Adenovirus-mediated overexpression of liver carnitine palmitoyltransferase I in INS1E cells: effects on cell metabolism and insulin secretion.
Lipid metabolism in the beta-cell is critical for the regulation of insulin secretion. Pancreatic beta-cells chronically exposed to fatty acids show higher carnitine palmitoyltransferase I (CPT I) protein levels, higher palmitate oxidation rates and an altered insulin response to glucose. We examined the effect of increasing CPT I levels on insulin secretion in cultured beta-cells. We prepared a recombinant adenovirus containing the cDNA for the rat liver isoform of CPT I. The overexpression of CPT I in INS1E cells caused a more than a 5-fold increase in the levels of CPT I protein (detected by Western blotting), a 6-fold increase in the CPT activity, and an increase in fatty acid oxidation at 2.5 mM glucose (1.7-fold) and 15 mM glucose (3.1-fold). Insulin secretion was stimulated in control cells by 15 mM glucose or 30 mM KCl. INS1E cells overexpressing CPT I showed lower insulin secretion on stimulation with 15 mM glucose (-40%; P<0.05). This decrease depended on CPT I activity, since the presence of etomoxir, a specific inhibitor of CPT I, in the preincubation medium normalized the CPT I activity, the fatty-acid oxidation rate and the insulin secretion in response to glucose. Exogenous palmitate (0.25 mM) rescued glucose-stimulated insulin secretion (GSIS) in CPT I-overexpressing cells, indicating that the mechanism of impaired GSIS was through the depletion of a critical lipid. Depolarizing the cells with KCl or intermediary glucose concentrations (7.5 mM) elicited similar insulin secretion in control cells and cells overexpressing CPT I. Glucose-induced ATP increase, glucose metabolism and the triacylglycerol content remained unchanged. These results provide further evidence that CPT I activity regulates insulin secretion in the beta-cell. They also indicate that up-regulation of CPT I contributes to the loss of response to high glucose in beta-cells exposed to fatty acids.
Rubí
Blanca
B
Department of Biochemistry and Molecular Biology, University of Barcelona, School of Pharmacy, Diagonal 643, E-08028 Barcelona, Spain.
Antinozzi
Peter A
PA
Herrero
Laura
L
Ishihara
Hisamitsu
H
Asins
Guillermina
G
Serra
Dolors
D
Wollheim
Claes B
CB
Maechler
Pierre
P
Hegardt
Fausto G
FG
D000255:Adenosine Triphosphate / Q000378:metabolism; D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D015153:Blotting, Western; D002334:Carnitine O-Palmitoyltransferase / Q000096:biosynthesis* / Q000235:genetics; D002460:Cell Line; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004852:Epoxy Compounds / Q000494:pharmacology; D005227:Fatty Acids / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D007004:Hypoglycemic Agents / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism*; D007515:Islets of Langerhans / Q000378:metabolism; D008099:Liver / Q000201:enzymology*; D008928:Mitochondria / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D019308:Palmitic Acid / Q000378:metabolism; D011189:Potassium Chloride / Q000378:metabolism; D020033:Protein Isoforms; D051381:Rats; D014280:Triglycerides / Q000378:metabolism; D015854:Up-Regulation
D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D004852:Epoxy Compounds; D005227:Fatty Acids; D007004:Hypoglycemic Agents; D007328:Insulin; D020033:Protein Isoforms; D014280:Triglycerides; D019308:Palmitic Acid; D011189:Potassium Chloride; D000255:Adenosine Triphosphate; D002334:Carnitine O-Palmitoyltransferase; D005947:Glucose; C054207:etomoxir; D010100:Oxygen