Thenoyltrifluoroacetone ttfa

-production The levels of intracellular NADPH-dependent O 2production was assessed by chemiluminescence of lucigenin (bis-Nmethylacridinium nitrate; Sigma Chemical) as described previously. 22 INS-1 cells were incubated with various concentrations of AOPP for indicated time. To verify the sources of O 2
-production, cells were pre-treated for 1 h at 37 °C with NADPH oxidase inhibitors (diphenyleneiodonium, DPI, 100 μmol/l; apocynin, 10 μmol/l), a nitric oxide synthase inhibitor (N 4 -nitro-L-arginine methyl ester, L-NAME, 10 μmol/l), a xanthine oxidase inhibitor (oxypurinol, 100 μmol/l), a mitochondria complex I inhibitor (rotenone, 250 μmol/l), a mitochondria complex II inhibitor thenoyltrifluoroacetone (TTFA, 10 μmol/l), a mitochondria complex III inhibitor (myxothiazol, 10 μmol/l) and a O 2 -scavenger (c-SOD, 200 U/ml) (all from Sigma Chemical), respectively. The experiments were then repeated as described above. Cell homogenate (100 μg/well) were added into a 96-well microplate. Immediately before recording, dark-adapted lucigenin (5 μmol/l) with or without NADPH (100 μmol/l) were added to cell homogenates. Light emission was recorded every minute for 30 min (VICTOR V Wallac 1420; PerkinElmer, Turku, Finland). Data were expressed as counts per second (c.p.s.).