Adenosine triphosphate regulates NADPH oxidase activity leading to hydrogen peroxide production and COX-2/PGE2 expression in A549 cells

Lin CC, Lee IT, Wu WL, Lin WN, Yang CM. Adenosine triphosphate regulates NADPH oxidase activity leading to hydrogen peroxide production and COX-2/PGE(2) expression in A549 cells. Am J Physiol Lung Cell Mol Physiol 303: L401-L412, 2012. First published July 6, 2012; doi:10.1152/ajplung.00090.2012.-Non-small cell lung carcinoma (NSCLC) accounts for most of all lung cancers, which is the leading cause of mortality in human beings. High level of cyclooxygenase-2 (COX-2) is one of the features of NSCLC and related to the low survival rate of NSCLC. However, whether extracellular nucleotides releasing from stressed resident tissues contributes to the expression of COX-2 remains unclear. Here, we showed that stimulation of A549 cells by adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S) led to an increase in COX-2 gene expression and prostaglandin E-2 (PGE(2)) synthesis, revealed by Western blotting, RT-PCR, promoter assay, and enzyme-linked immunosorbent assay. In addition, ATP gamma S induced intracellular reactive oxygen species (ROS) generation through the activation of NADPH oxidase. The increase of ROS level resulted in activation of the c-Src/epidermal growth factor receptor (EGFR)/phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/nuclear factor (NF)-kappa B cascade. We also found that activated Akt was translocated into the nucleus and recruited with NF-kappa B and p300 to form a complex. Thus, activation of p300 modulated the acetylation of histone H4 via the NADPH oxidase/c-Src/EGFR/PI3K/Akt/NF-kappa B cascade stimulated by ATP gamma S. Our results are the first to show a novel role of NADPH oxidase-dependent Akt/p65/p300 complex formation that plays a key role in regulating COX-2/PGE2 expression in ATP gamma S-treated A549 cells. Taken together, we demonstrated that ATP gamma S stimulated activation of NADPH oxidase, resulting in generation of ROS, which then activated the downstream c-Src/EGFR/PI3K/Akt/NF-kappa B/p300 cascade to regulate the expression of COX-2 and synthesis of PGE2 in A549 cells. Understanding the regulation of COX-2 expression and PGE2 release by ATP gamma S on A549 cells may provide potential therapeutic targets of NSCLC.