Marked over expression of uncoupling protein-2 in beta cells exerts minor effects on mitochondrial metabolism.
Evidence is conflicting as to the impact of elevated levels of uncoupling protein-2 (UCP-2) on insulin-producing beta cells. Here we investigated effects of a fourfold induction of UCP-2 protein primarily on mitochondrial parameters and tested for replication of positive findings at a lower level of induction. We transfected INS-1 cells to obtain a tet-on inducible cell line. A 48 h exposure to 1 μg/ml of doxycycline (dox) induced UCP-2 fourfold (424 ± 113%, mean±SEM) and 0.1 μg/ml twofold (178 ± 29%, n=3). Fourfold induced cells displayed normal viability (MTT, apoptosis), normal cellular insulin contents and, glucose-induced insulin secretion (+27 ± 11%) as well as D-[U-(14)C]-glucose oxidation (+5 ± 9% at 11 mM glucose). Oxidation of [1-(14)C]-oleate was increased from 4088 to 5797 fmol/μg prot/2h at 3.3mM glucose, p<0.03. Oxidation of L-[(14)C(U)]-glutamine was unaffected. Induction of UCP-2 did not significantly affect measures of mitochondrial membrane potential (Rhodamine 123) or mitochondrial mass (Mitotracker Green) and did not affect ATP levels. Oligomycin-inhibited oxygen consumption (a measure of mitochondrial uncoupling) was marginally increased, the effect being significant in comparison with dox-only treated cells, p<0.05. Oxygen radicals, assessed by dichlorofluorescin diacetate, were decreased by 30%, p<0.025. Testing for the lower level of UCP-2 induction did not reproduce any of the positive findings. A fourfold induction of UCP-2 was required to exert minor metabolic effects. These findings question an impact of moderately elevated UCP-2 levels in beta cells as seen in diabetes.
Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway. ingrid.hals@ntnu.no
Ingrid K
IK
Hals
Hirotaka
H
Ogata
Elin
E
Pettersen
Zuheng
Z
Ma
Anneli
A
Björklund
Frank
F
Skorpen
Kjartan Wollo
KW
Egeberg
Valdemar
V
Grill
D000818:Animals; D045744:Cell Line, Tumor; D004318:Doxycycline / Q000494:pharmacology; D005947:Glucose / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D007473:Ion Channels / Q000096:biosynthesis* / Q000235:genetics; D053078:Membrane Potential, Mitochondrial; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000096:biosynthesis* / Q000235:genetics; D009840:Oligomycins / Q000494:pharmacology; D010084:Oxidation-Reduction; D010101:Oxygen Consumption / Q000187:drug effects / Q000502:physiology; D051381:Rats; D014162:Transfection; D000071246:Uncoupling Protein 2
D007328:Insulin; D007473:Ion Channels; D024101:Mitochondrial Proteins; D009840:Oligomycins; C000604389:UCP2 protein, human; C000604391:Ucp2 protein, rat; D000071246:Uncoupling Protein 2; D005947:Glucose; D004318:Doxycycline