MITOCHONDRIAL UCP2 IS ASSOCIATED WITH UNCOUPLED RESPIRATION AND LOW RADIATION-INDUCED HYDROGEN PEROXIDE LEVELS IN DRUG RESISTANT LEUKEMIC CELLS

A. Antoniou¹, M.K. Newell² and M-E. Harper^1
1Department of Biochemistry, Microbiology, and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada K1H 8M5
²Department of Biology, University of Colorado at Colorado Springs, Colorado Springs, CO 80918

Based on evidence that mitochondrial metabolism is altered during cell death, we compared bioenergetic characteristics in the wild type and drug resistant leukemic cell lines, L1210 and L1210/DDP, respectively.  L1210/DDP cells are cell surface Fas negative.  Flow cytometry of permeabilized cells showed higher uncoupling protein 2 (UCP2) and bcl-2 levels in resistant versus wild type cells.  Western blotting of UCP2 confirmed this.  UCP2 is thought to cause a mitochondrial proton leak, and uncoupling of oxidative phosphorylation.  Increased UCP2 corresponded to increased proton leak dependent oxygen consumption in resistant cells which was inhibited by GDP.  Full analyses of oxidative phosphorylation reactions revealed lower mitochondrial membrane potentials, and dramatically higher leak dependent oxygen consumption in resistant versus wild type cells.  Hydrogen peroxide levels were similar despite higher oxygen consumption in resistant cells.  Radiation resulted in lower increases in hydrogen peroxide in resistant cells.  The artificial uncoupler, CCCP, decreased hydrogen peroxide levels in wild type but had no effect on the already uncoupled resistant cells.  Our findings are consistent with a role for UCP2-mediated uncoupling in the protection of cells from reactive oxygen species and cytotoxic therapies.

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