow Cytometry; D015870:Gene Expression; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D005822:Genetic Vectors; D005947:Glucose / Q000494:pharmacology; D005721:Glutamate-Cysteine Ligase / Q000235:genetics* / Q000378:metabolism; D005978:Glutathione / Q000032:analysis; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007375:Interleukin-1 / Q000494:pharmacology; D007515:Islets of Langerhans / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000032:analysis; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D007328:Insulin; D007375:Interleukin-1; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D048051:p38 Mitogen-Activated Protein Kinases; D005721:Glutamate-Cysteine Ligase; D005978:Glutathione; D005947:Glucose