T cell activation-induced mitochondrial hyperpolarization is mediated by Ca2+- and redox-dependent production of nitric oxide.
Activation, proliferation, or programmed cell death of T lymphocytes is regulated by the mitochondrial transmembrane potential (Deltapsi(m)) through controlling ATP synthesis, production of reactive oxygen intermediates (ROI), and release of cell death-inducing factors. Elevation of Deltapsi(m) or mitochondrial hyperpolarization is an early and reversible event associated with both T cell activation and apoptosis. In the present study, T cell activation signals leading to mitochondrial hyperpolarization were investigated. CD3/CD28 costimulation of human PBL elevated cytoplasmic and mitochondrial Ca(2+) levels, ROI production, and NO production, and elicited mitochondrial hyperpolarization. Although T cell activation-induced Ca(2+) release, ROI levels, and NO production were diminished by inositol 1,4,5-triphosphate receptor antagonist 2-aminoethoxydiphenyl borane, superoxide dismutase mimic manganese (III) tetrakis (4-benzoic acid) porphyrin chloride, spin trap 5-diisopropoxyphosphoryl-5-methyl-1-pyrroline-N-oxide, and NO chelator carboxy-2-phenyl-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide, mitochondrial hyperpolarization was selectively inhibited by carboxy-2-phenyl-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide (-85.0 +/- 10.0%; p = 0.008) and, to a lesser extent, by 2-aminoethoxydiphenyl borane. Moreover, NO precursor (Z)-1-[2-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate diethylenetriamine elicited NO and ROI production, Ca(2+) release, transient ATP depletion, and robust mitochondrial hyperpolarization (3.5 +/- 0.8-fold; p = 0.002). Western blot analysis revealed expression of Ca-dependent endothelial NO synthase and neuronal NO synthase isoforms and absence of Ca-independent inducible NO synthase in PBL. CD3/CD28 costimulation or H(2)O(2) elicited severalfold elevations of endothelial NO synthase and neuronal NO synthase expression, as compared with beta-actin. H(2)O(2) also led to moderate mitochondrial hyperpolarization; however, Ca(2+) influx by ionomycin or Ca(2+) release from intracellular stores by thapsigargin alone failed to induce NO synthase expression, NO production, or Deltapsi(m) elevation. The results suggest that T cell activation-induced mitochondrial hyperpolarization is mediated by ROI- and Ca(2+)-dependent NO production.
Nagy
Gyorgy
G
Department of Medicine, State University of New York, College of Medicine, Syracuse, NY 13210, USA.
Koncz
Agnes
A
Perl
Andras
A
D001565:Benzoates / Q000494:pharmacology; D001896:Boron Compounds / Q000494:pharmacology; D018106:CD28 Antigens / Q000378:metabolism / Q000494:pharmacology; D017252:CD3 Complex / Q000378:metabolism / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D015220:Calcium Channels; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects / Q000276:immunology; D004338:Drug Combinations; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007424:Intracellular Fluid / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology*; D008564:Membrane Potentials / Q000187:drug effects / Q000276:immunology; D008928:Mitochondria / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis* / Q000502:physiology; D019001:Nitric Oxide Synthase / Q000096:biosynthesis; D052248:Nitric Oxide Synthase Type I; D052250:Nitric Oxide Synthase Type III; D010084:Oxidation-Reduction / Q000187:drug effects; D017382:Reactive Oxygen Species / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors; D015398:Signal Transduction / Q000276:immunology; D013601:T-Lymphocytes / Q000187:drug effects / Q000201:enzymology / Q000276:immunology* / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects / Q000276:immunology
D001565:Benzoates; D001896:Boron Compounds; D018106:CD28 Antigens; D017252:CD3 Complex; D015220:Calcium Channels; D002614:Chelating Agents; D004338:Drug Combinations; C506926:ITPR1 protein, human; D007093:Imidazoles; D053496:Inositol 1,4,5-Trisphosphate Receptors; D017382:Reactive Oxygen Species; D018160:Receptors, Cytoplasmic and Nuclear; C079393:1,3-dihydroxy-4,4,5,5-tetramethyl-2-(4-carboxyphenyl)tetrahydroimidazole; D009569:Nitric Oxide; D015544:Inositol 1,4,5-Trisphosphate; D006861:Hydrogen Peroxide; C109986:2-aminoethoxydiphenyl borate; C496320:NOS1 protein, human; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D052250:Nitric Oxide Synthase Type III; D002118:Calcium