Oxidative stress increases internal calcium stores and reduces a key mitochondrial enzyme

Fibroblasts from patients with genetic and non-genetic forms of Alzheimer's disease (AD) show many abnormalities including increased bombesin-releasable calcium stores (BRCS), diminished activities of the mitochondrial alpha-ketoglutarate dehydrogenase complex (KGDHC), and an altered ability to handle oxidative stress. The link between genetic mutations (and the unknown primary event in non-genetic forms) and these other cellular abnormalities is unknown. To determine whether oxidative stress could be a convergence point that produces the other AD-related changes, these experiments tested in fibroblasts the effects of H2O2, in the presence or absence of select antioxidants, on BRCS and KGDHC. H2O2 concentrations that elevated carboxy-dichlorofluorescein (c-H2DCF)-detectable ROS increased BRCS and decreased KGDHC activity. These changes are in the same direction as those in fibroblasts from AD patients. Acute treatments with the antioxidants Trolox, or DMSO decreased c-H2DCF-detectable ROS by about 90%, but exaggerated the H2O2-induced increases in BRCS by about 4-fold and did not alter the reduction in KGDHC. Chronic pretreatments with Trolox more than doubled the BRCS, tripled KGDHC activities, and reduced the effects of H2O2, Pretreatment with DMSO or N-acetyl cysteine diminished the BRCS and either had no effect, or exaggerated the H2O2-induced changes in these variables. The results demonstrate that BRCS and KGDHC are more sensitive to H2O2 derived species than c-H2DCF, and that oxidized derivatives of the antioxidants exaggerate the actions of H2O2. The findings support the hypothesis that select abnormalities in oxidative processes are a critical part of a cascade that leads to the cellular abnormalities in cells from AD patients. (C) 2001 Elsevier Science B.V. All rights reserved.