The pro-oxidant chromium(VI) inhibits mitochondrial complex I, complex II, and aconitase in the bronchial epithelium: EPR markers for Fe–S proteins

Abstract

Hexavalent chromium [Cr(VI)] compounds (e.g. chromates) are strong oxidants that readily enter cells where they are reduced to reactive Cr species that also facilitate reactive oxygen species (ROS) generation. Recent studies demonstrated inhibition and oxidation of the thioredoxin system, with greater effects on mitochondrial thioredoxin (Trx2). This implies that Cr(VI)-induced oxidant stress may be especially directed at the mitochondria. Examination of other redox-sensitive mitochondrial functions showed that Cr(VI) treatments that cause Trx2 oxidation in human bronchial epithelial cells also result in pronounced and irreversible inhibition of aconitase, a TCA cycle enzyme that has an iron-sulfur (Fe-S) center that is labile with respect to certain oxidants. The activities of electron transport complexes I and II were also inhibited, whereas complex III was not. Electron paramagnetic resonance (EPR) studies of samples at liquid helium temperature (10 K) showed a strong signal at g = 1.94 that is consistent with the inhibition of electron flow through complexes I and/or II. A signal at g = 2.02 was also observed which is consistent with oxidation of the Fe-S center of aconitase. The g = 1.94 signal was particularly intense and remained after extracellular Cr(VI) was removed, whereas the g = 2.02 signal declined in intensity after Cr(VI) was removed. A similar inhibition of these activities and analogous EPR findings were noted in bovine airways treated ex vivo with Cr(VI). Overall, the data support the hypothesis that Cr(VI) exposure has deleterious effects on a number of redox-sensitive core mitochondrial proteins. The g = 1.94 signal could prove to be an important biomarker for oxidative damage resulting from Cr(VI) exposure. The EPR spectra simultaneously showed signals for Cr(V) and Cr(III) which verify Cr(VI) exposure and its intracellular reductive activation.

Damage to mitochondrial complex I during cardiac ischemia reperfusion injury is reduced indirectly by anti-anginal drug ranolazine

“…In our study, we observed no significant change in the g =2.023 signal representing the 3Fe-4S cluster, S3, of complex II with IR. In contrast, results of Myers et al [42] reported a decrease in the aconitase signal (with the same g value) following oxidative stress with chromium radicals. There was a significant increase in the amount of the EPR detectable UQ • radical during ischemia, and this was reduced by reperfusion as well as by Ran treatment.…”

“…EPR signals were obtained by scanning the samples in a high intensity magnetic field (3000-4000 G) at 10 Kelvin, 9.6 GHz, 5 mW microwave power and 5 G amplitude modulation. Each sample was scanned 9 times and spectra were averaged [41, 42]. Signal intensities were measured for g’ s of 2.023 (3Fe-4S attributed to cluster S3 of complex II, or to mitochondrial aconitase), 2.006 (attributed to semi-ubiquinone radical, UQ • ), 1.94 (2Fe-2S attributed to N1b of complex I, or to S1 of complex II), and 1.89 (4Fe-4S attributed to N4 of complex I, or to the Rieske center of complex III [43].…”

Potentially diagnostic electron paramagnetic resonance spectra elucidate the underlying mechanism of mitochondrial dysfunction in the deoxyguanosine kinase deficient rat model of a genetic mitochondrial DNA depletion syndrome

“…The spin-Hamiltonian parameters, midpoint potentials and relaxation behavior of these centers have been reasonably well characterized (69–87), along with some other tissue-specific signals from transferrin, ceruloplasmin, and catalase (88–90). Specific applications of EPR to mitochondria have included detection of an irreversible deficiency in Complex I FeS clusters in iron-deficient rats (91), heme-nitrosyl in substantia nigra of Parkinson’s diseased brain (92), chromium-dependent inhibition of Complexes I & II and aconitase (93), cardio- and neuro-protection against doxorubicin (80), prophylaxis against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine in a Parkinson’s mouse model (82), the differential sensitivity of aconitase and FeS clusters from Complexes I & III to oxidative and nitrosative stress in heart (94), and the sensitivity of Complex III FeS clusters in aging heart to ischemia (95). However, despite these successes in mechanistic studies, the authors are unaware of any direct application of EPR for functional pathophysiologic studies in humans or whole animal models with primary mitochondrial disease; the closest analog is a study in which a comparison of EPR signals from muscle biopsies of sepsis patients indicated significant depletion of Complex I FeS signals in those who died compared to survivors (86, 96).…”

Liver Aspartate Transaminase Isoenzymes as Biomarkers of Chronic Exposure to Chromium(VI)

“…Electrophoretic mobility of HMW-AST was greater than that of LMW-AST ( Figure 1B Liver LMW-AST isoenzyme activity dropped in rats treated for 15 days and increased in the 45-day and 60-day groups ( Figure 4A). We believe that these differences between the groups are related to lower mitochondrial oxygen consumption, attenuated gene expression in short-term exposure (15,16), and increased mitochondrial/microsomal lipid peroxidation, cellular induction, and failure in membrane integrity in longer exposures (17).…”