NADPH oxidase activation is required in reactive oxygen species generation and cell transformation induced by hexavalent chromium

Abstract

Hexavalent chromium [Cr(VI)] is a well-known human carcinogen associated with the incidence of lung cancer. Although overproduction of reactive oxygen species (ROS) has been suggested to play a major role in its carcinogenicity, the mechanisms of Cr(VI)-induced ROS production remain unclear. In this study, we investigated the role of NADPH oxidase (NOX), one of the major sources of cellular ROS, in Cr(VI)-induced oxidative stress and carcinogenesis. We found that short-term exposure to Cr(VI) (2μM) resulted in a rapid increase in ROS generation in Beas-2B cells, and concomitantly increased NOX activity and expression of NOX members (NOX1-3 and NOX5) and subunits (p22 phox, p47 phox, p40 phox, and p67 phox). Cr(VI) also induced phosphorylation of p47 phox and membrane translocation of p47 phox and p67phox, further confirming NOX activation. Knockdown of p47 phox with a short hairpin RNA attenuated the ROS production induced by Cr(VI). Chronic exposure (up to 3 months) to low doses of Cr(VI) (0.125, 0.25, and 0.5μM) also promoted ROS generation and the expression of NOX subunits, such as p47 phox and p67 phox, but inhibited the expression of main antioxidant enzymes, such as superoxidase dismutase (SOD) and glutathione peroxidase (GPx). Chronic Cr(VI) exposure resulted in transformation of Beas-2B cells, increasing cell proliferation, anchorage independent growth in soft agar, and forming aggressive tumors in nude mice. Stable knockdown of p47 phox or overexpression of SOD1, SOD2, or catalase (CAT) eliminated Cr(VI)-induced malignant transformation. Our results suggest that NOX plays an important role in Cr(VI)-induced ROS generation and carcinogenesis. © The Author 2011. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved.