Mitochondrial-derived free radicals mediate asbestos-induced alveolar epithelial cell apoptosis

Abstract

Asbestos causes pulmonary toxicity by mechanisms that in part involve reactive oxygen species (ROS). However, the precise source of ROS is unclear. We showed that asbestos induces alveolar epithelial cell (AEC) apoptosis by a mitochondrial-regulated death pathway. To determine whether mitochondrial-derived ROS are necessary for causing asbestos-induced AEC apoptosis, we utilized A549-rho(omicron) cells that lack mitochondrial DNA and a functional electron transport. As expected, antimycin, which induces an oxidative stress by blocking mitochondrial electron transport at complex III, increased dichlorofluoroscein (DCF) fluorescence in A549 cells but not in A549-rho(omicron) cells. Compared with A549 cells, rho(omicron) cells have less asbestos-induced ROS production, as assessed by DCF fluorescence, and reductions in total glutathione levels as well as less caspase-9 activation and apoptosis, as assessed by TdT-mediated dUTP nick end labeling staining and DNA fragmentation. A mitochondrial anion channel inhibitor that prevents ROS release from the mitochondria to the cytoplasm also blocked asbestos-induced A549 cell caspase-9 activation and apoptosis. Finally, a role for nonmitochondrial-derived ROS with exposure to high levels of asbestos (50 microg/cm(2)) was suggested by our findings that an iron chelator (phytic acid or deferoxamine) or a free radical scavenger (sodium benzoate) provided additional protection against asbestos-induced caspase-9 activation and DNA fragmentation in rho(omicron) cells. We conclude that asbestos fibers affect mitochondrial DNA and functional electron transport, resulting in mitochondrial-derived ROS production that in turn mediates AEC apoptosis. Nonmitochondrial-associated ROS may also contribute to AEC apoptosis, particularly with high levels of asbestos exposure.

Mitochondria-targeted Ogg1 and Aconitase-2 Prevent Oxidant-induced Mitochondrial DNA Damage in Alveolar Epithelial Cells

“…3) The mitochondria-regulated death pathway is the principal pathway mediating AEC apoptosis in vitro (18,34). 4) Antioxidants, including those targeted to the mitochondria, attenuate oxidative stress-induced endoplasmic reticulum stress and intrinsic apoptosis in AEC as well as other cell types (35)(36)(37).…”

The effects of N-acetyl cysteine, buthionine sulfoximine, diethyldithiocarbamate or 3-amino-1,2,4-triazole on antimycin A-treated Calu-6 lung cells in relation to cell growth, reactive oxygen species and glutathione

“…It is known that the collapse of MMP (Δae m ) is closely related to apoptosis (37). The increased intracellular H 2 O 2 played an important role in AMA-induced cell death in liver cells (38,39) and A549 human lung cancer cells (14). Likewise, AMA induced MMP (Δae m ) loss in Calu-6 cells.…”

“…The inhibition of electron transport causes a collapse of the proton gradient across the mitochondrial inner membrane, thereby breaking down the MMP (Δae m ) (11,13). This inhibition also results in the production of ROS (13,14). Evidence indicates that either the presence of ROS or the collapse of MMP (Δae m ) opens the mitochondrial permeability transition pore, which is accompanied by the release of proapoptotic molecules such as cytochrome c into the cytoplasm (15,16).…”

Mitochondrial Dok-4 Recruits Src Kinase and Regulates NF-κB Activation in Endothelial Cells

“…Of note, we found a putative mitochondrial targeting sequence with ␣-helix structure within the NH 2 -terminal domain of Dok-4 (amino acids [11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29] and Dok-5 (amino acids [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22], which is predicted by iPSORT (a subcellular localization site predictor for NH 2 -terminal sorting signals; website, biocaml.org/ipsort/iPSORT/). Therefore, we also generated a mutant containing a deletion of this mitochondrial targeting sequence, Dok-4-(⌬N11-29).…”