Pro-inflammatory cytokines increase reactive oxygen species through mitochondria and NADPH oxidase in cultured RPE cells

Reactive oxygen species (ROS) generated during inflammation are believed to play critical roles in various ocular diseases. However, the underlying mechanisms remain poorly understood. We investigated if pro-inflammatory cytokines, tumor necrosis factor TNF)-alpha, interleukin-1 beta (IL- 10), and interferon-gamma (IFN-gamma), induce ROS in human retinal pigment epithelial (RPE) cells. TNF-alpha, IL- 10 and IFN-y increased both intracellular and extracellular ROS production in a time- and dose-dependent manner. Thenoyltrifluoroacetone (TTEA), an inhibitor of mitochondrial respiratory chain, blocked TNF-alpha- and IFN-gamma-, but not IL-1 beta-induced ROS, whereas other two mitochondrial respiratory chain inhibitors, totenone and antimycin A, had no effect. NADPH oxidase inhibitor (diphenylene iodinium) abolished the ROS production induced by IL-1 beta or IFN-gamma, but not by TNF-alpha, whereas 6-aminonicotinamide (6AN), an inhibitor of the hexose monophosphate shunt (HMS), had no significant effects on the ROS induced by all three cytokines. ROS scavengers, pyrrolidinedithiocarbamate (PDTC) and N-acetyl-cysteine (NAC), reduced the levels of ROS induced by TNF-alpha, IL-1 beta and IFN-gamma (P < 0.05). Collectively, these results demonstrate that TNF-alpha, IL-1 beta and IFN-gamma increase mitochondrial- and NADPH oxidase-generated ROS in human RPE cells. Published by Elsevier Ltd.